Abstract: BACKGROUND: Teaching the principles of scientific research in a comprehensive way is important at medical and dental schools. In many countries medical and dental training is not complete until the candidate has presented a diploma thesis. The objective of this study was to evaluate the nature, quality, publication pattern and visibility of Finnish medical diploma theses. METHODS: A total of 256 diploma theses presented at the University of Oulu from 2001 to 2003 were analysed. Using a standardised questionnaire, we extracted several characteristics from each thesis. We used the name of the student to assess whether the thesis resulted in a scientific publication indexed in medical article databases. The number of citations received by each published thesis was also recorded. RESULTS: A high proportion of the theses (69.5%) were essentially statistical in character, often combined with an extensive literature review or the development of a laboratory method. Most of them were supervised by clinical departments (55.9%). Only 61 theses (23.8%) had been published in indexed scientific journals. Theses in the fields of biomedicine and diagnostics were published in more widely cited journals. The median number of citations received per year was 2.7 and the range from 0 to 14.7. CONCLUSION: The theses were seldom written according to the principles of scientific communication and the proportion of actually published was small. The visibility of these theses and their dissemination to the scientific community should be improved.

Abstract: ACuteTox is an integrated project under the EU-FP6 with the aim to develop a simple and robust in vitro testing strategy for prediction of human acute systemic toxicity, which could replace animal tests used for regulatory purposes. Studies show good correlation of over 70% between in vitro basal cytotoxicity and rodent LD50 values or human lethal blood concentrations. However, a number of discrepancies occur which result in misclassification. ACuteTox aims to identify factors that can eliminate these misclassifications. The outliers in the in vitro/in vivo correlation will be evaluated in order to introduce further parameters (ADE, metabolism and organ specificity), which might improve the correlation. Integration of alerts and correctors in a prediction algorithm, together with implementation of medium throughput approaches, would allow establishment of a new testing strategy to better predict toxic classification.

Abstract: In bone matrix, type I collagen is stabilised by covalent cross-links formed between adjacent collagen molecules; the majority of which is believed to be immature, divalent bonds. For studying these immature forms in detail, we have developed an immunoassay for a synthetic peptide SP 4 that is analogous with and detects a linear epitope within the C-telopeptide of alpha1-chain of type I collagen. The SP 4 assay, together with the ICTP assay, which is specific for the trivalently cross-linked C-telopeptide, was used for the isolation of the differently cross-linked C-telopeptide structures of the alpha1-chain of type I collagen present in mineralised human bone. Amino acid analysis, peptide sequencing and MALDI-TOF mass spectrometry were used to identify and characterise each of the isolated structures. The cross-link content of each isolated peptide was identified. In the trivalent ICTP peptide, only 40% was cross-linked with pyridinoline, the remainder of the cross-links being currently uncharacterized. The divalent peptides contained only previously characterised cross-linking structures. Most of the divalent cross-links were dihydroxylysinonorleucine (DHLNL), with minor amounts of hydroxylysinonorleucine (HLNL). The relative proportion of the HLNL cross-link was slightly higher in the divalent alpha1Calpha2H peptide. A substantial amount of uncross-linked telopeptide structures was also found. Previous studies, where direct chemical cross-link analyses have been used to assess the maturity of cross-linking, have inferred that bone contains more divalently than trivalently cross-linked C-telopeptides. The immunochemical peptide approach used in this study may help to detect presently uncharacterized, trivalent cross-links, the presence of which is strongly suggested in this study. It also provides additional information regarding the extent and maturity of tissue type I collagen cross-linking in health and disease.

Abstract: Chemotherapy-induced ovarian failure causes rapid bone loss in premenopausal women with early breast cancer. The aim of the present study was to investigate the effect of intravenous intermittent clodronate during adjuvant chemotherapy in prevention of this rapid bone loss. 45 premenopausal women with early stage breast cancer were treated with adjuvant chemotherapy. In addition, all women were randomly allocated to receive either seven cycles of intravenous clodronate infusions (1500 mg each) parallel to the chemotherapy or no further therapy. The mean bone loss in the lumbar spine at 6 months was -0.5% in the clodronate group and -1.4% in the control group (p = 0.22) and, at 12 months, -3.9% and -3.6%, respectively (p = 0.62). Type I collagen metabolite PINP levels at six months were significantly lower in the clodronate group than in the control group: 22.6 microg/l (range 15.7-55.8 microg/l) and 44.0 microg/l (range 12.5-91.9 microg/l), respectively (p = 0.0001). At 12 months, no difference between the PINP levels in clodronate and control groups were seen. In conclusion, in this small study a short-term intermittent intravenous clodronate treatment did not seem to prevent clinically significantly the bone loss related to chemotherapy-induced ovarian failure in premenopausal women with early stage breast cancer, even though a significant reduction of a biochemical marker of bone turnover (PINP) was seen during the therapy.

Abstract: OBJECTIVE: The structure and distribution of type I and type III collagens in the extracellular matrix of malignant endometrium was evaluated for their roles in the development and progression of this neoplasm. STUDY DESIGN: Collagen synthesis and deposition in endometrial adenocarcinomas was determined by immunohistochemical analysis of type I and type III procollagen and verified by computer-assisted morphometry and in situ hybridization. RESULTS: In the stroma of well-differentiated adenocarcinomas increased intracellular collagen synthesis was observed in fibroblastic cells as well as increased extracellular formation of newly synthesized type I and type III procollagen. Collagen maturation was also rapid. In moderately differentiated tumors, destruction and dissolution occurred around invading islets, concomitantly with decreased deposits of both collagens, despite increases in corresponding mRNAs. In poorly differentiated neoplasms, solid epithelial islets coexisted with sparse and distinctly collagen-positive stroma. Poorly differentiated neoplasms also contained tumor cells exhibiting intracellular collagen staining as well as in situ hybridization signals. In highly malignant papillary adenocarcinomas, the tumor cells induced distinctly increased collagen synthesis and deposition of newly synthesized collagen but not the mature cross-linked protein. CONCLUSIONS: In malignancy, compression of surrounding stroma and a fibroproliferative response with increased collagen synthesis and deposition may prevent tumor growth. In more advanced lesions, stromal dissolution may permit tumor spread and in highly malignant lesions an abnormal stroma may promote neoplasm progression.

Abstract: Carbohydrates are thought to function as tags that mark circulatory glycoproteins for rapid clearance. To examine the role of the mannose receptor (MR) in glycoprotein clearance, we generated mice genetically deficient in MR. MR-/- mice were defective in clearing proteins bearing accessible mannose and N-acetylglucosamine residues and had elevated levels of eight different lysosomal hydrolases. Proteomic analysis of MR-/- and control mouse sera showed that an additional 4 out of 52 proteins identified were elevated in MR-/- serum. Each of these is up-regulated during inflammation and wound healing. Thus, MR appears to operate as an essential regulator of serum glycoprotein homeostasis.

Abstract: OBJECTIVE: the extent of the processing of type III procollagen to type III collagen was determined in nine human abdominal aortic aneurysms (AAA), and compared with ten samples of aortoiliac occlusive disease (AOD). METHODS: the aminoterminal propeptide (PIIINP) and telopeptide (IIINTP) of type III procollagen and collagen, respectively, were immunologically measured in the soluble and insoluble fractions of the extracellular matrix. The assay for PIIINP in the insoluble matrix was further validated. RESULTS: the insoluble matrices of AAAs contained at least 12 times more incompletely processed type III pN-collagen than AOD specimens (0.74% and 0.061%, respectively). Also, the soluble extracts of AAAs tended to contain more non-processed type III pN-collagen than free, properly cleaved aminoterminal propeptide. CONCLUSIONS: the larger amount of type III pN-collagen suggests an alteration in the metabolism of type III collagen in AAAs. This may partially explain the decreased tensile strength of the aortic tissue.

Abstract: BACKGROUND, MATERIALS AND METHODS: The role of epithelial cell growth and neoplastic transformation on collagen formation and deposition in the extracellular matrix (ECM) was analyzed by culturing immortalized human epidermal cell lines and Ras-transformed benign and malignant clones on collagen gels as transplants. The lesions were analyzed for extent of growth and morphology of epithelial and mesenchymal components as well as synthesis and deposition of different collagens. RESULTS: Immortalized cell lines required up to 5 weeks of growth for a well-organized mesenchyme to develop; transplants of Ras-transformed benign clones needed 3 weeks and transplants of highly malignant clones only 2 weeks to form an organized stroma. In transplants of immortalized cells after 2 weeks of growth newly-synthesized collagen type I and type III were deposited in the mesenchyme adjacent to the muscle, forming a mature ECM, while ECM was absent adjacent to growing, differentiated, immortalized cells. In transplants of Ras-transformed benign clones the subepithelial ECM was immature at day 14, but it was forming fibers at the same time in transplants of malignant clones. These were seen as thin irregular fibers in immunohistochemistry, ultimately organized into fibrillar structures in similar locations to active synthesis detected by in situ hybridization. Depositions of crosslinked mature type I collagen occurred later in similar locations. Type III collagen synthesis and deposition was most prominent in transplants of malignant cell clones, with degradation and destruction of the extracellular matrix around invading islets of malignant cells. CONCLUSION: The development of mesenchyme was directly related to duration of growth of transplants and degree of malignancy; mesenchyme organization was inversely related to differentiation of the epithelial cells. The results showed the usefulness of the transplant model in studies on cell and tissue growth and organization.

Abstract: BACKGROUND AND PURPOSE: The effects of radiation therapy on the turnover and structure of type I collagen were studied in irradiated and contralateral skin of 18 breast cancer patients without clinically evident fibrosis. MATERIALS AND METHODS: The rates of on-going type I collagen synthesis and degradation were assessed by the aminoterminal propeptide of type I procollagen (PINP) and by two different assays (ICTP and SP4) for the carboxyterminal telopeptide of type I collagen in the soluble tissue extracts, respectively. Also, TIMP-1, TIMP-2 and the MMP-2/TIMP-2 complex were measured in the tissue extracts. Insoluble skin matrices, containing the cross-linked type I collagen fibres, were heat-denatured and digested with trypsin. Then, the variants of the carboxyterminal telopeptide of type I collagen were separated by high performance liquid chromatography (HPLC). The major histidinohydroxylysinonorleucine (HHL)-cross-linked variant was quantified by the SP4 assay, and the minor pyridinoline analogue (PA)-cross-linked telopeptide was quantified by the ICTP assay. RESULTS: Both the synthesis and degradation of type I collagen were increased (r=0.906; P<0.001) on the irradiated side, whereas the concentration of the MMP-2/TIMP-2 complex was decreased. In the insoluble tissue digests, the HHL-cross-linked telopeptides of type I collagen, also, when expressed/tissue hydroxyproline, were increased in the irradiated skin. TIMP-1, TIMP-2 or PA-cross-linked telopeptides of type I collagen showed no differences between the two sides. CONCLUSIONS: Radiotherapy induces a long-term increase in the turnover of type I collagen and leads to the accumulation of cross-linked type I collagen in skin.

Abstract: Polymorphism in the Sp1 binding site in the first intron of the COL1A1 gene has been related to increased risk of osteoporosis in several populations. To overcome the difficulties associated with the use of mismatch oligonucleotide primers in the original method for its detection, we developed a procedure involving PCR amplification of a 598-base pair sequence from the intron and its digestion with the restriction enzyme Van 91 I. The more frequent allele is recognized by the enzyme, whereas the reaction is abolished in the less frequent allele. Two convenience samples from the population in northern Finland, consisting altogether of 173 individuals, were studied. The overall frequencies were 0.864 for the G and 0.136 for the T allele, with a heterozygocity of 27.2%. The frequency of the T allele is towards the lower end of the range observed for other European populations.

Abstract: The aminoterminal propeptide (hotPINP) of type I homotrimer, a putative malignancy-associated type I collagen variant, was purified for the first time and a method was established for its detection in pleural fluid. Samples of 58 patients, with malignant or benign disease, were studied with specific immunoassays for the two propeptides of type-I procollagen (PICP and PINP) and with HPLC-DEAE chromatography to separate the two PINP variants. HotPINP was present in 64% of both benign and malignant pleural effusion fluids, with the exception of malignant mesotheliomas, none of which showed the presence of hotPINP. Also the PICP to PINP ratios were lower than normal in both benign and malignant samples (altogether in 69% of samples), although this deviation was greater in malignancy. These two phenomena were independent of each other. As synthesis of the alpha1-homotrimer-variant of type-I collagen seems to be relatively common during the formation of pleural effusion, it may be generally related to a fibroproliferative reaction in the pleural wall.

Abstract: Circulating C-terminal propeptide of type I procollagen (PICP), mostly originating from bone, is mainly cleared by mannose receptors (MRs) in liver endothelial cells (LECs). We hypothesized that skin macrophage MRs could also play a role in local (in situ) clearance of PICP originating from skin type I procollagen synthesis. We tested this hypothesis in a male subject with a genetic systemic clearance defect, apparently due to an abnormality in MR function in LECs (or in PICP structure). Since skin macrophages may express the same MRs as LECs do, the genetic defect could affect them as well; hence, if elevated PICP concentrations even in skin interstitial fluid (IF) were found in our subject, it would suggest a role for local MR-mediated PICP clearance in skin. Since glucocorticoids (GCs) upregulate MRs in vitro, we measured the effect of topical GC on suction blister fluid (SBF)-PICP of the test person as compared with normal subjects. SBF-PICP was elevated in the case, which was consistent with the hypothesis. Furthermore, the GC-induced decrease was accentuated. The results suggest that skin macrophage MRs can have a role in skin PICP clearance in situ.

Abstract: The assay for the cross-linked carboxyterminal telopeptide of type I collagen (ICTP) has been shown to reflect increased type I collagen degradation in such pathological conditions as bone metastases and rheumatoid arthritis, but to be rather insensitive to the changes in physiological bone collagen turnover (e.g., induced by estrogen or bisphosphonate treatment). To determine the reasons for this discrepancy we localized the antigenic determinant recognized by the ICTP assay and studied the effects of two major osteoclastic proteinases, cathepsin K (EC 3.4.22.38) and matrix metalloproteinase-9 (MMP-9; gelatinase B; EC 3.4.24.35), on immunoreactivity. The antigenic determinant was shown to reside within the hydrophobic phenylalanine-rich regions of the carboxyterminal telopeptides of the two alpha1 chains of human type I collagen, situated between the triple helical domain and the lysine-derived trivalent cross-link. This conclusion was based on differences between the amino acid sequences and cross reactivities of the corresponding human and bovine antigens before and after proteolytic treatments with chymotrypsin. A trivalent cross-link is necessary for providing such a structure, because the divalently cross-linked and monomeric natural and synthetic peptides from the same region, but containing only one phenylalanine-rich sequence, showed poor immunoreaction. Recombinant human cathepsin K cleaved the trivalently cross-linked ICTP structure at two sites between the phenylalanine-rich region and the cross-link, destroying the reactivity with ICTP antibodies. On the contrary, the treatment of isolated ICTP by the matrix metalloproteinases MMP-9 (gelatinase B), MMP-1 (collagenase 1), or MMP-13 (collagenase 3) had no effect on the immunoreaction. Our results indicate that the increased circulating concentrations of ICTP found in several clinical situations are most likely produced by matrix metalloproteinases, whereas cathepsin K-mediated, osteoclastic bone resorption destroys ICTP antigenicity.

Abstract: BACKGROUND: Collagens are major proteins in the extracellular matrix, providing tissues with tensile strength. They are also important for cell adhesion and the invasion of malignant tumours. METHODS: Thirty-nine samples of human colon (24 diverticulosis, 6 malignant tumours, 9 controls) were collected during elective surgery. Immunoassays for different domains of type I and III collagens and procollagens were used in soluble tissue extracts and trypsin digests of tissue samples. RESULTS: The contents of cross-linked type I and III collagen telopeptides and total collagen were similar in diverticulosis and healthy tissue, whereas in malignant tissue maturely cross-linked type III collagen was scarce. Furthermore, some of the cross-linked type I telopeptide antigens were exceptionally small in size, indicating that the cross-linking of type I collagen in collagen fibres is impaired in cancer. The rate of type I collagen synthesis was clearly increased in malignancy, but not significantly in diverticulosis. However, type III collagen synthesis was increased in diverticulosis, but not in malignancy. CONCLUSIONS: In colon malignancy, the collagen cross-linking process was aberrant and the synthesis of type I collagen increased. In diverticulosis, the synthesis of type III collagen was increased, suggesting only moderately increased metabolic activity.

Abstract: Activation of the coagulation system in the alveolar space plays an important role in the pathogenesis of interstitial lung disease (ILD) and pulmonary fibrosis. The protein C (PC) pathway is the main modulator of coagulation activation. This study evaluated whether dysfunction of the PC pathway is associated with increased collagen synthesis in the intraalveolar space of patients with ILD. This study comprised 22 patients with ILD; of these, five had idiopathic pulmonary fibrosis (IPF), nine had sarcoidosis-associated ILD, and eight had collagen vascular disease-associated ILD (CVD-ILD). Thrombin-antithrombin complex (TAT) was measured as a marker of coagulation activation. As markers of the PC pathway activity, the concentration of activated PC-PC inhibitor (APC-PCI) complex and the APC-PCI/PC ratio were measured and, as a marker of collagen synthesis, the concentration of aminoterminal propeptide of type III procollagen (PIIINP) was measured in bronchoalveolar lavage fluid (BALF) of ILD patients. TAT was significantly increased in BALF from ILD patients as compared to control subjects. The concentrations of PIIINP were significantly elevated in patients with ILD as compared to healthy subjects. In contrast, the concentration of APC-PCI and the values of APC-PCI/PC ratio were significantly decreased in BALF from patients with ILD. BALF concentration of PIIINP was significantly and inversely correlated with the concentration of APC-PCI and with the APC-PCI/PC ratio. These findings suggest that dysfunction of the protein C pathway may have important physiopathologic implications in the development of pulmonary fibrosis in ILD.

Abstract: PURPOSE: This study aimed to characterize the distribution of structural domains of type I and III collagens in the wall of abdominal aortic aneurysms (AAAs), by the use of undilated atherosclerotic aortas (aortoiliac occlusive disease [AOD]) and healthy abdominal aortas as controls. METHODS: Immunohistochemical staining was applied with antibodies for the aminoterminal propeptides of type I (PINP) and type III (PIIINP) procollagens, which represent newly synthesized type I and III pN-collagens. In addition, an antibody against the aminoterminal telopeptide of type III collagen (IIINTP) was used as a means of detecting maturely cross-linked type III collagen fibrils. RESULTS: The newly synthesized type III procollagen detected by means of PIIINP staining was concentrated in the media in aneurysmal aortas, whereas type I pN-collagen was localized in the intima in both AAAs and AODs. The healthy aortas showed no immunoreactivity for either PIIINP or PINP. The cross-linked type III collagen, detected by means of IIINTP staining, stained transmurally in all study groups, but appeared more abundant in the media in AAAs. CONCLUSION: Our results strongly suggest that the metabolism of type III collagen is enhanced in AAAs. Intensive type III pN-collagen staining was present mainly in the media layer in AAAs, suggesting a role of type III collagen in aneurysm formation, whereas type I pN-collagen was present in the intima in both AAAs and AODs, suggesting that type I collagen synthesis is a fibroproliferative response related to the atherosclerotic process. The increased type III pN-collagen in AAAs may result in impaired fibril formation and, thus, in decreased tensile strength of aneurysmal tissue.

Abstract: During malignant growth many changes take place in the metabolism of fibrillar type III collagens in the connective tissues. The aminoterminal propeptide of type III procollagen (PIIINP) has been found to be often elevated in ovarian cancer. In the present study the prognostic value of serum PIIINP concentration in epithelial ovarian cancer is evaluated in relation to serum CA125. Fifty-six women were enrolled in the study. Serial venous blood samples were taken preoperatively and 6, 9 and 12 months after operation for PIIINP and CA125 determinations. The results were correlated to the three-year survival. In Kaplan-Meier survival analysis the preoperative (P = 0.0422), 9-month (P = 0.0062) and 12-month (P = 0.0062) serum PIIINP concentration distinguished between the patients with good and poor prognosis while CA125 did so only at 9- (P = 0.0005) and 12-month (P < 0.0001) follow-up. In the multivariate analysis the independent predictors of prognosis were the preoperative PIIINP and 12-month CA125 concentrations. The percentage changes in serum PIIINP concentration did not differentiate the patients with good or poor prognosis at any time point, whereas the changes in CA125 concentration significantly divided the patients into two prognostic groups during the second half of the postoperative year. We found that PIIINP and CA125 are complementary to each other as predictors of prognosis in epithelial ovarian cancer as preoperative PIIINP was better than CA125 and 1-year CA125 better than PIIINP in this function.

Abstract: The extent of processing of type III collagen is assessed, and the proportions of type I and III collagens are estimated in atherosclerotic plaques obtained from the carotid artery, common femoral artery, and aorta. The fraction of type III collagen that had retained its amino-terminal propeptide (pN-collagen) was 42% in the soluble extract but only 0.0081% in the insoluble residue. Taken together, only 0.011% of the type III collagen in whole plaques was in the form of type III pN-collagen. Together with the small amounts of the free propeptides of type I procollagen, this finding indicates a low rate of collagen turnover. The amounts of solubilized telopeptides of type I and III collagens were measured, after heat denaturation and trypsin digestion of the collagenous helix, by specific immunoassays for the corresponding trypsin-generated antigens. The mean proportion of type III collagen was 61% (95% confidence interval, 58% to 65%) in the carotid and femoral artery plaques and 56% (95% confidence interval, 44% to 68%) in the aortic specimens. The completely processed and cross-linked type III collagen seems to be the major collagen type in atherosclerotic plaques.

Abstract: BACKGROUND: Components of the extracellular matrix play a fundamental role in the process of tumor invasion. The objective of this study was to evaluate the value of Type I collagen metabolites as tumor markers in patients with lung carcinoma. METHODS: In this study, the serum concentrations of the cross-linked carboxyterminal telopeptide of Type I collagen (ICTP) and the aminoterminal propeptide of Type I collagen (PINP) were measured in 59 consecutive patients with lung carcinoma. The blood concentrations of neuron-specific enolase (NSE), carcinoembryogenic antigen (CEA), sialyl Le-1 antigen (SLX), squamous cell carcinoma antigen (SCC), and D-dimer were also evaluated. Data obtained on 18 age-matched healthy subjects and 12 age-matched patients with benign lung disease were available for comparison. RESULTS: The serum concentrations of PINP were significantly higher in patients with lung carcinoma than in age-matched controls. Prechemotherapy values of PINP and ICTP were significantly increased in patients who did not respond to chemotherapy compared with those who did respond. PINP and ICTP were significantly correlated with clinical stage, extent of bone metastasis, survival time, and D-dimer. PINP was also significantly correlated with tumor size. ICTP was significantly correlated with CEA, SLX, SCC, and NSE. PINP also tended to correlate with these tumor-associated glycoproteins. PINP had a better receiver operating characteristic curve than ICTP. The specificity of PINP (79%) for the diagnosis of bone metastasis was superior to that of ICTP (58%). CONCLUSIONS: The results of this study showed that, in addition to the important information that ICTP and PINP provide about the tumor cell-extracellular matrix interaction, they appear to be of great value as adjunct tools for the diagnosis of bone metastasis and as markers of the clinical response to therapy, clinical progression, and prognosis in lung carcinoma patients. However, more studies must be conducted to evaluate further the utility of these markers before their application in clinical practice.

Abstract: The development of cancer involves epithelial-stromal interactions. Alterations in the synthesis and deposition of type I and III collagens are related to the tumor morphology. Skin carcinogenesis in experimental animals provides a reliable model for the development of neoplasia. Ultraviolet (UV) irradiation is the main etiological factor for epidermal dysplasia and malignant tumors in man, but also for dermal degeneration. Non-neoplastic dermal changes and skin tumors induced by ultraviolet irradiation and 7,12-dimethylbenz(a)anthracene were investigated in various mouse strains with different susceptibilities to tumor formation. UVB irradiation resulted in an increased immunoreactivity of collagens in the dermis, in comparison with 7,12-dimethylbenz(a)anthracene. Increased synthesis and deposition of type I and III collagens were found in the stroma adjacent to benign alterations. In well-differentiated squamous cell carcinomas, a similar induction of collagen synthesis and deposition was observed. The destruction of fibrillary structures was more pronounced during the decrease of differentiation from moderately to poorly differentiated squamous cell carcinomas. Anaplastic carcinomas with spindle cell morphology displayed a delicate meshwork of reticular fibers and collagen III, and abnormal expression of mRNA for collagens in some malignant cells with epithelial characteristics. The underlying stroma reacts to the development of epithelial tumors in a reproducible way, which is related to the carcinogenic agent involved.

Abstract: OBJECTIVES: To assess the predictive significance of synovial fluid (SF) analysis for progressive radiological knee joint destruction in arthritis. METHODS: Altogether 55 patients with arthritis and knee joint effusion were included in the study. The diagnosis was rheumatoid arthritis (RA) for 44 of them, chronic seronegative spondylarthropathy for seven and juvenile rheumatoid arthritis for four. The mean age of the patients was 51.8 (SD 14.9, range 19-82) years, and the mean duration of disease 10.9 (SD 9.2, range 0.5-37) years. In addition to the routine laboratory tests, different markers of collagen synthesis and breakdown in serum and SF were assessed. The radiological grade of the knee joint was assessed by Larsen's method at the baseline and after a three year follow up. RESULTS: During the follow up, Larsen's grade deteriorated in 22 (40%) patients. These patients had a significantly higher median level of cross linked carboxyterminal telopeptide of type I collagen (ICTP) in SF at entry than those who had a stable index (p = 0.035). Serum ICTP did not have any predictive value for a specific joint. The median levels of total SF leucocytes (p = 0.012) and the subgroup of polymorphonuclear leucocytes (p = 0.018) were higher in the patients with a stable Larsen's index. However, the relation of SF leucocyte level to radiological progression could not be confirmed in the RA group. CONCLUSION: It is concluded that SF analysis may help in the identification of patients with inflammatory arthritis who are at risk for progressive destruction in a particular joint. A high total SF leucocyte level is not necessarily associated with a poor prognosis. Instead, a high SF ICTP level seems to reflect accelerated bone degradation.

Abstract: BACKGROUND: Epithelial malignancies often induce an enhanced expression of interstitial collagens in the fibroblasts within the tumor tissue and the surrounding non-neoplastic stroma. In uterine carcinosarcomas (malignant mixed müllerian tumors [MMMTs]) both the stroma and the epithelium are malignant. METHODS: In this investigation, both in situ hybridization and immunohistochemical staining were applied with two different antibodies that were capable of distinguishing between newly synthesized and mature, trivalently cross-linked Type I collagen to define Type I procollagen mRNA expression and the synthesis and maturation of the corresponding protein in MMMTs. RESULTS: In the better differentiated parts of these tumors, in which anticytokeratins stained only clearly carcinomatous cells, Type I procollagen mRNA expression was limited to stromal fibroblasts; mature Type I collagen bundles were abundant and regular. In poorly differentiated areas, in which anticytokeratins stained only a few individual cells, Type I procollagen mRNA was expressed peculiarly by three morphologically different cell types. In addition to benign mesenchymal cells, Type I procollagen mRNA was present in atypical epithelial and mesenchymal cells. In these tumors, the collagen bundles close to the malignant cells were comprised of newly synthesized Type I collagen, with only little evidence of the presence of mature, fully cross-linked collagen. CONCLUSIONS: These results strongly suggest that the undifferentiated cells of MMMTs are capable of producing their own stroma with irregularly arranged collagen bundles.

Abstract: We evaluated the significance of biochemical tumor markers, ie, aminoterminal propeptide of type III procoliagen, trivalently cross-linked COOH-terminal telopeptide of type I collagen (ICTP), aminoterminal propeptide of type I procollagen, and CA 125 in the prediction of ovarian cancer outcome and compared them with several classical indicators of prognosis. The concentrations of biochemical markers were determined from the preoperative serum specimens of 55 patients with epithelial ovarian cancer. In the univariate analysis, all biochemical markers except PINP and all conventional prognostic indicators except histological subtype correlated significantly with survival. In the multivariate Cox analysis of biochemical markers, serum ICTP remained the only significant prognostic indicator of overall survival. Among all variables, clinical stage and ICTP were the only independent and significant determinants of prognosis. Because the content of trivalently cross-linked, mature type I collagen (the breakdown of which is detectable in the ICTP test) in malignant ovarian cancer tissue has been reported to be lower and that of bivalently cross-linked and non-cross-linked collagen has been reported to be higher than in benign tumors, the source of excess ICTP in the circulation of ovarian cancer patients is most likely the degradative damage of soft tissues surrounding the progressively growing malignant lesions. The serum ICTP concentration can thus be regarded as an indicator of the invasion of ovarian cancer. Such information is not available by conventional methods. Therefore, the ICTP test will improve the accuracy of predicting clinical outcome in this disease.

Abstract: Type I collagen synthesis (PINP, PICP) and degradation (ICTP) markers were analysed from preoperative serum samples of 138 women with breast cancer (BC), 94 women with benign breast disease (BBD) and 100 healthy controls to evaluate the levels of these markers and the stage of BC at the time of diagnosis. We also compared the clinical utility of these markers in detecting BC with CA15-3 and CEA. The mean value of ICTP was statistically significantly elevated in the BC group (p < 0.001), as compared with the control group, but the elevated values in BC group were due to stage IV disease. The sensitivity of ICTP in detecting BC was 0.23, which was equal with CA15-3(0.24) or CEA(0.23). The sensitivity of both PICP and PINP for diagnosing BC was poor, but a tendency to higher serum levels of PINP and low PICP/PINP ratio was detected in patients with advanced stage IV disease. These results indicate that high preoperative serum levels of ICTP are associated with advanced BC, but like CA15-3 and CEA, its clinical value in diagnosing purpose is poor.

Abstract: Malignant tumours often induce a fibroproliferative response in the adjacent stroma, characterized by increased expression of type I and type III procollagens. In normal tissues, fibrillar collagens normally undergo extensive intermolecular cross-linking that provides tensile strength to the tissue. Here we set out to characterize collagen cross-linking in human ovarian carcinoma tissue in vivo. Biochemical and immunochemical methods were used for cross-linked telopeptides of type I and III collagens in samples of benign and malignant serous tumours. The locations and staining patterns of these proteins were visualized immunohistochemically. The contents of both total collagen and the cross-linked type I and type III collagens in the malignant samples were only about 20% of those in the benign tumours. The cross-linked telopeptide antigens derived from the collagens were smaller and more heterogeneous in size in the malignant than in the benign tumours, indicating a defective cross-linking process scarcely leading to the formation of mature cross-links in the collagen fibres in malignancy. Immunostaining revealed disorganized type I and type III collagen bundles in carcinomas. These findings suggest that the collagen cross-linking process is aberrant in malignant tumours, possibly resulting in increased susceptibility of tumour collagens for the proteolysis often associated with tumour invasion.

Abstract: 52 patients with early rheumatoid arthritis (RA) were followed with regular measurements of bone mineral density (BMD) and serum markers of type I collagen metabolism in order to determine whether they develop osteoporosis during the first two years of the disease course and if the changes in type I collagen metabolites reflect the alterations in BMD. The mean percentage BMD change over the first year of follow-up was -0.91 for lumbar spine (LS) and -0.76 for femoral neck (FN); the corresponding figures from 0 to 24 months was -1.3 and -0.8, respectively. During the follow-up, only five patients developed osteoporosis by the Z-score definition (<-1). If defined by T-score (<-2.5) none of the patients developed osteoporosis. The BMD change correlated neither with the clinical parameters of disease activity nor with the markers of collagen metabolism. However, the BMD change in FN was associated with the cumulative corticosteroid dose (r=-0.31, p <0.05, 95% CI -0.54 to -0.04). Reasons for the lack of accelerated bone loss in our series are open to various interpretations. This series was community based and most of the patients had mild RA. The patients were also actively treated and their physical function did not deteriorate.

Abstract: We describe the clinical findings and biochemical features of a male child suffering from a so far undescribed lethal connective tissue disorder characterised by extreme hypermobility of the joints, lax skin, cataracts, severe growth retardation, and insufficient production of type I and type III procollagens. His features are compared with Ehlers-Danlos type IV, De Barsy syndrome, and geroderma osteodysplastica, as these disorders show some symptoms and signs shared with our patient. The child died because of failure of the connective tissue structures joining the skull and the spine, leading to progressive spinal stenosis. The aortic valve was translucent and insufficient. The clinical symptoms and signs, together with histological findings, suggested a collagen defect. Studies on both skin fibroblast cultures and the patient's serum showed reduced synthesis of collagen types I and III at the protein and RNA levels. The sizes of the mRNAs and newly synthesised proteins were normal, excluding gross structural abnormalities. These findings are not in accordance with any other collagen defect characterised so far.

Abstract: Change in the synthesis of type I collagen, the major extracellular matrix component of skin and bone, are associated with normal growth, tissue repair processes, and several pathological conditions. Expression of the COL 1A1 gene is regulated by transcriptional and post-transcriptional mechanisms. However, the hormonal regulation of type I collagen synthesis in human bone has not been well characterized. We have studied the influence of calcitriol, dexamethasone, retinoic acid, and estradiol on the COL 1A1 gene expression by determining the secretion of the C-terminal propeptide (PICP) and the levels of alpha 1(I) procollagen mRNA in cultured human MG-63 and SaOs-2 osteoblast-like osteosarcoma cells. Similar experiments were also performed with respect to expression of the nuclear proto-oncogenes, c-fos and c-jun, in MG-63 cells. In MG-63 cells, calcitriol stimulated the synthesis and secretion of PICP. The alpha 1(I) procollagen mRNA level was elevated with no effect on message stability, indicating a transcriptional mechanism of regulation. In contrast, dexamethasone treatment was accompanied by an accelerated rate of alpha 1(I) procollagen mRNA turnover, observed as decreased amounts of the message and the secreted PICP, implying a posttranscriptional regulation. Retinoic acid, in turn, decreased the levels of alpha 1(I) procollagen mRNA and secreted PICP by slowing down transcription of the COL1A1 gene without any effect on message stability. The ability of these hormones to regulate the alpha 1(I) transcripts was sensitive to puromycin treatment, suggesting an involvement of an induced mediator protein in the action of the hormones on the COL1A1 gene. Both dexamethasone and calcitriol rapidly but transiently increased the expression of the c-fos and c-jun proto-oncogenes. Neither proto-oncogene responded to retinoic acid treatment with significant changes in mRNA levels. Estradiol treatment was found to have no influence on type I procollagen synthesis. In SaOs-2 cells, which are not as well differentiated as the MG-63 cells, calcitriol and dexamethasone did not influence type I procollagen synthesis. Retinoic acid as well as estradiol reduced collagen gene expression in these cells. These findings suggest that hormonal effects on type I procollagen synthesis may depend on the maturational state of the osteoblastic cells that express different regulatory factors and receptors, resulting in, in each case, a finely adjusted rate of gene expression.

Abstract: The synthesis and degradation of type I and type III interstitial collagens releases several antigenic metabolites, whose measurement allows the metabolism of connective tissue to be evaluated under a variety of different conditions. In this study we investigated the influence of benign and malignant ovarian neoplasms on the metabolism of these collagens. The study population comprised patients with benign (n = 53), borderline (n = 6) or malignant (n = 36) ovarian neoplasms. We quantified the serum, cyst fluid and peritoneal/ascitic fluid concentrations of the amino-terminal propeptide of type I (PINP) and III (PIIINP) procollagens, indicators of the synthesis of type I and III collagen, respectively and the cross-linked carboxy-terminal telopeptide of type I collagen (ICTP), an indicator of type I collagen degradation. Macrophage colony-stimulating factor 1 (CSF-1) concentration was also assayed as its serum level is increased in ovarian cancer and CSF-1 may be involved in the regulation of collagen metabolism. The concentration of each antigen was significantly higher in patients with malignant tumour than with benign neoplasm in each comparison, except for ICTP in peritoneal fluid and for CSF-1 in cyst fluid. The high ascitic fluid concentration of PINP, PIIINP or CSF-1 correlated with malignancy, and the low cyst fluid concentration of any of the four markers was indicative of benign tumour. Levels of CSF-1 did not correlate with the levels of any of the markers of collagen turnover. The concentration of PINP in ascites was about 50 times higher and in cyst fluid about eight times higher than that in the serum from patients with malignant tumour, whereas the respective ratios for ICTP were only 2.5 and 1.3. In such patients, the ratio of ascitic fluid to serum concentration was also about 80-fold higher for PIIINP and about 20-fold higher for PINP than for ICTP. The different distributions of PIIINP, PINP and ICTP suggests dominance of synthetic processes or retarded elimination of PIIINP and PINP in ovarian cancer. In advanced malignancies, the accumulation of PINP and PIIINP in abdominal space, possibly due to increased synthesis and/or failed resorption, may promote ascites formation. This study shows that both accelerated synthesis and breakdown of fibrillar collagens are characteristic of ovarian malignancy, and suggests that measurements of cyst fluid or ascitic fluid concentrations of collagen metabolites or CSF-1 could be used in the differential diagnosis of benign and malignant ovarian neoplasms.

Abstract: OBJECTIVE: To investigate growth and markers of collagen and bone metabolism in prepubertal children with asthma. STUDY DESIGN: We measured growth velocity over 12 months and markers of collagen types I and III synthesis (PINP, PICP, PIIINP), collagen type I degradation (ICTP), and bone metabolism (bone-specific alkaline phosphatase and osteocalcin) on one occasion in 56 prepubertal children with stable asthma, 39 of whom were treated with inhaled budesonide or beclomethasone. Collagen data were compared with normal control values. RESULTS: Children treated with inhaled steroids had reduced collagen synthesis (PINP, PIIINP) compared with control subjects (p = 0.038, p = 0.045), although PICP was increased (p = 0.05). Carboxyterminal telopeptide of type I collagen was reduced in patients treated with inhaled steroids (p < 0.0005) compared with nonsteroid-treated patients. Serum osteocalcin but not bone-specific alkaline phosphatase was significantly reduced in children treated with inhaled steroids (p < 0.02). Significant correlation was observed between PIIINP and ICTP and growth velocity. CONCLUSION: Collagen turnover is reduced in children with asthma receiving long-term inhaled steroid treatment. Markers of collagen synthesis provide a more accurate reflection of growth disturbance than osteocalcin and bone-specific alkaline phosphatase.

Abstract: The distribution of type I collagen, the major component of human dermis, was characterized by immunohistochemistry in skin lesions of chromoblastomycosis, a chronic cutaneous mycosis, before and after a specific antifungal treatment with terbinafine to study the changes induced in the lesions by the treatment. Newly synthesized type I collagen was studied with an antibody directed against the aminoterminal propeptide of the molecule (PINP), whereas mature, cross-linked type I collagen was detected with an antibody against the carboxyterminal telopeptide of type I collagen (ICTP). The isopeptide N epsilon gamma-glutamyl lysine (N epsilon gamma GL), synthesized by transglutaminase and able to cross-link several components of the extracellular matrix, has also been investigated with two monoclonal antibodies to determine if it is involved in the stabilisation of the fibrotic cutaneous lesions. The degradative process involved in the remodelling has also been assessed by immunohistochemistry with anti-metalloproteinase (MMP-1 and MMP-9) and anti-tissue inhibitor (TIMP-1) antibodies. All tissue macrophages stained for CD68 and MMP-9, but not for MMP-1, while the polymorphonuclear neutrophils had an elastase and a weak MMP-9 phenotype. The fibroblasts of fibrotic areas stained constantly for N epsilon gamma GL and PINP. The immunostaining of extracellular matrix for ICTP and N epsilon gamma GL, and the number of PINP-positive fibroblasts, decreased significantly after one year of antifungal treatment. Terbinafine treatment decreases the synthesis of type I collagen and leads to a partial reversal of the cutaneous fibrotic lesions, independently of the cure of the fungal infection.

Abstract: The aim of this study was to determine the correlation between changes in collagen metabolites (ICTP, mature cross-linked carboxy-terminal telopeptide of type I collagen; PINP, the amino-terminal propeptide of type I procollagen) and bone mineral density (BMD) in 206 pre- and post-menopausal breast cancer patients with non-metastatic disease. All patients received adjuvant cancer treatment--premenopausal patients chemotherapy and post-menopausal patients anti-oestrogens. In addition, the patients were also randomized to receive oral clodronate 1600 mg daily for 3 years. BMD was measured at baseline and at 1 and 2 years, the collagen metabolites at baseline and at 1 year. There was a highly significant negative correlation between the changes in PINP and BMD in lumbar spine and femoral neck from baseline to 12 months in all patients (r(s) = -0.68, P < 0.0001, and -0.45, P < 0.0001, respectively), and in pre- and post-menopausal patients separately. The changes in PINP levels at 12 months predict further changes in BMD at 24 months (r = -0.70, P < 0.0001, and -0.51, P < 0.0001, respectively). ICTP and BMD changes correlated significantly only in lumbar spine of premenopausal patients who developed rapid bone loss due to chemotherapy-induced amenorrhoea (r(s) = -0.34, P = 0.0003). The PINP levels at 12 months were significantly lower in the clodronate group than in the control group (P < 0.0001). Our results indicate that PINP is a sensitive marker of bone turnover rate. Changes in PINP levels significantly predicted changes in BMD and correlated with the antiresorptive efficacy of clodronate treatment.

Abstract: Increased synthesis and degradation of extracellular matrix components are associated with breast cancer development. This study evaluated type I and type III procollagen mRNA expression and the corresponding protein synthesis and maturation, as well as the tissue distribution of these collagens, in benign breast lesions, infiltrating ductal carcinomas, and their metastases by in situ hybridization and immunohistochemistry. In the benign lesions, the type I and type III collagen bundles were regularly organized and the expression of the corresponding mRNA was weak, indicating a relatively slow collagen turnover. In the malignant tumours, increased expression of type I and type III procollagen mRNAs was observed in the fibroblastic cells of the stroma; the malignant epithelial cells did not participate. The staining of corresponding newly-synthesized pN-collagens showed aberrant bundles in the invasive front of the malignant tumours. Newly-synthesized type I and type III procollagens were occasionally observed in fibroblastic cells, particularly in grade 2 and grade 3 tumours. Metastases of breast carcinoma resembled poorly differentiated primary tumours with respect to their collagen synthesis and deposition. The increased synthesis of fibrillar type I and type III procollagens may serve as a pathway for tumour invasion. The enhanced synthesis is associated with the formation of aberrant collagen bundles, which may be more readily degradable and may thus facilitate breast tumour invasion.

Abstract: Systemic sclerosis (SSc; scleroderma) results in the excessive deposition of extracellular matrix components in affected organs. This is partly due to enhanced synthesis; however, the role of degradative processes in this disease is still poorly understood. Sera of 32 patients with SSc (22 with the diffuse, 10 with the limited form) and of six patients with morphoea were assessed using radioimmunoassays for the cross-linked carboxy terminal telopeptide of type I collagen (ICTP) and for the amino terminal propeptide of type I procollagen (PINP) reflecting type I collagen degradation and synthesis, respectively. In 27 of the 32 patients with SSc, the concentration of ICTP was above the upper limit of the normal value (4.6 micrograms/L) and the mean level was clearly elevated at 7.92 micrograms/L. The ICTP concentration correlated with the skin score measuring the extent of the lesions, whereas no such correlation was found for PINP. The ICTP antigen in serum, studied by immunoblotting, had a molecular weight of about twice that of the trypsin-generated fragment isolated from human bone collagen. The mean concentration of serum PINP was 43.9 micrograms/L and no patient exceeded the upper limit of the normal range (80 micrograms/L). We report here for the first time that the concentration of the type I collagen degradation product ICTP in serum shows a close correlation with the extent of skin fibrosis in patients with SSc. We conclude that the increased deposition of type I collagen in this disease is accompanied by an increased turnover of this molecule, indicating a more complex derangement of synthetic and degradative processes than previously acknowledged.

Abstract: BACKGROUND: The propeptides derived from type I and III procollagens, PICP, PINP and PIIINP, indicate the synthesis of the corresponding collagens. Their circulating concentrations reflect the growth velocity in infants and children METHODS: We measured these propeptides in 145 samples of amniotic fluid of normal pregnancy. In addition, we have analysed an amniotic fluid and serum sample from a mother with osteogenesis imperfecta, and later the infant's serum sample was also collected for procollagen propeptide analysis. RESULTS: High concentrations of propeptides, 100-1000 times higher than those in adult serum, were found in early second trimester, decreasing significantly towards term, reflecting the decreased foetal growth rate. Interestingly, the amino-terminal propeptide of type I procollagen, PINP, decreased more that the corresponding carboxy-terminal propeptide, PICP, although both are in principle derived from the same protein. At both stages of pregnancy, the discrepant ratio of PICP to PINP indicated a molar excess of PINP. Abnormally low concentrations of PICP and PINP with normal PIIINP concentrations measured in amniotic fluid and in the serum indicated decreased synthesis of type I procollagen in a foetus/infant with mild osteogenesis imperfecta. CONCLUSIONS: Our data show a decrease in collagen synthesis with the stage of pregnancy and lower values of type I procollagen propeptides in a case of OI.

Abstract: Cutaneous parasitic lesions, associated with a dense fibrous reaction, markedly improved under albendazole treatment in one case of supraumbilical skin localization of alveolar echinococcosis. Since collagen cross-linking increases during fibrogenesis and contributes to the stability of fibrotic lesions, we monitored the level of the cross-links pyridinoline and pentosidine in skin lesions from this patient to determine if they would reflect the changes occurring during treatment. We looked at the deposition of cross-linked type I collagen by immunohistochemistry and also measured the serum concentrations of pentosidine and of a fragment of type I collagen (ICTP), which contains a site of pyridinoline formation. Albendazole treatment did not affect either the collagen content of skin lesions or the serum concentrations of ICTP and pentosidine, but it led to a pronounced decrease in pyridinoline level concomitant with the disappearance, observed by immunohistochemistry, of extensively cross-linked fibrotic type I collagen. The follow-up of collagen cross-linking by pyridinoline in skin tissue thus appears to be useful in reflecting the improvement of fibrotic skin diseases during therapy.

Abstract: Fifty-seven patients with advanced prostate cancer resistant to first-line hormonal therapy were treated with estramustine and additionally randomized for treatment with clodronate or placebo. Clodronate treatment was started with 5 days intravenous administration (300 mg day[-1]) and followed by oral treatment (1.6 g day[-1]) for 12 months. Skeletal pain relief was only about 10% better in the clodronate than in the placebo group. The results do not support the superiority of combined intravenous and oral treatment with clodronate compared with oral administration only.

Abstract: A number of proteins that act as necessary catalysts for correct protein folding and oligomerization in the endoplasmic reticulum (ER) are known to be retained in the organelle via the KDEL-receptor mediated retrieval mechanism. However, a complementary system that may help to retain these proteins in the organelle lumen has been suggested to exist and likely involves physical protein-protein interactions at the level of endoplasmic reticulum (ER) itself. In this report, we provide both morphological and biochemical evidence in support of this proposal. We show that in collagen-secreting human skin fibroblasts, protein disulfide isomerase and newly synthesized procollagen chains exist predominantly in an "aggregated" state, and form a reticular-like matrix in the ER lumen in vivo. The size of the aggregates was found to be variable, and may exceed 1.5 million Da. Aggregate formation appeared to be transient and to involve multiple types of protein-protein interactions, including formation of aberrant disulfide bonds. Association of protein disulfide isomerase, on the other hand, was found to require at least partly function-related disulfide bonds. These results support the existence of a reticular-like matrix in the ER lumen, and suggest that aggregation may be part of the normal maturation pathway during collagen biosynthesis.

Abstract: The biochemical possibilities for developing specific assays for type I collagen metabolism are described. Type I collagen synthesis can be assessed either by the analysis of the carboxyterminal or aminoterminal propeptides, which are in principle produced in a molar ratio of 1:1. However, in clinical situations altered behaviour can be found, the reasons for which may be altered clearance or even the existence of variant forms of type I collagen. Type I collagen degradation can be specifically detected by analysis of either cross-linked carboxy- or aminoterminal telopeptides or by the cross-links themselves liberated during the degradation processes. The heterogeneity of the cross-links and the constituent chains of the cross-linked peptides in different tissues and possibly in different clinical situations introduce problems, which should be studied and resolved in the future.

Abstract: STUDY DESIGN: Open, prospective trial with patients participating in an active back restoration program. OBJECTIVES: To compare the concentrations of biochemical indicators of Type I and III collagen synthesis and Type I collagen degradation in the serum of patients with chronic low back pain and healthy control subjects and to evaluate the effect of active back rehabilitation based on vigorous exercise on collagen metabolism. SUMMARY OF BACKGROUND DATA: The aim of active back rehabilitation is to restore the physical function of low back pain patients falling into the so-called "deconditioning syndrome." The changes in functional muscle strength measurements during the restoration rehabilitation program always depend on motivation, learning phenomena, and fear of pain and injury, so that even more objective ways of showing changes in physical activity are needed. METHODS: Specific radioimmunoassays for the amino-terminal (PINP) and carboxy-terminal (PICP) propeptides of Type I procollagen, the amino-terminal propeptide of Type III procollagen (PIIINP), and the cross-linked carboxy-terminal telopeptide of Type I collagen (ICTP) were used for serum samples obtained from 41 patients before, during, and after an active back restoration program and from 16 age- and sex-matched healthy control subjects. RESULTS: The circulating concentrations of PINP and PICP were initially lower in the patients ([mean +/- SD] 35.3 +/- 12.5 micrograms/L and 119.0 +/- 32.6 micrograms/L, respectively) than in the control subjects (47.9 +/- 18.0 micrograms/L and 136.7 +/- 47.7 micrograms/L, respectively; P < 0.05 for PINP). Toward the end of the active back rehabilitation program, both PINP and PICP increased in the patients (P < 0.001 for the increase between the initial level and the end of rehabilitation). There was a significant difference in the time courses of these changes, with the circulating PICP concentration increasing earlier than that of PINP. In the intervention group, the PIIINP concentration also increased (P < 0.01), whereas the ICTP concentration remained unchanged, with a tendency to decrease. No changes occurred in the control subjects. CONCLUSIONS: Active back rehabilitation based on vigorous exercise increases. Type I collagen synthesis in patients with chronic low back pain; this may provide a means of objectively verifying the effects of such rehabilitation.

Abstract: The aminoterminal propeptide of type III procollagen (PIIINP) and the carboxyterminal propeptide of type I procollagen (PICP) and hyaluronan (HA) were measured in plasma and suction blister fluid from 13 systemic sclerosis patients and 11 healthy volunteers. Suction blisters and skin biopsies were from the transition zone between normal skin and scleroderma, and uninvolved abdominal skin of patients. The median value of suction blister PIIINP from the transition zone was 38% higher than suction blister PIIINP from uninvolved skin. PIIINP was localized to the dermis by immunohistochemical techniques. PICP and HA levels in blisters from the transition zone were 87% and 53%, respectively, above the levels measured in uninvolved skin. Furthermore, PICP and HA blister levels from the transition zone were 67% and 63%, respectively, higher than the levels measured in healthy volunteers. In plasma from scleroderma patients levels of PIIINP and HA were 38% and 127% higher, respectively, than in plasma of healthy volunteers. The plasma PICP level was not significantly higher in scleroderma patients. Finally, PICP, PIIINP and HA levels were several times higher in suction blister fluid than in plasma. The data indicate that a fibrogenetic process takes place in the transition zone of scleroderma. The method may be used to monitor the progression of scleroderma skin lesions in vivo.

Abstract: In children with acute lymphoblastic leukemia (ALL), the metabolism of type I collagen, the major collagen of bones, may be changed at diagnosis and during early chemotherapy. In the present study, bone formation and degradation rates were evaluated longitudinally in 35 children with ALL, using two serum markers of bone collagen formation: the amino-terminal (PINP) and carboxyterminal (PICP) propeptides; and a marker of degradation: the carboxyterminal telopeptide of type I collagen (ICTP). These serum markers were determined at diagnosis, during induction treatment (at 1, 4, and 6 weeks), and during consolidation treatment (at 8 and 12 weeks). The changes in the serum markers suggested that, at diagnosis, type I collagen turnover (i.e., both synthesis and degradation) was remarkably low. The median serum levels of PINP, PICP, and ICTP were -2.6 SDS (standard deviation score), -1.5 SDS, and -2.5 SDS, respectively. The PICP and PINP levels declined further during the first week of therapy (p < 0.001), whereas the ICTP levels had risen by end of the induction phase (p < 0.05). By the end of the 12 week interval, the concentrations of the formation and degradation markers had returned to normal (p < 0.01). Our findings suggest that ALL is accompanied by low turnover of bone collagen. The abnormalities are at first aggravated, but then corrected, by treatment.

Abstract: Postmenopausal hormone replacement therapy (HRT) lowers the turnover rate of the mineralized bone matrix, the predominant organic component of which is type I collagen. The effect of estrogen on bone metabolism has been monitored by measuring the circulating concentration of the carboxy-terminal propeptide of type I procollagen (PICP), which decreases during HRT. We have recently developed assays for the intact amino-terminal propeptide (PINP) of type I procollagen, a protein set free from the other end of the same gene product. PICP and PINP, both derived from the synthesis of type I collagen, but differing in their further metabolism, were assessed in 47 postmenopausal women, aged 45-66 years, undergoing postmenopausal HRT. Estradiol-gel applied daily was combined to a continuous progestin administered by three different routes. Serum samples obtained before the treatment and 6 and 12 months after its commencement were analyzed for PICP, PINP, PINP Col 1 (assay variant measuring also the degradation product of PINP) and PIIINP (amino-terminal propeptide of type III procollagen). During HRT the circulating concentration of PICP decreased by 20%, that of PINP by 42% and that of PINP Col 1 by 32% in 12 months. The correlation between the two propeptides, which was 0.676 before the treatment, increased to 0.851 in 6 months and to 0.815 in 12 months. The correlations between PINP and PINP Col 1 were 0.872 before the treatment and increased to 0.925 and 0.941 after 6 and 12 months of treatment, respectively. The serum concentration of PIIINP, which reflects the turnover of the soft tissue collagens, did not change remarkably. Our findings indicate that the intact PINP is a more dynamic marker of bone metabolism than PICP and can therefore be recommended as a marker reflecting the effect of estrogen on bone collagen formation during HRT.

Abstract: The aim of the study was to estimate the usefulness of measuring the circulating concentration of serum aminoterminal propeptide of type III collagen (S-PIIINP) in screening for hepatobiliary diseases in patients with ulcerative colitis. S-PIIINP was measured in 69 patients with ulcerative colitis and normal liver biochemistry, in 14 patients with ulcerative colitis and elevated catalytic concentration of alkaline phosphatases in serum (S-ALP, EC 3.1.3.1) but without primary sclerosing cholangitis (PSC), and in 20 patients with ulcerative colitis and PSC. The median S-PIIINP was 3.1 micrograms l-1 in patients with ulcerative colitis and normal liver biochemistry, 4.3 micrograms l-1 in patients with ulcerative colitis and hepatobiliary disorder other than PSC and 8.9 micrograms l-1 in those with ulcerative colitis and PSC. When the S-PIIINP cut-off level was set at 5.0 micrograms l-1, 1% of the patients with ulcerative colitis and normal liver biochemistry, 21% of those with hepatobiliary disorder, not PSC, and 90% of the patients with PSC had S-PIIINP values above that concentration. In conclusion, S-PIIINP above 5.0 micrograms l-1 in a patient with ulcerative colitis strongly suggests concomitant PSC.

Abstract: The propeptides PICP and PINP are derived from the synthesis of type I collagen, a major matrix protein of bone and soft tissues. The aim of this cross-sectional study was to investigate their value as indicators of the aggressivity of breast cancer. Serum PINP, PICP, and total alkaline phosphatase were determined from 89 breast cancer patients. Forty had major bone and/or soft tissue metastases with an aggressive disease course: the progressive disease (PD) group. Forty-nine had either none or minor bone and/or soft tissue metastases with a stable clinical course: the stable disease group (SD). The mean value of PINP in the PD group was 7.2 times higher than that in the SD group (276 +/- 79 microg/l versus 38 +/- 3 microg/l, respectively; P = 0.005), whereas PICP mean value was only 1.7 times higher in the PD group (174 +/- 20 microg/l versus 100 +/- 5 microg/l; P = 0.001). The ratio of PICP to PINP was 1.02 +/- 0.07 in the PD group and 3.07 +/- 0.18 in the SD group (P < 0.001). The correlation between PICP and PINP was linear in the SD group and nonlinear in the PD group. The results indicate that high serum PICP and PINP concentrations and a low PICP:PINP ratio are associated with a highly aggressive nature of breast cancer. Determination of PINP, in particular, may be valuable when evaluating the clinical status of a breast cancer patient.

Abstract: To evaluate the role of collagen metabolites in the prediction of the response to GH treatment we measured the serum concentrations of the C-terminal propeptide of type I procollagen (PICP) and the N-terminal propeptide of type III procollagen (PIIINP) with specific RIAs in 35 short children (16 boys) before and after 5 days, 5 weeks and 3 months of GH therapy. The mean age of the children was 10.3 years (range 1.9-16.4 years) and the bone age ranged from 1.2 to 12.5 years (mean 7.6 years). The initial mean relative height (RH) was -3.6 SDS (range -6.6 to -2.4 S.D.). Nineteen children were found to have GH deficiency (GHD; peak GH responses in two pharmacological tests < 10 micrograms/l), while the remaining 16 were considered to have undefined short stature (USS). The children were treated with recombinant human GH (0.1 U/kg given subcutaneously at bedtime 6-7 times/week). The increases in RHI over the first 6 and 12 months of therapy were used as response measures. There was already a significant increase (P < 0.001) in both the serum PICP and PIIINP levels at 5 days, and the concentrations continued to rise up to 3 months, PICP levels rising less than the PIIINP levels. In the whole group the RHI over 6 months correlated most strongly with the absolute PICP concentrations at 3 months (rS = 0.59; P < 0.05), while the absolute PIIINP concentrations at 3 months showed the strongest relation to the one year RHI (rS = 0.69; P < 0.001). In the GHD group the 6 month RHI was most strongly related to the absolute PICP concentration at 3 months (rS = 0.59; P < 0.05). In the USS group the absolute PICP concentrations at 3 months correlated most strongly with the one year RHI (rS = 0.82; P < 0.01). Significant correlations were also observed between the absolute PIIINP levels at 3 months and the 6 month RHI (rS = 0.60; P < 0.05) and 12 month RHI (rS = 0.76; P < 0.01) in this group. These results show that GH therapy results in an unequivocal increase in circulating concentrations of PICP and PIIINP. The serum PICP and PIIINP concentrations may be of value in the prediction of the long-term response to GH therapy.

Abstract: Simple bone marrow fibrosis is seen in 10-30% of multiple myeloma (MM) patients. We investigated the incidence and characteristics of the bone marrow stromal alterations, in order to characterize the collagens involved by immunohistochemistry, and to evaluate the use of serum aminoterminal propeptide of type III procollagen (PIIINP) as a marker of marrow fibrogenesis and disease activity in MM. 34 consecutive patients with newly diagnosed MM were included prospectively, and followed for 12-30 months. Compared with the findings in 15 normal individuals we found increased interstitial deposits of collagen III in 48% of MM patients, whereas deposits of collagen I were not increased. Interstitial fibrosis appeared to be restricted to areas of severe plasma cell infiltration, but it could also have a more dispersed presentation in the severely infiltrated marrow. There was a high co-distribution of collagen III fibrils and reticulin fibres. Serum PIIINP levels were elevated in most patients, and in the follow-up study serum PIIINP showed a good correlation with the response to treatment. Patients with resistant or progressive disease had continually elevated levels of PIIINP. In most patients with responsive disease serum PIIINP normalized, and we observed no relapses in patients who had normal serum PIIINP levels. Other patients who responded to treatment by reduced M-component level, but had persistently elevated serum levels of PIIINP, had either early relapses or developed progression of osteolytic lesions in spite of unchanged M-component levels. Therefore an elevated serum PIIINP during treatment might indicate an active malignant clone. Serum PIIINP does not simply follow the M-component, but gives further information of potential therapeutic value.

Abstract: Calcium deficiency is a major etiological determinant of rickets in Nigerian children and is accompanied by undermineralization of the developing bone matrix which is composed largely of type I collagen. We have assessed types I and III collagen metabolism by measuring the circulating concentrations of teh N- and C-terminal pro-peptides (intact PINP and PICP) and the C-terminal telopeptide (ICTP) of type I collagen, and the N-terminal pro-peptide (PIIINP) of type III collagen in 94 healthy Nigerian children and in 44 children aged 1-5 years with active calcium-deficiency rickets. In active rickets the mean levels of the four collagen metabolites were approximately twofold higher than in the healthy children, despite a wide variation of individual values. Mean intact PINP was 812 +/- 279 versus 403 +/- 189 microg/liter; PICP was 573 +/- 265 versus 348 +/- 299 microg/liter; PIIINP was 16.8 +/- 8.6 versus 10.8 +/- 3.6 microg/liter, and ICTP was 28.4 +/- 17.2 versus 11.9 +/- 4.1 microg/liter (all P < 0.001), in rachitic and healthy children, respectively. Healthy children younger than 3 years had higher levels of all the collagen metabolites than those between 3 and 5 years (all P < 0.05). Alkaline phosphatase was greater in rickets than in the healthy group (P < 0.001) whereas mean osteocalcin levels were slightly lower (P = 0.009). 1,25(OH)2D correlated with all the collagen propeptides, but not with ICTP in the healthy children. No such correlations were found in rickets, where there was a poor inverse correlation between 1,25(OH)2D and ICTP. These data suggest that collagen turnover is elevated in cases of calcium-deficiency rickets, where vitamin D status is adequate, possibly indicating increased turnover of undermineralized osteoid.

Abstract:

Abstract: To assess the mechanisms of osteopenia in inflammatory bowel disease (IBD), the serum markers of bone formation (osteocalcin and carboxyterminal propeptide of type I procollagen (PICP)) and bone degradation (carboxyterminal telopeptide of type I collagen (ICTP)), the bone mineral density (BMD) of the lumbar spine and the proximal femur and calcium intake of 150 unselected IBD patients and 73 healthy controls were investigated. The patients had higher ICTP values (3.69 (SD 1.40) microgram/l) than the healthy controls (3.25 (1.00) microgram/l, p = 0.035), but no differences in serum PICP and osteocalcin between these groups were detected. In the patients, the ICTP, PICP, and osteocalcin values did not have any significant correlation with BMD, but the patients with ICTP values above 3.6 microgram/l had significantly lower Z scores than those with lower ICTP. In the controls, however, a positive correlation between serum ICTP and BMD was found. The ulcerative colitis patients with total colitis had higher values of ICTP (3.96 (1.58) microgram/l) than those with a left sided disease (3.04 (0.86) micrograms/l, p = 0.009). The patients with a history of clinically active disease (n = 20) had higher ICTP (4.58 (1.55) microgram/l) and osteocalcin (12.56 (5.64) microgram/l) values than the patients (n = 130) with quiescent disease (ICTP 3.56 (1.33), p = 0.002, and osteocalcin 9.76 (3.62), p = 0.017). Increased serum osteocalcin, PICP, and ICTP concentrations and reduced BMD Z scores were found in a subgroup of Crohn's disease patients with a history of an active disease (n = 11). Raised serum ICTP and normal values of osteocalcin and PICP in IBD patients show increased breakdown of type I collagen without a compensatory increase in its synthesis suggesting an increased rate of bone degradation as a probable mechanism for osteopenia in IBD. Raised ICTP values are related to reduced bone mineral densities.

Abstract: Skin atrophy has been observed after prolonged use of inhaled corticosteroids. We therefore studied the effect of inhaled budesonide and nedocromil in patients with asthma on concentrations of procollagen propeptides in suction blister fluid reflecting skin collagen synthesis in vivo. Both types I and III procollagen propeptide concentrations decreased significantly after 6 wk of either 1,600 micro g/day (n=10) or 400 micro g/day (n=9) of inhaled budesonide but not in control subjects using inhaled nedocromil 16 mg/day (n=9). The reduction in mean propeptide concentrations ranged from 39 to 63%; the effects of the two budesonide doses did not differ significantly. Thus, even a low dose of inhaled corticosteroid represses skin collagen synthesis within a relatively short period.

Abstract: Malignant ovarian tumors induce a strong fibro-proliferative reaction characterized by the active production of type I and type III procollagen both locally in the ovary as well as more remotely in the peritoneal cavity. Our purpose was to determine the origin of the increased collagen production observed in serous ovarian tumors with different histological grades of malignancy, ie, whether the malignant cells or the stromal fibroblasts are responsible for the synthesis of collagen fibers. We visualized the mRNAs corresponding to the pro alpha 1(I) and pro alpha 2(I) chains of type I procollagen and the pro alpha 1(III) chain of type III procollagen by in situ hybridization. Strong signals for both chains of type I procollagen were seen in stromal fibroblasts next to tumor cell islets, whereas the reaction was weak or absent near benign ovarian cysts. In poorly differentiated tumors, the signals were particularly abundant and occasionally also seen in the neoplastic cells themselves. Type III procollagen mRNA expression was similar, although somewhat less distinct. These findings indicate that the production of interstitial procollagens is related to the degree of malignancy and neoplastic activity of tumors. The formation of collagen in well differentiated ovarian tumors is a function of stromal fibroblasts, whereas in poorly differentiated tumors, aberrant expression of one or several chains of type I and type III procollagens in the neoplastic cells is also likely to take place.

Abstract: Thirty-six patients with bone metastases included in a trial of supportive calcitonin on the treatment response to systemic therapy were monitored by conventional radiography, conventional indicators of bone metabolism [alkaline phosphatase (AP), osteocalcin (gla), urinary hydroxyproline excretion (OHP), urinary calcium (uCa), serum calcium (sCa)] and collagen metabolites (ICTP, the pyridinoline cross-linked carboxy-terminal telopeptide of type I collagen; PICP, the carboxy-terminal propeptide of type I procollagen; and PIIINP the amino-terminal propeptide of type III procollagen). All patients had been on the same systemic treatment for at least 3 months at the start of the trial. There was a positive correlation between the concentrations of ICTP and PICP at baseline (Spearman's rank-order correlation coefficient rs = 0.62). Both ICTP and PICP showed statistically significant correlations to the other markers of bone metabolism (except sCa and uCa) as well as to the number of bone metastases on bone scans. Reduction in ICTP correlated significantly with the treatment response at three months (rs = - 0.57). while PICP showed a borderline negative correlation to therapy response (rs = - 0.37). Of all the biochemical parameters studied the changes in ICTP showed the best correlation with the treatment response. PICP and ICTP changes in patients with progressive disease differed significantly from those in patients with responding and stable metastases, whereas no difference was found between responders and stable patients.

Abstract: Collagen plays a specific role in the maintenance of vascular integrity and in the thrombosis and scar formation processes. Therefore we found it interesting to study the changes in interstitial collagen metabolism during acute myocardial infarction treated with thrombolytic agents. Changes in collagen synthesis were evaluated by obtaining assays of the serum concentrations of the carboxyterminal propeptide of type I procollagen. Except fibrin plasmin is capable of degrading extracellular matrix components including collagen, and this capability was evaluated by monitoring the serum concentrations of the aminoterminal propeptide of type III procollagen. Twenty-four patients with suspected acute myocardial infarction and indications for thrombolytic therapy were randomized to receive either streptokinase (n = 11) or tissue plasminogen activator (n = 13). The patient groups were identical in their clinical characteristics. Serum levels of the aminoterminal propeptide of type III collagen increased rapidly on infusion of the thrombolytic agents, with the maximal mean increases of 44% and 16% in the streptokinase and TPA-treated groups, respectively. Levels of the carboxyterminal propeptide of type I collagen did not change during the thrombolytic therapy. A transient decrease occurred in the type I propeptide concentration at postinfarction day 2, and this decrease was followed by a secondary increase at days 4 to 6 in both patient groups studied. We conclude that thrombolytic agents stimulate the breakdown of interstitial collagen and that the collagen-degrading activity of TPA is lower than that of streptokinase. This factor may contribute to the relatively higher rethrombosis rate seen after TPA, because exposed collagen in the affected vascular wall stimulates thrombosis formation. On the other hand, increased collagen degradation followed by inhibition of collagen synthesis in the infarcted myocardium might increase the risk for cardiac rupture, especially after streptokinase treatment.

Abstract: Three biochemical markers of collagen metabolism were measured in 39 osteosarcoma patients. The pretreatment values did not predict outcome, and the markers showed no consistent change upon development of metastases. Both the age of the patients and the multimodality therapy affected the collagen metabolites. These findings emphasise the need for cautious interpretation of tumour-associated markers.

Abstract: AIMS: The purpose of this pilot study was to test whether the rate of collagen synthesis is measurable in the aqueous humour samples in reoperated and previously unoperated eyes. METHODS: The material consisted of 28 eyes of 27 patients, aged 5 to 82 years, in whom aqueous humour samples were obtained during eye surgery. Fifteen patients had no history of previous eye surgery (control group) while 12 patients were re-operated (study group). The carboxyterminal propeptide of type I procollagen (PICP) and the aminoterminal propeptide of type III procollagen (PIIINP) were measured by specific immunoassays in the aqueous humour samples. RESULTS: The mean concentration of PIIINP in the study group (8.4 (SD 12.5) micrograms/l) was statistically significantly larger than that of the control group (0.4 (0.4) micrograms/l) (p < 0.0037). The respective values for PICP were 98.8 (SD 177.7) micrograms/l in the study group and 0.7 (SD 2.8) micrograms/l in the control group (p < 0.0005). The eyes in the study group which were re-operated within 1 year showed values increased 20-fold compared with the eyes in the control group and those eyes in the study group which had had their previous operation more than a year ago. In three eyes aqueous humour samples were also obtained from the encapsulated Molteno bleb and showed values increased 12-fold compared with those from the anterior chamber. CONCLUSIONS: PICP and PIIINP immunoassays are suitable for measuring the rate of collagen synthesis in the aqueous humour and may be useful in studies on pharmacological modulation of wound healing in glaucoma surgery.

Abstract: We evaluated serum and urinary markers of bone turnover in 14 children with asthma during inhaled budesonide and nedocromil treatments. Both the markers of formation (serum carboxy- and amino-terminal propeptides of type I procollagen and serum osteocalcin) and the markers of degradation (serum carboxy-terminal telopeptide of type I collagen and urinary pyridinium cross-links) decreased (p < 0.05) during budesonide treatment for 6 months. During inhaled nedocromil treatment (for the following 6 months), the markers returned to the normal levels. These transient decreases in the markers of both formation and degradation of bone suggest that inhaled budesonide may slightly decrease the bone turnover rate. However, normal "coupling" between formation and degradation seemed to operate, e.g. a change in one resulted in a corresponding change in the other, so that net bone loss did not necessarily occur.

Abstract: We describe a developmental dentin disorder distinct from dentin defects characterized thus far. The proband was a 9-year-old boy who was the only family member known to be affected in five generations. The dental defect was not associated with any general disease or developmental disorder. The teeth appeared normal with the exception of the pink hue seen in some primary teeth. Radiographs showed pathological resorption of primary teeth and abnormally shaped pulp chambers and denticles in permanent teeth. Root canals were wide in developing teeth, but appeared thin in erupted teeth. Histological examination of two primary molars revealed canal-like defects in dentin. In the crown, the canals appeared as clusters, which alternated with columns of normal tubular dentin, and in the virtually atubular root dentin they were haphazardly distributed. Scanning electron microscopic examination confirmed the distribution pattern of the canals. In transmission electron microscopy, the defects were found to contain symmetrically banded, segmental collagenous structures. The canal contents immunostained with antibodies to the N-terminal propeptide of type I procollagen, suggesting retention of the propeptide extension in type I collagen. Whereas type III collagen reactivity was barely detectable in the canal region, staining for type V collagen and the non-fibril-forming type VI collagen was strong. The findings imply that the pathogenesis of the defect could be related to a local failure of odontoblasts to produce normal dentin matrix.

Abstract: We have developed quantitative immunoassays for the intact, trimeric amino-terminal propeptide of human type I procollagen (PINP) and its Col1 domain. Intact PINP was isolated from the pleural fluids of cancer patients by a combination of ion-exchange, gel-filtration, and reversed-phase chromatographies. The amino-terminal Col1 domain of PINP was isolated after bacterial collagenase treatment of the heat-denatured trimeric propeptide. For the intact PINP assay we used a polyclonal antibody with only 1.2% cross-reaction with the monomeric Col1 domain. In human serum, this assay detects only one peak of PINP antigenicity that has the size of known intact PINP. Under similar conditions, an assay for the Coll domain of PINP recognized two circulating antigens. The biological relevance was further verified in wound fluid. Interassay and intraassay CVs were 3.1-9.3% for values within the reference intervals (mean +/- 2SD) for intact PINP in serum, which were 19-84 microg/L for women and 20-76 microg/L for men.

Abstract: The use of inhaled corticosteroids in bronchial asthma has widened, but there is a lack of data on their effect on bone collagen synthesis and degradation. This paper reports the effect of three dose levels (200, 1000 and 2000 micrograms day-1, 3 weeks each) of inhaled beclomethasone on specific characteristics of bone collagen metabolism in seven postmenopausal women with new asthma without any previous corticosteroid therapy. Serum aminoterminal (PINP) propeptide of human type I procollagen was seen, after an initial increase, to decrease significantly (from 42.2 to 35.5 micrograms l-1, P = 0.001) with the higher doses of inhaled beclomethasone, but no statistical change was found in the carboxyterminal propeptide (PICP) or type I collagen crosslinked telopeptide (ICTP). This data shows that type I collagen synthesis may be disturbed when using high-dose inhaled corticosteroids. However, further studies are needed to assess the effects of inhaled beclomethasone on the ability of the osteoblasts to form bone matrix, and on the density of bone during a longer treatment period with inhaled corticosteroids.

Abstract: The minimum dosage of transcutaneous hormone replacement therapy (HRT) able to exert protective effects on postmenopausal bone mass, especially in older women, is uncertain. This study investigates the effects of transcutaneous HRT at two different doses of oestradiol [Estraderm 25 and 50 (E25, E50)] over 2 years in 44 postmenopausal women younger than 67 years and 27 of 67 years and older. Circulating biochemical markers of bone and connective tissue turnover, collagen type I (intact PINP, PICP) and type III (PIIINP) propeptides and type I telopeptide (ICTP), osteocalcin (OC) and alkaline phosphatase (AP) were measured. The responses of the biochemical markers in all the groups were very similar, and most of the observed changes occurred within the first year of treatment. E25 had an effect on the bone markers similar to that of E50, and there was little difference in response according to the patient's age. PINP fell markedly after 1 year in all groups to about half the pretreatment level, with a smaller drop in the second year. PICP responded more variably, and mean values were little changed. There was a slight fall at the higher dose in the younger women, and also in the older women (whose baseline level was higher) on the lower dose. The correlation between PINP and PICP was 0.52 at pretreatment and 0.84 after 2 years of treatment. PIIINP showed no changes. OC and AP both fell in all groups by the first year, but less markedly than PINP. Their response was slightly less pronounced in the older women. ICTP fell marginally in the younger women, and only after 2 years, regardless of dose. Postmenopausal serum oestradiol levels increased after HRT and were associated with decreased binding globulin (SHBG) levels in all groups. After E25 patch application individual serum oestradiol levels were variable and peaked between 13 and 36 h with a median value of 83.8 pmol L-1. Our data suggest that low-dose transcutaneous HRT restores circulating oestradiol levels in postmenopausal osteopenic women of all ages as effectively as conventional-dose HRT and is associated with decreased circulating markers of bone and connective tissue turnover.

Abstract: BACKGROUND & AIMS: Cross-linking participates in the increased stability of collagen towards proteolytic degradation. Liver collagen cross-linking by pyridinoline, from the lysyl oxidase pathway, and by pentosidine, issued from glycation, was investigated to determine their respective contribution to collagen stabilization in patients with an irreversible liver fibrosis caused by the parasitic granulomatous disease alveolar echinococcosis. METHODS: Liver pyridinoline and pentosidine were analyzed by high-performance liquid chromatography, and urinary pyridinoline was analyzed by immunoassay. Cross-linked type I collagen was localized by immunohistochemistry with an antibody against the C-terminal part of the molecule, involved in pyridinoline formation, that was measured in serum by radioimmunoassay. RESULTS: In contrast to pyridinoline, pentosidine decreased in fibrotic lesions. Cross-linked I collagen was located predominantly in collagen bundles in the periparasitic granuloma. Serum pentosidine and urinary pyridinoline levels did not differ significantly from controls, but the serum concentration of the C-terminal telopeptide of type I collagen increased significantly. CONCLUSIONS: Lysyl oxidase-mediated cross-linking is the major process contributing to the stabilization of collagen in granulomatous fibrosis, and glycation is not significantly involved in it. The changes induced by alveolar echinococcosis in liver collagen metabolism are associated with an increase in serum C-telopeptide of type I collagen.

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Abstract: In our previous double-blind trial, we reported that clodronate reduced the incidence of bone lesions, fractures, pain and hypercalcaemia in multiple myeloma. Recently, it has been assumed that the antiresorptive effect of bisphosphonates on the osteoclasts is mediated through the osteoblasts. We therefore determined, in 244 patients of the same trial, serum assays of aminoterminal propeptide of type I procollagen (PINP) and type I collagen degradation product (ICTP). PINP is an early synthesis product of proliferating osteoblasts, in comparison to the alkaline phosphatase (AP) which is secreted by differentiated osteoblasts during the maturation phase of collagen. ICTP circulates in serum when old bone is resorbed. Our results indicate that after 25 months, the PINP levels decreased in the clodronate group (from 68.9 +/- 4.4 micrograms/l to 37.2 +/- 3.5 micrograms/l; P < 0.001) but not in the control group (from 61.5 +/- 3.2 micrograms/l to 69.3 +/- 7.5 micrograms/l; P < NS). The fall in the ICTP levels was markedly steeper in the patients receiving clodronate (from 8.38 +/- 0.80 micrograms/l to 4.58 +/- 0.32 micrograms/l; P < 0.01) than placebo (from 7.84 +/- 0.53 micrograms/l to 6.45 +/- 0.95 micrograms/l; P = NS). A significant difference between the study groups was seen at 4 months in the PINP, at 7 months in the ICTP and at 13 months in the AP levels, suggesting that clodronate affected through the proliferating osteoblasts, the osteoclasts, and through the osteoclasts, the differentiated osteoblasts. High baseline ICTP, PINP and AP levels indicated a poor prognosis. The decrease of the markers by clodronate was more marked in survivors than in non-survivors.

Abstract: The effect of ovarian carcinoma on the circulating concentration of the cross-linked carboxyterminal telopeptide of type I collagen (ICTP) was studied in 17 patients before and during treatment. ICTP is a complex peptide released from type I collagen, which is the main organic component of bone matrix but also induced in ovarian carcinoma tissue and the peritoneal cavity by the tumour. CA 125 and the aminoterminal propeptide of type III procollagen (PIIINP) were also measured. The preoperative CA 125, PIIINP and ICTP concentrations were pathological in 16 (94%), nine (53%) and seven (41%) patients, respectively. There was a positive correlation between the PIIINP and ICTP levels in serum. After surgery and one course of cytotoxic chemotherapy, the median CA 125 concentration decreased whereas those of PIIINP and ICTP increased. During monitoring of the treatment response, patients with regressive disease showed lower CA 125 concentrations than those with stable or progressive disease. For PIIINP and ICTP there was no clear difference between these response categories, but altogether, simultaneously increasing serum CA 125, PIIINP and ICTP concentrations were consistently associated with poor prognosis.

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Abstract: Psoriasis is an inflammatory skin disease of unknown a etiology which also involves changes in dermal elements. Previous in vitro studies have shown an increased collagen synthesis rate in cultured fibroblasts. In this study collagen synthesis was studied in vivo in the uninvolved skin of psoriatic patients using a newly developed method in which collagen propeptides were measured in suction blister fluid. Both type I and type III collagen synthesis rates, as measured in terms of the carboxyterminal propeptide of type I procollagen (PICP) and the aminoterminal propeptide of type III procollagen (PIIINP), were increased about two-fold in uninvolved psoriatic skin as compared with controls, the mean level of PICP being 870 and 457 micrograms, respectively (P < 0.001), and of PIIINP being 294 and 124 micrograms, respectively (P < 0.01). The increased collagen synthesis rate was also confirmed by in situ hybridization using specific probes. Collagen mRNAs were found to be particularly abundant in psoriatic patients, who also demonstrated a high collagen synthesis rate when assayed by measuring collagen propeptides. The increased rate of collagen synthesis in the uninvolved psoriatic skin seemed not to be related to the severity of the disease or to various treatments such as UVB, PUVA, retinoids or cytostatic drugs, but seemed more likely to be due to the psoriasis itself. Interestingly, skin thickness was not increased in the patients with psoriasis, even though collagen synthesis was markedly elevated, perhaps suggesting that in psoriasis the turnover rate of collagen is enhanced.

Abstract: BACKGROUND. Type I collagen is a major constituent of the interstitial connective tissue. Although ovarian carcinoma is known to induce the expression of type I collagen in the peritoneal cavity, the distribution and metabolic activity of this collagen in ovarian tumor tissue are not known. METHODS. The distributions and staining intensities of different molecular forms of type I collagen in ovarian neoplasms were studied immunohistochemically with antibodies to the aminoterminal propeptide of type I procollagen (PINP) and the cross-linked carboxyterminal telopeptide of type I collagen (ICTP), reflecting the presence of newly synthesized and old, cross-linked type I collagen, respectively. RESULTS. A regular pattern of moderately staining, relatively uniform fibers was observed in the stroma of benign serous and mucinous cystadenomas, indicating limited participation in tumor growth. The staining was accentuated subepithelially in borderline epithelial neoplasms and in well differentiated cystadenocarcinomas, suggesting induction of the stromal collagen synthesis by the tumor cells. Fewer degraded collagen fibers were found in moderately differentiated carcinomas, most likely because of enzymatic degradation of the stroma surrounding the neoplasms during tumor spread. Strongly staining, irregular collagen fibers occurred closely around islets of tumor cells in undifferentiated malignant neoplasms and in metastases of ovarian carcinomas; also, intracellular staining was present in part of the malignant cells. In most cases, the staining reactions obtained with the two different antibodies were similar, probably indicating rapid processing of the newly synthesized type I collagen (indicated by PINP) to a maturely cross-linked form (indicated by ICTP). CONCLUSIONS. Synthetic and degradative processes are typical of the collagenous matrix in malignant ovarian tumors. Aberrant expression of type I collagen may occur in anaplastic ovarian carcinomas.

Abstract: Although osteosclerotic bone metastases are characteristic of prostate cancer, mixed metastases with a lytic component are not uncommon. Type I collagen is synthesised by osteoblasts and accounts for about 90% of the organic matrix of bone. We have used new specific immunoassays for PICP (carboxy-terminal propeptide of type I procollagen) and ICTP (cross-linked carboxy-terminal telopeptide of type I collagen) which allow simultaneous assessment of the synthesis and degradation of type I collagen respectively. Forty patients with bone metastases due to prostate cancer at the time of diagnosis were investigated with these methods. Twenty-three of them had sclerotic (S) and 17 had mixed metastases with sclerotic and lytic components (S + L) as assessed by radiographs. The concentrations of PICP and ICTP in serum as well as the activity of alkaline phosphatase (AP) were increased in all patients of the S + L group, who had more aggressive bone disease and a shorter survival than the S group (P < 0.017). The ICTP level was above the reference range in half of the patients in the S group, whereas the PICP and AP levels were elevated in 35%. Of the bone markers, only ICTP was of prognostic significance (P < .05). We conclude that ICTP and PICP give information about the type and activity of the skeletal metastases. In addition, ICTP predicts prognosis.

Abstract: OBJECTIVE--To assess the potential of markers of collagen metabolism to reflect disease processes in rheumatoid arthritis (RA). METHODS--Serum (S) and synovial fluid (SF) from 59 patients with RA, and a knee joint effusion and serum from 90 control subjects were studied with radioimmunoassays for the aminoterminal propeptides of type I and type III procollagens (PINP and PIIINP, respectively). The breakdown of type I collagen was quantified with a radioimmunoassay for the cross linked carboxyterminal telopeptide of type I collagen (ICTP). RESULTS--About 50% of the patients had increased S-ICTP and S-PIIINP values, whereas S-PINP was increased in only 20% of the patients. The mean SF:S ratios of these markers varied between 4 (for ICTP) and 340 (for PIIINP), indicating that markers of collagen metabolism are formed locally and then released into the circulation. SF-PINP and SF-PIIINP correlated with each other (rs = 0.86, p < 0.001) and with SF-ICTP (rs = 0.69, p < 0.001, and rs = 0.65, p < 0.001, respectively). SF-ICTP was clearly related to radiographic findings in the corresponding knee joint, patients with gross bone deformation having the greatest SF-ICTP concentrations. S-ICTP and S-PIIINP also correlated with conventional markers of disease activity, such as C reactive protein and joint swelling score. CONCLUSION--Markers of collagen metabolism both in serum and synovial fluid can be measured to provide an assessment of disease process in patients with RA. ICTP and PIIINP are the most informative.

Abstract: OBJECTIVE: We investigated the clinical utility of macrophage colony-stimulating factor 1 versus CA 125 and the aminoterminal propeptide of type III procollagen in endometrial carcinoma. STUDY DESIGN: Serum levels of the three substances were measured in 159 patients with untreated endometrial adenocarcinoma and in 24 patients treated with cytotoxic chemotherapy for recurrent endometrial adenocarcinoma. RESULTS: Initial concentrations of colony-stimulating factor 1, CA 125, and the aminoterminal peptide of type III procollagen were above the normal range in 73%, 11%, and 27%, respectively, of the patients. Colony-stimulating factor 1 levels correlated with those of the aminoterminal peptide of type III procollagen (r = 0.3, p = 0.002) and CA 125 (r = 0.20, p = 0.036) in the total group and with those of the aminoterminal peptide of type III procollagen in stage I and II patients (r = 0.3, p = 0.0023). Colony-stimulating factor 1 levels correlated significantly with tumor grade, whereas those of CA 125 and the aminoterminal peptide of type III procollagen correlated more closely with clinical stage. Mean colony-stimulating factor 1 levels (9.6 vs 7.7 ng/ml, p = 0.04) and the frequency of elevated CA 125 levels (31% vs 8%, p = 0.048) were higher in patients with poor prognosis than in those with good prognosis. Colony-stimulating factor 1, the aminoterminal peptide of type III procollagen, and CA 125 levels were useful in monitoring clinical behavior of the disease in 88%, 79%, and 63% of the cases, respectively. Levels of all three markers rose with disease progression, whereas colony-stimulating factor 1 and the aminoterminal peptide of type III procollagen fell with clinical responses to therapy. CONCLUSION: Elevated serum colony-stimulating factor 1 levels were the most accurate indicator of the presence and activity (progression, stabilization, or regression) of primary or recurrent disease. Accuracy was not further enhanced by measurement of CA 125 or the aminoterminal peptide of type III procollagen levels.

Abstract: Although portoenterostomy has greatly improved the prognosis of extrahepatic biliary atresia (EHBA), 10-20% of patients still die before 5 y of age, and the only treatment option is liver transplantation (LT). To investigate whether these patients may be identified at an early stage, when the changes of successful LT are optimal, we have measured serum concentrations of hyaluronic acid (HA), the amino-terminal propeptide of type III procollagen (PIIINP) and the carboxy-terminal and amino-terminal propeptides of type I procollagen (PICP, PINP) in 24 selected patients with EHBA, both before portoenterostomy and then every 6 mo until death (n = 10, age at death = 7-20 mo), LT (n = 6, age at LT = 1.1-4.8 y) or 5 y of age (n = 8). Raised serum HA above 200 micrograms/L before portoenterostomy identified those patients who would die or require LT in the first 5 y of life with a positive predictive value of 88%; after portoenterostomy, longitudinal changes in HA reflected clinical status in each patient. None of the other three markers was of prognostic value, and only PIIINP showed any relationship with clinical status, and then only up to 1.5 y. Interestingly, PINP (but not PICP) tended to be low in all patients before portoenterostomy and may reflect impaired bone collagen metabolism during early skeletal changes in EHBA. This study therefore suggests that measurement of serum HA may be a useful complementary test in EHBA, particularly in identifying, at an early stage, those patients who should be considered for LT.

Abstract: Liberation of the carboxy-terminal propeptide of type I procollagen (PICP) and the amino-terminal propeptide of type III procollagen (PIIINP) into body fluids reflects synthesis of the respective collagen types. Here, we followed PICP and PIIINP in serum with specific radioimmunoassays after hip surgery. Preoperative median of S-PICP was 112 micrograms/liter (range 87 to 154, n = 9), the 1-day median being 58 micrograms/liter (33 to 79). The corresponding medians for S-PIIINP were 4.4 micrograms/liter (3.5 to 7.0) and 3.3 micrograms/liter (2.0 to 3.5). The medians reached their maximums 14 days after surgery, 172 micrograms/liter (122 to 440) for S-PICP and 12.4 micrograms/liter (8.0 to 15.4) for S-PIIINP, after which the preoperative values were slowly approached over several months. Comparable results were found in a greater sample (n = 50). Our results indicate that the synthesis of structural collagen is inhibited immediately after surgery, but the inhibition is soon overcome by active collagen synthesis at the site of trauma. Collagen metabolism remains activated for several months after surgery.

Abstract: In this study, we investigated the relation between calcium kinetic indices of bone remodeling (resorption rate, r; and formation rate, m, respectively) and two serum markers of type I collagen turnover: the pyridinoline cross-linked carboxyterminal telopeptide domains of type I collagen (S-ICTP a marker of bone matrix degradation) and the carboxyterminal propeptide of human type I procollagen (S-PICP, a marker of bone matrix formation). We studied three groups: (i) healthy controls (n = 19), (ii) a mixed group of high and low-turnover bone diseases without mineralization defects (myxedema, thyrotoxicosis and primary hyperparathyroidism n = 38), and (iii) osteoporosis (n = 52). In healthy controls, a significant regression of S-PICP on m was obtained (R = 0.53, SEE/Y = 0.44, P < 0.02). Significant regressions were also demonstrable in high- and low-turnover bone disease (R = 0.50, P < 0.001), SEE/Y = 61%) and osteoporosis (R = 0.49, P < 0.001, SEE/Y = 50%). In controls the regression coefficient for the regression of S-ICTP on r was 0.19 (NS), in high and low turnover bone disease 0.66, (SEE/Y = 59%, P < 0.001) and in the osteoporotic group 0.40 (SEE/Y = 61%, P < 0.01). We conclude that S-PICP and S-ICTP reflect whole skeletal bone formation and resorption rates in a variety of metabolic bone diseases including osteoporosis.

Abstract: Circulating osteocalcin, a marker of bone formation, is under strong genetic influence, and this effect is related to the genetic influence on bone density. To examine genetic influences on bone turnover further, other markers of bone formation (serum carboxyterminal propeptide of type I procollagen, PICP), bone resorption (serum pyridinoline cross-linked carboxyterminal telopeptide of type I collagen, ICTP), and nonosseous connective tissue synthesis (serum aminoterminal propeptide of type III procollagen, PIIINP) were studied in 82 female twin pairs: 42 monozygotic (MZ) and 40 dizygotic (DZ) twin pairs (mean age, MZ; 48.4 yr; DZ; 45.6 yr). The intraclass correlation coefficients of MZ twin pairs, rMZ, for serum PICP (0.78) and serum ICTP (0.68) were significantly greater than the corresponding rDZ (0.31 and 0.36, respectively), but a genetic effect on serum PIIINP was not demonstrable. Within DZ twin pair differences in serum PICP predicted differences in lumbar spine bone density (r = -0.37); higher serum PICP levels indicating the twin with the lower lumbar spine bone density. Also within pair differences in serum ICTP and PICP predicted differences in bone density at the lumbar spine independent of serum osteocalcin. These data indicate that both synthesis and degradation of type I collagen are genetically determined and that this phenomenon is related to the genetic regulation of bone density.

Abstract: This study was undertaken to determine the fate of circulating NH2-terminal propeptide of type I procollagen (PINP) in rats. Radiolabeled PINP showed a biphasic serum decay curve after intravenous injection. 79% of the material disappeared from the blood during the initial alpha-phase (t1/2 alpha = 0.6 min), while the remaining 21% was eliminated with a t1/2 beta of 3.3 min. The major site of uptake was the liver, 78, 1, and 21% of its radioactivity being recovered in isolated liver endothelial cells (LEC), Kupffer cells, and parenchymal cells, respectively. In LEC, fluorescently labeled PINP accumulated in small (0.1 microns) peripheral and larger (> 0.1 microns) perinuclear vesicles within 10 min at 37 degrees C after a binding pulse at 4 degrees C. These grew in size with increasing chasing time, reaching a maximum diameter of 1 microns or more after 30 min, and taking the shape of rings that were stained only along their periphery. At chase intervals exceeding 30 min, the size of the vesicles decreased, and after 60 min the stain appeared in smaller, densely stained perinuclearly located vesicles. Degradation of 125I-PINP to free smaller fragments and 125I- was significant after 30 min. Only formaldehyde-treated albumin, acetylated LDL, polyinosinic acid and NH2-terminal propeptide of type III procollagen (PIIINP) competed with PINP for uptake. These findings indicate that clearance of PINP and PIIINP, which are normal waste products generated in large quantities, is a physiological function of the scavenger receptor in LEC.

Abstract: The value of serum procollagen peptide (PICP) as a non-invasive index of bone formation was studied in 18 patients established on haemodialysis. There was a significant correlation between PICP and serum alkaline phosphatase activity (ALP; r = 0.55, P < 0.05), and between PICP and osteocalcin (r = 0.53, P < 0.05). PICP also correlated significantly with histomorphometric indices of bone formation, particularly bone formation rates (BFR) as estimated by the tetracycline double-labelled technique (r = 0.74, P < 0.01), but not with those of bone resorption. There was a similar relationship between BFR and ALP. From the regression analyses, a normal BFR was associated with normal PICP values despite the absence of renal function, suggesting that the impact of renal function on serum concentrations of PICP may not be large. Seven patients had histochemical evidence for significant aluminum overload. In these patients the expected suppression in biochemical and histological indices of bone formation was associated with inappropriately raised PICP concentrations. The mechanism of this discrepancy is not clear, but caution is advocated in the interpretation of PICP in the presence of significant aluminium overload. Our findings otherwise suggest that PICP may be a useful non-invasive index of bone formation in patients on haemodialysis.

Abstract: A study was conducted to explore the effects of different sources of dietary carbohydrate on the altered metabolism of basement membrane proteins (type IV collagen and laminin) that occurs in the streptozotocin-diabetic rat. The substitution of barley for sucrose reduced the elevated plasma concentration of 7-S collagen and of laminin P2, peptide fragments that retain antigenicity of the parent proteins. Similar effects were noted in the liver and kidney content of these peptides. It is suggested that the beneficial property of barley is due to its high content of chromium, as has been demonstrated in the study of glucose metabolism.

Abstract: We describe a family with an apparently autosomal-dominant trait that caused extremely high circulating concentrations of the carboxyl-terminal propeptide of type I procollagen (PICP). All family members examined had normal values for other biochemical markers of bone formation and degradation and no related clinical abnormalities. Furthermore, their serum concentrations of the amino-terminal propeptide of type I procollagen (PINP) were normal. Although PINP and PICP are released from the same precursor molecule, PINP is cleared from the circulation via the scavenger receptor in liver endothelial cells, whereas PICP is cleared via the mannose receptor of these cells. We thus hypothesize that the clearance of circulating PICP is compromised in the affected subjects of this family, the result of either a defective mannose receptor function or an abnormal molecular structure of their PICP.

Abstract: We introduce a simple serum test to predict which patients will have bone problems after liver transplantation. The crosslinked part of collagen I (s-ICTP) was measured in 21 patients with primary biliary cirrhosis before transplantation. Those with postoperative fractures had increased pretransplant values of s-ICTP compared with those without fractures.

Abstract: The new assay of cross-linked carboxyterminal telopeptide of type I collagen (ICTP), a serum marker for bone collagen degradation, was evaluated in serial measurements of 66 patients with early RA during a 3-yr prospective study. Initially 51% of RA patients had elevated levels of serum ICTP compared to healthy controls. During the subsequent months after starting anti-rheumatic treatment, the mean ICTP levels decreased in parallel with the clinical and laboratory variables measuring disease activity. Despite marked clinical improvement with anti-rheumatic treatment, a steady increase in radiological progression of joints was observed. Throughout the follow-up serum ICTP levels correlated with inflammatory parameters and from the first year on with the radiological changes assessed annually. However, initial serum ICTP levels correlated better than the other variables of disease activity with the subsequent erosive progression of joints indicating that measurement of serum ICTP may serve as one of the prognostic markers for joint damage in early RA.

Abstract: We used transmission immunoelectron microscopy and polyclonal antibodies to study the reactivities of Types III and VI collagen in dentin of normal human permanent and primary teeth and in primary teeth from five patients with dentinogenesis imperfecta (DI) associated with osteogenesis imperfecta and occurring as a single trait. In the normal permanent tooth, reactivity of Type III collagen was occasional and, where present, peritubular. Staining of normal primary teeth was less occasional but still rare, whereas the abnormal dentin stained more uniformly. Atypical, non-striated fibrillar structures that also showed Type III collagen reactivity were observed in dentin of two of the three patients with DI as a single trait. Later, these two patients proved to be first cousins. Unlike antibodies to the N-terminal pro-peptide of Type I pro-collagen, antibodies to the C-terminal telopeptide of Type I collagen, used for comparison stained the affected dentin homogeneously. Reactivity of Type VI collagen, not detected in normal teeth, was seen in the dentin of all abnormal teeth, in association with non-fibrillar delicate material. This study also shows that although readily detectable in dentin affected by DI, Type III collagen is a minor constituent of normal human dentin matrix.

Abstract: OBJECTIVE. To evaluate the serum concentration of the mature, crosslinked carboxy terminal telopeptide of type I collagen (ICTP) in patients with early onset rheumatoid arthritis (RA) and to assess its correlation with clinically relevant features of the disease. METHODS. The serum ICTP concentration was measured in 99 adult patients with RA with a duration of disease less than one year. The main clinical outcome measures were joint count, modified Lansbury index, erythrocyte sedimentation rate (ESR), C-reactive protein (CRP), Larsen index and number of erosions in peripheral joints. RESULTS. There was a strong positive correlation (p < 0.001) between the serum ICTP concentration and the Lansbury joint index and a weaker but still significant correlation (p < 0.01) between the serum ICTP concentration and the number of eroded hand and foot joints. CONCLUSION. Serum ICTP may have value as a marker of tissue destruction in patients with early RA.

Abstract: The effects of systemic glucocorticoid and isotretinoin treatments on type I and type III collagen synthesis in intact skin were investigated by measuring the carboxyterminal and aminoterminal propeptides of type I procollagen, and the aminoterminal propeptide of type III procollagen, in suction blister fluid (SBF), in a study of 27 patients. All three parameters were significantly lower in the SBF of glucocorticoid-treated patients than in controls or patients undergoing treatment with isotretinoin, whereas the latter two groups did not differ significantly from each other. During glucocorticoid treatment, the concentrations of the procollagen propeptides were only about 20% of the corresponding control values, indicating that systemic therapy with prednisone at a dose of 0.48 mg/kg per day almost totally abolishes collagen synthesis in the skin. These results indicate that systemic glucocorticoid treatment suppresses the synthesis of both type I and type III collagen in the dermis, and suggest that many side-effects of these drugs, such as atrophy of the skin, are due to this inhibition. Systemic isotretinoin treatment did not stimulate skin collagen synthesis. Thus, its regenerative effect on connective tissue may be mediated by mechanisms other than direct stimulation of collagen synthesis.

Abstract: Chronic immobilization could markedly affect calcium and bone metabolism in elderly people. To investigate this, and to test the theory of 'type II' osteoporosis in bedridden elderly patients with low vitamin D status, 55 such subjects were examined. Serum concentrations of ionized calcium (Ca++), intact parathyrin (PTH) and two novel markers of bone collagen formation (carboxyterminal propeptide of type I procollagen; PICP) and resorption (carboxyterminal crosslinked telopeptide of type I collagen; ICTP) were measured. The effects on these parameters after 40 weeks of supplementation with vitamin D (1000 IU d-1) and/or calcium (1 g d-1) were subsequently prospectively evaluated. Despite low (mean 11.6 nmoll-1) serum 25-hydroxyvitamin D levels (25-OHD), those of 1,25-dihydroxy-vitamin D (1,25-(OH)2D) were mostly normal. Neither correlated with Ca++ or PTH. PTH correlated negatively not only with Ca++ (r = -0.328, P < 0.05) but also with ICTP (r = -0.306, P < 0.05). Mean PICP was normal but ICTP was elevated and tended to correlate positively with Ca++ (r = 0.268, P = 0.06). Vitamin D supplementation did not change PICP or ICTP considerably, despite slightly increased 1,25-(OH)2D and slightly decreased PTH. Ca++ values were normal and remained stable. In conclusion, Ca++ and PTH are poor indicators of vitamin D status in chronically immobilized elderly subjects. Furthermore, the results suggest that the increased bone resorption is not due to 'type II' secondary hyperparathyroidism; rather the resorption is primarily increased. Correction of vitamin D deficiency does not seem to benefit ageing bones unless adequate mechanical loading is provided.

Abstract: We investigated the effect of the menopause and postmenopausal hormone replacement therapy (HRT) on the serum concentration of carboxy-terminal pyridinoline cross-linked telopeptide of type I collagen (ICTP), a potential new biochemical marker of bone resorption. A group of 44 healthy postmenopausal women, aged 45-54 years, had about 19% higher serum ICTP than did a group of 42 healthy premenopausal women aged 35-50 years (3.6 +/- 0.8 micrograms/l v 3.0 +/- 0.7 micrograms/l (mean +/- SD); p < 0.01), although there was a large overlap in the values. The 44 postmenopausal women also participated in a longitudinal clinical study, in which 20 received HRT and 24 received a placebo. Compared with the placebo group, those who received HRT had a significant (p < 0.05) decrease in ICTP of about 12% at the end of 1 year of treatment, but again there was considerable overlap in the values. The menopause- and HRT-induced changes in ICTP were less than those seen in serum osteocalcin, serum total alkaline phosphatase, and fasting urinary excretion of hydroxyproline, calcium, pyridinoline and deoxypyridinoline. We conclude that the menopause increases and HRT decreases ICTP, although these changes are less pronounced than those seen in other biochemical markers of bone turnover.

Abstract: The effect of aging, sex, skin location and a short whole body ultraviolet B (UVB) phototherapy on type I and type III skin collagen synthesis were studied by measuring carboxy- and aminoterminal propeptides of type I and aminoterminal propeptide of type III procollagens (PICP, PINP and PIIINP, respectively) in suction blister fluid (SBF). The concentrations of PICP and PIIINP correlated negatively with age in the material of 30 men and 27 women (age range 23-86 years, mean age 51 years). As the subjects were divided into 3 age groups (23-34, 35-54 and 55-86 years) and the mean concentrations of the procollagens were compared between the groups, the youngest age groups in men and in the whole material had significantly higher concentrations than the two older age groups. These results confirm the previous results, which have shown by other methods that skin collagen synthesis decreases during aging. SBF was also obtained from 4 different body regions (back, arm, abdomen and leg) of 5 young male subjects and from 2 different body regions (abdomen and thigh) of 9 older subjects. Wide individual range in the concentrations of PICP, PINP and PIIINP was noticed but, in general, none of the body regions seemed to have more excessive collagen synthesis than the other ones. SBF was further collected from abdominal skin of 8 young male dermatological patients receiving UVB phototherapy. When the concentrations of PICP and PIIINP in SBF obtained before the treatment, after the treatment and from the covered control area were compared no statistical differences were noted, suggesting that short UVB treatment has no effect on skin collagen synthesis in vivo.

Abstract: Progressive ovarian carcinoma induces the synthesis of type I and type III procollagens both in the tumour tissue and in the peritoneal cavity. We studied the processing of these proteins by determining the different antigen forms related to their propeptide parts by gel filtration and subsequent immunological assays. Samples of ovarian cyst fluid and peritoneal ascitic fluid were obtained from patients with benign and malignant ovarian tumours. In both benign and malignant ovarian cysts, the predominant procollagen antigens were the free propeptides, with few or no larger components, indicating efficient processing of types I and III procollagens in the tumour tissue. In ascitic fluid the processing was more variable. The aminoterminus of type III procollagen was partially unprocessed in all samples studied, whereas that of type I procollagen was nearly always completely processed. There was a clear difference between malignant and benign tumours in the processing of the carboxyterminus of type I procollagen: a significant part of the carboxyterminal propeptide antigen was invariably associated with a collagenous domain in malignant tumours, whereas in benign tumours the free propeptide predominanted. The results indicate that interstitial procollagens are effectively processed in the tumour tissue during the fibroproliferative reaction typical of malignant ovarian tumours, whereas the processing of the procollagens released into peritoneal ascitic fluid is incomplete.

Abstract: OBJECTIVE: Simultaneous monitoring of total collagen synthesis as well as synthesis in intact skin and in the wound to verify the higher priority of wound healing after surgery. SUMMARY BACKGROUND DATA: Synthesis of acute phase proteins is stimulated by surgical trauma. At the same time, production of albumin is inhibited and there is a net catabolism of skeletal muscle proteins. Similarly, the authors have found a transient inhibition of total collagen synthesis after surgery. The authors hypothesized a lower priority in synthesis of structural and peripheral collagen for the benefit of wound healing. METHODS: The concentrations of the carboxyterminal propeptide of type I procollagen (PICP) and the aminoterminal propeptide of type III procollagen (PIIINP) were measured in suction blister fluid of intact skin and in wound fluid in ten surgical patients. PICP and PIIINP concentrations in serum were also measured. Specific radioimmunoassays were used. RESULTS: In peripheral skin, the median preoperative concentrations of PICP and PIIINP were 228 and 140 micrograms/L, respectively. On postoperative days 1, 2, 4 and 7, the median concentration of PICP was 145 (p = 0.01, Wilcoxon signed rank sum test), 102 (0.02), 159 (0.03), and 152 (0.06) micrograms/L, respectively. The postoperative medians of PIIINP were 68 (p = 0.17), 76 (0.04), 66 (0.06), and 56 (0.03) micrograms/L, respectively. At the same time, collagen synthesis in the wound increased dramatically from the second day on. After an initial decrease, propeptide concentrations in serum gradually increased from the fourth day on. CONCLUSIONS: Collagen synthesis is regulated for the benefit of the wound during the acute phase response.

Abstract: Electron microscopy of peripheral nerves obtained from two diabetic patients revealed large deposits of microfibrils and the presence of Luse bodies in the vicinity of perineurial cells. Microfibrils were found to accumulate also in the sciatic nerves of diabetic BB rats; these microfibrillar deposits were shown to contain type VI collagen by immunoelectron microscopy. Connective tissue cells cultured from rat sciatic nerves were exposed to high glucose concentrations. High glucose concentrations up-regulated the mRNA steady-state levels of alpha 1(VI), alpha 2(VI), and alpha 3(VI) chains of type VI collagen and caused accumulation of type VI collagen-containing fibrils in the cultures. Immunostaining and in situ hybridizations demonstrated that perineurial cells, Schwann cells, and fibroblasts expressed type VI collagen at the mRNA and protein levels. The results suggest that the turnover and supramolecular assembly of type VI collagen are perturbed in diabetic nerves and that glucose per se increases the expression of type VI collagen in vitro.

Abstract: OBJECTIVES--Previous reports indicate that serum osteocalcin (serum bone GLA protein (S-BGP)) and carboxyterminal propeptide of type I procollagen (PICP) can be used as indicators of bone formation and turnover. The purpose of this study was to assess the activity of bone formation in patients with rheumatoid arthritis (RA) using S-BGP and S-PICP. The biochemical data were compared with data obtained from bone histomorphometry. METHODS--Concentrations of S-BGP and S-PICP were measured in 119 women with RA aged 30-66 years and 47 healthy female controls matched for age. Bone histomorphometry of iliac crest samples was performed in 107 patients with RA. RESULTS--Weak to moderate correlations between the serum markers and histological bone formation parameters were found. Concentrations of S-BGP and S-PICP were significantly decreased in patients with RA compared with the controls (S-BGP 7.2 (2.3) v 8.7 (2.1) micrograms/l; S-PICP 105 (32) v 117 (38) micrograms/l. The lowest values were found in patients with recent onset RA. CONCLUSIONS--These findings suggest that bone formation and bone remodelling are generally reduced in patients with RA.

Abstract: In the present investigation, collagen synthesis and degradation were studied by measuring the carboxyterminal propeptide of type I procollagen (PICP), the aminoterminal propeptide of type III procollagen (PIIINP) and a type I collagen-specific degradation peptide (ICTP) in the sera of 43 male patients, treated for acne with isotretinoin or with tetracycline. The values were compared with those observed in 24 acne patients without treatment and in healthy controls. The treatment with isotretinoin did not seem to affect these parameters in a cross-sectional setting, whereas tetracycline treatment was associated with slightly decreased levels of ICTP. Since there were marked variations in the PICP, PIIINP and ICTP levels between individual subjects, a follow-up study, including male and female patients, others than in the first part of the study, was conducted. Two other biochemical markers of bone metabolism, osteocalcin, reflecting osteoblastic activity, and tartrate-resistant acid phosphatase (TRAP), reflecting osteoclastic activity, were also analyzed. In females, all these parameters were lower than in males. In addition, the changes in females were more pronounced; in particular, PIIINP and TRAP were significantly increased in females during retinoid treatment (p < 0.05 and p < 0.01, respectively). Importantly, no increase was found in the synthesis of type I collagen during retinoid treatment, suggesting that the commonly used retinoid dosages do not stimulate the synthesis of type I collagen in vivo.

Abstract: Clodronate relieves bone pain in patients with skeletal metastases. Since the pain relieving mechanism of clodronate may be associated with the antiosteoclastic activity, we have investigated whether the drug has simultaneous actions on bone resorption and pain. Although osteosclerotic metastases are characteristic of prostate carcinoma, bone resorption is also accelerated. The resorbing process can be investigated using a specific immunoassay for ICTP (cross-linked carboxyterminal telopeptide region of type I collagen) which allows the measurement of the degradation of type I collagen in serum samples. We have also determined serum concentration of PICP (carboxyterminal propeptide of type I procollagen) which reflects the synthesis of type I collagen (osteoid). Patients who have relapsed after first-line hormonal therapy, were randomised to receive estramustine phosphate (E) with or without clodronate (C) (E + C, n = 50; E, n = 49). The dose of E was 560 mg and that of C 3.2 g for the first month, thereafter 1.6 g. We saw elevated ICTP and PICP levels in the majority of the patients. A transient decrease in ICTP values occurred simultaneously with pain relief. The changes were more accentuated in the E + C than in the E group but the difference was not significant. In each group serum phosphate concentration decreased markedly (P = 0.001) whereas the activity of alkaline phosphatase remained increased, both indicating a development of osteomalacia during E therapy. The short-term antiosteoclastic effect of C may be explained by the dose reduction, hyperosteoidosis and osteomalacia which inhibit the binding of C on the crystal surfaces and by the late phase of disease.

Abstract: Serum concentrations of the markers of collagen synthesis and degradation, collagen I propeptide (PICP), collagen III propeptide (PIIINP) and the cross-linked telopeptide of type I collagen (ICTP) were measured in young male dermatological patients and in control subjects. No significant differences were noted between patients suffering from atopic eczema (n = 24), other eczemas (n = 11), acne (n = 8), psoriasis (n = 7) or tinea (n = 9) and the control subjects (n = 24). In the total study population representing patients with common skin diseases and control subjects there was a significant correlation between the serum concentrations of PICP and PIIINP and between the concentrations of PICP and ICTP. This suggests that synthesis of type I and III collagens in vivo is coordinated and that the degradation and synthesis of type I collagen is balanced. These markers were also measured in older patients suffering from psoriasis, eczema and various connective tissue diseases. It was noted that the degree of skin involvement in these diseases was not related to the serum concentrations of the markers of collagen metabolism. The highest levels of PICP and PIIINP were observed in a patient with systemic mastocytosis (PICP 309 micrograms/l and PIIINP 8.0 micrograms/l). Increased levels of PIIINP were also found in patients with a high alcohol consumption. We have previously demonstrated that systemic glucocorticoids reduce collagen propeptide levels in serum. In the present study we also proved that systemic glucocorticoids have no effect on collagen degradation.(ABSTRACT TRUNCATED AT 250 WORDS)

Abstract: We have previously shown that the serum aminoterminal propeptide of type III procollagen (PIIINP) is a prognostic factor for survival in localised soft-tissue sarcomas, and that elevated values are frequent in metastatic disease. In the present study PIINP is analysed during chemotherapy in 26 patients with advanced sarcomas. Non-responders had a significantly higher pretreatment level of PIIINP than responders (P = 0.05), when only patients with no recent therapeutic interventions were studied. However, during chemotherapy PIIINP followed the clinical course of the malignant disease in only a minority of patients. Patients with recent surgery or recently completed chemotherapy had an increased pretreatment PIIINP value (P = 0.03). In these patients PIIINP declined during chemotherapy irrespective of tumour response. A pretreatment PIIINP level within the reference range tended to increase with time irrespective of response. Moreover, the values taken during a chemotherapy infusion were significantly higher than those immediately preceding the corresponding cycle (P = 0.001). Our results suggest that pretreatment PIIINP is of value as a prognostic factor for chemotherapy response in patients with advanced sarcomas. During chemotherapy PIIINP is of minor importance in monitoring response because of the influence of chemotherapy and other therapeutic interventions on the level of PIIINP.

Abstract: Type I collagen makes up more than 90% of bone matrix. Therefore, analysis of antigens related to collagen formation and degradation in bone should provide good and specific estimates of both bone resorption and bone formation rates. In this study we measured serum levels of the pyridinoline cross-linked telopeptide domain of type I collagen (ICTP) as a marker of bone resorption and serum carboxy-terminal propeptide of type I procollagen (PICP) as a marker of bone formation. Serum levels of the two antigens were correlated to histomorphometric indices of bone resorption and bone formation calculated from iliac crest bone biopsies in a group of 18 individuals with high- and low-turnover bone disease (myxedema, primary hyperparathyroidism, and thyrotoxicosis). After logarithmic transformation the regression of S-ICTP on volume-referent resorption rate (BRs/R/BV) was significant (r = 0.61, p < 0.01, SEM/Y = 56%). S-ICTP also showed a significant regression on the volume-referent cancellous bone balance (r = -0.45, p < 0.05, SEM/Y = 412%). S-PICP was significantly correlated to the mineral appositional rate (r = 0.53, p < 0.05) and volume-referent bone formation rate (r = 0.61, p < 0.01, SEM/Y = 48%). The correlation to bone turnover as expressed in the activation frequency was also highly significant (r = 0.61, p < 0.01, SEM/Y = 51%). No significant correlation with wall thickness or bone balance was demonstrable per remodeling cycle. Thus, assays employing antigens that reflect collagen formation and degradation are useful instruments for the evaluation of rates of bone remodeling in metabolic bone disease.

Abstract: We analyzed the p53 protein immunohistochemically in bronchial dysplasias or squamous cell carcinomas in situ and in squamous cell lung carcinomas occurring in the same patients. The polyclonal antibody used (CM-1) is directed against the wild-type p53 protein, but also recognizes the mutated p53 in formalin-fixed and paraffin-embedded sections. To study the integrity of basement membranes (BMs) and the possible invasion of the dysplastic epithelium, immunostainings for the BM proteins laminin and type IV collagen were used. Nine of the 17 dysplasias showed p53 protein expression (53%); it was significantly more often seen in severe dysplasias and carcinomas in situ than in mild or moderate dysplasias (P = 0.04). The p53 antigenicity was generally located in the basal part of the epithelium. The BMs beneath mildly dysplastic epithelia were continuous. In contrast, those under moderately or severely dysplastic epithelia showed occasional disruptions. p53 protein expression was also found in dysplastic epithelium above a continuous BM suggesting an ominous process before signs of invasion. Twelve of the 17 squamous cell carcinomas showed p53 protein expression (71%). There was a significant concurrent p53 expression in bronchial dysplasias and their related squamous cell carcinomas (P = 0.009), so that all nine cases of p53 positive bronchial dysplasia also showed p53 positivity in the associated squamous cell carcinomas. These findings indicate that p53 protein expression is possible in premalignant bronchial lesions, and suggests that the p53 expression could, at least in some cases, be an early event in the development of a squamous cell carcinoma of the lung.

Abstract: We developed a radioimmunoassay (RIA) for the carboxy-terminal telopeptides of type I collagen (ICTP), cross-linked with the helical domain of another type I collagen molecule, after isolation from human femoral bone. The cross-linked peptide was liberated by digesting insoluble, denatured bone collagen either with bacterial collagenase or with trypsin, and purified by two successive reversed-phase separations on HPLC, with monitoring of pyridinoline-specific fluorescence. The purity of the peptide was verified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and its origin in the type I collagen fibers was determined by amino-terminal amino acid sequencing. Polyclonal antibodies and a separation reagent containing second antibody and polyethylene glycol are used in the RIA. An immunologically identical, somewhat larger antigen is present in human serum; its concentration increases in multiple myeloma and in rheumatoid arthritis. The ICTP antigen seems to be cleared from the circulation by the kidneys, because glomerular filtration rates that are two-thirds of normal or less are associated with increased circulating ICTP concentrations. The CVs of the method are between 3% and 8% for a wide range of concentrations. The analysis of 40 serum samples can be completed in 4 h.

Abstract: Increased serum concentrations of aminoterminal propeptide of type III procollagen occur in advanced ovarian cancer. To study their origin, we compared the expressions of type I and type III procollagens in ovarian tumor tissue and the peritoneal cavity with immunoassays for the propeptide domains of these procollagens. Samples of tumor cyst fluid, peritoneal ascitic fluid, tumor vein blood, and peripheral blood were obtained at operation from 50 women with malignant ovarian neoplasms and 61 women with benign neoplasms. The ascitic fluid concentrations of both type I and type III procollagen antigens were significantly higher in the malignant tumors than in the benign ones, but this difference was evident only for type I procollagen in the tumor cysts. The aminoterminal propeptide of type III procollagen concentration in the peripheral blood was higher in the patients with malignant tumors, whereas the concentrations were similar in the tumor veins. The enhanced type I procollagen synthesis in the malignant tumors did not affect the corresponding antigen in the blood. The findings suggest that progressive ovarian carcinoma invariably induces a fibroproliferative response, characterized by active expression of type I and type III procollagens. The increased circulating aminoterminal propeptide of type III procollagen is derived from the peritoneal cavity rather than from the tumor tissue via the ovarian vein.

Abstract: In order to determine whether bone turnover varies during the normal menstrual cycle, we measured biochemical markers of bone resorption (serum pyridinoline cross-linked carboxy-terminal telopeptide of type I collagen (sICTP), fasting urinary hydroxyproline/creatinine, fasting urinary pyridinoline/creatinine and fasting urinary deoxypyridinoline/creatinine) and bone formation (plasma osteocalcin, serum carboxy-terminal propeptide of type I procollagen and serum alkaline phosphatase) in ten healthy premenopausal women every two or three days for a complete menstrual cycle. A cyclic pattern was detected in sICTP, with its nadir during the follicular phase and its peak during the luteal phase, and an overall variation of 17% during the menstrual cycle (p = 0.004). No cyclic changes were observed in the urinary parameters of bone resorption or in the biochemical markers of bone formation. We conclude that sICTP, a new biochemical marker of bone resorption, undergoes small variations during a normal menstrual cycle in premenopausal women, whereas the biochemical markers of bone formation remain constant.

Abstract: OBJECTIVES--To assess the extent and clinical significance of type I collagen degradation in rheumatoid arthritis (RA). METHODS--Serum samples from 90 consecutive patients with RA from a cross-sectional population based study and 90 age- and sex-matched controls were analysed with the new assay of cross-linked carboxyterminal telopeptide of type I collagen (ICTP). RESULTS--Patients with RA had significantly higher concentrations of ICTP than the controls. ICTP correlated strongly with measures of impairment in RA, such as the erosive state of joint disease (ES) (r = 0.57, p < 0.001) and Keitel function test (KFT) (r = 0.49, p < 0.001), and more weakly with various disease activity markers. When erythrocyte sedimentation rate (ESR), ES or KFT were used as indicators of disease severity among the patients with disease duration over five years, ICTP distinguished the more serious RA from milder cases. CONCLUSIONS--Elevated serum concentrations of ICTP are common in RA and are associated with signs of aggressive disease.

Abstract: One hundred seventy-four children and adolescents with insulin-dependent diabetes mellitus were examined for joint contractures and skin manifestations in their hands. Joint contractures were found in 52 (29.8%) and skin manifestations in 29 (16.6%) patients. To eliminate the possible confounding effects of age and duration of diabetes on the variables to be studied, patients younger than 7 y and with a duration of diabetes shorter than 3 y were excluded from the subsequent analyses. Of the remaining 108 children, those with joint contractures had lower serum concentrations of the 7-S domain of type IV collagen and the P1 fragment of laminin than the other patients (p = 0.033) but higher mean glycated Hb levels (p = 0.048). A clear association was noted between the occurrence of joint contractures and skin changes (p = 0.007). Background retinopathy was found in six patients (5.6%), three of whom had stage II joint contractures (p = 0.064). The children with skin changes and those with combined joint and skin manifestations more often had insulin-dependent diabetes mellitus in their first-degree relatives (p = 0.038 and p = 0.043, respectively). No difference in relative height was found between the groups. No association could be seen between disease susceptibility antigens in the HLA-D locus and joint or skin manifestations. The lower levels of circulating collagen and laminin metabolites in the diabetic children with joint contractures suggest that these patients are characterized by a reduced turnover of basement membranes in tissues.(ABSTRACT TRUNCATED AT 250 WORDS)

Abstract: BACKGROUND. Abnormal interaction with the extracellular matrix is a basic property of malignant cells. Type III collagen is a major constituent of the extracellular matrix of soft tissues. METHODS. Deposition of the aminoterminal propeptide of type III procollagen (PIIINP) was studied in benign (n = 41), borderline (n = 4), and malignant (n = 32) human ovarian tumors using the avidin-biotin-immunoperoxidase technique and affinity-purified antibodies to human PIIINP. It was then compared with the serum PIIINP concentrations of the patients at the time of operation. RESULTS. In malignant tumors, the distribution of PIIINP was irregular both close to the epithelial cancer cells and further away, in the stroma. Another feature typical of malignant tumors was the varying staining intensity of the PIIINP-positive fibers. The benign tumors were characterized by a regular organization and an intensive staining of PIIINP. Borderline tumors showed a slightly decreased staining intensity and altered PIIINP distribution. A significant positive correlation was found between the PIIINP concentration in serum and the degree of irregularity in the distribution of PIIINP. CONCLUSIONS. These preliminary results indicate that malignant transformation in ovarian tumors is associated with disintegration of adjacent collagenous structures and with alterations in type III procollagen metabolism, which also leads to increased serum PIIINP levels. They suggest that biochemical or immunohistochemical detection of the PIIINP antigen could be clinically useful in ovarian tumors.

Abstract: Immunohistochemically detectable p53 protein using a polyclonal antibody (CM-1) was studied in 42 carcinomas of which 11 were grade I, 22 grade II and nine grade III carcinomas. Additionally 14 urothelial dysplasias were studied. In 11 of these a diagnosis of transitional cell carcinoma was established before and in one after the dysplasia diagnosis. Twenty-one out of 42 (50%) cases of transitional cell carcinoma were positive for the p53 protein. Eleven out of 14 (78%) dysplasias and 10/12 (83%) related carcinomas were p53 positive. One out of 11 grade I (9%), 12/22 grade II (55%) and 8/9 grade III (89%) tumours showed positivity for p53. There were significantly more p53 positive cases in grade II-III tumours than in grade I tumours (P = 0.004). There were significantly more p53 positive cases in stage T2-T4 tumours than in stage T1 tumours (P = 0.035). In only one case among the 11 dysplastic lesions following the treatment of a carcinoma the dysplastic lesion was p53 negative while the preceding carcinoma was p53 positive. All dysplasias and 28 carcinomas were also immunostained for laminin and type IV collagen to evaluate the continuity of basement membranes (BMs). Clearly disrupted BMs were observed only in grade III carcinomas. These cases showed the most p53 immunopositivity. The results show a strong association of p53 staining between dysplasias and transitional cell carcinomas of the urinary bladder indicating that these lesions might share similar p53 changes. The correlation to grade, clinical stage and to disrupted BM suggests that p53 mutations may be associated with the evolution of aggressive growth characteristics in transitional cell carcinomas or, alternatively, that p53 positive tumours of a more aggressive type from the start. Whether p53 staining can be used as an adjunct in the assessment and follow-up of epithelial changes of patients treated for a p53 positive bladder carcinoma deserves to be studied.

Abstract: Differential diagnosis of hepatic complications after bone marrow transplantation (BMT) is often difficult. To assess whether serum concentrations of the aminopropeptide of type III procollagen (PIIINP) could facilitate diagnosis, we measured serum PIIINP, corrected for age by conversion to standard deviation scores (SDS), serially after BMT in 27 children. A preliminary study of 11 patients showed that a PNIIINP-SDS cutoff of 8.0 was an optimum for diagnosis of veno-occlusive disease (VOD). PIIINP-SDS was increased above cut-off 1-25 days before the onset of clinical signs in the 12 patients (4 from the preliminary group, the others from a group of 16 studied prospectively) who developed VOD, with subsequent changes in PIIINP-SDS mirroring the course of VOD. By contrast, PIIINP-SDS remained below cut-off in all other patients, including 7 with liver graft-versus-host disease and 3 with drug hepatotoxicity. PIIINP-SDS values greater than 8.0 predict, diagnose, and monitor VOD after BMT.

Abstract: The concentrations of the carboxy-terminal propeptide of type I procollagen (PICP) and the amino-terminal propeptide of type III procollagen (PIIINP) in serum were followed prospectively as indicators of synthesis of the respective collagens in 266 healthy primiparas during the second and third trimesters of pregnancy. The PIIINP concentration increased 2-fold during the third trimester; the median value was 10.0 micrograms/L at gestational week 40, with a range from 4.6-32.7 micrograms/L (reference interval for nonpregnant women, 1.7-4.2 micrograms/L). A significant increase also took place in the PICP concentration. The frequency distributions of the two parameters were near normal at week 26. They started to broaden out at week 32 and were wide at week 38. In severe preeclampsia, PIIINP tended to increase more than in normal pregnancies of the same duration. The increase in PIIINP correlated significantly with maternal weight gain and the birth weight of the infant, but not with other parameters related to pregnancy or delivery. At gestational week 38, there was a significant correlation between the circulating concentrations of insulin-like growth factor-I and PIIINP or PICP, but not between human placental lactogen and PICP, and only a weak association between human placental lactogen and PIIINP. This suggests that insulin-like growth factor-I is involved in the regulation of type I and type III collagen synthesis during pregnancy.

Abstract: We compared the procollagen synthetic properties of MG-63 osteosarcoma cells with those of cultured human skin fibroblasts. In both cells, the expressions of type I and III procollagens are largely dependent on the constant presence of ascorbate and coordinately decreased by the neutral polymer dextran T-40. The amino-terminal propeptides of pro-alpha 1 and pro-alpha 2 chains of type I procollagen are phosphorylated and those of the pro-alpha 1 and pN-alpha 1 chains of type III procollagen both phosphorylated and sulfated, there being no difference in net charge in the propeptides between these cell types. The major differences between MG-63 and normal fibroblasts are the exceptionally high relative synthesis of type III procollagen by MG-63 cells, up to about 40% of the total of types I and III (6% in cultured skin fibroblasts), and the inability of ascorbate-supplemented MG-63 cells to deposit collagens into an insoluble pericellular matrix. A longer dextran treatment shifts up to one-fourth of the proline-labeled extracellular macromolecules into the matrix fraction within 4 days (in control 4%). Despite processing of the procollagens to the respective collagens in the matrix, neither control matrices nor those induced by dextran induced increased production of alkaline phosphatase. In cultures up to 4 days postconfluence the proportion of type III collagen produced tended to increase over that in early confluent cultures. With respect to collagen production, the MG-63 cell line is not a representative of the osteoblast lineage but rather resembles a proliferative wound fibroblast.

Abstract: Serum concentration of PIIINP (aminoterminal propeptide of type III procollagen), an indicator of connective tissue metabolism, is often increased in advanced gynecological carcinoma and correlates with the tumor growth. Because the healing wound also affects collagen metabolism we measured the serum PIIINP concentration in 15 patients operated on for endometrial carcinoma and in 28 patients operated on for benign gynecological diseases. Serial serum samples were collected for 35-51 days after surgery. In endometrial carcinoma, serum PIIINP level was increased in 53% of the patients before the operation. Postoperatively, the mean PIIINP concentration increased further, remaining at a pathological level throughout the study and with a peak at one week after the operation. In relative terms an abdominal operation for benign disease increased the serum PIIINP concentration as much as in endometrial carcinoma, but the mean level remained within the reference interval. After vaginal operation the increase in PIIINP concentration was not significant. The initially increased PIIINP concentration in endometrial carcinoma is probably caused by the effects of the malignancy on collagen metabolism. Because the relative response of PIIINP to abdominal operation was similar in both groups, the PIIINP response to surgical trauma in patients with malignant disease seems to depend only on the extent of the operation. In this group the transient increase in serum PIIINP concentration does not prevent the use of PIIINP as a tumor marker, since the follow-up examinations for malignancy do not start earlier than two months after operation.

Abstract: Sera of 85 patients with benign soft tissue lesions or sarcomas of soft tissues were investigated for a collagen metabolite, the aminoterminal propeptide of type III procollagen (PIIINP). Patients were divided into three groups: benign soft tissue lesions (n = 39), localised (n = 29) and metastatic (n = 18) soft tissue sarcomas (STS). Values of PIIINP above the reference range were found in 15%, 28% and 50% of the respective groups. The difference in the concentration of PIIINP was statistically significant between the benign lesions and the localised sarcomas; P = 0.05, and between the benign lesions and the metastatic sarcomas; P less than 0.001. In localised sarcomas there was a correlation between PIIINP and bone-involvement (r = 0.61, P = 0.002) and in metastatic disease between PIIINP and liver metastases (r = 0.77, P less than 0.001). In localised sarcomas the overall survival for patients with a value of PIIINP above the reference range was significantly poorer (P = 0.03) than for patients with values within the reference range, even after stratification for the histological malignancy grade of the tumours (P = 0.04).

Abstract: Schistosomiasis, in contrast to alcoholic liver disease, leads to presinusoidal hepatic fibrosis, which determines the prognosis of the disease. Because conventional liver function tests and liver biopsy specimens provide little information about the dynamics of the fibrotic process, we measured the serum concentrations of procollagen type III N-propeptide and procollagen type I C-propeptide, believed to mainly reflect collagen synthesis, and procollagen type IV C-propeptide and collagen type VI, two presumptive markers of collagen degradation. Determinations were performed in 15 healthy control subjects, 69 patients with various stages of infection with Schistosoma mansoni/Schistosoma haematobium (28 with an early active infection and no organ involvement, 27 with hepatosplenic involvement and 14 with complications of portal hypertension) and 16 patients with alcoholic cirrhosis. In addition, liver biopsy specimens were obtained from 30 schistosomal patients (18 with hepatosplenic involvement and 12 with complications of portal hypertension for histopathological grading and collagen histochemistry. Procollagen type III N-propeptide was significantly elevated in the three patient groups with schistosomiasis when compared with controls (p less than 0.01). Also, patients with higher histological grading showed significantly higher procollagen type III N-propeptide values (p less than 0.05). In alcoholic patients, procollagen type III N-propeptide was even higher and increased parallel to the severity of the disease, determined by using a combined clinical and laboratory index. Procollagen type I C-propeptide was only elevated in early infection (p less than 0.05) and steadily decreased with disease progression.(ABSTRACT TRUNCATED AT 250 WORDS)

Abstract: Collagen is synthesized as procollagen and large extra domains known as propeptides are cleaved off enzymatically. In the present study we have measured the carboxyterminal propeptide of type I collagen (PICP) and the aminoterminal propeptide of type III collagen (PIIINP) in blister fluids of human skin. High concentrations of PICP were found in the spontaneous blisters of patients with bullous pemphigoid, erysipelas, or erythema multiforme. Detectable amounts were also found in suction blisters induced on healthy skin. Because the concentrations in suction blisters were several times higher than in corresponding serum, most of PICP and PIIINP was derived from the underlying dermis. This method was used for assessing type I and type III collagen synthesis after topical glucocorticoid treatment. Clobetasol-17-propionate (CP) decreased the concentrations of PICP by 75% after 1 d of treatment, the maximum inhibition (92%) being found after 2 d treatment. PIIINP was also affected. Hydrocortisone and hydrocortisone-17-butyrate also decreased the concentrations of PICP and PIIINP, but less markedly than CP. Partial recovery was seen 3 d after stopping the treatment. Thus measurement of collagen type specific propeptides in suction blisters can be used as an estimate of collagen synthesis in vivo, avoiding both local anesthesia and skin biopsing. With radioimmunoassays for PICP and PIIINP a large number of samples can also be processed simultaneously.

Abstract: To investigate the lymphatic transport of the aminoterminal propeptide of type III procollagen (PIIINP) we established a thoracic duct-venous shunt in 6 pigs. Porcine PIIINP was purified, characterised, and compared with human PIIINP to ensure the suitability of the radioimmunoassay of human PIIINP for measurements in pigs. SDS-PAGE and radioimmunoinhibition assays show human and porcine PIIINP to be similar, thus indicating that the assay of human PIIINP is also reliable for determinations on pig serum and lymph. Intact PIIINP, as identified by gel filtration, accounted for 60% and 40% of the total PIIINP immunoreactivity in lymph and serum, respectively. The higher amount of total immunoreactivity and proportion of intact PIIINP in lymph compared with serum support the hypothesis that intact PIIINP is transported from peripheral tissue into the circulation by lymph. Two days after the shunt was established, the lymph was collected quantitatively hour-by-hour for 24 h. The flow was higher during the light periods than in the dark (p less than 0.01). The PIIINP concentration varied inversely with the flow, being higher in the dark hours (p less than 0.03). However, the total collected amount of PIIINP in lymph did not differ during the light and dark periods. Serum PIIINP remained unchanged over the 24 h. The lymphatic clearance of total PIIINP immunoreactive components was 6.2 ml serum/min and the lymphatic clearance of intact PIIINP was 9.1 ml serum/min, equal to 7 and 10 times the plasma volume/24 h, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Abstract: Type I and type III collagen are components of a healing wound, and major structural proteins. According to our previous study, wound fluid concentrations of the liberated propeptide extensions of procollagens can be used to monitor collagen synthesis in the wound. Serum concentrations of the carboxyterminal propeptide of type I procollagen (PICP), and the aminoterminal propeptide of type III procollagen (PIIINP) were studied here for up to half a year in 102 patients, admitted for major abdominal surgery. In a frequent follow-up (n = 9), one minimum and two maxima were found for S-PICP, occurring 1 day, 7 days, and 2 months after surgery, respectively. S-PIIINP had a minimum at 1 day and a peak at 10 days. Relative changes (follow-up result/pre-operative concentration) of the propeptides in 50 uncomplicated patients were compared. The 1-day minimum of S-PICP was 0.60 (SD 0.18), and that of S-PIIINP 0.89 (0.27), (P less than 0.0001, 95% CI for the mean difference 0.21 to 0.36). The 7-day peak of S-PICP was 1.4 (0.5), and that of S-PIIINP 2.5 (1.2), (P less than 0.0001, CI 0.81 to 1.42). The 2-month-peak of S-PICP was 1.6 (0.3), and at the same time the relative S-PIIINP was still 1.7 (0.3) without any separate peak. Major infectious (n = 8) and other (12) complications, exploratory procedures (22) and patients with abnormal pre-operative propeptide levels (8) were studied separately. Two early deaths were excluded. Only major infection had a remarkable effect on the responses of S-PICP (3/8) and S-PIIINP (5/8).(ABSTRACT TRUNCATED AT 250 WORDS)

Abstract: The effect of systemic glucocorticoid treatment on collagen synthesis in patients with various dermatoses was studied by measuring the carboxyterminal propeptide of type I procollagen (PICP) and the aminoterminal propeptide of type III procollagen (PIIINP) in serum. Changes in the propeptide concentrations were compared with those of osteocalcin, which reflects osteoblastic activity, and tartrate resistant acid phosphatase (TRAP), which reflects osteoclastic activity. The treatment caused significant decreases in levels of PICP, PIIINP and osteocalcin of 38, 34 and 49%, respectively (P less than 0.001). For TRAP, both increases and decreases were seen. The effects on PICP and PIIINP were evident 2-4 days after the onset of steroid therapy. The decrease in PICP was dose-related (r = 0.470, P less than 0.005) but even relatively small doses (0.1 mg of prednisone/kg/1 day) caused a significant reduction in PICP. After cessation of treatment, the levels of PICP returned to the pretreatment level in 1 week. The present study demonstrates that systemic glucocorticoid therapy in humans suppresses the synthesis of type I and III collagens and also non-collagenous bone matrix proteins.

Abstract: Type I collagen is the main collagen type found in mineralised bone. Specific immunoassays for PICP (carboxyterminal propeptide of type I procollagen) and ICTP (cross-linked carboxyterminal telopeptide region of type I collagen) allow simultaneous assessment of the synthesis and degradation of type I collagen in serum samples, respectively. Our aim was to find out whether these metabolites of type I collagen are useful markers for following bone turnover and evaluating treatment response in multiple myeloma, which is a good model disease of excessive osteolysis. Fifteen consecutive patients were studied before and throughout their treatment. Samples for serum PICP and ICTP were collected before starting each treatment course of melphalan and prednisolon. Response to treatment was evaluated by following the changes in M protein and bone roentgenograms. The disease was progressing in four and regressive in 11 patients, but in four of these a recurrence occurred. In nonresponders the ICTP concentration was permanently elevated despite treatment. In responders both increased or normal levels of ICTP were initially observed, but they returned to or remained in the reference interval during treatment. The ICTP concentration increased upon recurring disease. There was a strong correlation between the extent of bone lesions and ICTP. There was no correlation between ICTP and PICP, the latter mainly remaining within the reference range, a finding that suggests no change in bone formation. ICTP was a significant predictor for survival in this patient group (P less than 0.05). We conclude that ICTP is a specific and sensitive marker for bone resorption. Simultaneous use of serum ICTP and PICP offers an additional and easy means to follow bone turnover and evaluate the response to therapy in multiple myeloma.

Abstract: There are several indications that the functions of human osteoblasts and osteoclasts have circadian rhythms with peak activities occurring at night. It is not known, however, whether the principal function of these cells, namely synthesis and degradation of the organic matrix of bone, of which about 90% is type I collagen, also has a circadian rhythm. This was therefore investigated for both the formation of type I collagen and the degradation of type I collagen in bone using two newly developed serum markers: the serum concentration of the carboxy-terminal propeptide of type I procollagen (PICP) as a marker of formation and the serum concentration of the carboxy-terminal pyridinoline cross-linked telopeptide of type I collagen (ICTP) as a marker of degradation. PICP and ICTP were measured by RIA in samples taken every 3 h over a 24 h period in 12 healthy premenopausal women (age 32 +/- 5 years, mean +/- SD). Both PICP (p = 0.003) and ICTP (p = 0.00003) showed a significant circadian rhythm, with about 20% higher values at night than in the afternoon. We conclude that serum markers of both the formation of new type I collagen and the degradation of old type I collagen in bone exhibit a clear circadian rhythm, with increased activity of both osteoblasts and osteoclasts at night. The etiology of this circadian rhythm is still unknown.

Abstract: Immunocytochemistry, radioimmunoassay, immunoblotting, Northern analysis, and polymerase chain reaction (PCR) technique were applied to investigate the distribution of laminin and its neurite outgrowth domain in brains of neuropathologically verified cases of Alzheimer's disease and Down's syndrome. New antibodies against a neurite outgrowth domain of laminin were characterized and were used in localization of this peptide antigen in the human brain. Laminin was found as large punctate deposits in all plaques in the affected brains. Laminin synthesis was increased as assessed by RNA blotting and immunoblotting, and glial cells were heavily immunoreactive with antibodies for a neurite outgrowth-promoting peptide antigen of the B2 chain of laminin. This peptide antigen not only was produced by glial cells but also was deposited in the brain tissue. As this peptide antigen promotes neurite outgrowth at low concentrations, and is specifically neurotoxic at high concentrations, it may play a synergistic role with other molecules in inducing the sprouting and neurodegeneration occurring in brains of patients with either Alzheimer's disease or Down's syndrome.

Abstract: The effect of insulin on the serum levels of the amino-terminal propeptide of type III procollagen (PIIINP) was investigated in patients with non-insulin-dependent diabetes mellitus, whose disease was unsatisfactorily controlled by oral drugs. Before insulin therapy the PIIINP values of the patients (3.2 +/- 1.3 micrograms/l, n = 38) varied within the range of healthy subjects (3.1 +/- 0.6 micrograms/l, n = 50, NS). Insulin therapy (6-20 IU at bedtime plus the oral drugs) improved the glycemic control and increased the serum PIIINP during a 4 week (3.1 +/- 0.9 to 3.8 +/- 1.1 micrograms/l, P less than 0.01, n = 8) and an 8 week period (3.2 +/- 1.3 to 3.8 +/- 1.6 micrograms/l, P less than 0.001, n = 22). The values were still elevated after 6 months on insulin (3.5 +/- 1.5 to 4.0 +/- 1.7 micrograms/l, P less than 0.01, n = 12). Placebo-insulin did not alter the concentration of PIIINP (3.1 +/- 0.6 to 2.8 +/- 0.6 micrograms/l, NS, n = 8) whereas the glycemic control improved and body weight decreased. The PIIINP values correlated with fasting insulin before (r = 0.403, P less than 0.05, n = 30) and after the therapy (r = 0.452, P less than 0.001, n = 60). Insulin therapy, while correcting the hormone deficiency and restoring glucose and protein metabolism, seems to activate the synthesis of type III procollagen in patients with NIDDM. This may promote the atherosclerotic process.

Abstract: In this study , serum levels of classical serum markers of bone formation [carboxyterminal propeptide of procollagen type I (S-PICP), bone Gla protein (S-BGP)], and total alkaline phosphatase (S-AP)) were related to the calcium kinetic index of whole skeletal mineralization rate (m) by regression analysis in a variety of metabolic bone diseases. For each disease, the regression coefficient (r) as well as the fraction: standard error of estimate/mean dependent variable (SEE/Y) were determined. In a group of 19 normals, only the regression of S-PICP on m reached significance (r = 0.53, P < 0.02, SEE/Y = 0.44), whereas regressions of S-AP and S-BGP on m were nonsignificant. In a pooled material of high- and low-turnover bone diseases without mineralization defects or spinal fracture [myxedema, thyrotoxicosis, and primary hyperparathyroidism (n = 48)], a highly significant positive regression of S-PICP on m was demonstrable (r = 0.50, SEE/Y = 0.63, P < 0.001). The regression coefficients obtained for S-BGP and S-AP were 0.74 (P < 0.001, SEE/Y = 0.41) and 0.42 (P < 0.01, SEE/Y = 0.55), respectively. When analyzing individual diseases in this group, significant differences among the three markers were detectable. In a group of 52 osteoporotics, S-PICP correlated significantly to m (r = 0.49, P < 0.001, SEE/Y = 0.50). Corresponding r-values for S-BGP and S-AP were 0.21 (NS) and 0.48 (P < 0.001, SEE/Y = 0.61), respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Abstract: Bone marrow fibrosis is known in myelomatosis and depends on the extent of plasma cell infiltration. The serum concentration of the aminoterminal propeptide of type III procollagen (PIIINP) has previously been reported to reflect fibrogenesis in the marrow in myelofibrosis. Here we followed 15 consecutive patients with newly diagnosed multiple myeloma with repeated PIIINP measurements during treatment with intermittent courses of melphalan and prednisolone. PIIINP was found to change with clinical behaviour of the disease, nonresponders and patients with recurrent disease having elevated values and the values in responders decreasing to the normal level or remaining there. Collagen fibres in plasma cell infiltrates of biopsies from bone marrow or skeletal tumours of these patients stained heavily with antibodies against PIIINP. Our results suggest that PIIINP works as a noninvasive indicator of bone marrow fibrogenesis. In multiple myeloma PIIINP is a sensitive, but not specific marker of disease course.

Abstract: Seventy-nine osteoporotic (prior forearm or vertebral fracture), but otherwise healthy, postmenopausal women (aged 55 to 75 years) were allocated to two double-blind trials: (1) 39 women received either nandrolone decanoate (anabolic steroid) 50 mg as an intramuscular depot injection or a placebo injection every 3 weeks for 1 year; and (2) 40 women received either 2 mg 17 beta-estradiol plus 1 mg norethisterone acetate or placebo tablets daily for 1 year. Sixty-seven (85%) completed the 1 year of treatment. Serum concentration of type I procollagen carboxy-terminal propeptide (PICP) was measured before and at 3, 6, 9, and 12 months of therapy. In addition, 32 of the women had an iliac bone biopsy taken after double tetracycline labeling. Initial serum PICP correlated significantly with histomorphometrically measured rate of bone formation (r = .4; P less than .05) and plasma bone Gla protein (r = .6; P less than .001), but not with histomorphometrically measured bone resorption or biochemical estimates of bone resorption (fasting urinary hydroxyproline and calcium). Estrogen-progestogen therapy significantly decreased (P less than .001) serum PICP by about 30%, whereas anabolic steroid therapy hardly affected it. We conclude that serum PICP may be used as a noninvasive measurement of bone formation on a group basis. Whereas bone formation is clearly decreased during estrogen-progestogen therapy, it is not affected by long-term therapy with anabolic steroids.

Abstract: Osteoblast-like cells are commonly found in the vicinity of osteoclasts formed in long-term human bone marrow cultures, and they are believed to be derived from osteogenic cell precursors belonging to the stromal cell system. This paper describes a new culture method for human osteoblasts from the adherent cell population of long-term human mononuclear bone marrow cultures. The cells obtained exhibited all the classic characteristics of osteoblasts. They contained high intracellular concentrations of alkaline phosphatase and they secreted the osteoblast-specific marker bone Gla protein. Collagen production was mainly (95-98%) procollagen type I propeptide and only minute quantities of procollagen type III propeptide were detectable by radioimmunoassay in the conditioned medium. After eight weeks the cells formed a mineralized matrix on exposure to beta-glycerophosphate and ascorbic acid. This system provides a model for the study of osteoblast differentiation in vitro and may form the basis for the use of defined media in bone cell cultures due to the presence of high concentrations of osteoblast precursors.

Abstract: To quantify wound healing in surgical patients, samples of wound fluid were collected through a silicone rubber tube for 7 postoperative days and their concentrations of the carboxyterminal propeptide of type I procollagen (PICP) and the aminoterminal propeptide of type III procollagen (PIIINP) were measured with specific radioimmunoassays. The mean concentration of PICP in would fluid on day 1 was 207 +/- 92 (SD) micrograms/L, and on day 2 908 +/- 469 micrograms/L (p less than 0.001, signed rank test). On day 7, the mean concentration reached was 380 times higher than that of day 1 (79,330 +/- 54,151 micrograms/L). Only one peak of PICP antigenicity, corresponding to the intact propeptide as set free during synthesis of type I procollagen, was detected on Sephacryl S-300 gel filtration analysis of wound fluid samples. The mean concentration of PIIINP was 70 +/- 61 micrograms/L on day 1, 86 +/- 88 micrograms/L on day 2, and 180 +/- 129 micrograms/L on day 3 (p less than 0.001 when compared with day 1). Finally on day 7, a 250-fold concentration (17,812 +/- 9839 micrograms/L), compared with day 1, was reached. Methods described in the present paper allow separate and repetitive quantification of the synthesis of both type I and type III procollagen during human wound healing.

Abstract: The effects of modified protein sparing therapy (PSP) and total parenteral nutrition (TPN) on total and wound metabolism were studied for 96 hours after laparotomy and a small gastric excision in 40 rabbits starved for seven days. A further eight starved and eight non-starved animals served as controls for the blood variables. Normal healing up to day 14 was studied in 20 non-starved animals. The difference in deaths and animals in poor condition, 42.1 per cent in PSP and 18.6 per cent in TPN, respectively, was clear but statistically non-significant. PSP led to a lower mean serum albumin concentration than TPN, 25.7 +/- 3.7 (SD) and 28.7 +/- 3.0 (p = 0.02), respectively. The animals receiving PSP excreted significantly more 3-methylhistidine. TPN maintained a positive nitrogen balance, but PSP produced a negative one. The collagen content of the skin scar was lower after PSP (3.1 +/- 0.7 mg) than after TPN (4.5 +/- 1.3 mg) (p less than 0.05), the latter coming close to the level for normal 4-day healing, 4.5 +/- 1.2 mg. Prolyl 4-hydroxylase (PPH) activity showed no difference. No inter-group differences in collagen were found in the stomach. Both regimens totally reversed the starvation-induced decrease in PPH activity in the stomach, but only partially in skin. Thus TPN produced better total and skin wound metabolism after laparotomy and starvation than did PSP. No differences in visceral wound healing were observed.

Abstract: The epithelial cell tumor markers squamous cell carcinoma antigen, CA 125, CA 15-3, and TAG 72, and the aminoterminal propeptide of type III procollagen, an indicator of collagen metabolism, were evaluated in 111 cervical carcinoma patients. Squamous cell carcinoma antigen was pathologic in 47%, aminoterminal propeptide of type III procollagen in 40%, CA 125 in 13%, CA 15-3 in 30%, and TAG 72 in 9% of the 91 patients with squamous cell carcinoma. The squamous cell carcinoma antigen, aminoterminal propeptide of type III procollagen, and CA 125 correlated with the clinical stage. The predictive value of a pathologic squamous cell carcinoma antigen was 78% and that of a negative result 68%. Squamous cell carcinoma antigen and aminoterminal propeptide of type III procollagen further increased the detection rate by approximately 20% from that obtained by squamous cell carcinoma antigen alone. In 16 patients with advanced disease, squamous cell carcinoma antigen correlated with the behavior of the disease in eight, aminoterminal propeptide of type III procollagen in nine, and CA 125 in six patients. Pathologic squamous cell carcinoma antigen, CA 125, CA 15-3, TAG 72, and aminoterminal propeptide of type III procollagen appeared in 11, 32, 31, 31, and 47% of 19 patients with adenocarcinoma, respectively. Squamous cell carcinoma antigen is clinically useful in squamous cell carcinoma but poor in adenocarcinoma, for which the other markers are better. Squamous cell carcinoma antigen, CA 125, and aminoterminal propeptide of type III procollagen may be used for monitoring the behavior of advanced squamous cell carcinoma.

Abstract: We have examined the relationship between blood markers of fibrogenesis and basement membrane formation and alcohol intake in patients with a wide range of clinical and histological severity of alcoholic liver disease (Niemelä et al., 1990). While the patients with a mean of less than 8 mM alcohol in morning urines (mild or moderate drinkers) had a significant (p < 0.00001) decrease in serum aminoterminal propeptide of type III procollagen, type IV collagen and laminin in a period of 27 +/- 1 weeks, the patients with more than 8 mM of urinary alcohol (heavy drinkers) had no improvement. There was also a significant decrease in serum gamma glutamyl transferase activity in the group with lower, but not in that with the higher urinary alcohol concentrations. The Combined Clinical and Laboratory Index (CCLI) decreased in both groups, although the recovery was significantly (p < 0.03) greater in those with the lower urinary alcohol levels (66% +/- 6%) than in the group with a urinary alcohol level of > or = 8 mM (28% +/- 15%). We suggest that connective tissue metabolism in alcoholic liver disease is closely related to alcohol intake and thus affects the prognosis of the alcoholic patient.

Abstract:

Abstract: When subconfluent cultures of primary human skin fibroblasts are incubated for 20 h in the presence of 5% neutral dextran, the newly synthesized procollagens are shifted from the medium into the pericellular matrix fraction. This is accompanied by an overall decrease by 30-50% in the secretion rates of these proteins, as indicated by the incorporation of tritiated proline into collagenase-sensitive macromolecules and by radioimmunoassays for the propeptide regions of type I and III procollagens. Inhibition of tyrosine sulphation in newly formed type III procollagen by NaClO3 does not change the distribution of this protein between the medium and matrix fractions either in the presence or in the absence of the polymer. Processing of type III procollagen at its N-terminus is incomplete in this situation, irrespective of whether the protein is present mainly in the medium or in the pericellular matrix. The concentrations of the mRNAs for the pro alpha 1(I)- and pro alpha 1(III)-chains of procollagens and for actin were monitored for up to 20 h; in the dextran-treated cultures these are not different from the corresponding concentrations in control cultures, indicating that the rapid down-regulation of the synthesis of type I and III procollagens in response to the enhanced pericellular matrix deposition induced by dextran is not due to transcriptional mechanisms.

Abstract: Estrogen stimulates osteoblastic collagen production in vitro, but whether the same stimulation takes place in vivo is still unknown. To test the stimulatory effects of a combined estrogen-gestagen regimen in vivo we monitored serum levels of the carboxy-terminal propeptide of human type I procollagen (S-PICP) in a group of 12 osteoporotic women over a 150 week treatment period. Spinal bone mineral content (BMC) increased to a maximum of 5% over pretreatment values around week 90. Serum alkaline phosphatase (S-AP) and serum bone gla protein (S-BGP) both fell from initial values of 220 U/liter and 39 ng/ml, respectively, to 146 U/liter (p less than 0.01) and 27.2 ng/ml (NS) around week 60 and remained reduced over the remaining treatment period. S-PICP also fell from 117 to 68 micrograms/liter at week 60 and 70 micrograms/ml at week 150 (P less than 0.01). This is equal to a reduction to 32 +/- 10% pretreatment levels. The reduction in S-PICP was not significantly different from that of the other two markers of bone formation (S-AP and S-BGP). Thus, provided the metabolic clearance of PICP remains unaltered after hormone replacement therapy, no major stimulation of osteoblastic collagen type I synthesis was demonstrable during estrogen-gestagen treatment in this population of osteoporotic women. The changes in bone markers seen in this study are therefore consistent with an estrogen-mediated reduction in the frequency of remodeling activation. Because of the reduction in bone turnover and methodologic limitations of bone marker assays, however, smaller increases in the amount of bone formed per activation could remain undetectable.

Abstract: BACKGROUND. Plasmin is capable of degrading extracellular matrix components such as collagen in vitro. To evaluate the significance of this for in vivo conditions, we set out to study the effect of streptokinase, which acts by converting plasminogen to plasmin, on the serum concentrations of the amino-terminal propeptide of type III procollagen (PIIINP) and the carboxy-terminal propeptide of type I procollagen (PICP). METHODS AND RESULTS. Twenty-three patients with suspected acute myocardial infarction were included in the study; 17 of them received thrombolytic therapy, and six were treated conservatively. PIIINP and PICP were assayed with radioimmunoassays. Kinetics of creatine kinase-MB release were determined to differentiate reperfusers from nonreperfusers. Composite curves of creatine kinase-MB release were constructed for different patient subgroups. During streptokinase infusion the serum concentrations of PIIINP increased rapidly, with a maximum mean increase of 50% (from 2.2 +/- 0.2 to 3.3 +/- 0.3 micrograms/l) in 45 minutes. A similar increase was also observed in two patients who received thrombolytic therapy but did not subsequently develop any myocardial infarction determined on the basis of enzyme release. The relative increase in PIIINP during streptokinase treatment was higher in those acute myocardial infarction patients with probable reperfusion than those with nonprobable reperfusion. Corresponding changes in PIIINP were not seen in the control group. Two days later there was a second increase in serum PIIINP for both patient groups. This change coincided with a similar increase in PICP. CONCLUSIONS. We conclude that streptokinase, probably by activation of plasminogen to plasmin, stimulates the breakdown of type III collagen during thrombolytic therapy. This phenomenon may decrease the risk of rethrombosis of the affected artery if the exposed collagen is responsible for thrombosis formation, but it could also be involved in the development of hemorrhagic complications during thrombolytic therapy. The second increase in PIIINP levels probably indicates type III collagen synthesis of the infarcted area. This investigation represents a pilot study, and more studies on the effects of various thrombolytic agents on interstitial collagen metabolism are obviously needed.

Abstract: The reproducibility and specificity of a new, rapid, simple RIA for measuring the concentration of the soluble carboxypropeptide of type I procollagen (PICP) in serum was confirmed. Serum PICP was determined in 442 healthy Caucasian subjects ranging in age from 3 wk to 18 y. Highest PICP values (mean +/- SD: 2200 +/- 350 micrograms/L) occurred in infants less than 3 mo of age, falling by 70% at 2 y and by an additional 10% at 4 y. There was no significant change in serum PICP between 4 and 16 y of age (330 +/- 130 micrograms/L), but a decrease to adult levels of less than 160 micrograms/L occurred by 18 y. In 76 children with growth disorders, serum PICP was related to linear growth velocity (p less than 0.001), although there were no significant differences in PICP among the 38 children with growth hormone insufficiency, the 21 short children with no endocrinologic abnormality, or the 17 tall children. All 15 prepubertal children treated with growth hormone for 3 mo showed significant increases in both growth velocity and serum PICP, with a significant relationship (p less than 0.01) between the degree of increases. The rise in serum PICP at 3 mo (but not baseline PICP values) predicted the increase in growth velocity after 1 y of treatment. Similar changes were observed in the concentration of the aminopropeptide of type III procollagen, except that serum aminopropeptide of type III procollagen showed a definite increase during puberty and a wider spread of values in growth disorders. We conclude that measuring serum PICP by the new, reproducible assay reflects height velocity in prepubertal children and may be a useful biochemical means of monitoring growth rates.

Abstract: Analyses of the aminoterminal propeptide of type III procollagen (PIIINP) in the cerebrospinal fluid (CSF) of 55 children and five young adults without any structural central nervous system (CNS) lesion are reported. The concentration was age-dependent, in that infants and small children had quite high values, whereas the concentration remained relatively constant after the age of 1.5 years. The concentrations of PIIINP in the CSF of 44 children with acute lymphoblastic leukemia (ALL) were prospectively determined at the time of diagnosis and during treatment, since deposition of type III collagen is known to occur during fibroproliferative responses triggered by inflammation. Chemical arachnoiditis is known to be associated with intrathecal methotrexate therapy in children with leukemia. The mean concentration in these children at diagnosis (5.8 micrograms/l +/- SD 2.8 micrograms/l) did not differ from that in age-matched controls (6.7 micrograms/l +/- SD 3.2 micrograms/l). Depending on type of the disease, the children were treated according to two different protocols. PIIINP concentrations were significantly higher during the therapy phases which included intrathecally administered methotrexate (P less than 0.001) than at diagnosis of the disease. Corticosteroid treatments were always associated with a significant decrease in PIIINP concentrations (P less than 0.01 and P less than 0.001 in the two groups, respectively), irrespective of the therapy phase. The results suggest that an increase in PIIINP concentration in the CSF of children with ALL is an indicator of a fibroproliferative response in the arachnoid. Corticosteroids may repress this response and possibly also prevent the development of adhesions in the arachnoid.

Abstract: We determined the reference interval for the carboxyterminal propeptide of type I procollagen (PICP), an indicator of the synthesis of type I collagen, in cerebrospinal fluid (CSF) by studying 32 infants and children, ages less than or equal to 15 years. The concentration of PICP is age dependent, with particularly high concentrations occurring in children younger than 1.5 years. In older children the concentration is stable (reference interval 20-92 micrograms/L). We also investigated the possibility that PICP in CSF could reflect local fibroproliferative changes in the arachnoid in a cohort of 42 children with acute lymphoblastic leukemia who were monitored by repeated sampling in connection with intrathecal therapy. Initially, there was no difference in PICP between the children with newly diagnosed leukemia and the controls. PICP concentrations were significantly higher (P less than 0.01) during intrathecal methotrexate therapy, with median values above the reference interval. Continuous corticosteroid treatment was associated with a significant decrease in PICP (P less than 0.02 and P less than 0.01, respectively, in two groups treated according to different protocols), close to the lower limit of the reference interval. Intrathecally administered methotrexate and systemic corticosteroid treatment are known to be associated with the development of arachnoiditis and with general repression of collagen synthesis, respectively. We conclude that PICP in CSF is a sensitive indicator of local fibroproliferation and ongoing collagen synthesis.

Abstract: The concentrations of the amino terminal propeptide of type III procollagen, the 7S domain of type IV collagen, and the fragment P1 of laminin (PIIINP, 7S, and P1 respectively) and the activity of galactosylhydroxylysyl glucosyltransferase (GGT) in serum were evaluated as indicators of disease activity in a cross sectional study of 84 patients with progressive systemic sclerosis. The mean values of PIIINP, P1, and GGT were raised in progressive systemic sclerosis, 19-32% of patients having abnormal values of the various tests. PIIINP, measured with two different assays, and P1 were associated with active, acute, or subacute disease. GGT also correlated positively with some acute phase proteins in the whole group, without a clear association with the course of the disease. Arthritis was associated with increased PIIINP concentrations as well as with an increased activity of GGT. Kidney disease led to raised concentrations of the degradation products of PIIINP. Raynaud's phenomenon in the hands was related to increased PIIINP concentrations.

Abstract: This study investigated the relationships of the serum markers of fibrogenesis and basement membrane formation to the clinical and morphological severity of alcoholic liver disease and to the degree of alcohol abuse. The concentrations of the aminoterminal propeptide of type III collagen, type IV collagen, and laminin were measured from 87 samples representing a wide range of clinical and histological severities of the disease, which were assessed with indices that have been shown to correlate well with the risk of dying within 1 yr. Significant correlations (p less than 0.00001) were found between the markers of connective tissue metabolism and the Combined Clinical and Laboratory Index: (aminoterminal propeptide of type III collagen, rs = 0.82; type IV collagen, rs = 0.82; laminin, rs = 0.81), as well as between these markers and the Combined Morphological Index: (aminoterminal propeptide of type III collagen, rs = 0.70; type IV collagen, rs = 0.68; laminin, rs = 0.64). Whereas the patients with less than 8 mM of alcohol in their morning urine (mild or moderate drinkers) showed a significant (p less than 0.00001) decrease in these markers in a period of 27 +/- 1 wk, the patients with more than 8 mM of urinary alcohol (heavy drinkers) had no improvement. It is proposed that both fibrogenesis and basement membrane formation are associated with disease severity, degree of alcoholic hepatitis, and alcohol intake, which are important determinants of prognosis in alcoholic liver disease.

Abstract: We studied 28 lung carcinomas representing different histological types and three of their regional lymph node metastases immunohistochemically by using specific antibodies against two basement membrane proteins--the 7S domain of type IV collagen and the P1 fragment of laminin. One feature common to all peripherally growing tumors, regardless of the histologic type, was an intact basement membrane between the tumor and the unaffected lung tissue. At these locations, the basement membrane was organized into alveolar structures that did not differ from normal lung tissue. The fibrotic central areas of the tumors did not exhibit this phenomenon. Based on these findings, we believe that malignant tumors of the lung utilize preserved alveolar basement membranes for their local spread. This finding seems to represent a general property of all lung carcinomas, not only adenocarcinomas of the bronchiolo-alveolar type.

Abstract: Serum concentrations of CA 125 and the aminoterminal propeptide of type III procollagen (PIIINP) were measured in 50 patients with clinical stage I or II (N = 16) and stage III or IV (N = 34) ovarian carcinoma before and during cytotoxic chemotherapy. Initially pathological concentrations of CA 125 were found in 92% of all patients and 100% of those in clinical stages II, III and IV. The concentration of PIIINP was at pathological levels in 71% of patients. Serum concentrations of CA 125 (P = 0.04) and PIIINP (P = 0.005) were higher in stages III and IV than in stages I and II. Initial concentration of PIIINP, but not of CA 125, was significantly (P less than or equal to 0.001) higher in the 19 patients who died of the malignancy than in the 31 patients alive at the end of the follow-up period. There was a significant inverse correlation (P = 0.01) between the initial PIIINP values and the survival time among patients with a poor prognosis. Initial concentration of CA 125 was of no prognostic value. During the follow-up, the serum concentrations of CA 125 and PIIINP correlated closely with clinical changes in the disease. Either or both of the tumour markers increased or remained at pathological levels before clinical relapse in patients who had initially responded. PIIINP was a more accurate marker (84%) than CA 125 (63%) in this respect. The information obtained from CA 125 and PIIINP concentrations was identical in 65% and complementary in 33% of the patients.(ABSTRACT TRUNCATED AT 250 WORDS)

Abstract: Type I collagen is the most abundant collagen type in soft tissues and the only type found in mineralized bone. We established a rapid equilibrium radioimmunoassay for the carboxyterminal propeptide of human type I procollagen (PICP), to be used as an indicator of the synthesis of type I collagen. We isolated type I procollagen from the medium of primary cultures of human skin fibroblasts, digested the protein with highly purified bacterial collagenase, and purified PICP by lectin-affinity chromatography, gel filtration, and ion-exchange separation on HPLC. The purity of the protein was verified by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and by N-terminal amino acid sequencing of its component chains. The final radioimmunoassay was established with polyclonal rabbit antibodies. Material antigenically related to PICP is readily detected in human serum. There is only one form of the serum antigen, its molecular size and affinity to the antibodies being similar to those of the isolated propeptide. Intra- and interassay CVs are 3% and 5%, respectively. Preliminary reference intervals for healthy adults (18 to 61 years of age) are 38-202 micrograms/L for men and 50-170 micrograms/L for women: in men the concentration is inversely related to age. The serum antigen is stable during storage and after repeated thawing.

Abstract: The concentration of the aminoterminal propeptide of type III procollagen (PIIINP) in serum as an indicator of tissue repair was studied in 71 surgical patients undergoing minor, moderate abdominal, major abdominal, or hip surgery. An increase in serum PIIINP concentrations took place within the first week, and its magnitude was related to the extent of the soft-tissue operations. After the hip replacement, the maximum PIIINP concentration was reached somewhat later. Very high levels of serum PIIINP were seen in three patients with serious wound infections. The serum PIIINP antigenicity consisted both before and after surgery of equal proportions of two forms, one corresponding to the propeptide as set free during synthesis of new collagen and the other being larger and probably derived from turnover of type III collagen fibers. The proportion of the latter form was accentuated in infection. In contrast, in the wound the form derived from synthesis of type III collagen predominated. These results suggest that the postoperative increase in serum PIIINP levels is partly the result of tissue repair and partly the result of whole-body turnover of type III collagen.

Abstract:

Abstract: Serum CA 125 (a marker of coelomic epithelial cells) and aminoterminal propeptide of type III procollagen (PIIINP; an indicator of collagen metabolism) concentrations were measured in 148 patients with endometrial carcinoma. An initial serum concentration of CA 125 was pathologic in 17% of the patients, the frequency of abnormal values being higher (P = 0.0001) in advanced (63%) than in early disease (10%). The serum PIIINP concentration was increased in 35% of the patients and more often (P less than 0.05) so in advanced (63%) than in early disease (31%). Among all the patients, at least one of the tumor markers was increased in 43% of the cases. In early disease 12 of 108 patients contracted recurrent cancer. The accuracy of the pathologic CA 125 (9%) and PIIINP (18%) concentrations in their prediction was poor. In the total material, pathologic CA 125 and PIIINP concentrations appeared simultaneously in 11 patients, of whom eight had poor prognoses. In monitoring of treatment response of 24 patients, regression was accompanied by normal or decreasing CA 125 and PIIINP values. The persistence of pathologic CA 125 and/or PIIINP concentration predicted relapse of the malignancy. In progressive disease, CA 125 and PIIINP concentrations together or separately remained at a pathologic level or increased continuously. In clinically stable endometrial carcinoma, CA 125 gave false-negative results in 71% of the determinations and PIIINP only in 12%. The current results suggest the use of CA 125 and PIIINP, simultaneously, in monitoring the clinical course of advanced endometrial carcinoma.

Abstract: Sodium chlorate, a potent inhibitor of sulfation reactions, completely inhibits the formation of tyrosine-o-sulfate in type III procollagen in human fibroblasts, when used in concentrations that do not affect the incorporation of radioactive amino acids into protein. The unsulfated type III procollagen is secreted into the medium at a rate comparable, to those of sulfated type III procollagen and type I procollagen, which normally does not undergo sulfation. The enzymatic cleavage of the aminoterminal propeptide of type III procollagen is incomplete in fibroblast cultures, irrespective of the sulfation status of the protein.

Abstract: The fate of the circulating C-terminal propeptide of type I procollagen (PICP) was studied. Trace amounts of 125I-PICP administered intravenously to rats disappeared from the blood with an initial t1/2 of 6.1 min. After 45 min the radioactivity was distributed as follows: liver, 36%; blood, 23%; kidneys, 18%; urine, 20%; spleen, 1%; lungs, 2%; heart, 0.4%. To prevent escape of label from the site of uptake, PICP was labelled with 125I-tyramine cellobiose (125I-TC), which is trapped intralysosomally. With this ligand a serum t1/2 of 8.7 min was recorded, and 70% and 20% was traced in the liver and kidneys respectively. The uptake per liver endothelial cell (LEC) was 1000 times that per parenchymal cell and twice that per Kupffer cell. At 1 h and 6 h after addition of 125I-PICP to cultured LEC, 15% and 45% respectively, had been endocytosed. Only ligands for the mannose receptor could compete with PICP for endocytosis. To study whether the same specificity was operative in vivo, 125I-PICP was injected along with an excess of ovalbumin, which is known to be endocytosed by the mannose receptor of LEC. The serum t1/2 was prolonged from 6 to 16 min, signifying that terminal mannose residues are an important signal for clearance of PICP. In conclusion, these studies show that LEC constitute the main site of uptake of circulating PICP. The uptake is mediated by endocytic receptors which recognize terminal mannose residues.

Abstract: We have previously found the serum concentration of the aminoterminal propeptide of type III procollagen, an indicator of collagen metabolism, to be increased in advanced ovarian cancer. In this study we measured the serum aminoterminal propeptide of type III procollagen concentration in healthy women during the menstrual cycle and in patients with salpingo-oophoritis, leiomyomas, endometriosis, and benign ovarian tumors. The concentration was higher in the luteal phase than that in the follicular phase, suggesting an association of collagen metabolism with ovarian steroid hormones. Severe salpingo-oophoritis increased the serum level of the aminoterminal propeptide of type III procollagen with a decrease to normal during recovery. Elevated values were occasionally seen in endometriosis and leiomyomas. These findings indicate that the aminoterminal propeptide of type III procollagen is a relatively unspecific indicator of ovarian carcinoma.

Abstract: Tissue-binding specificity of the type-3 fimbriae of pathogenic enteric bacteria was determined using frozen sections of human kidney. A wild-type Klebsiella sp. strain and the recombinant strain Escherichia coli HB101(pFK12), both expressing type-3 fimbriae, as well as the purified type-3 fimbriae effectively bound to sites at or adjacent to tubular basement membranes, Bowman's capsule, arterial walls, and the interstitial connective tissue. Bacterial adherence to kidney was decreased after collagenase treatment of the tissue sections. Recombinant strains expressing type-3 fimbriae specifically adhered to type V collagen immobilized on glass slides, whereas other collagens, fibronectin or laminin did not support bacterial adherence. In accordance with these findings, specific binding of purified type-3 fimbriae to immobilized type V collagen was demonstrated. Specific adhesion to type V collagen was also seen with the recombinant strain HB101(pFK52/pDC17), which expresses the mrkD gene of the type-3 fimbrial gene cluster in association with the pap-encoded fimbrial filament of E. coli, showing that the observed binding was mediated by the minor lectin (MrkD) protein of the type-3 fimbrial filament. The interaction is highly dependent on the conformation of type V collagen molecules since type V collagen in solution did not react with the fimbriae. Specific binding to type V collagen was also exhibited by type-3 fimbriate strains of Yersinia and Salmonella, showing that the ability to use type V collagen as tissue target is widespread among enteric bacteria.

Abstract: We studied the ultrastructural distribution of laminin, type IV collagen, and the amino terminal pro-peptide of type III collagen (type III pN-collagen) in normal human lymph nodes. After fixation with freshly prepared 4% paraformaldehyde mixed with 0.1% glutaraldehyde, cryoultramicrotomy proved to preserve the antigenicity of these proteins better than embedding in Lowicryl K4M. Sections were treated with rabbit antibodies against the 7S domain of human type IV collagen, the fragment P1 of human laminin, and the amino terminal pro-peptide of human type III pro-collagen, followed by anti-rabbit IgG conjugated to 10-nm colloidal gold. Laminin and type IV collagen were seen in the basement membrane structures of the blood vessels and in the walls of sinuses. The amorphous material between the collagenous fibers in locations corresponding to reticular fibers also contained laminin and type IV collagen. The amino terminal pro-peptide of type III pro-collagen was present in the collagenous fibers in reticular fibers and in the walls of blood vessels and sinuses. Therefore, a significant number of the type III collagen molecules in these fibers must have retained their amino terminal pro-peptide. These results indicate that the basement membrane proteins laminin and type IV collagen are genuine components of reticular fibers, as suggested earlier by immunohistochemical studies at the light microscopic level.

Abstract: The relationship between serum concentrations of the amino-terminal propeptide of type III procollagen (PIIINP) and growth was assessed in 307 healthy subjects and 82 children with disorders of growth (41 with insufficient growth hormone, 23 with short stature and normal endocrinologic studies, 18 with tall stature) by means of a recently developed, simplified PIIINP radioimmunoassay. The PIIINP value appeared to be related to height velocity; in healthy children of each sex, the pattern of change with age mirrored the shape of the standard height velocity curve; in children with disorders of growth, there was a statistical correlation (p less than 0.001) between PIIINP concentration and height velocity. However, measurement of serum PIIINP alone had no diagnostic value because there was considerable overlap of PIIINP values in children with growth hormone insufficiency, short stature, normal stature, and tall stature. The most appropriate application of PIIINP may be in the monitoring of prepubertal children receiving exogenous growth hormone therapy; in these patients, increases in height velocity were reflected by increases in PIIINP, and early increases in PIIINP may have predictive value.

Abstract: Interaction of the basement-membrane binding O75X adhesin of uropathogenic Escherichia coli with various extracellular matrix proteins was studied. The adhesin showed strong binding to type IV collagen immobilized on microtitre plates, whereas other collagens, laminin and fibronectin, were only weakly recognized. Similarly, specific binding of [125I]-labelled type IV collagen to O75X-positive bacteria was shown. Interaction of the two proteins was also demonstrated by affinity chromatography of the O75X adhesin on immobilized type IV collagen. The adhesin bound strongly to the immobilized N-terminal 7S domain of type IV collagen, and the binding of [125I]-labelled type IV collagen to O75X-positive bacteria was inhibited by the soluble 7S domain. Binding of O75X to type IV collagen and to its 7S domain was specifically inhibited by chloramphenicol but was not affected by periodate or endoglycosidase-H treatment of the glycoproteins. Our results show that the 7S domain of type IV collagen is the basement membrane receptor for the O75X adhesin and suggest an interaction based on protein-protein recognition. Inhibition of the interaction by chloramphenicol favours the supposition that a modified tyrosine is involved in the binding site.

Abstract: To investigate the clinical usefulness of the amino-terminal propeptide of type III procollagen (PIIINP) as an indicator of ovarian cancer behavior, 30 patients with advanced epithelial malignancy were monitored with serial serum PIIINP and CA-125 determinations before and during treatment. Initially, PIIINP and CA-125 concentrations were each separately increased in 87% of the cases and, simultaneously, in 77% of the cases. In monitoring treatment responses, PIIINP and CA-125 were identical in 17 patients (57%), both being good predictors of the clinical behavior of the disease in 16 cases and poor predictors in one case. In 13 patients (43%) they were complementary to each other. In three cases PIIINP alone and in one case CA-125 alone were clinically useful prognosis indicators. During the period of complete clinical response to cytotoxic chemotherapy of 16 patients, the CA-125 concentrations decreased to normal before the clinical disappearance of the tumor in eight cases. PIIINP did so in only two cases, thus correlating more precisely with the presence of malignancy. In second-look laparotomies, PIIINP concentrations correlated with the presence of occult cancer better than those of CA-125. In predicting recurrent malignancy in patients with transient complete response, PIIINP and CA-125 were clinically equal. According to the present data, PIIINP concentrations often give information not obtainable by CA-125, thus being useful in monitoring the clinical behavior of ovarian cancer.

Abstract: Glial cells of mammalian brain transiently produce laminin, an extracellular matrix protein and a potent inducer of neurite outgrowth of cultured neurons. We demonstrate that the glial laminin is a variant with the 200-kDa B chains present and the 400-kDa A chain missing. Immunoprecipitation of cell extracts of purified cultured astrocytes, using antibodies to native laminin, revealed the B chains only, although the same antibody immunoprecipitated both A and B chains from the cell extracts of a basement membrane producing embryonal carcinoma cell line (PYS). This result was further obtained if cell extracts of embryonic mouse brain or the PYS cells were immunoblotted using antibodies to native laminin. Northern analysis of the poly(A)+ RNA from the mouse brain revealed no A chain transcript, although the same probe detected the A chain message in a basement membrane producing F9 cell line. No differences in the sizes of the B chain transcripts in different tissues were seen. Immunostaining of embryonic mouse optic nerve and other areas of the brain, using affinity-purified chain-specific antibodies to the major subunits of laminin, showed that the glial cells expressed the B chains only. It was apparent that the punctate, extracellular deposits of laminin, found in association with neuronal migration and nerve pathway formation in vivo were due to the glial expression of the B chains. The A chain immunoreactivity in the brain was confined to basement membranes.(ABSTRACT TRUNCATED AT 250 WORDS)

Abstract: Orally administered malotilate was studied as a protective antifibrotic agent with respect to experimentally induced pulmonary fibrosis in rats using immunohistochemical methods. Specific antibodies raised in rabbits against the aminoterminal propeptide of human type III procollagen and against two basement membrane proteins, the 7S domain of human type IV collagen and the P1 fragment of human laminin, were used for the immunohistochemical analysis, and the result was confirmed by morphometry. Intraperitoneally injected carbon tetrachloride significantly increased the volume densities of reticulin fibers, type III pN-collagen, type IV collagen, and laminin, whereas treatment with malotilate completely normalized these. Binding of the antibodies to rat antigens was also demonstrated by immunoelectron microscopy in which the collagen fibers with a typical periodic pattern were labeled positively with rabbit antitype III procollagen, whereas the amorphous basement membrane material reacted positively with rabbit antitype IV collagen and antilaminin, indicating good, specific cross-reactivity between these antibodies and the rat antigens. It is concluded that malotilate prevents the accumulation of type III pN-collagen and two basement membrane proteins, type IV collagen and laminin, in experimental pulmonary fibrosis, and can potentially be developed to provide a useful drug for preventing pulmonary fibrosis in humans.

Abstract: Immunohistochemical staining was used to demonstrate basement membrane (BM) laminin and type IV collagen in eight cases of Kaposi's sarcoma (KS). These KS were not associated with AIDS and represented different histological stages of the disease: patch (three cases), plaque (one case), and nodule (four cases). Nine cases of benign angiogenic lesions, five of blood vessel origin, and four of lymphatic vessel origin were also studied. An early event in vascular proliferation at the patch stage of KS was an intersection of dermal collagen bundles and the appearance of granular BM material around this space. With the increase of amount and linear arrangement of BM material, well-defined capillaries were formed. Two types of capillary were found in KS lesions. One showed morphological features of blood capillaries, with a round lumen; thick, continuous BM, and occasional pericytes in the wall. The other included irregularly shaped vessels with thin, often disrupted BMs; thus these capillaries morphologically resembled lymphatic capillaries. BM staining also clearly revealed the vascular nature of the nodular lesions of KS, which were composed of a network of slit-like spaces surrounded by BMs. The solid tumor cell areas were sparse; they were composed of spindle-shaped cells surrounded by thin, interrupted basal laminae. By using antibodies against human laminin and human type IV collagen, it was also possible to demonstrate thin, widely disrupted BMs subendothelially in normal dermal lymphatic capillaries. Typically, the BMs in lymphangiomas and lymphangiectasias were continuous and more clearly defined.

Abstract: A characteristic histological lesion in early primary biliary cirrhosis (PBC) is disruption of the basement membrane around small bile ducts, which at later stages of the disease is followed by fibrosis. To assess the significance of serum basement membrane- and type III procollagen-related antigens in reflecting such processes, we have measured radioimmunologically the concentrations of serum laminin, type IV collagen and the aminoterminal propeptide of type III procollagen in 22 patients with PBC, classified into four stages according to liver histology. The mean laminin concentration in PBC patients was twice that of the healthy control subjects. Increased concentrations were observed in all patients with stage III or IV of the disease and also in 60% (6/10) of the patients, with early stages (I or II). Elevated serum type IV collagen concentrations were found only in four patients, all in the late, fibrotic stages of the disease. The basement membrane protein changes in serum were in accordance with immunohistochemical findings obtained with the antibodies against these proteins. Neither of these serum parameters emerged, however, as a significant predictive factor for survival. The changes in serum aminoterminal propeptide of type III procollagen resembled those in laminin P1. Moreover, the propeptide was also significant as a predictive factor for survival.

Abstract: This is an equilibrium-type radioimmunoassay for the amino-terminal propeptide of type III procollagen (PIIINP), which overcomes the problem of nonparallelism between the standard and human serum samples encountered with earlier assays. Proper selection of antiserum and reaction conditions diminishes interference from degradation products of the propeptide in serum. Because a rapid solid-phase-bound second-antibody step is included, the assay takes only 3 h. The intra-assay and the interassay CVs are both about 5%. In infants and children the concentration of PIIINP in serum closely parallels the growth-velocity curve. For 88 presumably healthy adults, the PIIINP concentration was 1.7-4.2 micrograms/L, about a third that measured with the previously available commercial assay. This is because of lack of inhibition by small Col 1 domain-related degradation products.

Abstract: The changes in the biochemical composition of basement membrane (BM) in slow-twitch (m. soleus, MS) and fast-twitch (m. rectus femoris, MRF) skeletal muscles of rats were studied during aging and life-time endurance training (treadmill running). The concentrations of the 7S domain of type IV collagen and of the P2 fragment of laminin were determined with radioimmunoassays in the muscles of rats aged 1, 2, 4, 10, and 24 months. The concentration of type IV collagen was higher in MS than in MRF and increased significantly with age. At older ages, the concentration tended to be higher in the MS of trained than untrained rats. The concentration of laminin was significantly higher in MRF than in MS, especially in old age. Consequently, the ratio of type IV collagen to laminin was significantly higher in MS than in MRF, the difference increasing with advancing age. The results show that the BMs in slow muscle are much more collagenous than those in fast muscle which contains, on the contrary, a higher concentration of laminin. This difference seems only partly to be explained by the higher capillarization of slow twitch muscle fibers. Aging and training seem further to distinguish the composition of the BM in slow and fast muscle.

Abstract: The expression of laminin in embryonic kidneys growing in ovo is followed with mouse-specific, affinity-purified antibodies against the laminin A and B chains. In mouse kidneys growing on the chicken chorioallantoic membrane, the epithelium and nephrogenic mesenchyme are derived from mouse and the vasculature from chicken chorioallantoic vessels. Hence, with the mouse-specific antibodies, it is possible to analyze the deposition of laminin chains by the nephrogenic tissue, because laminin derived from the chicken vasculature remains unstained. In these chimeras, only the laminin B chain, but not the A chain, is expressed in the undifferentiated nephrogenic mesenchyme. The basement membrane around the ureter bud is labeled by the antibodies against both laminin A and B chains. In the mesenchyme, the laminin A chain appears when the mesenchyme converts into tubules. The results suggest that the laminin A and B chains are synthesized differentially in the embryonic nephrogenic tissue.

Abstract: In ovarian carcinoma, elevated serum concentrations of the aminoterminal propeptide of type-III procollagen are related to the clinical stage of the disease and to the presence of ascites, which contains very high concentrations of the propeptide. In a follow-up of patients with advanced disease, favorable clinical course was associated with normalizing propeptide values, whereas in progressing disease the values increased several weeks before clinical progression. In stable disease the concentrations were constantly above the reference range. Laminin and type-IV-collagen-related serum antigens were mostly within the reference range.

Abstract: The serum levels of aminoterminal type III procollagen peptide (S-PIIINP), immunoreactive prolyl 4-hydroxylase protein (S-IRPH), 7S domain of collagen type IV (S-Col IV, 7S), and fragment P1 of laminin (S-Lam), which are associated with the metabolism of extracellular interstitial collagens and basement membranes, were measured sequentially for two years in 14 rheumatoid arthritis (RA) patients undergoing disease modifying antirheumatic drug treatment. Elevated S-PIIINP, S-IRPH, and S-Col IV, 7S levels were demonstrated in active RA. In active disease the metabolites showed some correlation with clinical and serological signs of disease activity. A high average synovial fluid/serum concentration ratio of PIIINP and of Col IV, 7S supports the concept that the increased serum levels of PIIINP and Col IV, 7S originated from the diseased joints. After 2 years of treatment a decline was observed in S-PIIINP and S-Col IV, 7S in treatment responders. However, the median levels of S-PIIINP and S-IRPH were still above the upper limit of normal, suggesting smouldering, subclinical inflammatory processes. S-Lam remained within the normal range in active and inactive disease.

Abstract: Twenty-four psoriatic patients on methotrexate were studied with liver biopsies and serum measurements of aminoterminal propeptide of type III procollagen (PIII NP). All but one of nine patients with serum levels of PIII NP above the normal range had liver fibrosis or cirrhosis and no normal liver biopsies were obtained in this group. In contrast, nine normal liver biopsies and two biopsies with minimal fibrosis were found among the 15 patients with normal serum levels of PIII NP. The study indicates that aminoterminal propeptide of type III procollagen can be utilized as a valuable non-invasive marker of fibrogenesis in the liver. This analysis is not specific for the liver, but it seems that the number of liver biopsies probably can be reduced in psoriatics on methotrexate who have normal levels of PIII NP.

Abstract: The cellular heterogeneity of cutaneous tumors from nine patients with type 1 (von Recklinghausen's) neurofibromatosis was studied using several antigenic markers with special reference to focal heterotopic differentiation and interindividual variation. Furthermore, cells which actively express the genes for type I and III collagens and fibronectin, the major components of the abundant extracellular matrix of neurofibromas, were localized using in situ hybridizations. In eight of nine cases, the S-100 protein positive cells, i.e. Schwann-like cells, composed 60 to 80% of the total cell population. However, in one case, only about 40% of the cells were S-100 protein positive. The latter tumor was studied with respect to perineurial cell differentiation and stained with a mixture of two antibodies, directed against the S-100 protein and type IV collagen. In Schwann cells, the staining reaction for S-100 protein was observed in the nuclear region, whereas the staining reaction for type IV collagen was located peripherally, corresponding to the basement membrane zone covering the cells. The stromal cells which showed only the peripheral staining profile were considered to be neoplastic perineurial cells. Distinct structures with epithelial, endothelial, or smooth muscle cell differentiation were present within the benign tumors, as detected by immunostaining for cytokeratin, epithelial membrane antigen, factor VIII-related antigen and desmin, respectively. In situ hybridizations revealed a clearly detectable expression of type I procollagen genes in less than 10% and type III procollagen gene in less than 5% of the total cell population. Active synthesis of fibronectin was limited to the vascular walls, when examined by in situ hybridization, and antibodies to cellular fibronectin localized to the same areas. However, antibodies to plasma fibronectin produced a uniform staining reaction throughout the tumors suggesting that most of the fibronectin in neurofibromas is plasma-derived. The latter observation suggests that neurofibroma cells are freely accessible to various plasma proteins, including growth factors, which may influence the growth characteristics of these lesions.

Abstract: The circulating concentrations of insulin-like growth factor I (IGF-I) and immunoreactive aminoterminal propeptide of type III procollagen (PIIINP) were measured in 12 children with short stature (8 GH deficient and 4 non-GH deficient) before and after 1 week, 5 weeks, and 3, 6, and 12 months of treatment with biosynthetic hGH. Seven children had a growth response (increase in relative growth velocity greater than 1.5 SD during the initial 6 months) to GH therapy (responders), whereas 5 failed to respond (nonresponders). No relationship was found between the pretreatment plasma IGF-I levels or their changes during therapy and the growth response. Serum PIIINP levels increased considerably in all but 3 children, after as little as 1 week of GH administration. After 5 weeks, all responders had an increase in their serum PIIINP concentrations of 40% or more, whereas the nonresponders had less or no increments. There was a close correlation between the GH-induced increase in serum PIIINP levels at 5 weeks and growth velocity after 6 months of GH therapy (r = 0.77; P less than 0.01). The correlation was even stronger with the growth velocity at 12 months (r = 0.83; P less than 0.001). The serum PIIINP response to short-term GH administration could be an early predictor of the growth response to long-term GH therapy. In contrast to plasma IGF-I, the PIIINP response may be useful both in GH deficient and non-GH deficient children.

Abstract: Serum concentrations of the aminoterminal propeptide of type III procollagen and of the 7S domain of type IV collagen, presumed to reflect fibrotic activity in liver tissue, and of the glycosamonoglycan hyaluronan, were obtained from 40 alcohol abusers, at the time of liver biopsy. The serological results were related to morphological findings in liver tissue, i.e. no fibrosis, fibrosis without cirrhosis, micronodular cirrhosis and macronodular cirrhosis, and to ultrastructural indications of perisinusoidal fibrosis in the acinar zone 3. All patients with fibrosis and cirrhosis on light microscopy had elevated serum levels of the type III procollagen peptide as well as of the 7S domain of type IV collagen. However, due to a considerable overlap between the groups, no relations could be demonstrated to the severity of the fibrosis, supporting the assumption that these serological markers reflect the current fibrotic activity and not the amount of fibrotic tissue previously deposited. Among patients without fibrosis on light microscopy, a relation between the propeptide levels and ultrastructural perisinusoidal zone 3 fibrosis was observed, suggesting that type III procollagen peptide may be valuable in detecting very early liver fibrosis. A positive correlation was demonstrated between the serum concentrations of type III procollagen peptide and hyaluronan. As hyaluronan is degraded in the liver endothelial cells, it is suggested that the liver is involved, not only in the synthesis, but also in the degradation of the propeptide.

Abstract: For quantitative analysis of wound healing in surgical patients, samples of wound fluid were collected through a silicone rubber tube and their concentration of the aminoterminal propeptide of type III procollagen was measured with a specific radioimmunoassay. Peritoneal fluid, collected through an abdominal drain, and serum were also analyzed. At day 1 after operation, the mean concentration of the propeptide was 30 times higher than the mean preoperative serum level (2.5 micrograms/L). A significant increase (p less than 0.001) occurred at day 3 in the wound and at day 2 in peritoneal fluid. At day 5 the mean wound concentration (2670 micrograms/L) was 1000 times higher than the serum level. In serum a small but significant increase (p less than 0.05) was found at days 5 and 30. The increase in wound fluid resulted from the intact, liberated propeptide, indicating that the results reflect the synthesis of type III collagen deposited in the wound. This procedure offers a quantitative tool for wound healing studies. Other extracellular matrix components can also be measured, the sequential pattern of their appearance can thus be assessed, and disturbances and treatment effects in wound healing can be detected.

Abstract: The turnover of type III collagen, a major constituent of the myometrium and the uterine cervix, during pregnancy was evaluated by monitoring serum antigens related to the aminoterminal propeptide of type III procollagen. Their concentration increased markedly towards term in most uncomplicated pregnancies, while their size distribution throughout the pregnancy resembled that seen in the sera of normal healthy persons. In some patients, however, the level remained low, indicating interindividual variation in the release into serum and metabolism of the propeptide. There were no distinct changes during or immediately after vaginal delivery. Values exceeding the reference range for uncomplicated pregnancies were found during weeks 28-37 in patients with pre-eclampsia, essential hypertension, intrahepatic cholestasis of pregnancy or twin pregnancy. Thus, pregnancy should be taken into account when evaluating results of the serum assay for the aminopropeptide and the use of this assay as an indicator of pregnancy complications warrants further study.