Joseph W Carlson

Abstract: In recent years, evidence is accumulating that cancer cells develop strategies to escape immune recognition. HLA class I HC down-regulation is one of the most investigated. In addition, different HLA haplotypes are known to correlate to both risk of acquiring diseases and also prognosis in survival of disease or cancer. We have previously shown that patients with serous adenocarcinoma of the ovary in advanced surgical stage disease have a particularly poor prognosis if they carry the HLA-A02* genotype. We aimed to study the relationship between HLA-A02* genotype in these patients and the subsequent HLA class I HC protein product defects in the tumour tissue.

Abstract: Human papillomavirus (HPV) infection is the major cause of cervical cancer, but the prevalence of different HPV types varies depending on geographical location and may change dramatically after introduction of HPV vaccination. Here, we aimed to gain some information regarding the recent prevalence of different HPV types, in cancer of the uterine cervix in the Stockholm region, before the introduction of public HPV vaccination in Sweden.

Abstract: Serous tubal intraepithelial carcinoma (STIC) is detected in between 5% and 7% of women undergoing risk-reduction salpingooophorectomy for mutations in the BRCA1 or 2 genes (BRCA+), and seems to play a role in the pathogenesis of many ovarian and "primary peritoneal" serous carcinomas. The recognition of STIC is germane to the management of BRCA+ women; however, the diagnostic reproducibility of STIC is unknown. Twenty-one cases were selected and classified as STIC or benign, using both hematoxylin and eosin and immunohistochemical stains for p53 and MIB-1. Digital images of 30 hematoxylin and eosin-stained STICs (n=14) or benign tubal epithelium (n=16) were photographed and randomized for blind digital review in a Powerpoint format by 6 experienced gynecologic pathologists and 6 pathology trainees. A generalized kappa statistic for multiple raters was calculated for all groups. For all reviewers, the kappa was 0.333, indicating poor reproducibility; kappa was 0.453 for the experienced gynecologic pathologists (fair-to-good reproducibility), and kappa=0.253 for the pathology residents (poor reproducibility). In the experienced group, 3 of 14 STICs were diagnosed by all 6 reviewers, and 9 of 14 by a majority of the reviewers. These results show that interobserver concordance in the recognition of STIC in high-quality digital images is at best fair-to-good for even experienced gynecologic pathologists, and a proportion cannot be consistently identified even among experienced observers. In view of these findings, a diagnosis of STIC should be corroborated by a second pathologist, if feasible.

Abstract: Pathology provides a critical bridge between the patients, their physicians and the therapeutic and surgical interventions that can be provided to them. Clinicians caring for patients in resource poor settings may provide basic healthcare, which does not include access to pathologic services; however, the value of pathology in alleviating health disparities for underserved patients is substantial when implemented. Partners in health is a comprehensive, community-based healthcare organization with clinics in 7 countries-most among the poorest in the world-which has the ability to obtain surgical biopsies and, if a pathologic diagnosis can be rendered, provide treatment, and long-term follow-up. Over the past 5 years, pathologists from the Brigham and Women's Hospital have collaborated with clinicians from partners in health to meet this need which included 129 cases from Haiti and Rwanda and a range of pathology: 64 malignancies, 28 normal tissue or nondiagnostic specimens, 16 infectious or inflammatory cases, 8 benign lesions, and other rare entities. Providing pathology services in resource poor settings through collaboration with clinicians working on-site is only hindered by the establishment of a working collaboration; however, the benefits are enormous and include patient access to curative or tailored therapies, logistical management of treatment resources, and exposure of pathologists to unique and challenging cases.

Abstract: SUMMARY: Most serous adenocarcinomas involving both the endometrium and ovary are presumed to arise in the endometrium. Recently, serous tubal intraepithelial carcinoma (STIC) has been implicated in the pathogenesis of pelvic serous carcinoma. This study explored the potential relationship between STIC and uterine serous carcinoma. Twenty-two consecutive cases of serous carcinoma involving the endometrium were studied. In each case, fallopian tubes were submitted in toto according to the protocol for sectioning and extensive examination of the fimbriated end. Extent of the endometrial tumor and presence/absence of STIC were documented. Immunostaining for p53 and Wilms tumor-1 was performed on all cases with STIC. p53 mutation analysis was performed in a subset of matched STICs and endometrial tumors. Eleven cases showed concurrent endometrial and adnexal involvement, including 6 with endosalpingeal involvement; STIC was confirmed in 5. In all 5, the concurrent endometrial tumor was either noninvasive, or exhibited only superficial (<5%) myometrial invasion. In 2 cases, identical p53 mutations were shared by both tubal and endometrial lesions. This study shows that noninvasive, genetically related serous carcinomas may coexist in both tube and endometrium. As management of serous neoplasms is predicated on site of origin, we propose that the sectioning and extensively examining the fimbria protocol be applied to all endometrial serous carcinomas and that tumors with concurrent STIC be classified as a distinct subset of pelvic serous carcinomas pending a clearer understanding of tumor origin.

Abstract: AIMS: Endometrial intraepithelial neoplasia (EIN) is a monoclonal precursor to endometrioid endometrial adenocarcinoma characterized by a geographic cluster of crowded glands with epithelial cytology altered relative to the background. It may demonstrate epithelial metaplastic changes, or arise within polyps, but the frequencies of these features as encountered in practice is unknown. The aim was to report the epithelial differentiation state and polyp context of 83 sequential EIN lesions diagnosed over a 2-year period. METHODS AND RESULTS: EIN is a rare lesion, seen in only 1.4% of endometrial biopsy specimens in a busy hospital-based practice. Of 83 EIN cases, 39 contained metaplastic changes (18% squamous morular, 14% tubal secretory and 5% each of secretory, mucinous or ciliated change). Endometrial polyps were more likely (odds ratio 5.2, P < 0.001) to occur in the endometrial biopsy specimens of women with EIN lesions (43.3%), compared with the background polyp rate (12.9%) of comparable specimens from the same patient population. CONCLUSIONS: Non-endometrioid differentiation and occurrence within polyps are frequent presentations of EIN lesions. Possible mechanisms of polyp association with EIN include: non-shedding of polyp tissue creating a shelter for persistence of pre-existing neoplastic glands, or promotion of premalignant glandular clones by unique polyp stroma.

Abstract: In the past 50 years, the concept of serous ovarian cancer has been progressively refined, with the distinction of the borderline serous tumour, identification of a smaller subset of well-differentiated serous malignancies and, recently, closer attention to the pathogenesis of high-grade serous malignancies. High-grade serous carcinoma, traditionally presumed to arise within Müllerian inclusion cysts of the ovarian surface, cortex and peritoneum, has recently been linked to the distal fallopian tube. This review addresses the disparate forms of serous neoplasia, which reflect both different genetic abnormalities and stages of differentiation of Müllerian epithelium. The significance of these different origins is addressed in the context of ovarian cancer prevention.

Abstract: PURPOSE: A diagnosis of primary peritoneal serous carcinoma (PPSC) requires exclusion of a source in other reproductive organs. Serous tubal intraepithelial carcinoma (STIC; stage 0) has been described in asymptomatic women with BRCA mutations and linked to a serous cancer precursor in the fimbria. This study examined the frequency of STIC in PPSC and its clinical outcome in BRCA-positive women. PATIENTS AND METHODS: Presence or absence of STIC was recorded in consecutive cases meeting the 2001 WHO criteria for PPSC, including 26 patients with nonuniform sampling of the fallopian tubes (group 1) and 19 patients with complete tubal examination (group 2; sectioning and extensively examining the fimbriated end, or SEE-FIM protocol). In selected cases, STIC or its putative precursor and the peritoneal tumor were analyzed for p53 mutations (exons 1 to 11). Outcome of STIC was ascertained by literature review. RESULT: Thirteen (50%) of 26 PPSCs in group 1 involved the endosalpinx, with nine STICs (35%). Fifteen (79%) of 19 cases in group 2 contained endosalpingeal involvement, with nine STICs (47%). STIC was typically fimbrial and unifocal, with variable invasion of the tubal wall. In five of five cases, the peritoneal and tubal lesion shared an identical p53 mutation. Of 10 reported STICs in BRCA-positive women, all patients were without disease on follow-up. CONCLUSION: The fimbria is the source of nearly one half of PPSCs, suggesting serous malignancy originates in the tubal mucosa but grows preferentially at a remote peritoneal site. The generally low risk of recurrence in stage 0 (STIC) disease further underscores STIC as a possible target for early serous cancer detection and prevention.

Abstract: AIMS: Small cell carcinoma of the ovary, hypercalcaemic-type (SCCOH) is morphologically similar to small cell carcinomas from other sites. The aims of this study were to (i) determine if a biomarker panel would distinguish small cell carcinomas of the ovary, cervix (SCCCx) and lung (SCCLu) and (ii) potentially determine the histogenesis of SCCOH. METHODS AND RESULTS: Nine ovarian small cell carcinomas (seven hypercalcaemic type; two pulmonary type), eight SCCCx and 22 SCCLu were immunostained for thyroid transcription factor (TTF)-1, WT-1, p16, cKIT and OCT3/4; a subset of cases were tested for human papillomavirus (HPV). WT-1 was diffusely positive in 6/7 SSCOH versus two of 33 other small cell carcinomas (P <or= 0.001). TTF-1 was diffusely positive in 20/22 SCCLu and 1/8 SCCCx, and negative in all SCCOH. p16 and cKIT demonstrated variable patterns of immunoreactivity in all cases. HPV was identified in 5/6 SCCCx; SCCOH and SCCLu were negative for HPV. CONCLUSIONS: Combined staining with WT-1 and TTF-1 will distinguish SCCOH from SCCLu and SCCCx with a sensitivity of 86% and specificity of 97%. HPV is specific for tumours of cervical origin, but p16 immunohistochemistry is not useful for this purpose. The presence of diffuse WT-1 supports a Müllerian origin for SCCOH, whereas the absence of cKIT and OCT3/4 argues against a germ cell origin.

Abstract: Magnetic resonance imaging (MRI) has an evolving role in the evaluation of breast lesions and is currently being used for the screening of high-risk patients (eg, women with a personal or family history of breast cancer), for the evaluation of extent of disease in patients with a current diagnosis of cancer, and for patients with suspicious, but indeterminate, findings by other imaging modalities. If a suspicious lesion detected by MRI is not well visualized by another method, an MRI-directed core biopsy or breast excision may be performed. MRI cannot be used to verify the lesion in the specimen because MRI lesion detection is dependent on uptake of gadolinium after intravenous injection. Accordingly, these breast excisions present unique challenges to pathologists. The purpose of this report is to define the surgical pathology issues involved in processing MRI-localized excisions. Retrospective review of 85 consecutive MRI-directed breast excisions from 77 patients was performed. Malignant lesions were present in 20 (24%) of 85 excisions, including 10 cases of invasive carcinoma (median size, 0.4 cm), 9 cases of ductal carcinoma in situ, and 1 case of lymphoma. Most of the malignancies (85% or 17/20) had no associated gross finding and only 5 (25%) of 20 of these malignancies were associated with a definite finding on the specimen radiograph. This study demonstrates that gross examination and specimen radiography do not identify most of the malignancies in MRI-localized biopsies and, therefore, optimal processing requires complete microscopic examination of these specimens.

Abstract: BACKGROUND: Breast magnetic resonance imaging (MRI) has been implemented as a screening tool for early detection and as a diagnostic test in the management of breast cancer. Lesions identified by MRI but not amenable to conventional biopsy techniques require MRI wire-guided excisional biopsy (MRIbx). We hypothesized that more tissue would be resected with MRIbx compared to Mammobx. We also sought to evaluate factors that might predict the presence of breast cancer in patients undergoing MRIbx. METHODS: We reviewed consecutive cases of breast MRIbx from 2004 to 2006 performed by seven surgeons. MRI was performed in patients with either a synchronous breast cancer or significant risk factors. Lesions visualized only by MRI underwent diagnostic MRIbx. The control group was comprised of consecutive cases that underwent diagnostic Mammobx during the same time period. The volumes of tissue resected, overall and by pathologic outcome, were compared. RESULTS: Seventy-one patients, with a mean age of 48, underwent MRIbx. Eleven (15.5%) invasive breast cancers and eight cases (11.3%) of ductal carcinoma in situ (DCIS) were identified. The median volume of tissue resected was significantly greater than that in the Mammobx group (28.8 cm3 vs 21.1 cm3, respectively). DCIS-containing MRIbx specimens were significantly larger than benign or invasive cancer-containing specimens. There was no significant overall association between either the indication for MRIbx or the size of the MRI lesion and the frequency of cancer. CONCLUSIONS: In patients undergoing breast MRIbx, 27% were found to have DCIS or invasive breast cancer. MRIbx was associated with significantly larger specimen volumes than Mammobx.

Abstract: AIMS: Nuclear staining for beta-catenin by immunohistochemistry is being used increasingly to diagnose desmoid tumours (deep fibromatoses), especially where the differential diagnosis includes other abdominal spindle cell neoplasms. This study aimed to define the prevalence of beta-catenin positivity in desmoid tumours and other morphologically similar spindle cell neoplasms. METHOD AND RESULTS: Nuclear beta-catenin expression was evaluated by immunohistochemistry in 270 soft tissue tumours. Nuclear immunopositivity was detected in 80% of cases of sporadic desmoid fibromatosis (24/30) and in 67% of tumours in patients with familial adenomatous polyposis (8/12). Nuclear positivity was also present in 14/25 superficial fibromatoses (56%), 3/10 low-grade myofibroblastic sarcomas (30%), 5/23 solitary fibrous tumours (22%), 1/5 infantile fibrosarcomas (20%), 1/18 desmoplastic fibroblastomas (6%) and 1/21 gastrointestinal stromal tumours (5%). No nuclear immunoreactivity was present in neurofibromas (0/26), schwannomas (0/25), nodular fasciitis (0/19), leiomyosarcomas (0/16), inflammatory myofibroblastic tumours (0/12), fibromas of tendon sheath (0/9), lipofibromatoses (0/5), Gardner fibromas (0/4), calcifying aponeurotic fibromas (0/4) or fibromatosis colli (0/1). CONCLUSION: Nuclear staining for beta-catenin is supportive, but not definitive, of the diagnosis of desmoid fibromatosis. No significant difference in immunoreactivity was observed between sporadic and familial desmoid fibromatoses. beta-Catenin negativity does not preclude the diagnosis of fibromatosis.

Abstract: A nanolithography method for altering the composition of supported phospholipid bilayers has been developed. Although a number of lithography procedures for various chemical systems currently exist, these methods typically rely on covalent bonding to stabilize the resulting structure. The method described here is suitable for an entirely noncovalent system, where the pattern is maintained by specific hydrophobic and other noncovalent interactions. This lithography is achieved by the atomic force microscope, which is used to insert phospholipid molecules into a supported reconstituted high-density lipoprotein−phospholipid bilayer with 20 nm line widths. Lipid molecules are delivered to the supported bilayer through transient structural disruption, forcing the acceptance of new lipid monomers into the bilayer structure. Precoating the atomic force microscope tip with specific lipids is sufficient to deliver new lipid to the patterned areas. These results demonstrate a new method for controlling the structure of highly dynamic noncovalent surface assemblies.

Abstract: Atomic force microscopy can be used to determine the vertical dimension of biological molecules under native conditions with high resolution. Deformation of soft proteins by the scanning force, however, introduces error in the magnitude of the measurement. In this work, the force-dependent height of NADPH-cytochrome P450 reductase, an integral membrane protein, was measured by atomic force microscopy and analyzed to account for the contribution of deformation to the observed height of the molecule above a model membrane surface. Imaging of single reductase molecules was accomplished by reconstitution into 10 nm diameter phospholipid bilayer particles, which provides a way of adsorbing the protein−phospholipid complex on a surface in the proper orientation. The results show that the height of the reductase is drastically underestimated in contact imaging mode. An analysis of force curves taken on single reductase molecules provides a height that better matches the known dimensions of the protein. This technique should be generally useful for determining the vertical dimension of biological samples that are severely deformed by contact imaging forces.

Abstract: One of the primary difficulties of imaging under aqueous solution with the scanning force microscope (SFM) is a lack of reliable tip characterization. The requirements of imaging under solution greatly limit the utility of available standards. We report the synthesis of metal binding colloidal gold (MBG) which allows direct deconvolution of tip shape during sample imaging. Divalent metal ions have been used extensively for binding samples to mica for SFM in fluid, especially biological samples such as DNA. The MBG synthesized here can be imaged bound to mica, demonstrating their usefulness for fluid SFM. Their diameter has been measured and is consistent with the expected value. They are shown to be robust to repeated imaging and stable in a variety of conditions. Their ability to be imaged is inhibited by the presence of a metal chelator. Finally, these gold particles, in conjunction with computational image restoration methods, have been used to analyze plasmid DNA images obtained in aqueous solution. Enabling colloidal gold particles to bind metals through the attachment of a synthesized polypeptide allows simpler and faster tip calibration in aqueous solution than calibrations obtained by previously described methods.

Abstract: A phospholipid bilayer of nanometer dimension has been used as a support for the study of reconstituted functional single-membrane proteins. This nanobilayer consists of an approximately 10-nm-diameter circular phospholipid domain stabilized by apolipoprotein A1. As a demonstration of this methodology, we formed the nanobilayers in the presence of hepatic microsomal NADPH-cytochrome P450 reductase. Incubation of a solution of enzyme-containing nanobilayers with a freshly cleaved mica substrate resulted in the spontaneous formation of a fully oriented supported monolayer of discoidal phospholipid domains. The P450-reductase in the oriented monolayer retains its catalytic activity. Characterization by scanning force microscopy revealed isolated single-membrane proteins that could be stably imaged over time. These results define a novel technique for the study of single-membrane proteins in a bilayer environment.

Abstract: The atomic force microscope (AFM) has been used to image a variety of biological systems, but has rarely been applied to soluble protein-lipid complexes. One of the primary physiological protein-lipid complexes is the high-density lipoproteins (HDL), responsible for the transport of cholesterol from the peripheral tissues and other lipoproteins to the liver. We have used the AFM to directly image discoidal reconstituted HDL (rHDL) particles for the first time. The height of these particles is consistent with a phospholipid bilayer structure, but careful high resolution measurements of particle diameters has indicated that they fuse when adsorbed to mica. Furthermore, it has been demonstrated that the AFM can be used to initiate this bilayer fusion in a controlled manner, allowing the fabrication of stabilized, nanometer scale, phospholipid bilayer "domains."

Abstract: Continuous oscillation on the 2P3/2 → 4I11/2 transition of Nd3+ in a f luorozirconate glass (ZBLAN) fiber at room temperature has been observed. When pumped at ~590 nm, a Nd:ZBLAN f iber 39 cm in length lases in the violet at 412 nm and produces ~0.5 mW of power for 320 mW of pump power and a cavity output coupling of 0.4%. The breadth of the laser’s excitation spectrum is ~12 nm (581–593 nm).

Abstract: Lasing in the ultraviolet has been observed in neodymium-doped fluorozirconate glass (ZBLAN) fibres at room temperature. For a cavity output coupling of < 0.1%, 74 µW of power was obtained at 381 nm (4D3/2 --> 4I11/2 transition of Nd3+) when a 45 cm long fibre was pumped with 275 mW at a wavelength of nominally 590 nm.

Abstract: New approaches to care of patients with these tumors, specifically interdisciplinary diagnosis and treatment and a team approach to more holistic care, are reflected in the editors' and contributors' approach here. Each of these cases is therefore represented and commented upon by a team of specialists and includes a range of diagnostics and imagery. Contributors cover tumors of the ovary with cases of an optimally resected stage III ovarian cancer, suboptimally debulked ovarian cancer, platinum-sensitive recurrent ovarian cancer, young woman with ovarian cancer, ovarian germ cell tumors, low malignant potential tumors of the ovary, sex-cord stromal ovarian tumors, ovarian metastasis from a solid tumor, and hereditary ovarian cancer. Cancers of the uterine corpus include early uterine cancer, advanced endometrial cancer, metastatic endometrial cancer, uterine sarcomas, and gestational trophoblastic disease, while those on cervical cancer include early, metastatic are rare histologies. Also includes cases of vaginal and vulvar cancers.

Abstract: Nanostructured materials is one of the hottest and fastest growing areas in today's materials science field, along with the related field of solid state physics. Nanostructured materials and their based technologies have opened up exciting new possibilites for future applications in a number of areas including aerospace, automotive, x-ray technology, batteries, sensors, color imaging, printing, computer chips, medical implants, pharmacy, and cosmetics. The ability to change properties on the atomic level promises a revolution in many realms of science and technology. Thus, this book details the high level of activity and significant findings are available for those involved in research and development in the field. It also covers industrial findings and corporate support. This five-volume set summarizes fundamentals of nano-science in a comprehensive way. The contributors enlisted by the editor are at elite institutions worldwide.

Abstract: Molecular complexes play a number of key roles in medicine and biology. Understanding how these complexes behave at interfaces, as well as controlling their structure, is of fundamental interest to science and technology. Lipoproteins, a complex of protein and lipid, serve as the primary transporter of cholesterol and other lipids throughout the body. There is a direct correlation between the presence of certain lipoprotein species and the incidence of coronary artery disease. High density lipoproteins are responsible for the removal of cholesterol from the peripheral tissues and, as such, lead to a reduction in the risk of heart disease. Reconstituted high density lipoproteins have been studied with the atomic force microscope. This microscope has revealed new insight into the structure of these supramolecular aggregates and their interfacial behavior. In order to allow accurate quantitation of lateral size a new imaging standard was developed. This standard, peptide modified colloidal gold, has been used in solution to solve tip induced artifacts and to correct for tip changes simultaneous with sample imaging. A spontaneous fusion process has been observed and examined, revealing the important structural role of apolipoprotein A-I , even in an interfacial setting. These discoidal rHDL have been found to generate a new form of supported bilayer structure which has been particulary amenable to atomic force microscope study and manipulation. The microscope has revealed the bilayer structure of these dises through nanodissection. It has also been used to control the spontaneous fusion process of these dises, resulting in spatially defined bilayer domains of nanometer scale dimensions. By using a source of exogenous lipid in solution the AFM has been used to induce lipid exchange with these surfaces, resulting in a compositional change of the supported bilayer in a spatially controlled manner. A model of this process has been proposed based on defects introduced by the AFM tip binding vesicles, which subsequently fuse with the surface. Finally, the AFM has been used to direct proteins to spatially selected regions, allowing the fabrication of heterogeneous membrane protein containing surfaces. In separate studies, myoglobin has been used to study the fluorescence lifetime behavior of oriented biomolecular surfaces. These studies have revealed differences in lifetime between differentially oriented myoglobin molecules, consistent with differential accessibility of the heme pocket to quenching by oxygen. A computational study of the dependence of linear dichroism on the distribution of protein orientation angle has revealed a constrained degenerate solution, which has impact on ail attempts to describe protein orientation with this technique.

Abstract: