Abstract: Active avoidance by tumor cells from attack and elimination by immune cells is an emerging cancer hallmark that is achieved primarily through decreasing the levels of major histocompatibility complex class I (MHC-I) at the cancer cells' surface. Deficiencies in MHC-I antigen-restricted immunosurveillance may be intertwined with an altered, Warburg-like cancer cell-intrinsic metabolism, another emerging hallmark of cancer that involves a switch from mitochondrial respiration to glycolysis to efficiently support large-scale biosynthetic programs that are required for active cell proliferation. We recently envisioned that intervention strategies aimed at reversing the bioenergetic signature of cancer cells (e.g., the antidiabetic biguanide metformin) should correct oncogene (e.g., HER2)-driven MHC-I defects, thus preventing immune escape of oncogene transformants. First, we explored how metformin treatment impacted mitochondrial biogenesis in cultured breast cancer cells overexpressing the membrane tyrosine kinase receptor HER2, the best-characterized downregulator of MHC-I. Metformin exposure was found to dose-dependently increase the expression levels of cytochrome c oxidase I and mitochondrial succinate dehydrogenase, which are encoded by mitochondrial and nuclear DNA, respectively. Second, we explored whether metformin-enhanced mitochondrial biogenesis might significantly alter the MHC-I status in breast carcinoma cells. MHC-I expression, as assessed by flow cytometry using an anti-HLA-ABC monoclonal antibody, was fully restored (up to ∼25-fold upregulation) in MHC-I-negative HER2 gene-amplified carcinoma cells. These findings may help delineate a previously unrecognized mechanism through which metformin (and metformin-like drugs) may enable a cancer patient's own immune system to mount an efficient anti-metastasis response that can prevent or delay disease recurrence. Restored antigenicity and immunogenicity of tumor cells may represent a previously unrecognized primary mode of action underlying the cancer-preventive effects of metformin.
Abstract: Dietary polyphenols may exert their pharmacological effect via synergistic interactions with multiple targets. Putative effects of polyphenols in the management of obesity should be primarily evaluated in adipose tissue and consequently in well-documented cell model. We used Hibiscus sabdariffa (HS), a widely recognised medicinal plant, as a source of polyphenols with a number of salutary effects previously reported. We present here the full characterisation of bioactive components of HS aqueous extracts and document their effects in a model of adipogenesis from 3T3-L1 cells and in hypertrophic and insulin-resistant adipocytes. Aqueous extracts were up to 100 times more efficient in inhibiting triglyceride accumulation when devoid of fibre and polysaccharides. Significant differences were also observed in reactive oxygen species generation and adipokine secretion. We also found that, when polyphenols were fractionated and isolated, the benefits of the whole extract were greater than the sum of its parts, which indicated a previously unnoticed synergism. In conclusion, polyphenols have interactive and complementary effects, which suggest a possible application in the management of complex diseases and efforts to isolate individual components might be irrelevant for clinical medicine and/or human nutrition.
Abstract: The paraoxonase (PON) group of enzymes, composed of PON1, PON2, and PON3, play an important role in decreasing oxidative stress by degrading lipid peroxides. PON1 synthesis is upregulated by PPAR. Several pharmacological compounds (acting as antioxidants and, hence, atheroprotective) stimulate both PPAR activity and PON1 expression. Recent evidence suggests that PON1 and the monocyte chemoattractant protein-1 (MCP-1) are involved in coordinating the inflammatory response in damaged tissues; PPAR may be central in the regulation of these biochemical pathways. This article reviews the state of knowledge on PON1 biochemistry and function, the influence of genetic variation, and the regulation of PON1 expression by pharmaceutical compounds that increase PPAR activity. We also describe recent lines of evidence suggesting links between PON1 and MCP-1 and how their production may be regulated by PPAR.
Abstract: We explored whether the Asp42Gly polymorphism (rs12075) in the DARC gene represents a confounding factor in the interpretation of monocyte chemoattractant protein-1 (MCP-1) concentration in circulating blood.
Abstract: DING proteins constitute an interesting family, owing to their intriguing and important activities. However, after a decade of research, little is known about these proteins. In humans, at least five different DING proteins have been identified, which were implicated in important biological processes and diseases, including HIV. Indeed, recent data from different research groups have highlighted the anti-HIV activity of some DING representatives. These proteins share the ability to inhibit the transcriptional step of HIV-1, a key step of the viral cycle that is not yet targeted by the current therapies. Since such proteins have been isolated from humans, we undertook a comprehensive study that focuses on the relationship between these proteins and HIV-infection in an infectious context. Hence, we developed a home-made ELISA for the quantification of the concentration of DING proteins in human serum. Using this method, we were able to determine the concentration of DING proteins in healthy and HIV-infected patients. Interestingly, we observed a significant increase of the concentration of DING proteins in non treated and treated HIV-infected patients compared to controls. In addition, cell cultures infected with HIV also show an increased expression of DING proteins, ruling out the possible role of antiretroviral treatment in the increase of the expression of DING proteins. In conclusion, results from this study show that the organism reacts to HIV-infection by an overexpression of DING proteins.
Abstract: We investigated the influence of the HIV infection on serum paraoxonase-3 (PON3) concentration and assessed the relationships with lipoprotein-associated abnormalities, immunological response, and accelerated atherosclerosis. We studied 207 HIV-infected patients and 385 healthy volunteers. Serum PON3 was determined by in-house ELISA, and PON3 distribution in lipoproteins was investigated by fast-performance liquid chromatography (FPLC). Polymorphisms of the PON3 promoter were analyzed by the Iplex Gold MassArray(TM) method. PON3 concentrations were increased (about three times) in HIV-infected patients with respect to controls (P < 0.001) and were inversely correlated with oxidized LDL levels (P = 0.038). Long-term use of nonnucleoside reverse transcriptase inhibitor (NNRTI)-based antiretroviral therapy was associated with a decrease of PON3 concentrations. In a multivariate linear regression analysis, these relationships were still strong when the main confounding covariates were considered. PON3 was mainly found in HDL in HIV-infected patients, but a substantial amount of the protein was detected in LDL particles. This study reports for the first time an important increase in serum PON3 concentrations in HIV-infected patients that is associated with their oxidative status and their treatment with NNRTI. Long-term, prospective studies are needed to confirm the possible influence of this enzyme on the course of this disease and its possible utility as an analytical biomarker.
Abstract: Abstract The epithelial-to-mesenchymal transition (EMT) genetic program is a molecular convergence point in the life-threatening progression of organ fibrosis and cancer toward organ failure and metastasis, respectively. Here, we employed the EMT process as a functional screen for testing crude natural extracts for accelerated drug development in fibrosis and cancer. Because extra virgin olive oil (EVOO) (i.e., the juice derived from the first cold pressing of the olives without any further refining process) naturally contains high levels of phenolic compounds associated with the health benefits derived from consuming an EVOO-rich Mediterranean diet, we have tested the ability of an EVOO-derived crude phenolic extract to regulate fibrogenic and oncogenic EMT in vitro. High-performance liquid chromatography (HPLC) coupled to time-of-flight (TOF) mass spectrometry assays revealed that the EVOO phenolic extract was mainly composed (∼70%) of two members of the secoiridoid family of complex polyphenols, namely oleuropein aglycone-the bitter principle of olives-and its derivative decarboxymethyl oleuropein aglycone. EVOO secoiridoids efficiently prevented loss of proteins associated with polarized epithelial phenotype (i.e., E-cadherin) as well as de novo synthesis of proteins associated with mesenchymal migratory morphology of transitioning cells (i.e., vimentin). The ability of EVOO to impede transforming growth factor-β (TGF-β)-induced disintegration of E-cadherin-mediated cell-cell contacts apparently occurred as a consequence of the ability of EVOO phenolics to prevent the upregulation of SMAD4-a critical mediator of TGF-β signaling-and of the SMAD transcriptional cofactor SNAIL2 (Slug)-a well-recognized epithelial repressor. Indeed, EVOO phenolics efficiently prevented crucial TGF-β-induced EMT transcriptional events, including upregulation of SNAI2, TCF4, VIM (Vimentin), FN (fibronectin), and SERPINE1 genes. While awaiting a better mechanistic understanding of how EVOO phenolics molecularly shut down the EMT differentiation process, it seems reasonable to suggest that nontoxic Oleaceae secoiridoids certainly merit to be considered for aging studies and, perhaps, for ulterior design of more pharmacologically active second-generation anti-EMT molecules.
Abstract: We have tested the hypothesis that the antidiabetic biguanide metformin can be used to manipulate the threshold for stress-induced senescence (SIS), thus accelerating the onset of cancer-protective cellular senescence in response to oncogenic stimuli. Using senescence-prone murine embryonic fibroblasts (MEFs), we assessed whether metformin treatment modified the senescence phenotype that is activated in response to DNA damaging inducers. Metformin significantly enhanced the number of MEFs entering a senescent stage in response to doxorubicin, an anthracycline that induces cell senescence by activating DNA damage signaling pathways (e.g., ATM/ATR) in a reactive oxygen species (ROS)-dependent manner. Using WI-38 and BJ-1 human diploid fibroblasts (HDFs), we explored whether metformin supplementation throughout their entire replicative lifespan may promote the early appearance of the biomarkers of replicative senescence. Chronic metformin significantly reduced HDFs' lifespan by accelerating both the loss of replicative potential and the acquisition of replicative senescence-related biomarkers (e.g., enlarged and flattened cell shapes, loss of arrayed arrangement, accumulation of intracellular and extracellular debris and SA-β-gal-positive staining). Metformin functioned as a bona fide stressful agent, inducing monotonic, dose-dependent, SIS-like responses in BJ-1 HDFs, which are highly resistant to ROS-induced premature senescence. Metformin-induced SIS in BJ-1 fibroblasts was accompanied by the striking activation of several microRNAs belonging to the miR-200s family (miR-200a, miR-141 and miR429) and miR-205, thus mimicking a recently described ability of ROS to chemosensitize cancer cells by specifically upregulating anti-EMT (epithelial-to-mesenchymal transition) miR-200s. Because the unlimited proliferative potential of stem cells results from their metabolic refractoriness to SIS, we finally tested if metformin treatment could circumvent the stress (e.g., ROS)-resistant phenotype of induced pluripotent stem cells (iPSCs). Metformin treatment drastically reduced both the number and the size of iPSC colonies and notably diminished the staining of the pluripotency marker alkaline phosphatase. Our current findings, altogether, reveal for the first time that metformin can efficiently lower the threshold for SIS to generate an "stressed" cell phenotype that becomes pre-sensitized to oncogenic-like stimuli, including DNA damaging, proliferative and/or stemness inducers.
Abstract: There are no data on the relationship between serum paraoxonase-3 (PON3) concentration and atherosclerosis in humans. Our aim was to investigate possible associations, using recently developed methods, in patients with peripheral artery disease (PAD) or coronary artery disease (CAD).
Abstract: The recently observed association between the APOC3-related rs10892151 polymorphism and serum triglyceride levels has prompted us the possibility to explore whether this genetic variant may play a major role in human immunodeficiency virus (HIV)/antiretroviral therapy-induced dyslipidemia.
Abstract: Experimental studies showed that paraoxonase-3 (PON3) retards lipoprotein oxidation. Our objective was to describe a new assay to measure serum PON3 concentrations and report their reference values in a population-based study. The influence of PON3 promoter polymorphisms and their relationships with PON1 and lipid profile were also studied. We generated an anti-PON3 antibody by inoculating rabbits with a synthetic peptide specific to mature PON3. This antibody was used to develop an ELISA. The average regression line of standard plots (n = 8) was y = 0.9587 (0.3392) log(10)x + 1.9466 (0.0861) [r(2) = 0.924 (0.0131); P < 0.001]. There was no cross reaction with PON1. Detection limit was 0.24 mg/l. Imprecision was ≤ 13.2%. Reference interval (n = 356) was 1.00-2.47 mg/l. PON3 was observed in HDL particles containing apolipoprotein (apo)A-I and PON1, but not apoA-II or apoE. Serum PON3 concentrations showed a moderate influence (about 10% variation) by PON3 promoter polymorphisms. Our study describes for the first time a method to measure serum PON3 concentrations. This method offers new opportunities in the investigation of the properties and role of PON3 in cardiovascular disease, with possible implications in clinical practice.
Abstract: Mycobacterium poriferae was described by Padgitt and Moshier (6) in 1987, when reporting its isolation from cell suspensions of the marine sponge Halichondria bowerbanski.…
Abstract: Liver steatosis is frequent in patients with chronic hepatitis viral infections. Intracellular fatty acid synthase (FASN) seems to play a substantial role in its pathogenesis. FASN can also be found in circulation and is significantly increased in HIV-infected individuals, especially if they are co-infected with hepatitis C virus (HCV).
Abstract: Characterization of the molecular function of complex phenols naturally present in extra virgin olive oil (EVOO) against the HER2-gene amplified JIMT-1 cell line, a unique breast cancer model that inherently exhibits cross-resistance to multiple HER1/2-targeted drugs including trastuzumab, gefitinib, erlotinib and lapatinib, may underscore innovative cancer molecules with novel therapeutic targets because they should efficiently circumvent de novo resistance to HER1/2 inhibitors in order to elicit tumoricidal effects. We identified pivotal signaling pathways associated with the efficacy of crude phenolic extracts (PEs) obtained from 14 monovarietals of Spanish EVOOs. i) MTT-based cell viability and HPLC coupled to time-of-flight (TOF) mass spectrometry assays revealed that anti-cancer activity of EVOO PEs positively correlated with the phenolic index (i.e., total content of phenolics) and with a higher presence of the complex polyphenols secoiridoids instead of lignans. ii) Genome-wide analyses using 44 K Agilent's whole human arrays followed by Gene Set Enrichment Analysis (GSEA)-based screening of the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway database revealed a differential modulation of the JIMT-1 transcriptome at the level of the cell cycle and p53 pathways. EVOO PEs differentially impacted the expression status of stress-sensing, G2-M check-point-related GADD45 genes and of p53-related CDKN1A, CDKN1C and PMAIP-1 genes. iii) Cell cycle and fluorescence microscopy analyses confirmed that secoiridoid-rich EVOO PE inhibited mitosis to promote G2-M cell cycle arrest. This was accompanied with the appearance of diffuse, even DNA staining with γH2AX and pan-nuclear hyperacetylation of Histone H3 at Lysine 18. iv) Semi-quantitative Signaling Node Multi-Target ELISAs determined that secoiridoid-rich EVOO PE drastically activated the mitogen-activated protein kinases MEK1 and p38 MAPK, a GADD45-related kinase involved in Histone H3 acetylation. Secoiridoids, a family of complex polyphenols characteristic of Oleaceae plants, appear to permit histones to remain in hyperacetylated states and through the resulting alterations in gene regulation to reduce mitotic viability and metabolic competence of breast cancer cells inherently refractory to HER-targeting therapies ab initio. Oleaceae secoiridoids could provide a valuable phytochemical platform for the design of more pharmacologically active second-generation phytopharmaceutical anti-breast cancer molecules with a unique mode of action.
Abstract: Research on paraoxonase-3 (PON3) has been hampered by the lack of methods for measurement. This is a pilot study aimed at exploring whether chronic liver impairment is associated with changes in serum PON3 concentrations, and to know whether this measurement may provide useful information to investigate this derangement.
Abstract: Atherosclerosis in symptomatic peripheral arterial disease affects wide portions of numerous arteries in lower extremities. The resulting active inflammation in a considerable amount of arterial tissue facilitates systemic detection via measurement of inflammation-related variables. We reasoned that the combined assessment of defense against oxidative stress, in the form of paraoxonase-1 (PON1), and monocyte migration measured as circulating (C-C motif) ligand 2 (CCL2), may play a role in the evaluation of these patients. Plasma CCL2 and serum PON1-related variables, assessed by their interaction with functional genetic variants, were measured in a cross-sectional study in patients with symptomatic PAD. We found that PON1 activity and concentration were significantly lower and CCL2 concentration higher in PAD patients compared to controls, that the combination of plasma CCL2 and PON1- related values, especially PON1 concentration differentiated, almost perfectly, controls from patients and that the expression of CCL2 and PON1 generally co-localized in the atherosclerotic lesion. Since no association with genetic variants was found, such a relationship is probably the result of the disease. Our data suggest a coordinated role between CCL2 and PON1 that may be detected in blood with simple measurements and may represent an indicator of the extent of atherosclerosis.
Abstract: Eur J Clin Invest 2010 ABSTRACT: Background  The paraoxonase (PON) enzyme family comprising PON1, PON2 and PON3 are antioxidant enzymes that degrade bioactive oxidised lipids and are thus antiatherogenic. Materials and methods  We investigated the localisation of the PON proteins during the development of atherosclerosis by immunohistochemical analysis. Results  In normal aortas, PON1 and PON3 were localised to smooth muscle cells (SMC) and endothelial cells. PON3 staining was stronger than that of PON1. During atherosclerosis development, SMC staining for PON1 and PON3 was greatly reduced, while macrophage staining for both proteins increased with PON1 predominating. Macrophage staining for PON1 and PON3 was significantly and positively related to the amount of aortic inflammation (both P < 0·001). Conclusions  Our data add support to the growing body of evidence for a cellular protective effect of PON1 and PON3 against the proinflammatory/proatherosclerotic effects of lipid peroxidation.
Abstract: By activating the ataxia telangiectasia mutated (ATM)-mediated DNA Damage Response (DDR), the AMPK agonist metformin might sensitize cells against further damage, thus mimicking the precancerous stimulus that induces an intrinsic barrier against carcinogenesis. Herein, we present the new hypothesis that metformin might function as atissue sweeper of pre-malignant cells before they gain stem cell/tumor initiating properties. Because enhanced glycolysis (the Warburg effect) plays a causal role in the gain of stem-like properties of tumor-initiating cells by protecting them from the pro-senescent effects of mitochondrial respiration-induced oxidative stress, metformin's ability to disrupt the glycolytic metabotype may generate a cellular phenotype that is metabolically protected against immortalization. The bioenergetic crisis imposed by metformin, which may involve enhanced mitochondrial biogenesis and oxidative stress, can lower the threshold for cellular senescence by pre-activating an ATM-dependent pseudo-DDR. This allows an accelerated onset of cellular senescence in response to additional oncogenic stresses. By pushing cancer cells to use oxidative phosphorylation instead of glycolysis, metformin can rescue cell surface major histocompatibility complex class I (MHC-I) expression that is downregulated by oncogenic transformation, a crucial adaptation of tumor cells to avoid the adaptive immune response by cytotoxic T-lymphocytes (CTLs). Aside from restoration of tumor immunosurveillance at the cell-autonomous level, metformin can activate a senescence-associated secretory phenotype (SASP) to reinforce senescence growth arrest, which might trigger an immune-mediated clearance of the senescent cells in a non-cell-autonomous manner. By diminishing the probability of escape from the senescence anti-tumor barrier, the net effect of metformin should be a significant decrease in the accumulation of dysfunctional, pre-malignant cells in tissues, including those with the ability to initiate tumors. As life-long or late-life removal of senescent cells has been shown to prevent or delay the onset or progression of age-related disorders, the tissue sweeper function of metformin may inhibit the malignant/metastatic progression of pre-malignant/senescent tumor cells and increase the human lifespan.
Abstract: The property of many bacteria to form biofilms constitutes a major health problem. Bacteria living in biofilms have a very high resistance to antibiotics. Biofilms may develop at a certain locations with the participation of secreted molecules, termed quorum-sensing signals, when a sufficient density of bacterial growth occurs. In Gram-negative bacteria, acyl homoserine lactones (AHL) have been identified as major quorum-sensing signals. The paraoxonases (PONs) constitute a family of enzymes comprising 3 members (PON1, PON2, and PON3) that have lactonase activity and are able to hydrolyze AHL. In this minireview, we summarize some existing basic knowledge on PON genetics, biochemistry, and function and describe recent research that reports evidence of the important roles that they may play in the organism's defense against biofilm formation. Finally, we propose some lines of future research that could be very productive.
Abstract: The incidence of obesity and related metabolic diseases is increasing globally. Current medical treatments often fail to halt the progress of such disturbances, and plant-derived polyphenols are increasingly being investigated as a possible way to provide safe and effective complementary therapy. Rooibos (Aspalathus linearis) is a rich source of polyphenols without caloric and/or stimulant components. We have tentatively characterized 25 phenolic compounds in rooibos extract and studied the effects of continuous aqueous rooibos extract consumption in mice. The effects of this extract, which contained 25% w/w of total polyphenol content, were negligible in animals with no metabolic disturbance but were significant in hyperlipemic mice, especially in those in which energy intake was increased via a Western-type diet that increased the risk of developing metabolic complications. In these mice, we found hypolipemiant activity when given rooibos extract, with significant reductions in serum cholesterol, triglyceride and free fatty acid concentrations. Additionally, we found changes in adipocyte size and number as well as complete prevention of dietary-induced hepatic steatosis. These effects were not related to changes in insulin resistance. Among other possible mechanisms, we present data indicating that the activation of AMP-activated protein kinase (AMPK) and the resulting regulation of cellular energy homeostasis may play a significant role in these effects of rooibos extract. Our findings suggest that adding polyphenols to the daily diet is likely to help in the overall management of metabolic diseases.
Abstract: Study Type - Aetiology (case series). Level of Evidence: 4. What's known on the subject? and What does the study add? Oxidative stress seems to be one of the biochemical causes of defective sperm function. Paraoxonases are antioxidant enzymes that degrade lipid peroxides. There is a paucity of data on the possible role played by these enzymes in the pathophysiology of male sub-fertility. The present study shows that testicular tissue of sub-fertile patients clearly expresses paraoxonases-1, 2, and 3. These findings suggest a role for these enzymes in the protection against lipid peroxidation inside the cell. However, the concentration and activity of paraoxonase-1 in semen are negligible and are probably the result of cellular catabolism, with no significant biological function.
Abstract: The biguanide metformin, a widely used drug for the treatment of type 2 diabetes, may exert cancer chemopreventive effects by suppressing the transformative and hyperproliferative processes that initiate carcinogenesis. Metformin's molecular targets in cancer cells (e.g., mTOR, HER2) are similar to those currently being used for directed cancer therapy. However, metformin is nontoxic and might be extremely useful for enhancing treatment efficacy of mechanism-based and biologically targeted drugs. Here, we first revisit the epidemiological, preclinical, and clinical evidence from the last 5 years showing that metformin is a promising candidate for oncology therapeutics. Second, the anticancer effects of metformin by both direct (insulin-independent) and indirect (insulin-dependent) mechanisms are discussed in terms of metformin-targeted processes and the ontogenesis of cancer stem cells (CSC), including Epithelial-to-Mesenchymal Transition (EMT) and microRNAs-regulated dedifferentiation of CSCs. Finally, we present preliminary evidence that metformin may regulate cellular senescence, an innate safeguard against cellular immortalization. There are two main lines of evidence that suggest that metformin's primary target is the immortalizing step during tumorigenesis. First, metformin activates intracellular DNA damage response checkpoints. Second, metformin attenuates the anti-senescence effects of the ATP-generating glycolytic metabotype-the Warburg effect-, which is required for self-renewal and proliferation of CSCs. If metformin therapy presents an intrinsic barrier against tumorigenesis by lowering the threshold for stress-induced senescence, metformin therapeutic strategies may be pivotal for therapeutic intervention for cancer. Current and future clinical trials will elucidate whether metformin has the potential to be used in preventive and treatment settings as an adjuvant to current cancer therapeutics.
Abstract: Rooibos (Aspalathus linearis) is a rich source of polyphenols and used to make a mild-tasting tea containing no caffeine, is low in tannins compared to green or black teas, and has antioxidant and antimutagenic/antitumoral properties. In vivo results show that rooibos has beneficial effects upon the lipid profile by decreasing serum triglycerides and cholesterol. In this sense, we have developed a simple and rapid method to separate and characterize simultaneously the polyphenolic compounds in aqueous and ethanolic rooibos extracts using high-performance liquid chromatography coupled to electrospray ionization time-of-flight mass spectrometry (HPLC-ESI-TOF-MS) and ion trap multiple mass spectrometry (HPLC-ESI-IT-MS(2)). The phenolic compounds were separated on a C(18) column (4.6 × 150 mm, 1.8 μm) with 1% formic acid in water/acetonitrile 90:10 v/v and acetonitrile as mobile phases. The accuracy mass data generated by TOF-MS together with the fragmentation pattern obtained by IT-MS(2) experiments confirmed the presence of 25 and 30 phenolic compounds in the aqueous and ethanolic extracts, respectively.
Abstract: BACKGROUND: Paraoxonase-1 (PON1) is an antioxidant enzyme that attenuates the production of the monocyte chemoattractant protein-1 (MCP-1) in vitro. Although oxidation and inflammation are closely related processes, the association between PON1 and MCP-1 has not been completely characterised due, probably, to that the current use of synthetic substrates for PON1 measurement limits the interpretation of the data. In the present study, we explored the relationships between the circulating levels of PON1 and MCP-1 in human immunodeficiency virus-infected patients in relation to the multifunctional capabilities of PON1. METHODS: We measured selected variables in 227 patients and in a control group of 409 participants. Serum PON1 esterase and lactonase activities were measured as the rates of hydrolysis of paraoxon and of 5-(thiobutyl)-butyrolactone, respectively. Oxidised LDL and MCP-1 concentrations were determined by enzyme-linked immunosorbent assay. High-density lipoproteins cholesterol, apolipoprotein A-I, and C-reactive protein concentrations were measured by standard automated methods. RESULTS: There were significant relationships between PON1 activity and several indices of oxidation and inflammation in control subjects and in infected patients. However, these relationships varied not only with disease status but also on the type of substrate used for PON1 measurement. CONCLUSION: The present study is a cautionary tale highlighting that results of clinical studies on PON1 may vary depending on the methods used as well as the disease studied. Until more specific methods using physiologically-akin substrates are developed for PON1 measurement, we suggest the simultaneous employment of at least two different substrates in order to improve the reliability of the results obtained.
Abstract: Nonalcoholic fatty liver disease is considered to be the hepatic manifestation of metabolic syndrome and is usually related to high-fat, high-cholesterol diets. With the rationale that the identification and quantification of metabolites in different metabolic pathways may facilitate the discovery of clinically accessible biomarkers, we report the use of (1)H NMR metabolomics for quantitative profiling of liver extracts from LDLr(-/-) mice, a well-documented mouse model of fatty liver disease. A total of 55 metabolites were identified, and multivariate analyses in a diet- and time-comparative strategy were performed. Dietary cholesterol increased the hepatic concentrations of cholesterol, triglycerides, and oleic acid but also decreased the [PUFA/MUFA] ratio as well as the relative amount of long-chain polyunsaturated fatty acids in the liver. This was also accompanied by variations of the hepatic concentration of taurine, glutathione, methionine, and carnitine. Heat-map correlation analyses demonstrated that hepatic inflammation and development of steatosis correlated with cholesterol and triglyceride NMR derived signals, respectively. We conclude that dietary cholesterol is a causal factor in the development of both liver steatosis and hepatic inflammation.
Abstract: ABSTRACT: BACKGROUND: Fatty acid synthase (FASN) is an enzyme synthesized by the liver and plays an important role in lipogenesis. The present study aimed to investigate whether serum FASN concentration may provide a direct link between HIV and/or HCV viral infections and lipid metabolic disorders commonly observed in HIV/HCV-infected patients. METHODS: We evaluated serum FASN concentration in 191 consecutive HIV-infected patients in the absence or presence of HCV co-infection. For comparison, 102 uninfected controls were included. Metabolic and inflammatory phenotype was also compared with respect to the presence of HCV co-infection. RESULTS: Serum FASN concentration was significantly higher in HIV-infected patients than in healthy participants and HCV co-infected patients showed higher levels than those without co-infection. Levels were also affected by treatment regimen, but marginally influenced by virological variables. Insulin concentration was the sole variable among metabolic parameters that demonstrated a significant correlation with serum FASN concentrations. Serum alanine aminotransferase (ALT) values correlated significantly with serum FASN concentration and provided the best discrimination with respect to the presence or absence of HCV co-infection. In multivariate analysis, only ALT, monocyte chemoattractant protein-1 (MCP-1) and the presence of antiretroviral treatment regimen significantly contributed to explain serum FASN concentration in HIV/HCV co-infected patients. CONCLUSION: Serum FASN concentration is significantly increased in HIV-infected individuals. The release of FASN into the circulation is further enhanced in patients who are co-infected with HCV. Subsequent studies should explore the usefulness of this indicator to monitor the effect of viral infections on disease progression and survival.
Abstract: Oxidative stress and inflammation play a central role in the onset and development of liver diseases irrespective of the agent causing the hepatic impairment. The monocyte chemoattractant protein-1 is intimately involved in the inflammatory reaction and is directly correlated with the degree of hepatic inflammation in patients with chronic liver disease. Recent studies showed that hepatic paraoxonase-1 may counteract the production of the monocyte chemoattractant protein-1, thus playing an anti-inflammatory role. The current review summarises experiments suggesting how paraoxonase-1 activity and expression are altered in liver diseases, and their relationships with the monocyte chemoattractant protein-1 and inflammation.
Abstract: In this observational, case-control study in infertile women, we observed elevated serum oxidative stress that was significantly correlated with an increase in serum paraoxonase-1 activity and without any evidence of a proinflammatory reaction. These results suggest a protective role of paraoxonase-1 against inflammation in this clinical setting.
Abstract: BACKGROUND: Monocyte chemoattractant protein-1 (MCP-1) facilitates the recruitment of monocytes/macrophages into vascular intima, and it is probably involved in the regulation of other signaling pathways relevant to the pathogenesis of arteriosclerosis and metabolic disturbances. However, chemokines are redundant. Consequently, the protective effect of MCP-1 deficiency may be mediated by changes in other cytokine signals. METHODS AND RESULTS: Changes in the pattern of gene expression in the aorta were evaluated in LDLr(-/-) and MCP-1(-/-) LDLr(-/-) mice fed either chow or Western-style diet. Functional analyses were used to characterize the pathways affected and to identify biological processes in which MCP-1 may play an additional role. Some data also suggest that MCP-5 may act as a surrogate for MCP-1 deletion. Arteriosclerosis lesion and plaque composition are associated with enrichment in the cytokine-cytokine receptor interaction pathway. CONCLUSIONS: There is a complex network of interactions linking MCP-1 and other cytokines. The lack of MCP-1 limits the aortic response to atherogenic stimuli, but does not completely protect against neointima formation. Activation of alternative inflammatory pathways in the vascular wall in response to MCP-1 deficiency should be considered to fully understand the actual role of this chemokine.
Abstract: BACKGROUND: Paraoxonase-1 (PON1), a lactonase synthesized by the liver, circulates in blood bound to high-density lipoproteins (HDL). This enzyme is thought to degrade oxidized phospholipids and play an important role in the organism's antioxidant and anti-inflammatory system. Chronic liver diseases are characterized by decreased serum PON1 activity. The aim of the present study was to investigate the compositional changes in HDL that could influence PON1 activity in liver impairment. METHODS: The study was performed in samples from five patients with advanced liver cirrhosis and with preserved renal function, chosen on the basis of having low serum PON1 activity and high serum PON1 concentration. As a control group, we accessed five healthy volunteers from among our hospital staff. Lipid and protein compositional analysis of lipoprotein particles were done by high-performance liquid chromatography, gel electrophoresis, and Western-Blot. RESULTS: HDL particles from cirrhotic patients had an increased phospholipid content that was inversely correlated to PON1 activity. The HDL particles contained high levels of PON1 that corresponded, in part, to an immunoreactive protein of high molecular weight (55 kDa) not present in control subjects. This protein was identified as glycosylated PON1 and was also present in biopsies from patients with steatosis and from rats with CCl(4)-induced hepatic impairment. These changes were associated with an increased plasma concentration of markers of oxidative stress, inflammation and fibrogenesis. CONCLUSION: Abnormalities in the composition of lipids and proteins of HDL particles, including PON1 glycosylation, are associated with the decrease in serum PON1 activity in patients with liver disease. These alterations may adversely affect the protective role of HDL against oxidative stress and inflammation in these patients.
Abstract: BACKGROUND: Human immunodeficiency virus (HIV) infection is associated with abnormal high-density lipoprotein (HDL) particles. We evaluated whether HIV infection and antiretroviral treatment promotes changes in cholesterol distribution among subpopulations of HDL particles of defined sizes. METHODS: HDL particles were isolated from 78 HIV infected patients and fractionated by gel permeation chromatography to obtain five subpopulations. Thirty-six patients were antiretroviral treatment naïve, while 42 patients were treated with efavirenz or protease inhibitors. Uninfected individuals were also included as controls. RESULTS: The distribution of cholesterol across HDL particle sizes was affected by HIV infection itself. Antiretroviral therapy reduced these alterations; only minor changes in small and very small HDL particles were observed in treated patients (p=0.01). Untreated patients with low CD4+ T cell counts had less cholesterol in medium (p=0.006), small (p=0.04) and very small (p=0.03) HDL particles. Treated patients with high CD4+ T cell counts had less cholesterol in the largest HDL particles (p=0.04), with overall particle distributions resembling those observed in uninfected participants. CONCLUSIONS: HIV infection itself may promote major changes in cholesterol distribution among HDL subpopulations that could be partially attenuated by current antiretroviral treatments. Further studies in larger populations are necessary to confirm the impact of HIV on lipoprotein composition and distribution.
Abstract: BACKGROUND: Circulating CCL2 concentration has been implicated in promoting atherosclerosis in patients infected with HIV. We evaluated whether CCL2 gene variants are associated with metabolic disturbances and plasma CCL2 levels in HIV-infected patients. METHODS AND RESULTS: CCL2 genotypes and estimated haplotypes, plasma CCL2 levels and indicators of metabolic status in HIV-infected patients were compared with a representative group of the general population. We also performed a carotid/femoral artery ultrasonography to detect sub-clinical atherosclerosis in these patients. Six haplotypes were estimated in more than the 5% of individuals, and accounted for more than 98% of the population. In HIV-infected patients, carriers of H1, H2 and H5 haplotypes had higher CCL2 concentration than carriers of H3, H4 and H6 haplotypes. However, only carriers of H1 and H5 haplotypes presented higher insulin resistance as well as higher proportion of patients affected with sub-clinical. Conversely, carriers of H2 haplotype, which also showed high plasma CCL2 concentration, were associated with less deleterious metabolic disturbances. CONCLUSIONS: Our data are consistent with the hypothesis that the genetic background of the host is involved in CCL2 production and that this chemokine is implicated in promoting metabolic disturbances and sub-clinical atherosclerosis in HIV-infected patients.
Abstract: OBJECTIVE: The purpose of this study was to analyze the association between the dilatation of the aortic root and the diameters of the rest of the aorta and to identify some related factors that could be used to identify patients at higher risk of presenting with an aortic aneurysm. METHODS: In 71 consecutive patients with a dilated aortic root identified by transthoracic echocardiography, prospective helical computed tomography was performed. Aortic diameters were measured perpendicular to the flow at seven levels in the thoracic and abdominal aorta. RESULTS: Ascending aorta diameter showed a moderate correlation with aortic indexed diameters at the thoracic and abdominal level in tricuspid aortic valve patients (r ranging from 0.37-0.56), whereas in patients with a bicuspid aortic valve, a moderate correlation between the ascending aorta diameters and the thoracic descending aorta diameters was observed (r 0.51-0.53). In a multivariate analysis, age was independently related to indexed diameter at all aortic sites (beta ranging from 0.06-0.12 per year), whereas aortic regurgitation was independently related only to thoracic aorta diameter (beta ranging from 1.17-1.84). Age (P < .0001), body surface area (P < .0001), and grade of aortic valve regurgitation (P = .001) independently predicted aortic volume. CONCLUSION: Different patterns of aortic diameters were observed in patients with dilated aortic root, depending on age, aortic valve morphology, and function. When a dilated aortic root is detected in older patients with a tricuspid aortic valve, an accurate cardiovascular survey that includes the entire aorta is needed. These results provide further evidence about the systemic nature of aortic dilatation.
Abstract: Research on the molecular basis of the hepatic alterations associated to obesity is dependent on the availability of suitable animal models. Apolipoprotein E deficient mice (ApoE(-/-)) and LDL-receptor deficient mice (LDLr(-/-)) develop steatosis and steatohepatitis when given pro-atherogenic diets. However, previous data suggest that these two models are not completely interchangeable, and that their metabolic phenotype may partially differ in response to nutrient stimuli. The present study further investigates this question, by comparing changes in hepatic inflammation, lipoprotein metabolism, and their related gene expressions. LDLr(-/-) mice were more susceptible to the development of obesity and hepatic steatosis, while the ApoE(-/-) model increased the amount of macrophages and inflammatory nodules in the liver. These changes were accompanied by a differential expression of selected members of the MAPK family and PPARs in the liver.
Abstract: We have studied the distribution of mRNA for paraoxonases (PON) 1 and 2 in 24 human tissues using Gene Expression Panels. PON1 mRNA was restricted to adult kidney, liver, and colon as well as fetal liver, whereas PON2 mRNA was more widely distributed in adult human brain, heart, kidney, spleen, liver, colon, lung, small intestine, muscle, stomach, testis, placenta, salivary, thyroid and adrenal glands, pancreas, skin, and bone marrow, as well as fetal brain and liver. PON2 mRNA was not found in ovary, uterus, or plasma leukocytes using the panels. However, using real time PCR, we found PON2 mRNA expression in human plasma leukocytes. There were differences between the tissue distribution of mRNAs found in this study and the immunohistochemical localization of the PON1 and PON2 proteins reported previously. In particular, PON1 protein is much more widely distributed than its mRNA, possibly indicating the delivery of PON1 to various tissues by HDL. In addition, differences between PON2 mRNA and protein distributions could be due to missence mutations in the PON2 gene, causing nontranslation of mRNA to protein in some tissues.
Abstract: To maintain homeostasis under diverse metabolic conditions, it is necessary to coordinate nutrient-sensing pathways with the immune response. This coordination requires a complex relationship between cells, hormones, and cytokines in which inflammatory and metabolic pathways are convergent at multiple levels. Recruitment of macrophages to metabolically compromised tissue is a primary event in which chemokines play a crucial role. However, chemokines may also transmit cell signals that generate multiple responses, most unrelated to chemotaxis, that are involved in different biological processes. We have reviewed the evidence showing that monocyte chemoattractant protein-1 (MCP-1 or CCL2) may have a systemic role in the regulation of metabolism that sometimes is not necessarily linked to the traffic of inflammatory cells to susceptible tissues. Main topics cover the relationship between MCP-1/CCL2, insulin resistance, inflammation, obesity, and related metabolic disturbances.
Abstract: A vicious cycle between oxidation and inflammation leads to complications in a growing number of disease states. Knowledge on tissue distribution of chemokines, mediators of inflammatory response, and paraoxonases, with antioxidant and anti-inflammatory actions, may be relevant. Using immunohistochemistry and quantitative real-time PCR we have investigated the distribution of PON1, 2 and 3, CCL2, 7, 8 and 12 and the chemokine receptor CCR2 protein and mRNA in 23 tissues from C57BL/6J mice. As expected, PON1, 2 and 3, CCL2, 7, 8 and 12 and CCR2 proteins were present in the vast majority of tissues investigated. Surprisingly, mRNA for these proteins was also expressed in most of these tissues suggesting local production and the ability to respond in situ to inflammatory stimuli. The wide distribution and expression of mRNA for paraoxonases and CC-chemokines suggest a systemic, probably coordinated, role in the overall inflammatory response.
Abstract: BACKGROUND: Oxidative stress is associated with human immunodeficiency virus (HIV) infection. Paraoxonase-1 (PON1) is an antioxidant enzyme that is bound to high-density lipoproteins (HDLs). We evaluated whether PON1 gene haplotypes influence the metabolic disturbances, presence of subclinical atherosclerosis, and virologic outcome associated with the infection. METHODS: DNA from blood samples collected from 234 HIV-infected patients and 633 healthy control subjects had single-nucleotide polymorphisms of PON1(192), PON1(55), PON1(-162), PON1(-832), PON1(-909), PON1(-1076), and PON1(-1741) analyzed using the Iplex Gold MassArray method. Subsequently, the influence of these single-nucleotide polymorphisms on measured biochemical and clinical variables was assessed. RESULTS: We observed significant differences in the haplotype distribution between the control subjects and the HIV-infected patients. Haplotype H10 (GTCCGTC) was more prevalent in the HIV-infected patients (6.41% vs 0.64%; P < .001), and haplotype H5 (GACCGTC) was less prevalent in HIV-infected patients (27.7% vs 42.9%; P = .001). In HIV-infected patients, haplotype H7 (AATTCCT) was associated with better CD4(+) cell count recovery, higher levels of HDL cholesterol (P = .048) and apolipoprotein A-I (P = .019), lower levels of triglycerides (P = .004), and lower rates of subclinical arteriosclerosis (P < .001). CONCLUSIONS: PON1 haplotypes segregate with HIV infection, HDL metabolism, the presence of subclinical atherosclerosis, and CD4(+) cell recovery after treatment.
Abstract: OBJECTIVES: HIV infection and its treatment are associated with dyslipidaemia and increased risk of cardiovascular disease. Accurate high-density lipoprotein (HDL) cholesterol values are necessary for the management of these abnormalities, but current methods have not been properly assessed in these patients. The aim of this study was to assess in HIV-infected patients the consistency and accuracy of a synthetic polymer/detergent homogeneous assay used to measure HDL cholesterol concentrations and to evaluate the impact of storage. METHODS: HDL cholesterol was measured using a synthetic polymer/detergent homogeneous method in samples from HIV-infected patients and healthy subjects for each of the storage regimens: baseline, after 1 week at 4 degrees C, and after 12 months at -80 degrees C. The ultracentrifugation and precipitation assays were used for comparison. RESULTS: Three out of every 20 samples from HIV-infected patients had discrepant HDL cholesterol values with respect to the ultracentrifugation method. Overestimation was associated with high C-reactive protein concentrations and underestimation with plasma gamma-globulin concentrations, an effect that was amplified by any of the storage conditions tested. CONCLUSIONS: Caution is needed when using the synthetic polymer/detergent homogeneous method for direct measurement of HDL cholesterol concentrations in HIV-infected patients. This assay is of limited use in clinical trials in which frozen samples are analysed.
Abstract: Transforming Growth Factor-b (TGFb) is a major driving force of the Epithelial-to-Mesenchymal (EMT) genetic program, which becomes overactive in the pathophysiology of many age-related human diseases. TGFb-driven EMT is sufficient to generate migrating cancer stem cells by directly linking the acquisition of cellular motility with the maintenance of tumor-initiating (stemness) capacity. Chronic diseases exhibiting excessive fibrosis can be caused by repeated and sustained infliction of TGFb-driven EMT, which increases collagen and extracellular matrix synthesis. Pharmacological prevention and/or reversal of TGFb-induced EMT may therefore have important clinical applications in the management of cancer metastasis as well as in the prevention and/or treatment of end-state organ failures. Earlier studies from our group have revealed that clinically-relevant concentrations of the biguanide derivative metformin, the most widely used oral agent to lower blood glucose concentration in patients with type 2 diabetes and metabolic syndrome, notably decreased both the self-renewal and the proliferation of trastuzumab-refractory breast cancer stem cell populations. Given that: a.) tumor-initiating cancer stem cells display a significant enrichment in the expression of basal/mesenchymal or myoepithelial markers, including an increased secretion of TGFb; b.) metformin treatment impedes the ontogeny of generating the stem cell phenotype by transcriptionally repressing key drivers of the EMT genetic program (e.g. ZEB1, TWIST1, SNAIL2 [Slug], TGFbs), we recently hypothesized that prevention of TGFb-induced EMT might represent a common molecular mechanism underlying the anti-cancer stem cells and anti-fibrotic actions of metformin. Remarkably, metformin exposure not only impedes TGFb-promoted loss of the epithelial marker E-cadherin in MCF-7 breast cancer cells but it prevents further TGF-induced cell scattering and accumulation of the mesenchymal marker vimentin in Madin-Darby canine kidney (MDCK) cells. We now propose that metformin, by weakening the ability of TGFb signaling to fully induce mesenchymal cell states in a variety of pathological processes including fibrosis (e.g. chronic renal disease, non-alcoholic steatohepatitis, heart failure or sclerosis) and malignant progression (and likely by reducing TGFb-regulated inflammation and immune responses -inflamm-aging-), molecularly behaves as a bona fide anti-aging modality.
Abstract: Diet supplementation and/or modulation is an important strategy to significantly improve human health. The search of plants as additional sources of bioactive phenolic compounds is relevant in this context. The aqueous extract of Hibiscus sabdariffa is rich in anthocyanins and other phenolic compounds including hydroxycitric and chlorogenic acids. Using this extract we have shown an effective protection of cultured peripheral blood mononuclear cells from the cellular death induced by H(2)O(2) and a significant role in the production of inflammatory cytokines. In vitro, the extract promotes the production of IL-6 and IL-8 and decreases the concentration of MCP-1 in supernatants in a dose-dependent manner. In humans, the ingestion of an acute dose of the extract (10g) was well tolerated and decreased plasma MCP-1 concentrations significantly without further effects on other cytokines. This effect was not due to a concomitant increase in the antioxidant capacity of plasma. Instead, its mechanisms probably involve a direct inhibition of inflammatory and/or metabolic pathways responsible for MCP-1 production, and may be relevant in inflammatory and chronic conditions in which the role of MCP-1 is well established. If beneficial effects are confirmed in patients, Hibiscus sabdariffa could be considered a valuable traditional herbal medicine for the treatment of chronic inflammatory diseases with the advantage of being devoid of caloric value or potential alcohol toxicity.
Abstract: OBJECTIVES: HIV-infected patients show an increased cardiovascular disease (CVD) risk resulting, essentially, from metabolic disturbances related to chronic infection and antiretroviral treatments. The aims of this study were: (1) to evaluate the agreement between the CVD risk estimated using the Framingham risk score (FRS) and the observed presence of subclinical atherosclerosis in HIV-infected patients; (2) to investigate the relationships between CVD and plasma biomarkers of oxidation and inflammation. METHODS: Atherosclerosis was evaluated in 187 HIV-infected patients by measuring the carotid intima-media thickness (CIMT). CVD risk was estimated using the FRS. We also measured the circulating levels of interleukin-6, monocyte chemoattractant protein-1 (MCP-1) and oxidized low-density lipoprotein (LDL), and paraoxonase-1 activity and concentration. RESULTS: There was a weak, albeit statistically significant, agreement between FRS and CIMT (kappa=0.229, P<0.001). A high proportion of patients with an estimated low risk had subclinical atherosclerosis (n=66; 56.4%). In a multivariate analysis, the presence of subclinical atherosclerosis in this subgroup of patients was associated with age [odds ratio (OR) 1.285; 95% confidence interval (CI) 1.084-1.524; P=0.004], body mass index (OR 0.799; 95% CI 0.642-0.994; P=0.044), MCP-1 (OR 1.027; 95% CI 1.004-1.050; P=0.020) and oxidized LDL (OR 1.026; 95% CI 1.001-1.051; P=0.041). CONCLUSION: FRS underestimated the presence of subclinical atherosclerosis in HIV-infected patients. The increased CVD risk was related, in part, to the chronic oxidative stress and inflammatory status associated with this patient population.
Abstract: The contribution of metabolic factors to the severity of liver disease is not completely understood. In this study, apolipoprotein E-deficient (ApoE-/-) mice were evaluated to define potential effects of hypercholesterolemia on the severity of carbon tetrachloride (CCl4)-induced liver injury. Under baseline conditions, hypercholesterolemic ApoE-/- mice showed increased hepatic oxidative stress (SOD activity/4-hydroxy-2-nonenal immunostaining) and higher hepatic TGF-beta1, MCP-1, and TIMP-1 expression than wild-type control mice. After CCl4 challenge, ApoE-/- mice exhibited exacerbated steatosis (Oil Red O staining), necroinflammation (hematoxylin-eosin staining), macrophage infiltration (F4/80 immunohistochemistry), and fibrosis (Sirius red staining and alpha-smooth muscle actin immunohistochemistry) and more severe liver injury [alanine aminotransferase (ALT) and aspartate aminotransferase] than wild-type controls. Direct correlations were identified between serum cholesterol and hepatic steatosis, fibrosis, and ALT levels. These changes did not reflect the usual progression of the disease in ApoE-/- mice, since exacerbated liver injury was not present in untreated age-paired ApoE-/- mice. Moreover, hepatic cytochrome P-450 expression was unchanged in ApoE-/- mice. To explore potential mechanisms, cell types relevant to liver pathophysiology were exposed to selected cholesterol-oxidized products. Incubation of hepatocytes with a mixture of oxysterols representative of those detected by GC-MS in livers from ApoE-/- mice resulted in a concentration-dependent increase in total lipoperoxides and SOD activity. In hepatic stellate cells, oxysterols increased IL-8 secretion through a NF-kappaB-independent mechanism and upregulated TIMP-1 expression. In macrophages, oxysterols increased TGF-beta1 secretion and MCP-1 expression in a concentration-dependent manner. Oxysterols did not compromise cell viability. Taken together, these findings demonstrate that hypercholesterolemic mice are sensitized to liver injury and that cholesterol-derived products (i.e., oxysterols) are able to induce proinflammatory and profibrogenic mechanisms in liver cells.
Abstract: Research into the paraoxonase (PON) gene family has flourished over the past few years. In the 1970s and 1980s, only PON1 was known, and the investigations were conducted, essentially, by toxicologists focusing on protection against organophosphate poisoning. Since then, two new members of the family, PON2 and PON3, have been identified, both being shown to play antioxidant and anti-inflammatory roles. Evidence exists indicating that the PON family is central to a wide variety of human illnesses such as cardiovascular disease, diabetes mellitus, metabolic syndrome, obesity, non-alcoholic steatohepatitis, and several mental disorders. However, research is hampered considerably by the methods currently available to measure the activity of these enzymes. In this review, we summarize the state of knowledge on PON biochemistry and function, the influence of genetic variations, and the involvement of PON in several diseases. The problems associated with PON measurement, such as sample acquisition, lack of reference methods, and variety of substrates, will be presented. Also, we cover some of the present lines of research and propose some others for future progress in this field.
Abstract: BACKGROUND: Paraoxonase-1 (PON1) is an antioxidant enzyme synthesized by the liver. It protects against liver impairment and attenuates the production of the pro-inflammatory monocyte chemoattractant protein-1 (MCP-1). We investigated the relationships between hepatic PON1 and MCP-1 expression in rats with liver disease and explored the possible molecular mechanisms involved. METHODS: CCl4 was administered for up to 12 weeks to induce liver damage. Serum and hepatic levels of PON1 and MCP-1, their gene and protein expression, nuclear transcription factors, and histological and biochemical markers of liver impairment were measured. RESULTS: High levels of PON1 and MCP-1 expression were observed at 12th week in the hepatocytes surrounding the fibrous septa and inflammatory areas. CCl4-administered rats had an increased hepatic PON1 concentration that was related to decreased gene transcription and inhibited protein degradation. Decreased PON1 gene transcription was associated with PPARdelta expression. These changes were accompanied by increased hepatic MCP-1 concentration and gene expression. There were significant direct relationships between hepatic PON1 and MCP-1 concentrations (P = 0.005) and between PON1 and the amount of activated stellate cells (P = 0.001). CONCLUSION: Our results from this experimental model suggest a hepato-protective role for PON1 against inflammation, fibrosis and liver disease mediated by MCP-1.
Abstract: Paraoxonase-1 (PON1) is an esterase and lactonase synthesized by the liver and found in the circulation associated with high-density lipoproteins. The physiological function of PON1 seems to be to degrade specific oxidized cholesteryl esters and oxidized phospholipids in lipoproteins and cell membranes. PON1 is, therefore, an antioxidant enzyme. Alterations in circulating PON1 levels have been reported in a variety of diseases involving oxidative stress including chronic liver diseases. Measurement of serum PON1 activity has been proposed as a potential test for the evaluation of liver function. However, this measurement is still restricted to research and has not been extensively applied in routine clinical chemistry laboratories. The reason for this restriction is due to the problem that the substrate commonly used for PON1 measurement, paraoxon, is toxic and unstable. The recent development of new assays with non-toxic substrates makes this proposal closer to a practical development. The present editorial summarizes PON1 biochemistry and function, its involvement with chronic liver impairment, and some aspects related to the measurement of PON1 activity in circulation.
Abstract: Paraoxonase-1 is a lactonase and an esterase and it plays a protective role in toxicity as well as in diseases involving oxidative stress. Recently, insights into how it may be modulated by environmental factors have acquired clinical relevance. This article reviews the state-of-the-art evidence regarding PON1 modulation by pharmacological products as well as nutritional and lifestyle factors.
Abstract: The phenolic fraction and other polar compounds of the Hibiscus sabdariffa were separated and identified by HPLC with diode array detection coupled to electrospray TOF and IT tandem MS (DAD-HPLC-ESI-TOF-MS and IT-MS). The H. sabdariffa aqueous extract was filtered and directly injected into the LC system. The analysis of the compounds was carried out by RP HPLC coupled to DAD and TOF-MS in order to obtain molecular formula and exact mass. Posterior analyses with IT-MS were performed and the fragmentation pattern and confirmation of the structures were achieved. The H. sabdariffa samples were successfully analyzed in positive and negative ionization modes with two optimized linear gradients. In positive mode, the two most representative anthocyanins and other compounds were identified whereas the phenolic fraction, hydroxycitric acid and its lactone were identified using the negative ionization mode.
Abstract: Fenofibrate changed the expression of chemokine genes in circulating leukocytes of HIV-infected patients with hypertriglyceridemia. The data suggest that fenofibrate when effective in the treatment of lipoprotein abnormalities, may act as a modulator of systemic inflammation. This particular action, therefore, may also influence the clinical course of the disease.
Abstract: To assess the role of monocyte chemoattractant protein-1 (MCP-1/CCL2) in the development of fatty liver, we have used LDLr(-/-) mice as an animal model of high-fat, high-cholesterol diet-induced liver steatosis. The rapid dietary induction of hepatic mRNA MCP-1 expression was paralleled by a concomitant increase in plasma MCP-1 that was strongly associated with the degree of liver steatosis. Hepatocytes showed an intense immunoreactivity for MCP-1 that was mainly located surrounding the hepatic lipid droplets. The intake of cholesterol also increased the concentration of MCP-1 in liver homogenates. This was accompanied by a differential expression of members of the PPAR family. Additionally, complete MCP-1 deficiency prevents the development of liver steatosis in LDLr(-/-) mice and partial deficiency is accompanied by a certain protective effect. Our data also suggest that MCP-1 may be important in the regulation of hepatic insulin resistance and may represent a link between inflammation and metabolic diseases. We conclude that dietary cholesterol upregulation of hepatic MCP-1 may help to understand the role of circulating MCP-1 in conditions where liver derangements are clinically important and in the association of liver steatosis with the metabolic syndrome.
Abstract: BACKGROUND: Fatty acid synthase (FASN) is an enzyme synthesized by the liver and plays an important role in lipogenesis. The present study aimed to assess whether serum FASN concentrations are altered in patients with chronic liver disease, and to investigate whether its measurement may be a useful tool in the clinical evaluation of this derangement. METHODS: We investigated 93 patients with chronic liver disease (14 minimal change disease, 79 steatohepatitis) and 100 control subjects. Serum FASN concentrations were measured using ELISA. RESULTS: Patients had a significant increase in serum FASN concentration (p<0.001), which was specifically associated with the hepatic Knodell sub-index III of portal inflammation (p=0.019). In addition, serum FASN concentrations were significantly correlated with the circulating levels of the monocyte chemoattractant protein-1 (MCP-1) (Spearman rho=0.375; p<0.001) and type III procollagen-N-peptide (P-III-P) (Spearman rho=0.297; p<0.001). CONCLUSIONS: Serum FASN concentrations are increased in patients with chronic liver impairment, and are associated with specific histological alterations and biochemical markers of portal inflammation. These data suggest that FASN measurement may contribute significantly to the evaluation of these patients.
Abstract: AIMS: We investigated the analytical performance of a new assay of the lactonase activity of paraoxonase-1 and its efficacy in the assessment of liver damage. DESIGN AND METHODS: Serum lactonase activity was determined by the hydrolysis of 5-thiobutyl butyrolactone in 633 healthy individuals and 369 patients with chronic liver disease. Paraoxonase-1, 2, and 3 gene polymorphisms were analyzed by the MassArray method. RESULTS: Linearity was up to 10 U/L. Detection limit was 0.12 U/L. Imprecision was < or = 17.7%. Lactonase values in our normal population were 5.99 (3.29-13.61) U/L. Lactonase activity showed a lower influence of genetic polymorphisms than the classical assay using paraoxon. Both measurements showed a similar efficiency in testing for liver dysfunction. CONCLUSION: We report a reliable assay using a non-toxic substrate for the measurement of serum lactonase activity. The influence of genetic variability is low. The assay could be a useful addition to tests evaluating liver impairment.
Abstract: Monocyte chemoattractant protein-1 (MCP-1) plays a relevant role in macrophage migration but recent findings suggest an additional role in lipid and glucose metabolism. We report the use of (1)H NMR spectroscopy as a useful complementary method to assess the metabolic function of this gene in a comparative strategy. This metabonomic analysis was rapid, simple, quantitative and reproducible, and revealed a suggestive relationship between the expression of the MCP-1 gene and hepatic glucose and taurine concentrations. This approach should be considered in genetically modified mice when a metabolic alteration is suspected, or in routine assessment of metabolic phenotype.
Abstract: Under certain clinical circumstances, folic acid can have undesirable effects. We investigated the following: (i) the effects of moderately high folic acid supplementation on the course of liver impairment in CCl(4)-treated rats and (ii) the influence of folic acid supplements on the hepatic recovery following the interruption of the CCl(4)-induced toxic injury. Four experimental groups of rats were used: CCl(4)-treated rats (0.5 ml of CCl(4) twice a week i.p.) fed standard chow for up to 12 weeks (Group A); treated rats fed chow supplemented with 25 mg/kg folic acid from weeks 6 to 12 (Group B); treated rats fed a standard diet but with CCl(4) discontinued after 6 weeks to allow for tissue recovery over 4 weeks (Group C); rats as Group C but fed a diet supplemented with 25 mg/kg folic acid from weeks 6 to 10 (Group D). Liver and blood samples were obtained for biochemical, histological, and gene expression analyses. Animals that received the supplement had a higher content of collagen, activated stellate cells, and apoptotic parenchymal cells in biopsy tissue at weeks 8 and 10 of treatment and more extensive alterations in serum albumin and bilirubin concentrations (Group B vs. Group A). In some of the time periods analyzed, alterations were observed in the expression of genes related to apoptosis (B-cell leukemia/lymphoma 2, inhibitor of apoptosis 2) and to fibrosis (procollagen I, matrix metalloproteinase 7). In the recovery period (Groups C and D), folic acid administration was associated with increased hepatic inflammation and apoptosis and with a decrease in the tissue inhibitor of metalloproteinase-3 expression following 1 week of recovery. We conclude that folic acid administration aggravates the development of fibrosis in CCl(4)-treated rats. Follow-up studies are needed to determine whether folic acid treatment would be contraindicated in patients with chronic liver diseases.
Abstract: A method for selective extraction using SPE, electrophoretic separation at basic condition and the identification by using exact masses and fragmentation patterns has been developed in order to know the anthocyanins in dried calyces of Hibiscus sabdariffa L. A detailed and comparative study of several extraction procedures has been carried out to obtain the maximum number of anthocyanidins from the calyces and then a CE-TOF-MS method in positive mode using ESI has been developed for the separation and rapid identification of anthocyanins in H. sabdariffa L. Delphinidin-3-sambubioside, cyanidin-3-sambubioside have been detected as main components and cyanidin-3-O-rutinoside, delphinidin-3-O-glucoside and cyanidin-3,5-diglucoside, and chlorogenic acid as minor constituents. The confirmation of the anthocyanidins and chlorogenic acid was carried out using fragmentation ions with the IT-mass spectrometer (IT-MS).
Abstract: The paraoxonase (PON) enzyme family, comprising PON1, PON2, and PON3, are antioxidant enzymes that degrade oxidised phospholipids. We describe the immunohistochemical localisation of the PON proteins in the normal mouse. Antibodies were obtained by inoculating rabbits with peptides derived from specific sequences of mature PONs. PON1 and PON3 were detected in the skin external epithelium, acini of the sebaceous glands, tongue epithelium, acini of the submandibular gland, surface epithelia of the stomach and the intestine, hepatocytes, exocrine pancreas acini, fibre tracts of the encephalon and the spinal cord, skeletal and cardiac muscle, eye lens epithelium and retinal layers, adipocytes, chondrocytes, epithelial cells of the trachea and bronchiole, ovary follicular fluid, seminiferous tubules, spermatozoa, and kidney proximal tubules. PON2 expression was weaker than that of PON1 and PON3, and was absent in some of the tissues studied, such as submandibular gland, nerve cells, and adipocytes. In muscle cells, PON2 expression was restricted to the endomysium. Apolipoprotein A-I did not colocalise with PONs, suggesting local synthesis. This study provides an experimental model to investigate the role played by these enzymes as antioxidants and their relationship with the development of a variety of diseases.
Abstract: Oxidative stress has a primary role in the pathogenesis of severe acute pancreatitis. Then, the antioxidant capacity is a critical factor in the progression of this disease. Serum paraoxonase-1 (PON1) is an esterase associated with high-density lipoprotein, which clinical interest resides in its ability to prevent or limit the lipid oxidation. The aim of this study was to investigate changes in PON1 activity in the early stages of acute pancreatitis and to find out if its alteration is related with the severity of the disease. To this purpose, we used an experimental model of taurocholate-induced mild and severe acute pancreatitis. Our results showed that serum activity and PON1 concentration decreased 18 h after the induction of a severe acute pancreatitis. In vitro analysis revealed that incubation with oxidized lipids obtained from pancreatitis samples results in the inactivation of the enzyme in a concentration-dependent manner. In addition to oxidative inactivation, we observed by Western blot, an immunoreactive band suggestive of proteolytic degradation of the enzyme, altogether indicating that during severe acute pancreatitis, there is a significant decrease in serum PON1 activity. This decrease is related with inactivation of the enzyme by oxidized lipids, probably followed by proteolytic degradation of the enzyme.
Abstract: Chronic liver diseases are accompanied by changes in the biochemical pathways related to the regulation of apoptosis and extra-cellular matrix deposition. The present study was designed to investigate, using low density arrays, changes in the hepatic gene expression together with hepatic biochemical and histological alterations in rats that had liver impairment induced by chronic exposure to CCl(4). Further, we examined the possible recovery of genetic and pathological changes following the cessation of the hepatotoxic injury. Experimental fibrosis was induced in male Wistar rats by CCl(4) administration. Animals were subdivided into two groups. One group was given CCl(4 )and animals were killed at 8 and 12 weeks of treatment. The other group was treated with CCl(4) for 6 weeks, the CCl(4 )was then stopped and, subsequently, subgroups of animals were killed after 1 and 2 weeks of recovery. CCl(4) administration over 12 weeks was associated with significant changes in B-cell leukemia/lymphoma 2, procollagen type I alpha 2, matrix metalloproteinases 3 and 8, tissue inhibitors of metalloproteinases 1, 2, and 3 and the inhibitor of apoptosis 4 gene expressions. Recovery after CCl(4) cessation was associated with changes in procollagen type I alpha 2, matrix metalloproteinase 7, tissue inhibitors of metalloproteinases 1 and 2, inhibitor of apoptosis 4, and survivin gene expressions. This study shows an association between changes in the expression of several genes regulating hepatic cell apoptosis, the fibrosis process, and the recovery of the hepatic function after removal of the toxic injury.
Abstract: BACKGROUND AND PURPOSE: The initial steps of atherosclerosis and the entry of HIV into the cell share similar biological mechanisms. Therefore, our hypothesis is that the progression of atherosclerosis in patients with HIV infection can be influenced by variations in genes implicated in both processes. METHODS: The progression of atherosclerosis over a 2-year follow-up period was measured as the combined carotid and femoral intima media thickness (IMT) in 141 patients with HIV infection. The DeltaIMT (IMT(follow-up)-IMT(baseline)) values were used to segregate patients as minimal progressors or regressors (lowest DeltaIMT tertile), slow progressors (mid DeltaIMT tertile), and rapid progressors (highest DeltaIMT tertile). Mutations CCR-5Delta32, CCR-2 64I, MCP-1-2518G, SDF1-3'A, and CX3CR-1 (T280 mol/L and V249I) in the host DNA were determined. Mean age of the patients was 38.96 (SEM: 0.61) and 68.8% were male. The mean DeltaIMT was 0.045 mm (0.01) per year, which represented a significant progression (P<0.001) with respect to baseline values. Patients with minimal progression or regression had a significantly (P=0.01) higher CD4 cell count than slow progressors and rapid progressors. Multivariate analyses indicated that age and total cholesterol were positively associated with IMT progression. In contrast, the CD4 cell count, the SDF1-3'A, and the CX3CR-1 249 I mutated alleles were associated with lesser IMT progression. CONCLUSIONS: The course of atherosclerosis in patients with HIV infection is influenced by polymorphisms in the SDF1 and CX3CR1 genes by metabolic variables and by the CD4 cell count. These data would be of help in assessing therapeutic needs of these patients.
Abstract: We describe the effect of MCP-1 deficiency in mice rendered hyperlipemic by the concomitant ablation of the LDL receptor. The MCP-1(-/-)LDLr(-/-) mice in comparison with LDLr(-/-) mice showed a decreased lipoprotein clearance, derangements in free fatty acids delivery and less glucose tolerance when fed a regular chow, and they showed a partial resistance to alterations in glucose and lipid metabolism induced by dietary fat and cholesterol. They also were less prone to the development of diet-induced obesity. Our results suggest that the role of MCP-1 in metabolism is relevant and that, although new hidden complexities are evident, the function of MCP-1/CCL2 extends far beyond the monocyte chemoattractant effect. Therefore, the regulatory mechanisms influenced by MCP-1 should be fully ascertained to understand the metabolic consequences of inflammation and before considering MCP-1 as a therapeutic target.
Abstract: The determinant factors for the development of atherosclerosis in response to dietary cholesterol were examined in two animal models to assess the comparability of results. We studied 128 male Apo E(-/-) and 128LDLr(-/-) mice randomly assigned to baseline (n=8) and 5 groups (n=24 each) that differed only in their dietary fat and cholesterol supplements. At 10, 16, 24 and 32 weeks of age, 8 animals from each group were sequentially sacrificed and the variables analyzed. The lesion sizes changed at different rates but they were predictable and did not differ in complexity. We observed, however, significant differences between strains, particularly in the constitutive expression of liver genes, their metabolic response to dietary cholesterol, their feeding behaviour, their glucose tolerance and the gain in body weight. Both strains presented characteristics that resemble steatohepatitis but manifestations were more severe in LDLr(-/-) mice. The divergent responses indicate that the choice of the diet and the model should be carefully considered in atherosclerosis studies and extrapolations interpreted with caution.
Abstract: Atherosclerosis is an inflammatory disease in which several chemokines are implicated. The roles of these molecules extend from the recruitment of circulating inflammatory cells to the activation of inflammatory and pro-thrombotic cascades, which ultimately leads to an atherosclerosis-related event. One of the most studied chemokines is monocyte chemoattractant protein-1 (CCL2), which has been strongly linked to atherosclerosis in both animal and human studies. The higher the expression of either the CCL2 gene or its receptor CCR-2, the higher the likelihood of developing atherosclerosis in genetically-modified animals. Conversely, the deletion of either CCL2 or its receptor is followed by a significant reduction in the development of atherosclerotic plaques. Studies in humans yield controversial results. Most of these studies linked the plasma CCL2 concentration to the occurrence of atherosclerosis or related events; however, this relationship does not seem to be independent of the classical, known risk factors. Currently, there are no suitable analytical tools to reach strong conclusions with respect to the value of plasma CCL2 concentration as a biomarker of atherosclerosis, but experimental evidence suggests that the CCL2/CCR2 pathway should be further explored as a diagnostic, prognostic and therapeutic target.
Abstract: 1. Patients with advanced chronic renal disease and anaemia have decreased serum paraoxonase-1 (PON1) activity and an increased degree of oxidative stress compared with normal subjects. The present study investigated the effects of treatment of anaemia with exogenous recombinant erythropoietin (EPO) beta and iron on levels of antibodies against oxidized low-density lipoproteins (ox-LDL), as well as on serum PON1 activity and concentration, in predialysis patients with chronic renal disease. 2. Forty-nine patients with chronic renal failure and haemoglobin (Hb) < 11 g/dL were treated over a period of 6 months with EPObeta (80-120 U/kg per week, s.c.) and variable doses of iron. Selected biochemical variables were determined before and after treatment. 3. Treatment with EPObeta and iron was associated with a significant increase in mean (+/-SD) blood Hb concentration compared with pretreatment values (12.8 +/- 1.5 vs 9.9 +/- 0.6 g/dL, respectively; P < 0.001). The average dose of EPObeta was 6160 +/- 3000 U/week. After 6 months of treatment, compared with pretreatment values, the median levels (95% confidence intervals) of antibodies against ox-LDL were decreased (17.5 (10.6-24.4) vs 24.8 (11.5-38.1) U/mL, respectively; P < 0.001), serum PON1 activity was slightly but significantly increased (123.6 (76.1-343.6) vs 101.0 (50.0-332.5) U/L, respectively; P = 0.016) and the concentration of PON1 was significantly decreased (37.3 (11.8-76.2) vs 46.7 (24.6-98.0) mg/L, respectively; P < 0.001). There were no significant changes in total cholesterol, triglycerides or cholesterol fraction concentrations before and after treatment. 4. We suggest that EPObeta and iron treatment of anaemia promotes significant changes in serum PON1 activity and concentration and has a beneficial effect on oxidative stress in predialysis patients with chronic renal disease.
Abstract: OBJECTIVES: To investigate the relationship between serum paraoxonase-1 and liver damage in chronic alcoholic patients. To assess the diagnostic accuracy of paraoxonase-1 plus standard biochemical tests in the assessment of liver damage in alcoholics. DESIGN AND METHODS: We studied 328 chronic alcoholics and 368 healthy individuals. RESULTS: Paraoxonase-1 activity was decreased and the concentration was increased in alcoholics (P<0.001). The enzyme activity was correlated with albumin (r=0.45; P<0.001) and prothrombin time (r=0.49; P<0.001). Addition of paraoxonase-1 activity measurement to a battery of biochemical tests increased the sensitivity in differentiating between patients and controls up to 96.6% but did not improve the sensitivity in differentiating between subgroups of alcoholics. CONCLUSIONS: Paraoxonase-1 was related to the severity of alcoholic liver disease. Its measurement was useful in discriminating between patients and healthy subjects, but did not add any valuable information in subgroups of alcoholics.
Abstract: BACKGROUND: Higher high-density lipoprotein concentrations are associated with a better disease course in HIV-infected patients. Paraoxonase-1, an enzyme contained within high-density lipoproteins, is thought to hydrolyse oxidised lipids. The aim of the present study was to investigate the relationships between HIV infection and the circulating activity and concentration of paraoxonase-1, and the concentration of high-density lipoproteins, apolipoprotein A-I and oxidised low-density lipoproteins. METHODS: We studied patients with HIV infection (n=212) and healthy subjects (n=409). In all the participants we measured the relevant biochemical and genetic variables. The statistical associations between these variables and paraoxonase-1 activity and concentration were assessed using multiple linear regression analysis. RESULTS: Serum paraoxonase-1 activity was decreased (P<0.001) and its concentration was increased (P=0.017) in HIV-patients compared to the controls. HIV infected patients had lower HDL-cholesterol and apolipoprotein A-I concentrations. Multivariate regression analysis showed that serum paraoxonase-1 activity was associated with the CD4+ T lymphocyte count (P<0.05), apolipoprotein A-I (P<0.001), and paraoxonase-1 genetic polymorphisms (P<0.001). Paraoxonase-1 concentration was associated with that of serum beta-2-microglobulin (P<0.001). CONCLUSIONS: Both, paraoxonase-1 activity and concentration were influenced by HIV-infection and these were related to alterations in HDL composition and the immunological status of the patients.
Abstract: BACKGROUND: Antiretroviral drug efficacy has been widely studied in relation to viral factors. Mutations in the HIV co-receptors and their natural chemokines, however, may be critical in HIV infection and treatment response. We compared the efficacy of protease inhibitor (PI) treatment among PI-naïve patients grouped according to whether they carried the chemokine CC motif receptor 2 (CCR-2) 64I and monocyte chemoattractant protein 1 (MCP-1)-2518G alleles. METHODS AND RESULTS: HIV-infected patients who were PI-naive were selected for the study (n=164) but there was no restriction on lymphocyte CD4 count or plasma HIV viral load. Follow-up was for the first 24 months of treatment. Clinical and laboratory data were obtained every 3 months. All the participants were genotyped for the MCP-1-2518G, CCR-2 64I, CCR-5Delta32 and stromal derived factor 1 (SDF1) 3'A mutated alleles. The results indicated that patients carrying the mutated allele of MCP-1 had a higher mean CD4 cell count throughout the follow-up period than those with the common allele (P=0.01). Also, patients with the MCP-1 and CCR-2 mutated alleles were more likely to continue to have an undetectable viral load following treatment (P=0.05). CONCLUSION: A better response to PI treatment appears to be conferred by mutations in the host MCP-1 and CCR-2 genes, and may be related to the cellular axis-of-entry used by the retrovirus.
Abstract: The aim of this study was to investigate the longitudinal changes in serum paraoxonase-1 (PON1) activity from preconception throughout normal pregnancy and their relationships with maternal dietary vitamin C and E intake. The study was performed in 35 women (studied at preconception, at 8, 20 and 32 weeks of pregnancy, and at labour). PON1 activity decreased significantly from 145.8 (109.8-198.8) U/L at preconception to 111.1 (85.3-179.9) U/L (p<0.01) at 32 weeks and 100.4 (54.7-171.4) U/L (p<0.001) at labour. There was a direct association between vitamin C intake and PON1 at week 32 (p=0.018). We conclude that adequate vitamin C intake in pregnant women may merit consideration, since vitamin C supplementation has proved beneficial in the prevention of preeclampsia in women at increased risk of this condition.
Abstract: Individuals with HIV-1 infection are at increased risk for cardiovascular events, and lipodystrophy is generally associated with pro-atherogenic metabolic disturbances. We conducted a case-control study to assess the presence of sub-clinical atherosclerosis in HIV-1-infected patients with or without lipodystrophy (LD) and to evaluate the influence of monocyte chemoattractant protein-1 (MCP-1) on the development of both carotid atherosclerosis and LD. The study population consisted of 43 patients with LD and 86 patients without LD. We determined carotid intima-media thickness (IMT), MCP-1 concentrations in plasma, and MCP-1 genotype (presence or absence of the -2518G allele). HIV-1-infected patients with LD showed increased risk (OR=3.71, 95% CI=1.10-12.47, p=0.03) for sub-clinical atherosclerosis, and MCP-1 plasma concentration was significantly correlated with IMT in these patients (Pearson=0.31, p=0.03). Furthermore, presence of LD was a determinant for MCP-1 plasma concentration (beta=0.18, p=0.05). In summary, HIV-1-infected patients with clinically manifest LD are at higher risk for atherosclerosis and our observations support the relationship between inflammation and atherosclerotic disease.
Abstract: There are increasing evidences showing that inflammation participates in atherosclerosis. Therefore, the therapeutic use of anti-inflammatory agents should be considered. We have induced chronic, aseptic inflammation upon the injection of turpentine and tested the effect of dexamethasone on lipoprotein metabolism and, consequently, atherosclerosis in apolipoprotein E-deficient mice. Aseptic inflammation caused a significant decrease in hyperlipidemia. Treatment with dexamethasone elicited the opposite effect increasing hyperlipidemia through mechanisms related to the increase in the synthesis of triglyceride-rich lipoproteins. Changes in plasma lipids correlated with those observed in the size of atherosclerotic lesions. Our data suggest the presence of a common mechanism present in both observations and which is probably related to the cytokine secretion. Among the candidates, we chose to test the effect of interleukin-6 because it is involved in both processes, atherosclerosis and inflammation, and its expression is efficiently repressed by corticosteroids. The injection of recombinant interleukin-6 in our mice elicited the same effects observed in our model of inflammation. We conclude that manipulation of inflammation-related mechanisms modulates lipid homeostasis and development of atherosclerotic plaque in rodents.
Abstract: Highly active antiretroviral therapy in Human Immunodeficiency Virus (HIV) has been associated with lipodystrophy, insulin resistance and atherosclerosis. We investigated the effects of rosiglitazone or metformin on fasting and postprandial inflammatory and antioxidant variables in HIV-infected males with lipodystrophy. Thirty-one patients were randomly assigned to receive either rosiglitazone (4 mg twice daily) or metformin (1 g twice daily) for 26 weeks. At baseline and after treatment, standardized 10-h oral fat loading tests were performed. Before treatment, inflammatory variables remained unchanged but there was a postprandial decrease in high density lipoprotein (HDL)-cholesterol and paraoxonase (PON1) activity. Rosiglitazone and metformin reduced homeostasis model assessment index (HOMA) similarly (-34% and -37%, respectively, P<0.05 for each). Both treatments increased fasting and postprandial PON1 activity and decreased postprandial monocyte chemoattractant protein 1 (MCP-1) concentrations. However, plasma C-reactive protein (CRP) and Interleukin-6 (IL-6) concentration did not change throughout the study. To decrease insulin resistance results in a higher anti-oxidant and consequent lower pro-inflammatory action of HDL. This may confer protection against accelerated atherosclerosis in these patients.
Abstract: 1. In the present study, we have explored the effect of rosiglitazone on post-prandial paraoxonase (PON)-1, an enzyme with potent anti-oxidant properties that may protect against atherosclerosis because increased post-prandial lipaemia, although sometimes understated, is part of the diabetic dyslipidaemia. 2. A randomized, cross-over, placebo-controlled, double-blind clinical trial was performed. Participants (19 type 2 diabetic patients on oral antihyperglycaemic agents) were randomly assigned to receive either placebo or rosiglitazone 4 mg twice daily for 8 weeks. After a 6 week wash-out, the alternative treatment was implemented. Standardized 6 h oral fat-loading tests were performed after each treatment period. 3. Patients assigned to rosiglitazone had increased fasting PON-1 activity (from 331 +/- 29 to 362 +/- 32 U/L before treatment vs after treatment, respectively; P = 0.015), although the PON-1 mass did not change (68.8 +/- 21.1 vs 64.2 +/- 25.4 mg/L before treatment vs after treatment, respectively). In addition, rosiglitazone significantly decreased fasting plasma peroxides compared with placebo (162 +/- 25 vs 214 +/- 28 mmol/L, respectively; P = 0.019). The post-prandial fall in PON-1 activity, expressed as area under the curve, was attenuated by rosiglitazone (-97 +/- 14 vs-161 +/- 24 Uh/L for rosiglitazone vs placebo, respectively; P = 0.02) and the increase in PON-1 activity caused by rosiglitazone correlated with reductions in fasting plasma glucose (r = -0.42; P < 0.05), homeostatic model assessment index (r = -0.59; P < 0.01) and peroxides (r = -0.40; P = 0.07). 4. The present data indicate that rosiglitazone may convey increased protection against the oxidative modification that represents increased post-prandial lipaemia.
Abstract: OBJECTIVES: We investigated functional and structural markers of atherosclerosis in human immunodeficiency virus (HIV)-infected patients in relation to the presence of the metabolic syndrome (MS). BACKGROUND: Antiretroviral combination therapy in HIV has been associated with cardiovascular risk factors that cluster in the MS. METHODS: Thirty-seven HIV-infected patients underwent assessment of flow-mediated vasodilation (FMD), aortic pulse-wave velocity (PWV), and carotid intima-media thickness (IMT). Age-matched type 2 diabetic patients (n = 13) and healthy controls (n = 14) served as reference groups. RESULTS: Fifteen HIV-infected patients (41%) fulfilled the National Cholesterol Education Program criteria of the MS. The FMD was similarly impaired in HIV-infected patients without the MS (MS- group) and the diabetic patients (5.1 +/- 0.4% and 4.9 +/- 0.6%, respectively) compared with controls (8.8 +/- 0.7%). The HIV-infected patients with the MS (MS+ group) had even more impaired FMD (2.5 +/- 0.3%). Carotid IMT was similarly increased in the MS+ group and the diabetic patients compared with the other groups. Aortic PWV was increased in the diabetic patients only. In HIV-infected patients, FMD was related to metabolic parameters, whereas aortic PWV and IMT were related to parameters of HIV infection, time on antiretroviral combination therapy, inflammatory (C-reactive protein and leukocytes) and metabolic parameters. CONCLUSIONS: The data of the present study suggest an increased cardiovascular risk in HIV-infected patients, even in the absence of clustering of metabolic risk variables. The presence of the MS in HIV is associated with even more advanced atherosclerotic changes. Presumably, both HIV infection and antiretroviral therapy may promote atherosclerosis through mechanisms involving endothelial cells, either directly or indirectly via metabolic risk factors.
Abstract: In humans, hypercholesterolemia, steatohepatitis, and risk for arteriosclerosis are associated. Apolipoprotein E-deficient mice, a widely used animal model, show both arteriosclerosis and steatohepatitis in response to high-fat and cholesterol diets. We have found a relationship between these conditions and a higher mRNA aortic and hepatic monocyte chemoattractant protein-1 (mcp-1) gene expression. Both tissues respond in a similar way when dietary cholesterol is provided for a few weeks but differently if the conditions persist for a protracted period of time. After 8 months of treatment, the mcp-1 gene expression in the aorta continues increasing but in the liver decreases. This coincides with a significant increase in hepatic ppar-delta anti-inflammatory gene expression. Apparently, the arterial wall cannot prevent the deleterious effects of higher mcp-1 expression by increasing ppar-delta gene expression and the lesion progress. However, in the liver, the activation of anti-inflammatory genes may reduce the hepatic mcp-1 expression which significantly decreases the inflammatory response. This differential inflammatory gene expression in aorta and liver may support the idea that anti-inflammatory transcription factors are involved in the response to diet and inflammation. Therefore, the use of cholesterol-enriched diets should be carefully considered in the apolipoprotein E-deficient mice because they may trigger different stimuli and seriously hinder the interpretation of possible findings.
Abstract: BACKGROUND: Monocyte chemoattractant protein-1 (MCP-1) plays a crucial role in atherosclerosis and it has been recently proposed as a surrogate biomarker of long-term clinical outcomes in patients with acute myocardial infarction. Little is known of the factors that may influence plasma MCP-1 concentrations. METHODS: We studied 384 healthy volunteers and 226 HIV-infected patients as a model of chronic inflammatory condition that predisposes to sub-clinical atherosclerosis. RESULTS: In healthy participants there were significant associations between plasma MCP-1 concentration and age, smoking status, and serum triglyceride concentrations that were not observed in the HIV-infected patients. The plasma concentration of MCP-1 was significantly associated with the polymorphism at position -2518 of the MCP-1 gene and, in patients, with the carotid artery intima-media thickness. There were also significant correlations indicating a close association between MCP-1 and HIV disease activity. However, in a multiple regression model, only age, the MCP-1 genotype and smoking status showed significant, and independent, associations with plasma MCP-1 concentrations. CONCLUSION: Plasma MCP-1 concentration is genetically determined and associated with age and smoking habit and it also correlates with subclinical atherosclerosis in HIV-infected patients.
Abstract: BACKGROUND/AIMS: We previously reported that paraoxonase-1 activity measurement may be useful for the evaluation of liver diseases. Because oxidative stress plays a role in liver apoptosis, and lipid peroxides are hydrolyzed by paraoxonase-1, we have extended our studies to explore the relationships between this enzyme and oxidative stress, fibrosis and apoptosis. METHODS: We measured paraoxonase-1 activity and concentration, soluble FAS concentration, serum fibrosis markers, and total peroxides in a group of patients with minimal hepatic changes (n=25), chronic hepatitis (n=51), or liver cirrhosis (n=17). We also measured the Knodell activity index in liver biopsies and performed FAS and PON1 immunostaining. RESULTS: Patients with liver diseases showed an increase in soluble FAS, fibrosis markers and paraoxonase-1 concentrations, as well as a decrease in PON1 activity. Paroxonase-1 activity and concentration were correlated with soluble FAS (r=-0.43, P<0.001 and r=0.27, P=0.007, respectively). Paraoxonase-1 concentration showed a significant inverse association with FAS immunostaining (P=0.013) and a direct association with PON1 immunostaining (P<0.001). CONCLUSIONS: These results suggest an active role of PON1 in the regulation of oxidative stress, fibrosis and hepatic cell apoptosis in chronic liver diseases.
Abstract: The impaired mitochondrial function hypothesis in schizophrenia is based on evidence of altered brain metabolism, morphology, biochemistry and gene expression. Mitochondria have their own genome, which is needed to synthesize some of the subunits of the respiratory chain enzymes. Mitochondrial DNA (mtDNA) is maternally inherited and we observed an excess of maternal transmission of schizophrenia in a set of parent-offspring affected pairs. We therefore hypothesized that mutations in the mtDNA may contribute to the complex genetic basis of schizophrenia. The entire mtDNA of six schizophrenic patients with an apparent maternal transmission of the disease was sequenced and compared to the reference sequence. We have identified 50 variants and among these six have not been previously reported. Three of them were missense variants: MTCO2 7750C>A, MTATP6 8857G>A and MTND4 12096T>A. These were maternally inherited because they were also present in the mtDNA of their respective schizophrenic mothers and none of them were found in 95 control individuals. The MTND4 12096T>A (Leu446His) is a heteroplasmic variant present in five of the six mother-offspring patient pairs that triggers a non-conservative substitution in the ND4 subunit of complex I. Sequence alignment of 110 ND4 peptides from all eukaryotic kingdoms shows that only hydrophobic amino acids are found in this position. Moreover, leucine was conserved or substituted by an isoleucine in all mammalian species. This indicates that the presence of histidine could affect complex I activity in patients with schizophrenia.
Abstract: OBJECTIVE: We postulated that in type 2 diabetes, the postprandial phase is a pro-inflammatory state that can be modulated by PPAR-gamma agonists. For this purpose, we determined the effects of rosiglitazone (8 mg/d) on postprandial leukocyte counts and pro-inflammatory cytokines (IL-6 and IL-8) in patients with type 2 diabetes. METHODS AND RESULTS: A randomized, 8-week, cross-over, placebo-controlled, double-blind clinical trial was performed in 19 patients with type 2 diabetes. Standardized 6-h oral fat-loading tests were performed after each treatment period. During placebo treatment, blood leukocytes increased to a maximum 6-h postprandially, due to significant increases in neutrophils and lymphocytes. Concomitant postprandial increases were observed for IL-6 and IL-8, the major chemokines responsible for leukocyte recruitment. Rosiglitazone reduced the incremental area under the curves (dAUCs) for IL-6 (-63%, p<0.01) and IL-8 (-16%, p<0.05). The dAUC for leukocytes decreased with 37% (p<0.05), due to a specific reduction of neutrophils (-39%, p<0.05). CONCLUSIONS: Rosiglitazone attenuated the postprandial increases of neutrophils, IL-6 and IL-8 in patients with type 2 diabetes. Since inflammation is a major force driving atherosclerosis, and man lives in a postprandial period most part of the day, a reduced inflammatory response after a meal may delay progression of atherosclerosis. CONDENSED ABSTRACT: We postulated that in type 2 diabetes, the postprandial phase is a pro-inflammatory state that can be modulated by PPAR-gamma agonists. Rosiglitazone attenuated the postprandial increases of neutrophils, IL-6 and IL-8 in patients with type 2 diabetes. These effects may contribute to cardiovascular risk reduction.
Abstract: C-reactive protein (CRP) has been proposed as an independent risk factor for cardiovascular disease. In this study we sought to investigate the association between several nutritional and lifestyle factors and serum CRP concentration in a population-based study. We studied 359 individuals (172 women, 187 men; age range 18-75 years) randomly selected from the town hall's registers and assessed their daily dietary intake using a 3-day estimated-food record. The median serum CRP concentration was 1.40 mg/L (range <0.10-47.48 mg/L; geometric mean 1.20 mg/L). We noted significant and independent direct associations between CRP and age, body-mass index, female sex, and serum triglyceride concentration. Bivariate analysis showed a significant inverse association between CRP and many nutrients (e.g., carbohydrates, proteins, lipids, thiamine, pyridoxine, tocopherol, and folate), but multiple-regression analysis indicated that only the effect of dietary folate intake was not dependent on other factors. Differences in folate intake did not produce changes in plasma homocysteine concentration, and we detected no negative correlation between dietary folate intake and log homocysteine (r = .02, P = .711). Strong positive correlations between the intake of folate and numerous other nutrients were found. This population-based study shows that a higher folate intake, in addition to other known constitutive and lifestyle factors, is significantly associated with a lower serum CRP concentration.
Abstract: C-Reactive protein has been associated with several complications of pregnancy. The aims of the present study were: (1) to evaluate a turbidimetric immunoassay for the measurement of C-reactive protein; and (2) to investigate the chronological changes of the levels of this protein from preconception throughout normal pregnancy and its relationship with variables associated with preconception and pregnancy outcome. Inter-assay imprecision was <5% for C-reactive protein >1 mg/L and 18% at a mean value of 0.33 mg/L. The limit of detection was 0.10 mg/L. The method was linear between 0.10 and 30 mg/L. There were no observed interferences from jaundice, hemolysis, lipemia or paraproteinemia at the levels studied. There was good agreement with the nephelometric method. A total of 39 women were studied at preconception, at 8, 20 and 32 weeks of pregnancy, and in labor. Preconception C-reactive protein concentration was 1.17+/-0.18 mg/L and increased (p<0.001) throughout pregnancy up to 5.69+/-0.82 mg/L. Body mass index at preconception and weight gain during pregnancy were the main factors associated with this increase in C-reactive protein.
Abstract: Efavirenz treatment has been associated with increases in HDL-cholesterol concentrations, and the circulating levels of the drug have been related to the multidrug resistance gene 1 (MDR-1) C3435T polymorphism. The changes in the measured lipid parameters were evaluated in 59 HIV-infected patients initiating efavirenz-based treatment at baseline and at 12 months of follow-up. Efavirenz treatment increased HDL-cholesterol. The changes in concentrations appeared to be influenced by the MDR-1 gene polymorphism, in which CC > CT > TT.
Abstract: BACKGROUND: The measurement of soluble transferrin receptor (sTfR) has been proposed as a valuable marker of erythropoietic activity and iron status. However, the possibility that mutations in HFE and/or transferrin genes have a direct effect on this parameter has not been sufficiently investigated. The present report addresses this point in the general population. METHODS: Serum sTfR, ferritin, iron and transferrin, as well as the H63D and the C282Y polymorphisms of the HFE gene and the TF C1/C2 polymorphism of the transferrin gene, were analysed in 348 subjects. RESULTS: We observed significant and independent associations of serum sTfR with sex (2.68+/-1.27 mg/L in men vs. 2.25+/-1.33 in women; P=0.002), H63D polymorphism (2.61+/-1.34 in wild type homozygotes vs. 2.28+/-1.25 in carriers of one or two mutated alleles; P=0.009), and serum iron concentration (r=-0.17; P=0.002). CONCLUSION: The H63D mutation of the HFE gene has a moderate but significant influence on sTfR concentration in the general population, the presence of one or two mutated alleles being associated with an average of 0.27 mg/L less sTfR than nonmutated homozygotes.
Abstract: We aimed to investigate the effect of turpentine-induced inflammation in an atherosclerosis-prone murine model. We have induced a chronic aseptic inflammation in apolipoprotein E-deficient mice, with or without a dietary supplement of aspirin (n = 10, each), by the injection of a mixture (1:1) of turpentine and olive oil in the hind limb twice weekly for a period of 12 weeks. Control animals were injected with olive oil alone (n = 10). The control mice did show any alteration neither in plasma nor at the site of injection. Turpentine-treated mice showed a significant increase in plasma TNF-alpha and SAA concentrations which indicated a systemic inflammatory response that was not substantially affected by aspirin. Also, turpentine injections significantly reduced the plasma cholesterol concentration, probably decreasing intestinal cholesterol re-absorption, and attenuated the size of atherosclerotic lesion. Both effects were minimally influenced by aspirin. The burden of atherosclerosis correlated with plasma lipid levels but not with plasma inflammatory markers. Finally, there was a concomitant decrease in the expression of the hepatic mdr1b gene that correlated with the decrease in plasma cholesterol concentration. Therefore, we conclude that mdr1 is an additional factor to consider in the complexity of alterations in cholesterol metabolism that occur in this model.
Abstract: The present study was aimed (1) to investigate the effect of cholesterol and fat enriched diets on the development of steatohepatitis in apolipoprotein E-knockout mice, and (2) to study the chronological relationships between the development of hepatic alterations, hypercholesterolemia and atherosclerotic lesions in this experimental model. The study consisted of two protocols. Protocol 1 was used in 90 mice subdivided in groups of 18. For 10 weeks, each group was given a diet with different fat and cholesterol contents. Protocol 2 was used in 42 mice, subdivided in four groups. Each group was given a diet enriched with cholesterol and palm oil and they were sacrificed at 8, 13, 18 and 24 weeks of age. Results were as following. (1) Mice given high fat/high cholesterol diets developed an impairment of liver histology consisting of fat accumulation, macrophage proliferation, and inflammation. (2) These effects were modulated by the type of fat: olive oil was mainly associated with macrovesicular steatosis and cholesterol plus palm oil with severe steatohepatitis. (3) There was a chronological and quantitative relationship between liver impairment and the formation of atheromatous lesions. We conclude that apolipoprotein E-knockout mice may be a useful model for investigating the mechanisms of diet-induced steatohepatitis.
Abstract: The IRIS IQ 200 automated urine analyser is a microscope associated to digital imaging software, and is able to classify urine particles into 12 categories and report quantitative results. We evaluated this analyser with respect to linearity and precision, and compared results with microscopic examination of the urine sediment, and with the measurement of urine strips in a CLINITEK 500 analyser. The assay was linear between 10 and 1030 particles/microL. The detection limit was 6 particles/microL. Intra- and inter-assay coefficients of variation were 1.9% and 2.3%. Results by the IQ 200 analyser showed highly significant correlations with those of the urine sediment (erythrocytes: rho=0.68; leukocytes: rho=0.60; epithelial cells: rho=0.66; p<0.001), and urine strips (erythrocytes: rho=0.67; leukocytes: rho=0.66; p<0.001). These results indicate that the IRIS IQ 200 analyser may play a useful part in the automated examination and measurement of urine specimens.
Abstract: BACKGROUND: HIV-infected patients have higher rates of subclinical atherosclerosis. The chemokine stromal derived factor 1 (SDF-1) is the natural ligand for the CXCR4 HIV co-receptor, is highly expressed in atherosclerotic plaques, and the plasma concentration is lower in individuals homozygous for the mutant allele (SDF1-3'A). We tested the influence of SDF1-3'A on atherosclerosis in HIV-infected patients. METHODS: We performed carotid ultrasonography and determined the SDF1-3'A DNA polymorphism in 183 HIV-infected patients. Classical cardiovascular risk factors and antiretroviral therapy were also recorded. From these patients, we selected a group of 134 patients taking protease inhibitor-based antiretroviral therapy and in whom the lipid profile over an 18-month follow-up was collated. RESULTS: We found atherosclerosis in 113 (61.7%) and a lower number of patients with the SDF-1 mutated allele in the group with carotid atherosclerosis compared to those without (41.6% versus 57.1%; P = 0.04). Using a logistic regression analysis, age and dyslipidaemia were significantly associated with atherosclerosis but the SDF1-3'A allele exerted a protective effect on the development of atherosclerosis (odds ratio, 0.45; 95% confidence interval, 0.14-1.02; P = 0.05). Further, we observed that, in the selected group of patients there were lower plasma low-density lipoprotein cholesterol concentrations [mean +/- SEM, 2.06 +/- 0.34 mmol/l] throughout follow up in those patients without carotid lesions and who also carried the mutated SDF1-3'A allele (P = 0.04). CONCLUSION: The SDF1-3'A allele is associated with a lower presence of subclinical carotid atherosclerosis in an HIV-infected population.
Abstract: BACKGROUND: Hepatitis C virus infection is associated with hepatic free radical formation and enhanced lipid peroxidation and an individual's antioxidant status may play an important role. Paraoxonase-1 is an esterase that degrades oxidised lipids. In the present study, we investigated the genetic association of the most important paraoxonase-1 gene polymorphisms and the susceptibility to HCV-related chronic hepatitis. METHODS: Paraoxonase-1 polymorphisms at positions -107, 55 and 192 were analysed from the genomic DNA of 186 patients and 386 healthy volunteers, as well as the serum concentration of total peroxides and standard biochemical tests. RESULTS: Patients with chronic hepatitis had a higher frequency of the RR isoform of the 192 polymorphism than healthy subjects (13% vs. 7%, P<0.05). There were no significant differences with respect to the -107 and 55 polymorphisms. The plasma concentration of peroxides was higher in patients with chronic hepatitis [349.5 (246.2-479.8) vs. 115.4 (95.7-172.3) mumol/L; P<0.001]. CONCLUSIONS: The present study suggests that the paraoxonase-1 192 polymorphism contributes, together with other polymorphisms, to the variations in the host response to HCV infection.
Abstract: BACKGROUND: The use of antiretroviral combination therapy in HIV has been associated with lipodystrophy and cardiovascular risk factors. OBJECTIVE: To compare the effects of the peroxisome proliferator-activated receptor-gamma agonist rosiglitazone and metformin for treating HIV lipodystrophy. DESIGN: An open, randomized, 6-month clinical trial. SETTING: University Medical Center, Utrecht, the Netherlands. PATIENTS: 39 HIV-infected men with lipodystrophy. INTERVENTION: Rosiglitazone, 8 g/d, or metformin, 2 g/d [DOSAGE ERROR CORRECTED] MEASUREMENTS: Insulin sensitivity estimated by the oral glucose tolerance test, subcutaneous and visceral abdominal fat measured by single-slice computed tomography, endothelial function measured by flow-mediated vasodilation, and fasting plasma measurements. Two patients in the metformin group withdrew from the study. Complete case analysis was performed. RESULTS: Compared with metformin, rosiglitazone increased subcutaneous abdominal fat (between-treatment change from baseline, 27 cm2 [95% CI, 7 cm2 to 46 cm2]) and visceral abdominal fat (between-treatment change from baseline, 24 cm2 [CI, 6 cm2 to 51 cm2]). The area under the curve for insulin after the oral glucose tolerance test decreased similarly with both agents, but only rosiglitazone increased adiponectin levels. Metformin showed greater benefits on fasting lipid profile than rosiglitazone. Flow-mediated vasodilation statistically significantly increased with metformin (mean change, 1.5% [CI, 0.4% to 3.3%]) and not with rosiglitazone (mean change, 0.7% [CI, -1.1% to 2.7%]). The metformin versus rosiglitazone increases did not statistically differ. Rosiglitazone and metformin did not change C-reactive protein levels. LIMITATIONS: This small trial was not blinded or placebo-controlled and did not measure clinical outcomes. CONCLUSIONS: The findings emphasize the importance of individualized care in HIV-infected patients. Although rosiglitazone may partly correct lipoatrophy, metformin improves visceral fat accumulation, fasting lipid profile, and endothelial function.
Abstract: It has been suggested that total plasma homocysteine (tHcy) concentrations and methylenetetrahydrofolate reductase (MTHFR) gene polymorphisms are risk factors for schizophrenia. We conducted a case-control study to investigate whether tHcy levels and MTHFR C677T and A1298C variants are associated with schizophrenia, giving special consideration to confounding factors. Logistic regression analysis showed that neither tHcy nor MTHFR polymorphisms were associated with schizophrenia. Homozygosity for MTHFR C677T was associated with higher tHcy concentrations in control and schizophrenia groups (P<0.01), which was mainly driven by the male group. The A1298C variant did not show any association with tHcy concentrations. In conclusion, these results do not confirm an independent relationship of tHcy and MTHFR genotype with risk of schizophrenia.
Abstract: We investigated the relationships between plasma monocyte chemoattractant protein-1, serum C-reactive protein, and the degree of hepatic inflammation in patients with chronic liver disease. Monocyte chemoattractant protein-1 concentration was correlated with the histological hepatic inflammation (estimated by the Knodell index) and with standard liver function tests (P<0.01). C-reactive protein was not correlated with any of the variables studied. These results underline the role of monocyte chemoattractant protein-1 in the pathogenesis of liver impairment and suggest that this chemokine may be a reliable marker of inflammation in hepatic derangements.
Abstract: The interaction of blood with the arterial tree may play an important role in the development of atherosclerotic lesions. The aims of this study were (1) to determine how anemia or increased hematocrit affect the development of atherosclerosis and (2) to find relationships between hematologic and hemorrheologic variables in apolipoprotein (apo) E-deficient mice. Forty-two mice were randomly divided into 3 groups of 14 mice each. There was no further manipulation in the control group. To induce anemia, the mice from one of the groups were repeatedly bled, drawing approximately 250 microL blood from each mouse twice a week. To increase the hematocrit levels in another group of mice, we injected 20 U recombinant human erythropoietin every other day. The development of lesions and the main variables involved in atherogenesis were compared among groups. Our results show that atherosclerosis was attenuated in the mice that were bled, and this was not accounted for by changes in plasma lipid levels, the distribution of lipoprotein particles, the body iron distribution, or oxidation parameters. Moreover, atherosclerosis was enhanced in the mice treated with the continuous administration of erythropoietin. To ascertain the relationship between hematocrit and whole blood viscosity, we measured both variables in pooled blood from 24 additional mice, which were manipulated to ensure a wide range of values. We found a direct and significant correlation between hematocrit and blood viscosity and between hematocrit and lesion size. Our data support in vivo the idea that hemorrheology has an important role in atherogenesis in this particular animal model.
Abstract: Aspirin has potent antiinflammatory properties and attenuates atherosclerosis in apolipoprotein-E-deficient mice fed a high-fat, high-cholesterol diet. In an attempt to clarify the contradictory results obtained with normal chow, we studied the effect of aspirin for a prolonged period of time. The mice were fed a commercial chow until the experiment began at 8 weeks of age. Blood samples were then obtained and several mice (n=8) were sacrificed. The diet of the remaining 48 animals was supplemented with 200 g/kg palm fat and 1 g/kg cholesterol. They were then randomly divided into 2 groups, one of which received 0.5 mg/day of aspirin. The aspirin had a time-dependent effect. First, the extent of lesion decreased; then the effect was neutral; and, finally, after longer periods of being fed the atherogenic diet and receiving aspirin, the extent of the lesion increased. The transitory effect of aspirin should be elucidated in the absence of high dietary lipids.
Abstract: BACKGROUND: Patients infected with HIV present with premature atherosclerosis, and the 2 diseases share common pathogenic pathways. We investigated mutations in the monocyte chemoattractant protein-1 (MCP-1) and CCR-2 genes, which are known to control aspects of these pathways, to ascertain whether they are involved in atherogenesis in these patients. METHODS AND RESULTS: We performed carotid and femoral artery ultrasonography to detect subclinical atherosclerosis in patients infected with HIV (n=183). MCP-1-2518G and CCR-2 64I polymorphisms were determined in the HIV group and in a population-based control group (n=348). We also determined MCP-1 circulating levels in the HIV group. The presence of MCP-1-2518G in the group of patients with subclinical atherosclerosis was significantly higher than in patients without atherosclerotic lesions (47.5% versus 18.2%, respectively; P<0.001). Furthermore, the patients with atherosclerotic lesions had higher MCP-1 plasma concentrations than did patients without lesions (74.15 [4.03] versus 57.81 [3.67] pg/mL, respectively; P=0.03). When adjusted for known cardiovascular risk factors, the MCP-1-2518G allele was associated with subclinical atherosclerosis (OR 5.72, 95% CI 1.74 to 18.80, P=0.004). Compared with measurements conducted approximately 2.5 years earlier in a subset of 40 patients, intima-media thickness (IMT) in the carotid artery progressed at a mean rate of 0.06 mm/y more rapidly in patients bearing the MCP-1-mutated allele (P=0.08). CONCLUSIONS: HIV-infected patients with the MCP-1-2518G allele have a 5-fold increased risk for atherosclerosis, as assessed by ultrasonography.
Abstract: OBJECTIVES: Lipoprotein(a) has been proposed as an independent risk factor for cardiovascular disease. This lipoprotein possesses a marked size polymorphism that makes difficult to measure accurately its concentration in plasma. The International Federation of Clinical Chemistry recently recommended to carefully evaluate new commercial methods for lipoprotein(a) measurement to discard the possible influence of lipoprotein(a) isoforms on immunoreactivity. They also recommended to perform population-based studies for different ethnic and geographic groups. Therefore, in the evaluation of a fully automated, particle-enhanced turbidimetric immunoassay for the measurement of lipoprotein(a) we have determined its reference interval in the Spanish population, an area with the lowest incidence of cardiovascular disease in Europe. DESIGN AND METHOD: We evaluated a commercial kit of reagents calibrated against the Proposed Reference Material and determined the effect of lipoprotein(a) size polymorphism on the measurements. A population-based study was carried out in two different villages on the Mediterranean coast of Spain. RESULTS: Imprecision at different lipoprotein(a) concentrations ranged between 3.0 and 15.4%. Recovery was 98.5 +/- 2.1. Detection limit was 4.8 nmol/L. There were no significant interferences from lipemia, jaundice, hemolysis, paraproteinemia, apolipoprotein B or plasminogen. We did not observe any effect of the lipoprotein(a) size polymorphism on the measurements. Mean (and SD) values for plasma lipoprotein(a) (n = 369) were 53.6 (65.3) nmol/L, the median was = 25.3 nmol/L and range varied between <4.8 and 356.0 nmol/L. CONCLUSION: The present article presents an accurate and practical assay for measuring plasma lipoprotein(a) concentrations and describes its reference values in a population of Spanish Caucasians. Our results are similar to those obtained in other Caucasian populations (between 10 and 25% higher than in participants of the CARDIA study).
Abstract: BACKGROUND: Wolfram syndrome is a neurodegenerative disorder that is inherited in an autosomal recessive mode and characterized by the presence of diabetes mellitus and optic atrophy. Patients and heterozygote carriers are at an increased risk of suffering psychiatric disorders. Mutations in the Wolfram gene (WFS1 ) (4p16.1) are responsible for the development of the disease, and mRNA and protein expression of WFS1 have recently been found in areas of the rat brain that can be related to the psychiatric symptoms. OBJECTIVE: To test the hypothesis that WFS1 mutations in heterozygote carriers or other variants of WFS1 can predispose to mental illness. METHODS: Stage 1: Exons 2, 4 and 8 of that harbour mutations in Spanish Wolfram syndrome families were examined by Single Strand Conformation Polymorphism and sequencing analysis in 43 patients with affective disorder to identify variants and mutations. Stage 2: two variants identified in stage 1 were analysed in 152 psychiatric patients (118 schizophrenia and 34 affective disorder) and 177 control subjects. RESULTS: Six variants (I333V Ile-->Val, F341, N500, R708, K774, K811) and a WFS1 mutation (R818C, Arg-->Cys) were found in the 43 patients analysed in stage 1 of the study. In stage 2, the R818C mutation was not found in the group of psychiatric patients but it was present in one control subject. The association study conducted with the I333V variant did not find significant differences in allele or genotype frequencies between patients and control subjects. CONCLUSIONS: Our results suggest that WFS1 is not a major susceptibility gene for the development of psychiatric disorders in our population.
Abstract: The newly recognised apolipoprotein (apo) AV gene (APOAV) has been linked to fasting plasma triglyceride (TG) concentrations with some polymorphisms associated with elevated fasting TGs. Since fasting plasma TGs are mainly determined by the hepatic production of TG-rich particles (very low density lipoprotein; VLDL), and fasting TGs are the major determinants of postprandial lipaemia, we have evaluated the effects of an APOAV polymorphism on postprandial triglyceridaemia, which is largely determined by the intestinal production and clearance of chylomicrons. For this purpose, diurnal capillary triglyceridaemia (reflecting postprandial lipaemia) was determined in a cohort of 88 healthy volunteers (48 males and 40 females) in relation with a -1131T>C variant in the promoter of APOAV. Thirteen of these subjects (7 males and 6 females) were carriers of the -1131C allele, which has been associated with higher fasting plasma TG levels. The carriers had higher fasting capillary TG concentrations, although plasma TGs were not significantly different from non-carriers in this cohort. Surprisingly, total diurnal triglyceridaemia calculated as the area under the capillary TG curve was similar in carriers compared to non-carriers but after correction for fasting capillary TG levels, incremental diurnal triglyceridaemia was significantly lower in carriers (1.74 (5.27) mmol/h/l) than in non-carriers (4.91 (4.90) mmol/h/l; p = 0.036). The same trends were found for both males and females when analysed separately. Since dietary intake, which is a major determinant of incremental diurnal triglyceridaemia, did not differ between the two groups, we believe that these differences are at least partly explained by the APOAV. In summary, the APOAV assessed by means of the -1131T>C variant seemed to have a paradoxical effect on postprandial lipaemia when compared to fasting TG levels.
Abstract: BACKGROUND: Paraoxonase may protect lipoproteins and cell membranes from peroxidation, and alterations in the activity of this enzyme have been associated with some chronic diseases. Serum paraoxonase appears to be mainly under genetic control, but some studies suggest that environmental factors may also modulate its activity. The aim of the present study was to investigate whether diet and lifestyle affect serum paraoxonase activity. METHODS: We studied a population-based sample of 388 individuals (194 women and 194 men; age range, 18-75 years) and assessed their daily dietary intake using a 3-day estimated food record. The variables studied included serum paraoxonase activity, paraoxonase polymorphisms at positions 55 and 192, age, gender, smoking status, physical exercise, body mass index, energy consumption, nutrient intake (total lipids, saturated fatty acids, beta-carotenes, vitamins C and E), and serum lipid concentrations. RESULTS: Multiple linear regression analysis showed that only genetic polymorphisms, serum cholesterol, HDL-cholesterol concentrations, and cigarette smoking were significant predictors of serum paraoxonase activity. HDL-cholesterol concentrations were also related to body mass index, daily energy consumption, and saturated fatty acid intake. CONCLUSIONS: The between-individual variability of serum paraoxonase activity is regulated mainly by genetic determinants. Although HDL-cholesterol and tobacco smoking may contribute to the modulation of this enzyme, the other nutritional and lifestyle factors do not seem to play a significant role.
Abstract: The aim of the present study was to analyze, on a double-blind basis, the relationships between the apolipoprotein(a) (apo(a)) gene and protein size polymorphisms in healthy volunteers (n = 99) and patients with premature myocardial infarction (n = 91). Apo(a) genotypes were determined by pulse-field electrophoresis and phenotypes were separated by sodium dodecyl sulfate-agarose gel electrophoresis. Results showed that phenotyping overestimated apo(a) size with respect to genotyping (mean (SD) = 3.7 (3.4) kringle units; p < 0.001) in subjects with a double-band genotype, although both measurements were highly correlated (r = 0.83; p < 0.001). We also observed that the protein band in subjects with a single-band phenotype was related more closely to the smallest allele than to the largest allele band. The correlation of plasma lipoprotein(a) (Lp(a)) concentration was stronger with the phenotype than with the genotype. We hypothesize that post-translational modifications in the apo(a) molecule may be the most plausible explanation for the discrepancies observed. In conclusion, the present study highlights the dissimilarities between phenotyping and genotyping methods for the measurement of apo(a) size and suggests that laboratories should carefully consider these relationships and the transfer of results between such methodologies.
Abstract: Exactly how apolipoprotein a [APO(a)] isoform size affects the degree of cardiovascular risk associated with high lipoprotein a [LP(a)] levels is not fully understood. Using a sodium dodecyl sulfate-agarose APO(a) & LP(a) phenotyping method, we assessed the role of APO(a) size heterogeneity according to the number of kringle 4 repeats and the differential APO(a) protein expression in 91 male Spanish patients with premature coronary heart disease (CHD) compared with 99 healthy Spanish men. CHD patients had significantly increased median plasma LP(a) levels (0.31 g/L) and a higher percentage of subjects with LP(a) levels of 0.30 g/L or greater (51%) than controls (0.15 g/L and 23%, respectively). Patients with the double-band phenotype had significantly higher plasma LP(a) levels (median 0.37 g/L) compared with those expressing a single-band phenotype (median 0.20 g/L; P =.018) and with their corresponding controls (median 0.15 g/L; P <.001). The double-band phenotype and LP(a) values of 0.30 g/L or greater had a significant association with CHD (odds ratio [OR] 6.47, 95% confidence interval [CI] 2.51-16.7), stronger than that observed for the entire group (OR 4.19, 95% CI 1.97-8.90). The adjusted OR for the APO(a) protein pattern that equally expressed both isoforms indicates an independent association with premature CHD (OR 3.33; 95% CI 1.08-10.3). These results suggest that APO(a) phenotyping might be used in subjects with hyperlipoproteinemia a as a powerful marker to assess the risk of premature CHD because heterozygous status, mainly when both isoforms are equally expressed, is associated with higher cardiovascular risk.
Abstract: BACKGROUND: Paraoxonase 1 (PON1) is an ester hydrolase present in serum and in the liver. The aims of the present study were to investigate the following: (a) the relationship between serum PON1 activity alterations and the degree of liver damage in patients with chronic liver disease; (b) the influence of genetic variability on serum PON1 activity; and (c) the efficacy of serum PON1 activity measurement, alone and in combination with standard liver function tests, in the assessment of liver damage. METHODS: We studied 68 patients with liver cirrhosis, 107 patients with chronic hepatitis, and 368 apparently healthy volunteers. Baseline and salt-stimulated PON1 activities were measured by the hydrolysis of paraoxon. PON1 genotyping at positions 55 and 192 was analyzed by PCR and restriction isotyping. RESULTS: Baseline and stimulated PON1 activities were decreased (P <0.001) in chronic hepatitis and in liver cirrhosis. PON1 activity was significantly correlated with serum total proteins, albumin, and bilirubin in patients but not in controls. There were no significant differences with respect to allele and genotype frequencies between patients and controls. The combination of baseline serum PON1 with five standard biochemical tests had a higher classification accuracy (94% of patients; 96% of controls) than the five standard tests alone (75% of patients; 96% of controls). ROC plots demonstrated a high diagnostic accuracy for baseline serum PON1 [area under the curve, 0.89 (95% confidence interval, 0.86-0.93) in chronic hepatitis and 0.96 (95% confidence interval, 0.94-0.99) in cirrhosis]. Baseline PON1 provided the highest ROC area for cirrhosis vs controls. CONCLUSIONS: The significant decrease of PON1 activity in chronic liver diseases is related to the degree of hepatic dysfunction and not to allelic or genotypic differences. Addition of serum PON1 activity measurement to the current battery of tests may improve the evaluation of chronic liver diseases.
Abstract: Vitamin E is a lipid-soluble vitamin and an important antioxidant that protects lipoproteins and cell membranes from lipid peroxidation. The aims of the present study were to investigate, in patients with parenchymal liver cirrhosis, the following: (1) nutritional and vitamin E status in relation to compositional changes in lipoproteins; and (2) the effects of these alterations on the patients' plasma susceptibility to copper-mediated oxidation. Patients (n = 55) with liver cirrhosis and 25 healthy volunteers had vitamin E in serum and in isolated lipoprotein fractions analyzed by high-performance liquid chromatography (HPLC). Plasma susceptibility to peroxidation was measured by incubation with Cu(2+). Nutritional status was assessed by anthropometry. Vitamin E concentration was significantly decreased (P <.001) in the serum and in very-low-density lipoprotein (VLDL) and high-density lipoprotein (HDL) in cirrhotic patients. The decrease was related to the degree of liver impairment. There were significant correlations between cholesterol and vitamin E concentrations in serum and in all the lipoprotein fractions (r between 0.72 and 0.89; P <.001) in cirrhotic patients, but there were no significant relationships between vitamin E and any of the anthropometric indices of nutritional status. The plasma maximal oxidation rate was significantly increased in cirrhotic patients (P <.01) and was inversely related to the serum concentration of vitamin E (P <.05). We conclude that lipoprotein alterations and not nutritional factors should be regarded as major factors explaining serum vitamin E reduction in patients with parenchymal liver cirrhosis, and that vitamin E depletion is associated with an increased plasma susceptibility to oxidation.
Abstract: OBJECTIVES: To assess whether paraoxonase (PON1) polymorphisms at positions 55 and 192 and/or their phenotypic expressions influence the risk of myocardial infarction (MI) in Spanish population. DESIGN AND METHODS: Two hundred and fifteen male survivors of a MI and their age-matched controls were included in the study. Lipids, apolipoproteins (apo) A-I and B, PON1 activity on paraoxon and phenylacetate and PON1 polymorphisms were determined. RESULTS: Genotype distribution was similar in patients and controls. Enzyme activities were lower in patients, but multiple logistic regression analysis did not show any independent association with a higher risk of MI. CONCLUSION: None of the PON1 polymorphisms or their corresponding measured activities are independent risk factors for MI in our population.
Abstract: The present study describes the effects of several high-fat low-cholesterol antiatherogenic diets on the hepatic lipid peroxidation and hepatic antioxidant systems in apolipoprotein E-deficient mice. Eighty mice were distributed into five groups and fed with regular mouse chow or chow supplemented with coconut, palm, olive and sunflower seed oils. After ten weeks, they were sacrificed and the livers were removed so that lipid peroxidation and alpha-tocopherol concentrations, and superoxide dismutase, glutathione peroxidase and glutathione reductase activities could be measured. The size of the atherosclerotic lesions in the aortas was also measured. Results showed that the diets supplemented with olive oil, palm oil or sunflower seed oil significantly decreased the size of the lesion. However, there was an association between those mice that were on diets supplemented with palm or coconut oils and a significant increase in hepatic lipid peroxidation. This association was not found in animals fed with olive or sunflower seed oils, the diets with the highest content of vitamin E. The dietary content of vitamin E was significantly correlated (r = 0.98; p < 0.05) with the hepatic concentration of this compound. Our study suggests that the high content of vitamin E in olive oil or sunflower seed oil may protect from the undesirable hepatotoxic effects of high-fat diets in apo E-deficient mice and that this should be taken into account when these diets are used to prevent atherosclerosis.
Abstract: The epsilon 4 allele of APOE is generally accepted to be a risk factor in Alzheimer's disease and it has been related to other neuropsychiatric disorders, including schizophrenia. The results of several case-control studies have been inconclusive. To shed more light on this issue we carried out an association study that compared the APOE common variant in a group of 365 schizophrenia patients and 584 controls. We found no differences in the genotype distributions and allele frequencies of patients and controls. In the group of patients, we also analysed the possible influence of the epsilon 4 allele in the clinical variables. The most important findings are that the age at onset (AAO) of epsilon 4+ schizophrenic women, those that have one or two epsilon 4 alleles, is 4 years earlier than that of epsilon 4- women and their risk of suffering a negative syndrome subtype is four times greater. This was not found in schizophrenic men. Our results show that the APOE variant is not a risk factor for developing schizophrenia but that it may modulate its phenotypic expression in a sex-dependent manner.
Abstract: Ever since oxidation has been known to be involved in atherogenesis, antioxidants have received considerable attention as potential antiatherogenic agents. The lipid-soluble vitamin E is the main antioxidant carried by lipoproteins. Zinc is a water-soluble trace element that acts as a cofactor of superoxide dismutase (SOD) and has an antioxidant role in several oxidative processes. To test the hypothesis that zinc could adjuvate the antioxidant activity of vitamin E and diminish atherogenesis, we explored how supplementing diet with vitamin E and/or zinc would affect an atherosclerosis-prone animal like Apo E-deficient mice. The increased plasma concentrations of both vitamin E and zinc showed that absorption was high. They had a significant hypolipidemic effect and the supplemented animals had 25% less plasma cholesterol and triglyceride than controls. The SOD activity was significantly higher in washed erythrocytes from mice supplemented with zinc. The plasma of supplemented animals was also significantly more resistant to oxidation. The size of lesions in the proximal aortic region did not differ among groups. Therefore, dietary supplementation resulted in the expected antioxidant effects but there was no substantial attenuation of atherosclerosis in this particular model.
Abstract: Clinical studies have shown that there is a genetic contribution to the pathogenesis of schizophrenia. The molecular mechanisms of effective antipsychotic drugs and recent advances in neural development suggest that several dopamine receptor, serotonin receptor and neurotrophic factor genes might be involved in the disorder. In this study, we assessed the associations between schizophrenia and polymorphisms in the D2 and D3 dopamine receptor (DRD2, DRD3), the serotonin 2A receptor (5HTR2A), the brain-derived neurotrophic factor (BDNF), the ciliary neurotrophic factor (CNTF) and the neurotrophin-3 (NT-3) genes. Our results suggest that the polymorphisms at the DRD3, 5HTR2A, CNTF and BDNF gene loci are unlikely to make our sample more genetically susceptible to schizophrenia. However, we found significant differences in microsatellite allele frequencies between schizophrenic and control groups for DRD2 in the whole sample and for DRD2 and NT-3 only in women. Therefore, clinical differences in the presentation of schizophrenia between gender might be related to genetic factors.
Abstract: Elevated plasma Lp(a) is an independent risk factor for cardiovascular disease. Unique to Lp(a) is the apoprotein, apo(a) which can vary from 250 to 800 kDa in molecular weight. Small isoforms are also associated with the risk of cardiovascular disease. The purpose of this study was to examine the association of Lp(a) concentration, apo(a) size, and Lp(a) lysine-binding site(s) (LBS) function in patients with early onset heart disease, and age-matched controls. Mean values of Lp(a) were significantly higher in the patients than for the age-matched group. The smallest molecular weight isoform for each subject had significantly fewer kringles for the patients than the age-matched controls. There was a significant correlation between LBS activity and kringle number in the single-banded phenotypes of the patients, but not the controls. LBS activity was significantly higher in patients with small isoforms (< or =18 kringles) compared to controls. The odds ratio for coronary artery disease for high LBS activity and high Lp(a) concentration was 4.4 (p = 0.002) and for high LBS activity and small isoforms was 10.1 (p = 0.002). In the patients, Lp(a) concentration was higher, apo(a) size was smaller, and LBS activity higher in the small isoforms compared to the controls. This study suggests an association of high LBS activity in small isoforms of Lp(a) with disease in humans.
Abstract: Relationships between hepatic antioxidant paraoxonase (PON1) activity, lipid peroxidation, and liver injury were investigated in rats with CCl(4)-induced cirrhosis. The study was performed in 60 CCl(4)-treated rats and 60 control animals receiving a standard diet or one supplemented with zinc. Subsets of 10 animals each were killed at weeks 1, 5, and 7 of the study. Results showed that PON1 significantly decreased in rats given CCl(4) alone compared with control animals. This effect was partially reversed in animals receiving zinc. Conversely, lipid peroxides were significantly increased in rats given CCl(4) alone and returned to approximately normal values in animals receiving zinc supplement. PON1 was inversely correlated with lipid peroxidation in all the animals studied. These alterations coincided with changes in serum alanine aminotransferase activity. In vitro incubation of isolated microsomes with CCl(4) or malondialdehyde did not produce any significant changes in PON1, indicating that the decrease in PON1 in CCl(4)-treated animals was not secondary to a direct inhibitory effect of lipid peroxidation products. These data show a time course and quantitative relationship between PON1 activity and lipid peroxidation in rats with CCl(4)-induced cirrhosis and suggest that this enzyme plays a significant role within the antioxidant systems in liver microsomes.
Abstract: The aim of the present study was to investigate the time-course of changes in hepatic lipid peroxidation, cytochrome P450 and metallothionein concentrations, and superoxide dismutase and catalase activities in relation to the onset and development of cirrhosis in CCl4-treated rats. Further, the effects of oral zinc administration on these parameters were assessed. Cirrhosis was induced in 120 rats by intraperitoneal injections of CCl4 twice weekly over 9 weeks. Controls were 120 additional animals. Both groups were further subdivided to receive either a standard diet or one supplemented with zinc. Subsets of 10 animals each were euthanized at weeks 1, 2, 3, 5, 7 and 9 from the start of the study. Results indicated that zinc administration delayed the cirrhotic process and the increase in lipid peroxidation. These changes, consistently maintained during the first 5 weeks of the study, were associated with a significant decrease in the hepatic concentration of cytochrome P450 and an increase in the hepatic concentration of metallothioneins. Zinc supplementation did not produce any significant change in superoxide dismutase and catalase activities. These results suggest that cytochrome P450 and metallothioneins may play an important role in the hepato-protective effects of zinc against lipid peroxidation in experimental cirrhosis.
Abstract: Elevated plasma levels of lipoprotein(a) [Lp(a)] represent a major independent risk factor for the development of atherosclerosis. The kringle IV type 10 of apolipoprotein(a) [apo(a)] is the primary lysine binding site (LBS) of Lp(a) and is associated with lesion formation in transgenic mice. The purpose of this study was to search for mutations in the apo(a) kringle IV type 10 which could alter the LBS activity of Lp(a) from patients with coronary artery disease. We found the DNA region of kringle IV type 10 of apo(a) to be mutable but relatively well preserved in the Spanish population. We identified a novel mutation which probably leads to a truncated form of apo(a) in a patient heterozygous for the mutation and with low lysine binding activity and low plasma Lp(a) concentration. Two other mutations have been previously identified in humans, the substitutions W81R and M75T. The W81R was not found in our sample, but the M75T mutation was present in 43% of patients with coronary artery disease and 23% of age-matched controls. The genotype TT conferred a significant risk for myocardial infarction (odds ratio 2.53). This association was not due to linkage disequilibrium with kringle IV repeats. The M75T polymorphism was not associated with the LBS function of apo(a), but it influenced plasma Lp(a) concentration.
Abstract: BACKGROUND: There is considerable evidence to suggest that plasma lipoprotein(a) [Lp(a)] concentration is a cardiovascular risk factor. Confusing results in epidemiologic studies, however, suggest that the effects of storage should be further investigated. The influence of the assay method, the initial plasma Lp(a) concentration, and the apolipoprotein(a) [apo(a)] genotype are all factors that should be considered. METHODS: Blood was obtained from 65 survivors of premature myocardial infarction and 95 age-matched controls. The plasma samples were stored in sterile conditions at -70 degrees C for 5 years in the presence of antioxidant and antiproteolytic substances. Plasma Lp(a) was measured by immunoturbidimetry, and apo(a) alleles were determined by pulsed-field gel electrophoresis and Southern blotting. RESULTS: Plasma Lp(a) was significantly higher in patients. The mean kringle number for the smallest isoform was also lower in patients than in controls, but no differences were found in the distribution of the largest isoform. All patients and controls were heterozygotes. During storage, mean Lp(a) decreased significantly in samples from patients (-23%; P <0.001) but not in samples from controls (-9%; P, not significant). This was not related to the kringle number and was limited to samples with initial plasma Lp(a) concentrations between 41 and 345 mg/L. CONCLUSIONS: Plasma Lp(a) from patients is less stable than Lp(a) from controls, and the difference is not related to distribution of apo(a) genotypes but may be concentration-dependent. Differential sample stability may complicate the interpretation of several studies.
Abstract: Aspirin reduces the incidence of thrombotic occlusive events. Classically this has been thought to be due to the platelet inhibitory action of aspirin but it has recently been shown that inflammation plays a predominant role in the initiation and progression of lesions in atherosclerosis. In humans, treatment with aspirin reduces cardiovascular risk and slows carotid plaque growth in a dose-dependent fashion. We have explored this issue in Apo E-deficient mice on a high-fat, high cholesterol diet which provided these animals with a continuous administration of 500 microg/day of acetylsalicylic acid in the drinking water. After 10 weeks of treatment, the size of the atherosclerotic lesion at the aortic sinus had reduced by 35%. At the end of the trial there were no significant changes in either plasma lipids or in the quantitative distribution among lipoproteins. Likewise, the total antioxidant status and the resistance of plasma to oxidation in vitro was similar and there was no change in the distribution of iron deposits and in the relative composition of plasma pro-oxidants and antioxidants, or in the concentration of plasma in ferritin. Therefore, it is our hypothesis that the antiinflammatory effect is responsible for the reduction in lesion size. We propose that antiinflammatory molecules which do not cause gastrointestinal complications should be tested in humans to determine long-term efficacy in the attenuation of atherosclerosis.
Abstract: The ability of the apolipoprotein E-deficient mouse to develop spontaneous atherosclerosis, which resembles the human process, is an excellent model in which to assess the impact of dietary factors. This review discusses the role of several nutrients in the development of atherosclerosis and the mechanisms through which they act.
Abstract: Hyperhomocyst(e)inemia is an independent risk factor for atherothrombosis in several clinical settings in which renal function is impaired, but its prevalence in the nephrotic syndrome has not been investigated in detail, even though this syndrome provides an excellent model in which to study a possible link between albuminuria, proteinuria, and hyperhomocyst(e)inemia. We obtained plasma and urine from 27 patients with biopsy-confirmed membranous glomerulonephritis presenting nephrotic syndrome and 27 matched controls and determined the concentrations of homocyst(e)ine and proteins considered putative markers of glomerular and tubular function. Hyperhomocyst(e)inemia, defined as the mean +SD of the plasma homocyst(e)ine concentration of the controls [plasma homocyst(e)ine concentration >10.8 micromol/l] was present in 26% of the patients with nephrotic syndrome but in only 7.4% of the controls. Furthermore, the degree of hyperhomocyst(e)inemia was more severe in the nephrotic patients than in the controls. The existence of renal failure, tubular damage, and, interestingly, relatively well conserved glomerular function barrier were the main predictors of increased levels of plasma homocyst(e)ine. In conclusion, hyperhomocyst(e)inemia is a frequent cardiovascular risk factor present in patients with nephrotic syndrome and renal failure, but it is not directly associated with proteinuria.
Abstract: 1. The aim of the present study was to assess the effectiveness of repeated subcutaneous low-dose recombinant human erythropoietin (rHuEPO) on parameters associated with improved procurement of autologous blood; a procedure regularly used to preclude the need for homologous blood transfusion at the time of elective surgery. 2. Three groups of three volunteers each (n = 9) were administered one of three low doses of rHuEPO (30, 60 or 100 IU/kg bodyweight, s.c.) on days 1, 4 and 8. The plasma pharmacokinetic profile of rHuEPO was studied after the first and third injections. Statistical evaluations were intragroup and intraindividual comparisons. 3. There was a linear relationship between maximum plasma concentration (Cmax) and dose. In the overall study group, Cmax and area under the curve (AUC) were significantly decreased, while the mean residence time (MRT) and elimination half-life (t1/2beta) were significantly increased on day 8 relative to day 1. Significant and sustained increases in reticulocytes were observed after rHuEPO administration, which were maintained above the predose values throughout the study period. 4. In conclusion, rHuEPO, by subcutaneous repeat-dose, was eliminated more slowly and remained longer in the circulation, despite lowered plasma concentrations. Repeated low rHuEPO administration at doses > or = 60 IU/kg bodyweight stimulated modest but sustained reticulocyte concentrations, suggesting that cost may be substantially decreased in autologous blood donation or perioperative treatment programmes.
Abstract: OBJECTIVES: To evaluate a turbidimetric immunoassay for the measurement of ferritin, and to assay this method in a group of patients undergoing an autologous blood transfusion program. DESIGN AND METHODS: We used an ILab 900 analyzer. This instrument automates a particle-enhanced immunoturbidimetric assay with an analysis time of 9 min. This technique was compared with a microparticle immunoassay. The turbidimetric assay was used to measure ferritin in a group of 30 patients undergoing an autologous blood transfusion program. RESULTS: The assay was linear in the range 3-1400 microg/L (r = 0.9999). The intra- and inter-assay imprecision (CV) at 20, 97 and 469 microg/L were <3.0 and <5.0%, respectively. Recovery was 88. 7 to 97.4%. The detection limit was 3 microg/L. Hemoglobin (</=4 g/L), mild hyperbilirubilinemia (bilirubin </=50 micromol/L), triglycerides (</=10 mmol/L) and myeloma paraproteins did not interfere with the assay. The assay showed good correlation with a microparticle enzymoimmunoassay (r = 0.994) with a mean difference between methods of -6 +/- 16 microg/L. This method was sensitive, accurate, and fast enough for an efficient follow-up of autologous blood transfusion patients. CONCLUSIONS: The new automated serum assay for ferritin is an attractive alternative that avoids the need for dedicated instrumentation.
Abstract: 1. The aims of the present study were to assess: (i) the temporal relationships between hepatic lipid peroxidation, changes in the glutathione detoxification system and the onset/development of cirrhosis in CCl4-treated rats; and (ii) the effects of oral zinc administration on these parameters. 2. Cirrhosis was induced in 120 rats by intraperitoneal injections of CCl4 twice a week over 9 weeks. One hundred and twenty additional animals were used as controls. Both groups were further subdivided to receive either a standard diet or one supplemented with zinc. Subsets of 10 animals each were killed at weeks 1, 2, 3, 5, 7 and 9 from the start of the study. 3. Induction of cirrhosis produced a decrease in the components of the hepatic glutathione anti-oxidant system: glutathione transferase activity decreased from week 1, the concentration of reduced glutathione (GSH) decreased from week 5 and glutathione peroxidase (GPx) activity decreased from week 7. This impairment was chronologically related to an increase in free radical generation. Hepatic lipid peroxidation was significantly correlated with GPx activity (r = -0.47; P < 0.001) in CCl4-treated rats. Zinc administration did not produce any significant improvement of the hepatic glutathione system. 4. In conclusion, cirrhosis induction in rats by CCl4 administration produced a decrease in the hepatic glutathione antioxidant system that was related to an increase in free radical production. Furthermore, zinc supplementation produced a reduction in the degree of hepatic injury and a normalization of lipid peroxidation, but not an improvement of the hepatic GSH anti-oxidant system.
Abstract: With the rationale that a disease that presents with anticipation could be associated with expansion of trinucleotide repeats, we selected parent-offspring pairs of schizophrenia patients with earlier age at onset in the filial generation to measure the expansion of CAG repeats using the repeat expansion detection (RED) method. Intergenerational comparisons were made for age at onset, length of CAG repeats, and clinical variables. Although the patients from the filial generation became affected 13 years earlier than the parents (P < 0.0005), we did not find larger CAG repeats in the offspring. No association was found between size of CAG repeat and age at onset or with any other clinical variable. Overall, the frequency of patients with CAG repeats longer than 40 was 32%, which was similar to that observed in control subjects (27%). It is particularly noteworthy that in 86% of the pairs, the mother was the affected parent. In this Spanish sample with parent-offspring pairs presenting schizophrenia with clinical anticipation and apparent female bias of transmission, neither the phenomenon of anticipation nor disease status was associated with the expansion of CAG repeats.
Abstract: BACKGROUND: The measurement of immunoglobulin E (IgE) in serum is widely used in the diagnosis of allergic reactions and parasitic infections. We describe here a fully automated assay for human IgE suitable for routine application in a general chemistry analyzer. METHODS: We used an ILab 900 analyzer. This instrument automates a particle-enhanced immunoturbidimetric assay with an analysis time of 9 min. RESULTS: The assay was linear in the range 4-1000 kIU/L (r = 0.9998). The intra- and interassay CVs at 57, 235, and 434 kIU/L were <3.5% and <7.4%, respectively. The detection limit was 4 kIU/L. Hemoglobin (</=16 g/L), bilirubin (</=250 micromol/L), and myeloma paraproteins did not interfere with the assay. The assay showed good correlation with a microparticle enzyme immunoassay (r = 0.998) with a mean difference between methods of -6 +/- 26 kIU/L. CONCLUSION: The new automated serum assay for IgE is an attractive alternative that avoids the need for dedicated instrumentation.
Abstract: BACKGROUND/AIMS: The aims of this study were to ascertain: 1) whether hepatic cell DNA fragmentation is increased in rats with early stages of liver disease induced by carbon tetrachloride; 2) whether the inhibition of DNA cleavage is involved in the hepatoprotective effects of zinc; and 3) if relationships exist between DNA fragmentation and the onset of fibrosis in this experimental model. METHODS: Twenty-one treated rats and 23 controls were divided into two groups to receive either a standard diet or one supplemented with zinc. All the animals were sacrificed 1 week later for histological and biochemical assessments, which included a DNA fragmentation index, hepatic zinc and metallothionein concentrations, fibrosis measured by hepatic hydroxyproline concentration and plasma alanine aminotransferase activity. RESULTS: Hepatic cell DNA fragmentation was increased in rats with early hepatic fibrosis and the increase was independent of hepatocytolysis, as measured by alanine aminotransferase activity. Oral zinc administration inhibited hepatic cell DNA fragmentation in the treated rats and was proportional to the hepatic concentration of the metal. The mechanism of the zinc-mediated decrease in DNA cleavage was related to an increase in the hepatic metallothionein concentration. Hepatic cell DNA fragmentation was related to hydroxyproline concentration. CONCLUSIONS: Our results suggest that apoptosis may be involved in the early transformations occurring in the liver and which can lead to the initiation of cirrhosis. As such, the potential therapeutic use of zinc supplementation would warrant further investigation.
Abstract: The aim of this study was to identify apolar aldehydes in liver homogenates from rats with CCl4-induced cirrhosis and, as a corollary, the antioxidant effect of zinc administration. The study was performed in five control rats and in ten cirrhotic rats which were further sub-divided into two groups to receive either a standard diet or one supplemented with zinc. The percentage of hepatic fibrosis, plasma malondialdehyde concentration and alanine aminotransferase activity were measured as well as the following aldehydes: hexanal, octanal, decanal, 2-hexenal, 2-octenal, 2-nonenal, 2,4-heptadienal and 2,4-decadienal. Of the 10 cirrhotic rats, 4 had elevated concentrations of the highly toxic 2,4-dialkenals which coincided with a higher percentage of fibrosis and plasma alanine aminotransferase activity. These aldehydes were not observed in the control group. Zinc administration was associated with a reduction of the hepatic malondialdehyde concentration and an amelioration on the degree of hepatic injury. In conclusion, this study demonstrates the presence of the highly toxic 2,4-dialkenals in hepatic tissue of rats whith CCl4-induced cirrhosis. Results obtained would suggest that these particular aldehydes may be related to the severity of the hepatic injury.
Abstract: We have investigated the effect of most common oils used in human nutrition on the development of atherosclerosis in apoE-knockout mice. Seven groups of animals, separated according to sex, were fed for 10 weeks either chow diet or the chow diet 10% (wt/wt) enriched with different oils (palm, coconut, 2 types of olive oil, and 2 types of sunflower oil) without addition of cholesterol. At the end of this period, plasma lipid parameters were measured and vascular lesions scored. None of the diets induced changes in plasma cholesterol concentrations, whereas plasma triglycerides were uniformly reduced in all diet groups. Some diets caused significant reductions in the size of atherosclerotic lesions in males and others in females; males responded most to sunflower oils and females to palm oil and one olive oil (II). The lesion reduction in males consuming sunflower oils was associated with the decrease of triglycerides in triglyceride-rich lipoproteins, whereas the decrease in females consuming olive oil II or palm oil was accompanied by an increase in plasma apoA-I. The increase in plasma apoA-I in the latter condition, is mainly due to overexpression of hepatic message elicited by a mechanism independent of apoE ligand. The data suggest that the different diets modulate lesion development in a gender specific manner and by different mechanisms and that the development of atherosclerosis, due to genetic deficiencies, may be modulated by nutritional maneuvers that may be implemented in human nutrition.
Abstract: Epidemiological and experimental studies suggest that circulating erythrocytes play a role in the incidence of coronary heart disease. We investigated the influence of phenylhydrazine (PHZ)-induced anemia on the formation of atherosclerotic lesions in apo E-deficient mice on regular chow and on a high-fat, high-cholesterol diet during 10 weeks. The repeated doses of PHZ caused sustained anemia throughout the study, changes in the physical characteristics of erythrocytes and increased reticulocyte count. The lesions of the anemic animals were smaller than in the controls and this was even more evident in mice fed with the atherogenic diet. A positive correlation was found between circulating red blood cells at the end of the experiment and the area of aortic lesion. There was also a negative association between the lesion and the reticulocyte count. This reduced progression of atherosclerotic lesions is independent of nutritional status or the lipoprotein cholesterol distribution. The results suggest that mechanisms related to the number of circulating red blood cells may have a significant influence on the development of atherosclerosis.
Abstract: Disturbances in methyl-carbon metabolism, which result in hyperhomocysteinemia, have been associated with schizophrenia. Homozygosity for the T677 allele of the methylenetetrahydrofolate reductase (MTHFR) gene, which encodes for a thermolabile enzyme associated with hyperhomocysteinemia, has been found to be increased in schizophrenic patients. We have investigated whether plasma homocysteine concentration and the frequency of C677T MTHFR variant were increased in schizophrenic inpatients of a psychiatric hospital (n=210) compared with controls (n=218). There were no significant differences in plasma homocysteine concentrations between the schizophrenia and the control group. The distributions of T allele and TT genotype frequencies were similar in both groups (40% and 15%). These results show that impaired homocysteine metabolism is unlikely in schizophrenia.
Abstract: Platelet glycoprotein IIb/IIIa may be involved in the pathogenesis of myocardial infarction as the key element in platelet aggregation and as the binding site of lipoprotein(a) to platelets, inhibiting plasminogen binding and activation. Recently, a strong association between the P1A2 polymorphism of the glycoprotein IIIa gene and acute coronary thrombosis has been reported. although this has not been confirmed. In an associated study, we determined plasma lipoprotein levels, the apo E genotype and the P1A genotype in 250 males under 55 years with myocardial infarction and they were compared with 250 age- and sex-matched controls. Patients showed an over-representation of the epsilon3/4 genotype with respect to the control group. We found that there were no differences in the allelic frequency of P1A2 between case patients and age-matched controls (chi2 = 0.05, P = 0.92) and that subjects bearing the P1A2 allele showed higher plasma lipoprotein(a) concentration than p1A1/P1A1 individuals. Therefore, in this population there is no association between carriage of p1A2 allele and increased risk of myocardial infarction but the carriage of P1A2 is associated with higher plasma Lp(a) concentration.
Abstract: BACKGROUND: Clinical data suggest that autologous blood donation may prevent postsurgical venous thrombosis. If confirmed, this is probably due to beneficial effects in rheologic and hematologic variables which may be changed in patients as a result of repeated bleeding. STUDY DESIGN AND METHODS: To ascertain this point, we studied variations in hematological, hemorheological, coagulative and fibrinolytic parameters in 30 patients undergoing autologous blood donation. RESULTS: Whole blood viscosity (WBV), plasma viscosity and blood viscosity adjusted to 40% hematocrit, progressively and substantially decreased throughout the successive bleeding at all the shear rates considered. WBV was further reduced by presurgical hemodilution with autologous plasma which decreased the platelet and leukocyte count. The hemostasis and fibrinolysis variables, however, underwent no clinically significative changes. CONCLUSION: Repeated bleedings change most hemorheological variables. By decreasing cytocrit and viscosity, reducing aggregability and increasing blood cell deformability an optimal milieu to help prevent thrombosis is artificially created.
Abstract: We evaluated the performance of a homogeneous assay for the automated measurement of high-density lipoprotein cholesterol (HDL-C) and compared it with a conventional precipitation technique in the following groups of people: control subjects (group A), clinically-healthy elderly (group B), myocardial infarction patients (group C), nephrotic syndrome patients (group D), and liver cirrhosis patients (group E). The performance of the technique was acceptable with respect to precision, accuracy, linearity, and detection limit. Triglycerides up to 40 mmol/L and bilirubin up to 150 micromol/L did not cause interferences. Hemoglobin decreased HDL-C measurements. Samples were stable at -20 degrees C for up to four months. Bland-Altman plots showed a good agreement between both techniques in the control group but with a progressive divergence in the patient groups B to E. Results indicate limitations of the technique in certain clinical conditions and, coincidentally, the need for reliable calibration materials.
Abstract: The concentrations of 25 plasma proteins were measured in 22 patients with membranous nephropathy. For some large proteins, the plasma concentrations were increased; there were also large proteins with low plasma concentrations, but small or medium-sized proteins showed uniformly lower plasma concentration than the controls. Plasma colloid osmotic pressure (pi) and viscosity (eta) were not interrelated but showed positive and significant correlations with plasma concentrations of small and medium-sized proteins (pi) and plasma concentrations of large proteins (eta), respectively. Nephrotic plasma is not efficient in maintaining plasma pi but highly efficient in maintaining plasma eta. High plasma fibrinogen concentrations and low antithrombin III concentrations may predispose to thrombosis, and low IgG concentrations may account for the higher predisposition to bacterial infection. The relative composition of nephrotic plasma is heavily dependent on the size of the different proteins. Plasma pi and eta are also maintained by the relative preponderance of different plasma proteins.
Abstract: In the normal population, the usual effect of the epsilon 2 allele is to decrease plasma cholesterol and to increase plasma triglyceride. We report here the association of nephrotic syndrome and the apo epsilon 2 epsilon 2 genotype in which we observed a hyperlipidemia characterized by very low levels of lipoprotein lipase activity, chylomicronemia, high levels of plasma apo B, C III, E and lipoprotein(a), very low levels of high density lipoprotein cholesterol and concentrations of cholesterol and triglyceride that are higher than expected in all the other lipoprotein fractions. When proteinuria was partially resolved and plasma albumin levels returned to normal, a residual type III hyperlipidemia was still present. These findings suggest that the combination of apo epsilon 2 homozygosity and massive proteinuria may cause considerable changes in the clearance of triglyceride rich particles probably mediated by the almost complete absence of lipolytic enzymes and a low interaction of lipoproteins with specific receptors. The apo E genotype should be investigated in nephrotic patients with chylomicronemia.
Abstract: The behavior of apolipoprotein (apo) A-I in lipoprotein (Lp) AI and LpAI:AII was studied in 11 postmenopausal females and 11 males matched for plasma triglyceride and total cholesterol levels. Subjects consumed a baseline diet [35% fat (14% saturated, 15% monounsaturated, and 7% polyunsaturated), 15% protein, 49% carbohydrate, and 147 mg cholesterol/1000 kcal] for 6 weeks before the start of the kinetic study. At the end of the diet period, using a primed-constant infusion of [5,5,5-2H3]leucine, residence times (RT) and secretion rates (SR) of apoA-I were determined in 2 subpopulations of high-density lipoprotein (HDL) particles, LpAI and LpAI:AII. Plasma total cholesterol, low-density lipoprotein cholesterol, and triglyceride concentrations were similar in males and females. The mean plasma HDL cholesterol concentration in males (1.14 +/- 0.23 mmol/L; mean +/- SD) was lower than in females (1.42 +/- 0.18 mmol/L; P =. 0034). Similarly, the mean plasma concentration of apoA-I in males (130 +/- 21 mg/dL) was lower than that in females (150 +/- 19 mg/dL; P = .0421). The RT of apoA-I in either LpAI or LpAI:AII was similar between men and women. Despite the higher plasma apo A-I levels in female compared with male subjects, total apoA-I and apoA-I in LpAI and LpAI:AII pool sizes were similar between the two groups, attributable to the lower body weight of the female subjects. The mean SR of total apoA-I in males (8.5 +/- 2.7 mg.kg-1.d-1) was 22% lower than in females (10.9 +/- 2.3 mg.kg-1.d-1; P = .0389). The SR of both apoA-I in LpAI and LpAI:AII was lower in males than females, although the differences did not reach statistical significance. These data suggest that the difference observed in HDL cholesterol concentration between males and females is attributable to SR of apoA-I and not the catabolic rate.
Abstract: BACKGROUND/AIMS: To investigate whether physicochemical alterations in plasma lipoproteins are associated with changes in plasma oncotic pressure and viscosity in liver cirrhosis. METHODS: The study included 66 patients with cirrhosis (confirmed by liver biopsy) and 58 healthy volunteers. The constituents measured were: the concentration, density and composition of plasma lipoproteins; plasma oncotic pressure and viscosity; and the concentrations of albumin, total protein, haptoglobin, transferrin, immunoglobulin M and alpha2-macroglobulin. RESULTS: Step-wise multiple regression analysis indicated that, in the patients with cirrhosis, plasma oncotic pressure was significantly correlated with plasma albumin+viscosity (r=+0.85; p<0.001) and with plasma total protein+the density of low density lipoprotein (r=+0.74; p<0.001). The inclusion of viscosity and the density of low density lipoprotein in the regression equations significantly improved the observed correlation between albumin and plasma oncotic pressure (r=+0.70; p<0.001). Similarly, plasma viscosity was significantly correlated with the sum of plasma total protein and cholesterol (r=+0.68; p<0.001). The inclusion of cholesterol in the regression equation significantly increased the observed correlation between total protein and plasma viscosity (r=+0.59; p<0.001). CONCLUSIONS: These results suggest that lipoprotein alterations associated with liver cirrhosis may play a role in determining plasma oncotic pressure and viscosity, and thus could influence the progression of the disease.
Abstract: The repeat expansion detection (RED) method was described to detect expansions of trinucleotide repeats of unknown chromosomal location. We have improved the RED method by the use of 8-mer oligonucleotides and assessed its usefulness in 30 samples from patients with spinocerebellar ataxia type 1 (SCA1), Huntington's disease (HD), and Machado Joseph's disease (MJD), for which the number of CAG/CTG repeats was determined by sequencing. There was a good correlation between the number of repeats detected by sequencing and those identified by RED. However, in 17% of samples, the RED gave additional fragments for ligation products of different size than the CAG/CTG repeat expansion detected in the sample by sequencing. The same was observed in a group of control subjects (n = 78) without known clinical abnormalities in which products of more than 40 repeats were detected in 27% of them, indicating that CAG/CTG repeat expansions are common in the general population. Wether this corresponds to unidentified loci with expansions deserves further investigation.
Abstract: BACKGROUND: Clinical and pharmacokinetic data suggest that very low doses of subcutaneous recombinant human erythropoietin (rHuEPO) may be effective in a preoperative autologous blood deposit program. STUDY DESIGN AND METHODS: Fifty-two patients, scheduled for orthopedic surgery, were enrolled in a double-blind and placebo-controlled study. Patients were randomly assigned to the placebo group or to receive 30, 60, or 100 IU per kg of rHuEPO subcutaneously twice a week for 2 weeks before surgery. The dose of rHuEPO that was effective in facilitating the collection of 4 units of blood in the 2 weeks before surgery and that prevented a sharp decrease in hematocrit was determined. RESULTS: Only in patients receiving 100 IU per kg of rHuEPO did the outcome measurements differ significantly from those in the placebo group. With a higher (p < 0.01) cumulative increase in red cell volume during the study period (297 +/- 127 vs. 121 +/- 44 mL), 64 percent of those receiving 100 IU per kg of rHuEPO were able to donate 4 units of blood for autologous use, as compared with 23 percent of the placebo group (p < 0.05). Allogeneic transfusion was avoided, and the preoperative hematocrit and reticulocyte count were significantly higher in the patients receiving 100 IU per kg of rHuEPO (p < 0.05 and p < 0.01, respectively). CONCLUSION: Subcutaneously administered rHuEPO at a dose of 100 IU per kg twice a week for 2 weeks is effective in facilitating the collection of blood for autologous use and may improve the cost-benefit ratio of blood conservation interventions. Doses < or = 60 IU per kg are ineffective in facilitating such collections in this surgical setting.
Abstract: Lipoprotein measurements in a group of 29 patients with massive proteinuria and without hypoalbuminemia, were compared with those observed in matched controls and patients with overt nephrotic syndrome to assess the influence of plasma albumin concentration and proteinuria in modulating blood lipid levels. Plasma apoprotein B and apo B containing lipoproteins were not increased in proteinuric normoalbuminemic patients. There was a good correlation between plasma albumin and oncotic pressure (r = 0.937; P < 0.001). Plasma oncotic pressure was inversely correlated with plasma apoprotein B in nephrotic patients (r = -0.44, P = 0.017) but not in normoalbuminemics (r = 0.17, P = 0.369), suggesting that plasma albumin affects apoprotein B secretion. Other findings, however, indicate that multiple processes are ocurring simultaneously in these patients. There was an accumulation of very low- and intermediate density lipoproteins in normoalbuminemics, suggesting a residual defect in the lipoprotein removal. Also, raised (P < 0.05) lipoprotein(a) levels respect to controls (median, 0.15 g/l) were noted in both, normoalbuminemics (median, 0.72 g/l) and hypoalbuminemics (median, 0.84 g/l) with similar degree of proteinuria (6.4 vs. 6.6 g/24 h), suggesting that other mechanisms may be operative in lipoprotein(a) derangements. Our findings suggest that there is no unique mechanism in the pathogenesis of nephrotic hyperlipidemia but that both hypoalbuminemia and proteinuria can have a distinct contribution, individually or in combination.
Abstract: Type III hyperlipoproteinemia (HLP III) is characterized by the reduced catabolism and accumulation of chylomicron and very low density lipoprotein (VLDL) remnants. Most HLP III patients are homozygous for the apolipoprotein E2 (Cys112, Cys158) allele; however, several other mutations at this gene locus have been associated with this HLP. In order to assess the presence of rare apo E variants in our population, we have examined apo E phenotypes by isoelectric focusing (IEF) and genotypes by restriction enzyme analysis of polymerase chain reaction (PCR) amplified DNA in 15 patients with HLP III. Lack of concordance between these two methods was observed in 11 subjects (73.3%). DNA sequencing analysis of the receptor binding domain of the apo E gene in the 11 HLP III patients with discrepancies demonstrated the presence of six carriers of the epsilon 3(Arg136-->Ser) allele and three carriers of the epsilon 2(Gly127-->Asp) allele. Five HLP III patients were apo E2/E2 using IEF, but only 2 of them were epsilon 2 homozygous using PCR. Two patients were E3/E3 homozygous with normal DNA sequence in the low density lipoprotein receptor binding domain of apo E. In conclusion, our results show that a number of different apo E genotypes are associated with HLP III in this population. More specifically, mutations at positions 127 and 136 might be frequent in Spain and occur in patients with HLP III.
Abstract: Hepatic lipid peroxidation, metallothioneins, collagen and proline hydroxylase activity were investigated in 16 ethanol-fed rats and in 16 control animals. The rats were further divided into three groups to receive either a standard diet, a zinc-deficient diet or a zinc-supplemented diet. The animals were sacrificed at week 12 of the experiment for histological and biochemical assessments. Hepatic tissue examination indicated that oral zinc supplementation was associated with a decrease in lipid peroxidation, collagen deposition and proline hydroxylase activity together with an increase in metallothionein concentration in alcoholic rats. There were no significant differences in lipid peroxidation in the control group in relation to the diet. Zinc supplementation was associated with increased concentrations of hepatic metallothioneins together with decreased concentrations of proline-hydroxylase and collagen but to a lesser degree than in alcoholic animals. These results indicate that zinc is an efficient hepato-protective agent against lipid peroxidation in alcoholic rats and its effect may be, in part, mediated by the activation of metallothionein synthesis. Also, lipid peroxidation may be related to changes in hepatic collagen synthesis.
Abstract: Although lipoprotein abnormalities of the nephrotic syndrome are assumed to be related to the presence of proteinuria, this topic has not been investigated extensively. We measured lipoproteins from 19 nonuremic patients during and after remission of the nephrotic syndrome in an effort to determine the extent of their putative atherogenicity. As expected, disturbances involved primarily the apoprotein B-containing lipoproteins. No patient showed serum lipoprotein(a) [Lp(a)] < 300 mg/L during the acute phase. Lp(a) concentrations correlated significantly with those of apoprotein B, and both values decreased dramatically with the remission of the nephrotic syndrome. Surprisingly, despite the resolution of proteinuria, concentrations of intermediate-density lipoproteins and Lp(a) remained above normal in hypertriglyceridemic patients, suggesting a residual effect of nephrosis in the overall lipoprotein transport. Accumulation of atherogenic remnants should be considered a characteristic of the hyperlipidemia of the nephrotic syndrome, and aggressive treatment to reduce proteinuria is mandatory.
Abstract: A considerable demand for convenient, rapid, inexpensive assays of ferritin in serum has been generated in recent years in hospital laboratories and blood banks. We describe a simple and rapid particle-enhanced turbidimetric immunoassay suitable for routine application in a Monarch 2000 centrifugal analyzer with commercially available reagents. This fully automated assay (y) requires no pretreatment of sample, and correlation with a two-step sandwich ELISA (x) is excellent (y = 1.018x + 0.397, Sy/x = 0.027). The analytical range extends from 5 to 900 micrograms/L. Intraassay imprecision (CV) ranged from 1.1% to 5% for various specimen concentrations. Interassay imprecision ranged from 2.2% for above-normal concentrations (755 micrograms/L) to 9.5% for low concentrations (39 micrograms/L). No specimen-related carryover was detected. The method has been useful in our predeposit autologous blood transfusion program for rapid assessment of iron status in patients undergoing repeated phlebotomies.
Abstract: The accuracy of the Friedewald formula in estimating low-density lipoprotein (LDL) cholesterol was investigated in 47 alcoholic patients with liver disease (21 minimal-change, 26 cirrhotic) by comparing the results with those obtained by sequential preparative ultracentrifugation. In 14% of subjects with minimal-change disease, the error in the estimated LDL cholesterol was 50% +/- 9% (mean +/- SD; range 40-59%) and was related to the degree of attendant hypertriglyceridemia (r = 0.98; P < 0.001). A similar degree of error was observed in patients with cirrhosis, despite the absence of hypertriglyceridemia; an abnormal VLDL cholesterol: triglyceride ratio was the contributory factor in the discrepancy. We conclude that, as is the case in other clinical pathologies in which abnormalities of lipoprotein composition have been described (e.g., diabetes), the Friedewald formula to estimate LDL cholesterol may be inappropriate in chronic alcoholics, particularly those in whom a degree of hepatic dysfunction may be suspected.
Abstract: Regardless of type, uncontrolled diabetes represents a serious disruption of fuel homeostasis with consequences throughout the body. This may hamper the applicability of predeposited autologous blood transfusion in diabetic patients because metabolic changes are expected as a consequence of repeated bleeding. We undertook this study to determine whether the presence of non-insulin-dependent diabetes mellitus (NIDDM) influences the erythropoietin (EPO) response to repeated phlebotomies with respect to normal subjects. We included 22 consecutive patients scheduled for major surgery during a 2-year period in which clinical and metabolic complications were excluded and renal and liver function was considered unaffected. Selected biochemical and hematologic variables were serially measured during donation of several units of blood in a 12- to 29-day period. Bleeding produced a significant decrease in serum glucose, cholesterol, triglyceride, and apoprotein B concentration in diabetic patients. Except for glucose, this effect was not observed in controls. Both groups were comparable with respect to initial hemoglobin concentrations and all hematologic variables measured. The decrease in hemoglobin concentration did not produce clinical symptoms in these patients, and recovery was regarded as normal in both groups. Serum EPO levels in diabetic patients were negatively influenced by the initial hemoglobin A1c (HbA1c) proportion. Moreover, three nonrespondent diabetic patients with poor glycemic control responded normally 6 to 13 months later, in a second operation, when glycemic control had improved significantly. In conclusion, NIDDM may limit the donation of requested units for major surgery only if poor glycemic control is present. When possible, phlebotomies should be delayed and metabolic control reinforced.
Abstract: Cholesterol and triglyceride in plasma and lipoprotein fractions and serum apoprotein concentrations were measured in 51 chronic alcoholic subjects; 23 had minimal or mild hepatic changes (steatosis and/or fibrosis) and 28 had cirrhosis. Of the latter, 16 had stopped alcohol consumption at least 3 months before the study, while the other 12 and all the mildly affected patients had continued drinking. None of the patients presented with cholestasis or alcoholic hepatitis. The control group was composed of 15 healthy, non-drinking volunteers selected from the hospital staff with an age- and sex-distribution similar to that of the alcoholic group. Patients with minimal hepatic changes had plasma total cholesterol concentrations within the ranges of the normal population but with increased high density lipoprotein and decreased low density lipoprotein fractions. Total plasma triglyceride values were not significantly elevated but the distributions in the low density lipoprotein and high density lipoprotein fractions were significantly increased in patients compared to controls. This alteration was accompanied by a consistent increase in serum apolipoprotein C-III concentration. Conversely, in patients with cirrhosis, serum concentrations of apolipoproteins A-I and B were significantly lower and were reflected in the cholesterol concentrations in the lipoprotein fractions. Comparisons between abstainers and non-abstainers within the group with cirrhosis indicated that cessation of alcohol intake was not sufficient to rectify lipoprotein dysfunction following damage from cirrhosis.
Abstract: BACKGROUND: The prevalence of biliary lithiasis was studied in a sample of rural female population by calculation of the risk factors associated and the correlation between biliary lithiasis and serum lipoprotein concentrations. METHODS: Two hundred forty-nine women between the ages of 20-75 years in whom complete anamnesis and calculation of the Quetelet index (QI) were performed, were the subjects of this study. The investigation of biliary lithiasis was carried out by echography. Serum lipoprotein concentrations were determined by sequential ultracentrifugation. RESULTS: The prevalence of biliary lithiasis in the population studied was of 10.4%. A tendency to increase by age was observed and a peak between 50-60 years of age (p < 0.05) was registered. Biliary lithiasis was more frequent among the obese population (p < 0.05). With respect to lipoprotein parameters, all the lipid values of the different fractions demonstrated to be higher in the lithiasic population, although only the differences in cholesterol vehicled by very low density lipoproteins (C-VLDL) achieved statistical significance) (p < 0.001). CONCLUSIONS: The prevalence of biliary lithiasis in the female population studied is similar to that reported in other Western populations, and an increasing tendency in the prevalence of biliary lithiasis with age and obesity was observed. The levels of C-VLDL are higher in women with biliary lithiasis than in those without.
Abstract: This study was designed to further ascertain the presence in plasma of lipoprotein lipase (LPL) bound to circulating lipoproteins. Lipoprotein lipase mass and activity values in preheparin plasma from 20 volunteers were 69.8 +/- 6.6 ng.ml-1 and 1.54 +/- 0.15 mU.ml-1, respectively, and no significant correlation between mass and activity was observed. Fifteen min after heparin injection, LPL mass had increased to 536 +/- 60 ng.ml-1 and LPL activity to 261 +/- 34 mU.ml-1 and a highly significant correlation between the increments in mass and activity was observed. The released material had a specific activity of 0.57 +/- 0.03 mU.ng-1. The LPL mass in preheparin plasma eluted early from heparin-Sepharose, in the position expected for inactive LPL monomers. Western blot analysis showed that the eluted material had the size expected for the LPL subunit (55 kDa). The increment of mass and activity after heparin eluted later from heparin-Sepharose, in the position expected for active LPL dimers. It is concluded that preheparin plasma contains substantial amounts of inactive LPL protein, and that heparin releases mainly active LPL into circulation. On gel filtration LPL activity and mass in postheparin plasma eluted mainly in the positions of LDL and HDL. Electron microscopy of immunostained fractions showed reaction for LPL and apolipoprotein B, or apolipoprotein A-I, on the same particles. LPL mass in preheparin plasma eluted in a similar pattern, associated with LDL and HDL. In postprandial plasma substantial amounts of LPL protein eluted with the triglyceride-rich lipoproteins. When 125I-labeled bovine LPL was added to plasma or to ultracentrifugally isolated lipoproteins and then analyzed by gradient gel electrophoresis, the labeled lipase moved with the lipoproteins. The presence of substantial amounts of inactive LPL protein associated with lipoproteins in plasma may have important implications for the metabolism of the particles in view of recent reports on avid binding of LPL-lipoprotein complexes to cell surfaces and receptors.
Abstract: Fifteen patients on chronic maintenance hemodialysis without any additional known cause for dyslipidemia were arbitrarily divided into two groups based on fasting plasma triglyceride levels. The hypertriglyceridemic patients (plasma triglyceride levels above 170 mg/dl, N = 7) also had decreased high density lipoprotein (HDL) cholesterol levels and decreased post-heparin plasma lipoprotein lipase activity compared to the normotriglyceridemic patients (N = 8). All lipoprotein fractions collected by density gradient ultracentrifugation were triglyceride-enriched in the hypertriglyceridemic patients. Both groups of patients had elevated intermediate density lipoprotein levels, heterogeneity in the distribution of low density lipoproteins (LDL) and apoprotein-specific HDL subpopulations, and abnormalities in the size and composition of both LDL and HDL. The described alterations tended to be more marked in hypertriglyceridemic patients and are not detected by the usual laboratory evaluation of lipoproteins. These lipoprotein abnormalities have been shown to be atherogenic in patients without renal disease and are likely to contribute to the high prevalence of premature atherosclerosis in end-stage renal disease.
Abstract: Forty-two patients with the nephrotic syndrome were grouped according to the absence or presence of renal failure and/or diabetes mellitus. All patients had a similar degree of hypoalbuminemia and urinary protein losses. A lipid and apoprotein pattern was generated in serum and ultracentrifugally isolated lipoproteins. Low-density lipoprotein composition was essentially normal in uremic patients while in the other patients with the nephrotic syndrome, a considerable lipid enrichment was noted. The very-low-density lipoprotein content in lipids was uniformly increased in nephrotic patients irrespective of the presence of complications. High-density lipoprotein cholesterol and serum apolipoprotein A I and E concentration was significantly reduced in uremic patients with respect to normal subjects and to the other groups considered. Serum apolipoprotein A II and B levels were also decreased in uremics. All patients had increased serum apoprotein C II and C III concentration. We conclude that diabetes mellitus does not affect the pattern of hyperlipoproteinemia of nephrotic syndrome while the characteristic lipoprotein and apoprotein pattern of uremia is present irrespective of nephrosis in uremic, nondiabetic patients.
Abstract: 1. Lipid peroxidation and hepatic fibrogenesis were investigated in 25 carbon tetrachloride-treated rats and in 25 control animals. Rats were further divided into two groups to receive either a standard diet or one supplemented with zinc. From each group, animals were killed at weeks 3 and 18 of the experiment for histological and biochemical assessments which included hepatic lipid peroxide and collagen concentrations and plasma zinc concentration as well as the hepatic activities of proline hydroxylase and collagenase. 2. Results indicated that oral zinc supplementation was associated with a decrease in lipid peroxidation (mean 51%; P < 0.05), collagen deposition (mean 32%; P < 0.001) and proline hydroxylase activity (mean 30%; P < 0.05) at week 18, together with an increase in collagenase activity (mean 208%; P < 0.01) at week 3, in carbon tetrachloride-treated rats. 3. There was a significant direct correlation between lipid peroxidation and proline hydroxylase activity in carbon tetrachloride-treated rats (r = 0.52; P < 0.01) and also a significant inverse correlation between lipid peroxidation and plasma zinc concentration in these animals (r = -0.62; P < 0.001). 4. These findings are consistent with the hypothesis that hepatic lipid peroxidation plays an important role in the aetiology of hepatic fibrogenesis and that zinc mitigates the process.
Abstract: We have investigated the influence of variation of the concentrations of serum albumin and immunoglobulins on serum fructosamine concentration in 33 patients with nephrotic syndrome, and 18 patients with cirrhosis of the liver. Protein alterations were evident in these patients and they were compared with 109 normal subjects, 43 patients with type II diabetes mellitus and nine diabetic patients with nephrotic syndrome. The mean serum fructosamine concentration in diabetic patients (2.76 +/- 0.53 mmol/L) was significantly increased (P less than 0.001) by comparison with normal subjects (1.93 +/- 0.20 mmol/L) and the other patients studied. Patients with diabetic nephropathy had higher (P less than 0.01) serum fructosamine concentrations (2.23 +/- 0.54 mmol/L) than non-diabetic patients with the nephrotic syndrome (1.57 +/- 0.37 mmol/L) but remained with the normal range. Positive correlations were observed between fructosamine and immunoglobulins G and M in nephrotic and cirrhotic patients. Serum immunoglobulin A was also directly correlated with serum fructosamine in patients with cirrhosis of the liver. An inverse correlation between albumin and fructosamine in serum of patients with cirrhosis of the liver was also noted. We conclude that the fructosamine assay is not useful in the assessment of glycemic control in patients with cirrhosis of the liver, nephrotic syndrome or in any other clinical situation in which protein metabolism is altered.
Abstract: Oxidation of low density lipoprotein (LDL) has been demonstrated in vivo and directly implicated in the process of foam cell formation. Consequently, a considerable research effort has been devoted to the assessment of the metabolic behaviour of oxidized LDL. We have developed a simple and reproducible model to obtain oxidized LDL consisting of the dialysis of LDL (4 g/l) contained in a cellulose bag against 5 litres of 0.15 M NaCl, 5 microM CuSO4, 0.6 mM FeCl3, pH 7.4, 37 degrees C with constant oxygen bubbling. While the resulting particles have a number of physicochemical properties suggesting lipid oxidation, neither apo B fragmentation nor modification in the size and shape were observed. This oxidized LDL showed internalization into cells through both the apo B,E and the scavenger receptors and the rate of removal from the plasma in injected rats was faster than that observed for normal LDL. We suggest that these particles may represent an equivalent to the circulating oxidized LDL postulated in humans.
Abstract: To assess its hypolipidemic predictive value, maximally induced low density lipoprotein receptor activity was measured in vitro in peripheral blood lymphocytes from 20 polygenic hypercholesterolemic patients prior to their recruitment into a rigorous dietary and pharmacological treatment program. The patients in the diet program demonstrated significant beneficial changes in plasma, LDL and HDL cholesterol concentrations. After 12 weeks, those patients who had not had reductions in LDL cholesterol to within the reference range, had cholestyramine and/or bezafibrate supplements to the diet for a further six weeks. The beneficial trends continued. The LDL receptor values correlated well with pre-treatment LDL cholesterol and plasma apoprotein B concentrations (r = -0.472, P less than 0.01 and r = -0.526, P less than 0.01, respectively) and with the percentage change induced by diet (r = 0.510, P less than 0.01 and r = 0.480, P less than 0.01, respectively) and by diet supplemented with bezafibrate and cholestyramine (r = 0.634, P less than 0.05 and r = 0.629, P less than 0.05, respectively). In polygenic hypercholesterolemic patients in whom LDL receptor activity is high, treatment with diet alone may be sufficient whereas those with low receptor activity would require specific pharmacological intervention.
Abstract: The effects on plasma lipoproteins of four fat-modified diets were assessed in 11 nuns in a contemplative order in the Mediterranean region of Spain. Diet 1 [high polyunsaturated fatty acid (PUFA), low monounsaturated fatty acid (MUFA), low ratio of PUFAs to saturated fatty acids (P:S)] and diet 3 (low PUFA, high MUFA, low P:S) induced significant, directly comparable reductions in total plasma (12% and 13%, respectively) and low-density-lipoprotein (LDL) cholesterol (24% and 19%, respectively). Diet 2 [high PUFA, high MUFA, low saturated fatty acid (SFA), high P:S] induced greater decrements (23% and 30% in total plasma and LDL cholesterol, respectively). Diet 4 (low PUFA, low MUFA, high SFA, low P:S) induced a significant increase in LDL cholesterol of 11%. No significant changes in high-density-lipoprotein cholesterol were observed with these diets. Because the effects of PUFAs and MUFAs are comparable, no recommendations on modifying the habitual, high-MUFA-containing Mediterranean diet need be made other than, perhaps, a reduction in the overall intake of SFAs.
Abstract: The effect of lovastatin on the hyperlipidemia induced in rats with experimental nephrotic syndrome was investigated; toxicity and the effects on common blood chemistry parameters were also assessed. Hyperlipoproteinemia in this particular model is associated with an increase in hepatic synthesis of lipoproteins, and treatment with lovastatin could be the most suitable, since the drug inhibits cellular cholesterol synthesis. Lovastatin treatment resulted in a considerable reduction in plasma cholesterol and triglyceride levels. The decrease in cholesterol levels with treatment was mainly confined to the low-density lipoproteins (LDL) although there was a reduction in the nephrotic-syndrome-induced incremental level of high-density lipoprotein cholesterol. Other lipoprotein fractions were unaffected by lovastatin. LDL apoprotein B was increased in both groups of rats, but to a lesser degree in the lovastatin-treated group, suggesting a double effect, inhibition of both, cholesterol and apoprotein synthesis. Both groups of rats showed a certain degree of renal impairment as shown by significant elevations in plasma urea and creatinine levels. Hepatic damage was also observed, chemically and microscopically, in both groups of rats, being more pronounced in those rats treated with lovastatin in which a 50% mortality ensued after 2 weeks of treatment. At the dosage used this may have some implications in its therapeutic use in certain conditions.
Abstract: To assess its hypolipidemic predictive value, maximally induced low density lipoprotein receptor activity was measured in vitro in peripheral blood lymphocytes from 20 polygenic hypercholesterolemic patients prior to their recruitment into a rigorous dietary and pharmacological treatment program. The subjects in the diet program demonstrated significant beneficial changes in plasma, LDL and HDL cholesterol concentrations. After 12 weeks, those patients who had not had reductions in LDL cholesterol to within the reference range had cholestyramine and/or bezafibrate supplements to the diet for a further 6 weeks. The beneficial trends continue. The initial LDL receptor values correlated well with pre-treatment LDL cholesterol and plasma cholesterol and apoprotein B concentrations, and with the percentage change induced by diet and by diet supplemented with bezafibrate and cholestyramine. During the trial the only significant variations in plasmatic hormones were observed in TSH and T3 concentrations, but always within the reference range. In polygenic hypercholesterolemic patients in whom LDL receptor activity is high, treatment with diet alone may be sufficient whereas those with low receptor activity would require specific pharmacological intervention.
Abstract: Serum cholesterol, triglyceride, apoprotein Al and B, low-density and hig-density lipoprotein (LDL, HDL) cholesterol levels were quantitated, in two or more consecutive sample collections, in 289 middle aged males to assess the potential for misclassification of dyslipemia, according to the Sociedad Española de Arteriosclerosis and the USA National Cholesterol Education Program guidelines. They were grouped according to serum cholesterol values (less than 5.17 mmol/l, between 5.17 mmol/l and 6.17 mmol/l, and greater than 6.17 mmol/l) and misclassification was arbitrarily identified with different levels of the considered parameters. With cholesterol less than 5.17 mmol/l, misclassification was identified in 18.2% of the subjects, mainly with hypertriglyceridaemia or low HDL-cholesterol values. With cholesterol greater than 6.17 mmol/l there is a clear misclassification although obviated partially by the usual performance of complete routine lipoprotein analysis. Potential for misclassification is more evident when cholesterol was between 5.17 mmol/l and 6.17 mmol/l. To avoid such misclassification, HDL and LDL-cholesterol and serum cholesterol and triglyceride values should be measured in all subjects. Some data on the total cost in a general population screening are also presented; cost that on the author's opinion might be better invested under the control of a centralized institution.
Abstract: We describe the results of a study of asymptomatic Listeria monocytogenes serum prevalence in a population of pregnant women in the Reus area. The study includes newborn exam and correlation with laboratory data obtained by an indirect immunofluorescence method recently developed. The incidence of anti-Listeria antibody carriers was high (12%) but in no cases spontaneous abortions or fetal malformations were associated to high titers.
Abstract: The efficiency of an autotransfusion program was studied over two groups of patients undergoing an elective surgery. The A group (n = 41) was on dietotherapy program associated with a supplement of ferrous sulfate and the B group (n = 35) on dietary intervention only. Red cell volume (RCV), haemoglobin (Hb) and haematocrit (Ht) were determined previously to the autotransfusion, during the same and before the surgical intervention. Serum ferritin levels were measured in the B group patients previous to the autotransfusion and before the surgical intervention. No statistically significant differences were observed among the two groups with regard to the RVC variations, Hb or Ht during the autotransfusion process. Although 85% of the A group patients showed secondary digestive effects due to oral ferrous therapy, no patients from the B group showed any sign of these effects whatsoever. The results of the present study suggest that a diet rich in easily bioavailable iron could be an alternative therapy during the autotransfusion procedure.
Abstract: BACKGROUND AND METHODS: Patients with the nephrotic syndrome characteristically have multiple abnormalities of lipoprotein metabolism, but the cause and exact nature of these abnormalities are uncertain. In this study, we measured serum lipids and apoproteins in 57 patients with the nephrotic syndrome. We also determined the kinetic indexes of low-density lipoprotein (LDL) metabolism in six patients, and again in three of the six after recovery. RESULTS: The patients with the nephrotic syndrome had elevated serum concentrations of cholesterol, triglycerides, and phospholipids, which were confined to the lipoproteins containing apoprotein B. The serum concentrations of high-density lipoproteins and the associated A-I and A-II apoproteins were similar in the patients with the nephrotic syndrome and normal subjects. The relative proportions of lipids and their positive association with the increased serum concentrations of apoproteins B, C-II, C-III, and E suggest quantitative rather than qualitative differences in the lipoproteins. All the patients had lipiduria, which probably reflected the excretion of high-density lipoproteins, although no intact immunoreactive apoprotein A-I was found in urine. Serum albumin concentrations were inversely related to serum lipid concentrations in the patients, the severity of the hypoalbuminemia corresponding to the degree of change in serum lipoprotein concentrations. The kinetic studies of lipoprotein metabolism revealed an overproduction of LDL apoprotein B that returned to normal after recovery. CONCLUSIONS: The elevated serum concentrations of LDL cholesterol, other lipids, and apoprotein B in patients with uncomplicated nephrotic syndrome are due to reversible increases in lipoprotein production.
Abstract: Six patients (four women and two men) with mild to moderate hypercholesterolemia, but with no clinical evidence of the disease being monogenic familial hypercholesterolaemia and who, over the previous 3 months on a rigidly controlled hypolipidaemic diet therapy, showed no reduction in plasma cholesterol levels, were recruited into a study to assess the metabolic effects of Pirozadil, a new nicotinic acid derivative. After a 3 month treatment period, a significant reduction in plasma cholesterol from 299.8 +/- 31.2 mg/dl (mean +/- SD) to 256.8 +/- 18.1 mg/dl (P less than 0.02) and Low Density Lipoprotein (LDL) cholesterol from 211.7 +/- 44.9 mg/dl to 168.8 +/- 19.0 mg/dl (P less than 0.05) was observed. Although there was a trend toward decreased plasma and Very Low Density Lipoprotein (VLDL) triglyceride, the differences did not reach statistical significant. High Density Lipoprotein (HDL) cholesterol was unchanged. The drug was well tolerated with no side effects noted. To assess the mode of action, autologous125I-labelled LDL was injected and apoprotein B (apo B) kinetic parameters were measured; production rate (PR) and fractional catabolic rate (FCR). An in vitro measurement of the in vivo catabolism (LDL-apo B receptor activity in freshly isolated lymphocytes) was also measured pre- and post-treatment. The pharmacological intervention resulted in a significant decrease of 19.9% in PR from 10.5 +/- 1.81 mg/kg/d to 8.41 +/- 1.13 mg/kg/d (P less than 0.05) while the FCR remained relatively unchanged (0.260 +/- 0.042 vs 0.248 +/- 0.040 pools/d) as did the LDL receptor activity (78.2 +/- 20.9 vs 69.3 +/- 21.4 ng LDL/mg cell protein/hr).(ABSTRACT TRUNCATED AT 250 WORDS)
Abstract: The triglyceride and cholesterol content of total, very-low-, intermediate-, low-, and high-density lipoproteins, and of apolipoproteins (apo) Al, All, B, Cll, Clll, and E were determined in plasma from 107 patients with clinically well-controlled diabetes and from 66 age- and weight-matched healthy normal subjects. The diabetic patients were separated into two groups: those with insulin-dependent diabetes mellitus (IDDM, type 1, n = 24) and those with non-insulin-dependent diabetes mellitus (NIDDM, type 2, n = 83). The latter group contained two subgroups: those treated by diet (type 2d, n = 42) or by insulin (type 2i, n = 41). High-density lipoprotein cholesterol was increased in IDDM patients, and decreased in NIDDM patients relative to control subjects. Mean apo Al values in IDDM patients were higher than in their respective controls and in NIDDM patients. Concentrations of apo B, Clll, and E were higher in all diabetic patients than in the healthy controls, but those of apo Cll did not differ statistically between diabetics and nondiabetics. Although total plasma cholesterol and triglyceride concentrations were apparently near normal values in patients with good glycemic control, we found a persistent increase of intermediate-density lipoproteins (remnants) in all the diabetic groups studied. This factor may be related to the perceived increased cardiovascular risk in these individuals.
Abstract: With puromycin aminonucleoside-induced nephrotic syndrome (NS) in rats, twofold elevated levels of lipoproteins were observed. These levels were not related to proteinuria or to plasma albumin levels. Ultrastructural lesions induced in the kidneys by puromycin aminonucleoside were consistent with NS, while there was little or no hepatic involvement. Apolipoprotein B (apo B) kinetic measurements using homologous 125I-labeled low density lipoproteins (LDL) demonstrated a higher synthetic rate in nephrotic rats relative to controls (6.18 +/- 1.86 micrograms x g-1 x d-1 v 3.94 +/- 0.66 micrograms x g-1 x d-1 respectively, P less than .005), while the fractional catabolic rate was only marginally reduced (1.64 +/- 0.28 pools x day-1 in NS v 1.83 +/- 0.37 pools x day-1 in controls, P less than 0.4). These results indicate that in rats with experimentally induced NS, the expanded apo B-LDL pool results from increased synthesis of this apoprotein while no significant role can be ascribed to alterations in its catabolism. These data are consistent with our preliminary findings in NS in humans.
Abstract: Serum apoprotein A-I and lipid composition of high-density lipoproteins in 14 adults and 9 children with idiopathic untreated nephrotic syndrome without renal failure were studied. Serum apoprotein A-I in adults was significantly raised compared to a control group (1.72 +/- 0.49 and 1.45 +/- 0.15 g/l, Mean +/- SD, respectively); a similar trend was observed in children (1.88 +/- 0.62 and 1.52 +/- 0.21 g/l, respectively). High-density lipoprotein cholesterol was normal in both groups of patients (1.19 +/- 0.30 and 1.50 +/- 0.48 mmol/l in adults and in children, respectively), whereas high-density lipoprotein phospholipids were decreased in adults and increased in children. In addition, apoprotein A-I: high-density lipoprotein cholesterol ratios in both groups of patients were significantly increased with respect to their control groups. These alterations suggest the existence of qualitative changes in high-density lipoprotein composition. Despite other associated risk factors, and particularly from the point of view of the 'high-density lipoprotein hypothesis', our patients with untreated nephrotic syndrome without renal failure seem not to be at a higher coronary risk.
Abstract: We report two cases of ascending muscular weakness progressing to areflexic quadriplegia caused by severe derangement of potassium homeostasis. The first patient presented with a 17-alpha-hydroxylase deficiency and severe hypokalemia. The second case had primary adrenocortical deficiency (Addison's disease) and extreme hyperkalemia. Complete recovery ensued after correction of the metabolic disorder in both cases. The role of potassium in the pathophysiology of neuromuscular excitation is discussed. We conclude that when neuromyopathy is present, metabolic causes should be considered and the serum potassium determined.
Abstract: Cholesterol content in high-density lipoprotein (HDL) subfractions has been studied in 108 patients at different evolutive stages of chronic renal failure (CRF) under conservative treatment. Results have been compared with healthy control subjects, patients receiving hemodialysis, and renal graft recipients. Significant low levels of total HDL and HDL2 cholesterol are observed in men with CRF. The more severe the CRF, the more likely that total HDL and HDL2 cholesterol will be low. Moreover, a significant inverse correlation is found between HDL2 cholesterol and serum creatinine levels. In women, although a decrease in total and HDL2 subfraction is observed, no significant differences are found across the severity of CRF. Serum HDL2 cholesterol levels are decreased in men and women receiving hemodialysis, while raised total HDL and HDL2 cholesterol levels are observed in normally functioning renal grafts. These results indicate that according to the "HDL hypothesis," despite other associated risk factors, the high cardiovascular mortality rates noted mainly in men with CRF under conservative treatment and in patients receiving hemodialysis could be explained, at least in part, by the sustained and progressive decrease in total HDL and HDL2 values. From this point of view, our study suggests the need to promote early kidney transplantation.
Abstract: Serum gonadal hormones, gonadotrophins and zinc levels were studied in thirteen men aged 29-62 yr with chronic renal failure undergoing haemodialysis. All patients had decreased libido and impotence. Serum testosterone levels in patients (18.5 +/- 1.3 (SEM) nmol/l) were significantly lower (p less than 0.05) than in the control group (24.1 +/- 2.2 (SEM) nmol/l) although salivary testosterone levels were strictly within the normal range. Mean serum 17-beta-oestradiol and luteinizing hormone levels (0.19 +/- 0.03 (SEM) nmol/l, and 57.4 +/- 13.1 (SEM) IU/l, respectively) were significantly higher (p less than 0.05 and p less than 0.005, respectively) than in the control group (0.11 +/- 0.02 (SEM) nmol/l and 14.8 +/- 1.9 (SEM) IU/l, respectively). Mean progesterone and follicle-stimulating hormone levels in patients were not significantly different from those of control subjects. Mean prolactin values in patients (1,019 +/- 285 (SEM) mIU/l) were significantly higher (p less than 0.01) than in the control group (211 +/- 24 (SEM) mIU/l). Serum prolactin levels in five patients were extremely high (above 1,200 mIU/l). There was no statistically significant difference in serum zinc levels between patients and controls. As salivary testosterone is normal, it seems that hyperprolactinaemia and raised serum 17-beta-oestradiol levels may be responsible, at least in part, for sexual dysfunction in male patients with chronic renal failure receiving haemodialysis.
Abstract: Serum concentration of apoprotein A, high density lipoprotein (HDL)-cholesterol and HDL-phospholipids has been studied in thirteen consecutive episodes of diabetic ketoacidosis. In three patients with type I diabetes mellitus HDL2 and HDL3 subfractions were also measured. Patients with type I diabetes showed greatly decreased HDL-cholesterol concentration on admission which increased into the normal range after insulin treatment, while HDL-phospholipids decreased during treatment and apoprotein A remained almost unmodified. In three patients with type I diabetes a virtual absence of HDL2-cholesterol subfraction was observed, which rose to normal values during recovery. Conversely, in type II diabetes mellitus HDL-cholesterol was slightly reduced on admission, and tended to decrease during recovery. These findings imply the existence of abnormalities in the qualitative composition of HDL, and indicate that HDL-cholesterol can fluctuate much more rapidly than previously thought.
Abstract: Serum concentration of apoprotein A-I (apo A-I) and cholesterol content in high density lipoprotein (HDL) subfractions have been studied in 19 men and 11 women at the end stage of chronic renal failure undergoing hemodialysis. HDL2 cholesterol concentration was decreased in males [0.33 +/- 0.12 (mean +/- SD) mmol/l, controls 0.45 +/- 0.09 mmol/l; p less than 0.001]; in females HDL2 cholesterol was also decreased although without statistical significance (0.45 +/- 0.15 vs. 0.55 +/- 0.10 mmol/l). HDL3 cholesterol was significantly decreased in men (0.65 +/- 0.11 vs. 0.77 +/- 0.04 mmol/l; p less than 0.001) and also in women (0.61 +/- 0.12 vs. 0.82 +/- 0.07 mmol/l; p less than 0.005). However, serum concentration of apo A-I was within the normal range (1.13 +/- 0.16 milligram in males and 1.25 +/- 0.17 milligram in females; controls 1.24 +/- 0.17 and 1.35 +/- 0.19 milligram, respectively). The raised apo A-I/HDL2 cholesterol ratio in both men and women suggests the existence of qualitative changes in HDL subfractions as has been proposed in previous studies measuring total apo A and total HDL cholesterol in patients with chronic renal failure receiving hemodialysis. These abnormalities in the relative composition of HDL subfractions could play an important role as a vascular risk factor in patients with chronic renal failure undergoing hemodialysis.
Abstract: Isolated and washed cryoglobulins obtained from 30 patients with different diseases were studied for the presence of fibronectin by immunoelectrophoresis. Two of the isolated cryoglobulins were of the type I, 3 were of the type II, while the other 25 were of type III. Despite extensive washing, fibronectin was detected in all the cryoprecipitates. Our results indicate the constant presence of fibronectin in the cryoprecipitates from patients with different diseases and types of cryoglobulin. The possible mechanisms by which fibronectin could act in the precipitation of immunoglobulins are discussed.
