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S Marcovina

Publications and source records attributed to S Marcovina.

At least 37 records · Page 2Linked to original sources
PubMed1996

Cholesterol levels among Japanese Americans and other populations: Seattle Nikkei Health Study.

The purpose of this study was to compare average cholesterol levels between Seattle based Japanese Americans and three other populations: U.S. population, native Japanese population and native Japanese urban workers. A total of 1,466 Japanese Americans (724 men and 742 women) participated in cardiovascular disease screening in the Seattle area during 1989 94. Data sources for comparisons are from the Third National Health and Nutrition Examination Survey for 1988-91, the results of the National Cardiovascular Disease Examination Survey in Japan for 1990, and cardiovascular disease screening conducted by the Epidemiological Arteriosclerosis Research Institute in Japan for 1989. Total cholesterol and triglyceride levels of Seattle Japanese American men and women were highest among the four populations. Among men, high density lipoprotein cholesterol (HDL-C) levels for Seattle Japanese Americans and native Japanese were similar and fell between those of urban Japanese workers and the U.S. population. In women, the average HDL C levels were highest in the Japanese urban workers, second highest in Seattle Japanese Americans, and lowest in both the U.S. population and native Japanese population. These differences in lipid levels may be caused by both genetic and environmental factors, which are now under investigation.

Adult↗
PubMed1996

Localization of apolipoprotein A-I epitopes involved in the activation of lecithin:cholesterol acyltransferase.

Eight murine monoclonal antibodies (Mab) to apolipoprotein A-I were characterized for their epitopes and for their ability to interfere with lecithin:cholesterol acyltransferase (LCAT) activation mediated by apo apoA-I using a synthetic substrate. Using overlapping synthetic peptides we have identified six continuous epitopes that span amino acids 1-10 (Mab A-I-19), 96-101 (Mab A-I-15), 133-141 (Mab A-I-5), 140-145 (Mab A-I-9), 144-148 (Mab A-I-8), and 167-174 (Mab A-I-57). Furthermore, antibodies A-I-11 and A-I-16 recognized discontinuous epitopes, namely amino acids 124-128 and 144-148. When antibodies were tested for their ability to inhibit LCAT activation, an inhibitory effect was observed with those whose epitopes covered the area of apoA-I encompassing amino acids 96-174. From these data we conclude that several areas of apoA-I spanning the middle region of the apolipoprotein act in concert to stimulate LCAT activity, possibly by cooperative interaction with the enzyme.

Animals↗
PubMed1995

In vitro expression of structural defects in the lecithin-cholesterol acyltransferase gene.

Classic LCAT deficiency (CLD) and fish eye disease (FED) are two clinically distinct syndromes, associated with defects in the lecithin-cholesterol acyltransferase (LCAT) gene resulting in total (CLD) or partial (FED) enzyme deficiency. In order to investigate the underlying molecular mechanisms that lead to different phenotypic expression in CLD and FED, LCAT mutants associated with either CLD (LCAT147, LCAT156, and LCAT228) or FED (LCAT10, LCAT123, LCAT158, LCAT293, LCAT300, and LCAT347) were expressed in vitro in human embryonic kidney 293 cells and characterized with respect to LCAT expression and enzyme activity. Evaluation of mutant LCAT gene transcription by Northern blot analysis demonstrated LCAT mRNA of normal size and concentration. Although all constructs gave rise to similar intracellular LCAT mass, the amount of enzyme present in the media for LCAT147, LCAT156, and LCAT300 was reduced to less than 10% of normal, suggesting that these mutations disrupted LCAT secretion. Western blot analysis of cell culture media containing wild type or mutant LCAT demonstrated the presence of a single normal-sized band of 67 kDa. The ability of the different enzymes to esterify free cholesterol in high density lipoprotein-like proteoliposomes (alpha-LCAT-specific activity) was reduced to less than 5% of normal for CLD mutants LCAT147 and LCAT228 and FED mutants LCAT10, LCAT123, LCAT293, and LCAT347, whereas that of LCAT156, LCAT158, and LCAT300 ranged from 45 to 110% of control. Although most FED mutant LCAT enzymes retained the ability to esterify free cholesterol present in alpha- and beta-lipoproteins of heat-inactivated plasma, esterification was undetectable in all CLD mutants (LCAT147, LCAT156, and LCAT228). In contrast, all mutant enzymes retained the ability to hydrolyze the water soluble, short-chained fatty acid substrate p-nitrophenolbutyrate. In summary, our studies establish the functional significance of nine LCAT gene defects associated with either FED or CLD. Characterization of the expressed LCAT mutants identified multiple, overlapping functional abnormalities that include defects in secretion and/or disruption of enzymic activity. All nine LCAT mutants retained the ability to hydrolyze the water-soluble PNPB substrate, indicating intact hydrolytic function. Based on these studies we propose that mutations in LCAT residues 147, 156, 228 (CLD) and 10, 123, 158, 293, 300, and 347 (FED) do not disrupt the functional domain mediating LCAT phospholipase activity, but alter structural domains involved in lipid binding or transesterification.

Blotting, Northern↗
PubMed1995

Relative contributions of apolipoprotein(a) and apolipoprotein-B to the development of fatty lesions in the proximal aorta of mice.

Transgenic mice expressing transgenes for both human apolipoprotein B-100 (h-apoB) and apolipoprotein(a) [apo(a)] were fed a high-fat, atherogenic diet for 14 weeks to examine the effect of lipoprotein(a) [Lp(a)]on the development of aortic fatty lesions. The extent of lesions in the proximal region of the aorta of Lp(a) mice was measured by use of a computer-assisted image analysis of 20 sections per animal and compared with that of nontransgenic mice as well as mice expressing either the apo(a) or h-apoB transgene. The control (n = 23) and apo(a) (n = 22) transgenic mice had very small mean lesions areas (607 versus 128 microns2 per section). The h-apoB-expressing mice (n = 20) had significantly higher mean lesion areas (3288 microns2 per section) than either the control or apo(a) transgenic animals. Coexpression of apo(a) and h-apoB transgenes resulted in only a modest increase in lesion area (4678 microns2 per section, n = 19). Thus, the expression of human apo(a) in C57BL/6/SJL hybrid mice fed an atherogenic diet failed to significantly potentiate the development of aortic fatty lesions in the absence or presence of high levels of h-apoB.

Animals↗
PubMed1995

Testosterone enanthate at a dose of 200 mg/week decreases HDL-cholesterol levels in healthy men.

The concept that androgen alone can provide an effective male contraceptive has been tested in a multicentre, multiphase trial by the World Health Organization. Results from this trial showed that an ester of testosterone, testosterone enanthate (TE), administered at a dose of 200 mg/week, has a very high contraceptive efficacy, and suggested that, at least in some populations, androgen alone might provide a viable option for the control of male fertility. It has been claimed that testosterone represents one of the gender-related risk factors for coronary artery disease (CAD) in men. Epidemiological and interventional studies have failed to establish a convincing relationship between testosterone and high density lipoprotein cholesterol (HDL-C). Therefore, there is concern about possible negative effects on lipoprotein asset of an androgen-alone male contraceptive. In this study we analysed the effects of long-term (12 months) administration of TE (200 mg/week) in normal healthy men. Blood samples (six men > 10 h fast = Group 1; 30 men > 4 h fast = Group 2) were drawn from 36 men, monthly before the beginning of the injections (control), every 3 months throughout the study period (treatment), and 1 month after stopping TE injections (recovery). Total cholesterol (chol), triglycerides, HDL-C and LDL-C levels were measured in these samples. Biochemical parameters were also monitored. TE administration induced a significant decrease (15-20%) in HDL-C levels that was of comparable magnitude in men from both groups (fasting and non-fasting) and occurred regardless of basal HDL-C levels. No statistically significant effect on other lipoproteins was detected. Considering all men together, HDL-C levels were decreased in 78% of the men by month 3, 83% by month 6, 94% by month 9 and 97% by month 12 of treatment. In all men the HDL-C decrease was reversible within 1 month of stopping TE administration. It is concluded that: (1) injection of 200 mg TE/week causes a 15-20% decrease in HDL-C in normal men with no effect on other lipoproteins, (2) the suppressive effect of TE is maintained throughout the 1-year-injection period, and a direct relationship between the duration of TE administration and the proportion of men showing decreased HDL-C levels, was observed. (3) The HDL-C decrease was reversible within 1 month of stopping TE administration. These data will be important in designing further studies on male contraception, and in interpreting the relationship between testosterone levels, HDL-C levels and potential cardiovascular risk.

Adult↗
PubMed1994

Unique epitope of apolipoprotein A-I expressed in pre-beta-1 high-density lipoprotein and its role in the catalyzed efflux of cellular cholesterol.

The ability of mouse anti-apolipoprotein A-I (apo A-I) monoclonal antibodies to recognize pre-beta-HDL species in native plasma was determined. An antibody identifying residues 137-144 of the mature protein uniquely recognized pre-beta-1 HDL, an HDL species of low molecular weight implicated in early cholesterol transport from cell membranes to plasma [Castro, G. R., & Fielding, C. J. (1988) Biochemistry 27, 25-29]. Incubation of plasma with this antibody significantly inhibited the efflux of labeled cholesterol from cultured fibroblast monolayers. A second antibody, binding to residues 93-99 of apo A-I, recognized a second pre-beta-HDL species (pre-beta-2 HDL) but not pre-beta-1 HDL and did not inhibit cholesterol efflux. Several other antibodies had broad specificity for HDL (including pre-beta-1 HDL). This research suggests that apo A-I residues 137-144 are adjacent to or part of a structural site in pre-beta-1 HDL active in promoting the efflux of cellular cholesterol and that this site is not exposed in other HDL species.

Amino Acid Sequence↗
PubMed1993

Immunoreactivity of apo B towards monoclonal antibodies that inhibit the LDL-receptor interaction: effects of LDL oxidation.

We studied the immunochemical stability of the epitopes for six monoclonal antibodies to human apolipoprotein B-100 upon Cu(2+)-mediated (20 microM) oxidation of LDL. The antibodies used in this study, some of which are known to interfere with the interaction of LDL with their cellular receptors, recognize epitopes in the amino terminal region (Mb 19), in the middle part (6B, 2A, 7A, and 9A) and near aa 3500 (Mb 47) of native apo B. All antibodies except one (7A) recognized native and oxidized LDL (OxLDL) equally well; the immunoreactivity of the epitope for Ab 7A was markedly reduced upon LDL oxidation. Since antibodies 2A, 7A, 9A, and Mb 47 inhibit the LDL-receptor interaction and OxLDL poorly interact in vitro with the LDL receptor we conclude that: (1) various epitopes for monoclonal antibodies against native apo B are spared upon LDL oxidation; and (2) the epitopes for antibodies 2A, 9A, and Mb 47 do not define a unique domain of apo B directly involved in the binding of LDL to their receptor.

Antibodies, Monoclonal↗
PubMed1993

Comparative study of the activity and composition of HDL3 in Russian and American men.

Previous studies conducted within the framework of the Lipid Research Clinics Program showed a strong inverse correlation between high-density lipoprotein cholesterol (HDL-C) level and coronary heart disease (CHD) risk in American male populations, whereas in Russian populations such a correlation was less pronounced. It was assumed that HDL was less protective of CHD in Russian than in American males. This study compared the functional activity and lipid composition of HDL3 isolated from the blood plasma of men with low, normal, and high HDL-C levels from Moscow (Russia) and Seattle (United States) populations. Results obtained showed that American HDL3, irrespective of the plasma HDL-C level, had higher activity in stimulating both [3H]cholesterol and cholesterol mass efflux from cholesterol-loaded fibroblasts and in suppressing cellular cholesterol esterification when compared with Russian HDL3. American HDL3 remained more active than Russian HDL3, even when apolipoprotein E-containing particles were removed from HDL3 by heparin-Sepharose affinity chromatography. Russian and American 125I-HDL3 had similar binding to high-affinity cell-surface sites, but Russian HDL3 had a higher nonspecific binding component compared with American HDL3. This study demonstrates for the first time potential functional differences between HDL particles isolated from Russian and American populations. The lower activity of Russian HDL3 in promoting cellular cholesterol efflux may partly explain the higher CHD risk in the Russian population compared with the American one.

Adult↗
PubMed1993

Fish eye syndrome: a molecular defect in the lecithin-cholesterol acyltransferase (LCAT) gene associated with normal alpha-LCAT-specific activity. Implications for classification and prognosis.

We have identified the molecular defect in two siblings presenting with classical clinical and biochemical features of Fish Eye disease (FED), including corneal opacities, HDL cholesterol < 10 mg/dl, normal plasma cholesteryl esters, and elevated triglycerides. In contrast to previously reported patients with FED who are unable to esterify HDL-associated cholesterol, our patients' plasma lecithin-cholesterol acetyltransferase (alpha-LCAT)-specific activities assayed using an HDL-like proteoliposome substrate were 12.7-25.7 nmol/micrograms (19.5 +/- 1.8 in controls). In addition, significant residual cholesterol esterification was present in VLDL/LDL-depleted plasma, confirming the presence of HDL-associated alpha-LCAT activity. DNA sequence analysis of the proband's LCAT gene identified deletion of the triplet coding for leu300, which resulted in the loss of a restriction site for MlnI. Digestion of PCR-amplified DNA using MlnI established that both siblings are homozygous for this defect. Expression of LCAT300-del. in human embryonic kidney-293 cells revealed normal mRNA and intracellular LCAT concentrations. However, reduced amounts of LCAT300-del., which had a normal specific alpha-LCAT activity, were present in the media. In summary, we report the first case of FED associated with a mutant enzyme that has a normal alpha-LCAT-specific activity. The functional significance of this LCAT gene defect has been established in an in vitro expression system, which demonstrates that very small amounts of this functional LCAT mutant enzyme accumulate in the media. Characterization of LCAT300-del. established that selective alpha-LCAT deficiency is not a prerequisite for the development of FED. On the basis of our combined results, we propose that the residual amounts of total plasma LCAT activity and not its distribution on lipoproteins primarily determines the heterogeneity in phenotypic expression observed in familial LCAT deficiency syndromes.

Base Sequence↗
PubMed1992

Monoclonal antibody-based enzyme-linked immunosorbent assays (ELISA) for the measurement of vitamin K-dependent protein S: the effect of antibody immunoreactivity on plasma protein S antigen determinations.

Two monoclonal antibodies (Mabs) specifically directed to human protein S (PS) - named 5E9E9 and 3B10.25 - were produced and their properties compared to those of 2 previously characterized anti-PS-Mabs (HPS-2 and S10). 3B10.25, similar to S10, was directed to the calcium-free conformation of PS and had virtually identical affinity for free and C4b-binding protein (C4b-BP)-bound PS; 5E9E9 similar to HPS-2, had no calcium-dependency and was selectively directed to free PS. All Mabs were equally reactive to freshly purified and thrombin-cleaved PS. To evaluate the influence of C4b-BP bound PS on PS antigen determinations, ELISA systems employing the four Mabs individually as capture antibody (Ab) and peroxidase-conjugated polyclonal anti-PS IgG as detecting Ab were developed and compared to immunoelectrophoresis (EIA) and to an ELISA employing polyclonal anti-PS IgG as capture and detecting Ab, in the determination of PS in purified systems and in plasma. With all the ELISAs there was parallelism of dilution curves obtained with normal plasma and purified PS; however, supplementation of plasma with purified C4b-BP resulted in loss of parallelism when employing the Mabs directed to free PS as capture Ab. Influence of high C4b-BP on PS antigen determinations was confirmed in a series of plasma samples from patients with C4b-BP levels ranging from 70% to over 200%. Compared to the values obtained with the S10- or 3B10.25 - based ELISAs - which were similar despite a 10-fold difference in sample dilution - plasma PS was underestimated by the ELISAs employing 5E9E9 or HPS-2 while it was overestimated by EIA.(ABSTRACT TRUNCATED AT 250 WORDS)

Antibodies, Monoclonal↗
PubMed1992

Monoclonal antibodies to human low density lipoprotein identify distinct areas on apolipoprotein B-100 relevant to the low density lipoprotein-receptor interaction.

We have characterized the epitopes for ten murine monoclonal antibodies (Mabs) to human low density lipoprotein (LDL) and studied their ability to interfere with the LDL-receptor interaction. The epitopes for the antibodies were defined by using the following approaches: 1) interaction with apoB-48; 2) interaction with apoB-100 thrombolytic fragments; and 3) interaction with beta-galactosidase-apoB fusion proteins spanning different areas of the apoB-100 sequence. The results obtained are consistent with the following map of epitopes: Mab 6E, amino acids (aa) 1-1297, Mabs 5A and 6B, aa 1480-1693, Mabs 2A, 7A, 3B, and 4B, aa 2152-2377, Mabs 8A and 9A, aa 2657-3248 and 3H, aa 4082-4306. Four Mabs (2A, 5A, 7A, and 9A) whose epitopes are located in three different areas of apoB, dramatically reduced (up to 95%) the LDL-receptor interaction on cultured human fibroblasts; Fab fragments were as effective as the whole antibodies. Mab 3H, on the other hand, increased LDL binding up to threefold. These findings are consistent with the hypothesis that several areas of apoB-100 are involved independently or in concert in modulating the apoprotein B conformation required for interaction with the LDL receptor.

Antibodies, Monoclonal↗
PubMed1992

Immunohistochemical distribution of lipoprotein epitopes in xanthomata from patients with familial hypercholesterolemia.

Human xanthomas derived from four subjects with familial hypercholesterolemia (3 homozygotes and 1 heterozygote) were studied by immunohistochemical methods to determine the presence and distribution of lipoproteins, which have been implicated in the pathogenesis of atherosclerosis. Oxidatively modified low-density lipoprotein (OxLDL) epitopes detected with anti-OxLDL monoclonal antibodies, appeared to have a similar distribution in xanthomata to that of macrophages, detected by a cell-specific monoclonal antibody. Double antibody labeling with both an anti-macrophage antibody and an anti-OxLDL antibody demonstrated that OxLDL epitopes are associated with macrophages and occurred intracellularly. Low-density lipoprotein (LDL) epitopes were detected extracellularly, with a distribution that was different from that of OxLDL. In addition, apo(a) epitopes detected by an apo(a) specific monoclonal antibody, had a distribution similar to that of LDL in the dermis and subcutaneous tissues. The observed epitope distribution of LDL, OxLDL, or apo(a) was the same regardless of the method of treatment of the patients from whom the xanthomas were obtained (probucol, simvastatin, LDL apheresis). These findings suggest that OxLDL is likely to play a pathogenetic role in the lipid accumulation by macrophages in xanthomas, and suggest that Lp(a) also may play a role in their pathogenesis.

Adolescent↗
PubMed1990

Effects of intraperitoneal versus subcutaneous insulin administration on lipoprotein metabolism in type I diabetes.

In order to compare the effects of intraperitoneal (IP) versus subcutaneous (SC) insulin delivery on plasma lipoproteins, lipoprotein cholesterol, triglycerides, and very-low-density lipoprotein (VLDL) metabolism were compared in five type I diabetic patients while they were receiving continuous IP insulin (CIPII) or continuous subcutaneous insulin infusion (CSII). Each therapy regimen was of at least 1 month duration, and patients were treated in random order. Mean daily plasma insulin was lower on CIPII compared with CSII. CIPII was associated with lower VLDL triglycerides and VLDL apolipoprotein (apo) B, and higher high-density lipoprotein (HDL) and HDL3 cholesterol. The decreased VLDL on CIPII appeared to be the result of both decreased production and increased clearance of VLDL apo B. The results suggest that the more physiologic route of insulin therapy (CIPII) is associated with lipoprotein profiles of lower atherogenic potential.

Adult↗
PubMed1990

Immunochemical characterization of six monoclonal antibodies to human apolipoprotein A-I: epitope mapping and expression.

We have produced and characterized six murine monoclonal antibodies to human apolipoprotein A-I named A-I-9, A-I-12, A-I-15, A-I-16, A-I-19, and A-I-57. All monoclonal antibodies were specific for apolipoprotein A-I and bound between 55% and 100% of 125I-labeled high density lipoproteins (HDL) in a fluid phase radioimmunoassay. All antibodies possessed a higher affinity to apoA-I in HDL than to free, delipidated apoA-I. Two of them, particularly A-I-12 and A-I-15, which were directed to the same or very close epitopes on the molecule, recognized very poorly the delipidated protein. Binding of apoA-I to phospholipid restored the immunoreactivity of the monoclonal antibodies to the protein suggesting that lipids play an important role in determining the immunochemical structure of apoA-I. Using CNBr fragments and synthetic peptides, the epitopes for the antibodies were mapped as follows: A-I-19, CNBr fragment 1; A-I-12 and 15, CNBr fragment 2; A-I-9 and A-I-16, CNBr fragment 3; A-I-57, CNBr fragment 4. Antibody A-I-57 failed to recognized a mutant form of apoA-I, A-IMilano (Arg173----Cys) by immunoblotting and by competitive radioimmunoassay demonstrating that substitution of a single amino acid in human apoA-I may cause the loss of an antigenic determinant.

Animals↗
PubMed1989

Monoclonal antibodies directed to the calcium-free conformation of human protein S.

Four mouse hybridomas secreting monoclonal antibodies specific for human protein S (PS) have been generated. The antibodies, all of the IgG1 subclass, were designated S2, S3, S8, and S10. In a fluid phase radioimmunoassay, the binding of monoclonal antibodies to PS was about 30% greater in the presence of EDTA and totally inhibited in presence of Ca2+. Using the same technique, we performed displacement curves of 125I-labeled PS by purified PS, thrombin-cleaved PS, normal plasma, plasma from a patient on warfarin therapy, and plasma from a patient with no free PS and only PS bound to C4b-binding protein. The slopes of the curves show that the monoclonal antibodies reacted equally with all the tested forms of PS indicating that the antigenic site(s) to which the monoclonal antibodies are directed are present and exposed in free and bound PS, in thrombin-cleaved PS, and in the coumarin form of the protein. Each EDTA-dependent antibody, immobilized on Sepharose 4B-CNBr was used to purify PS from the barium citrate-absorbed, ammonium sulphate-soluble fraction of plasma. The fraction eluted from the immunoabsorbent with a buffer containing 4 mmol/l CaCl2 and analysed by SDS-PAGE, contained two bands, one migrating with conventionally purified PS and the other with purified C4b-binding protein. Homogeneous PS was obtained by chromatography of the barium citrate adsorbate on a DEAE-Sephadex column. The protein peak containing the bulk of PS was subsequently applied to the immunoadsorbent and eluted with 4 mmol/l CaCl2.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗