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Biomedical subjects

K Cianflone

Publications and source records attributed to K Cianflone.

At least 73 records · Page 4Linked to original sources
PubMed1995

Response of plasma ASP to a prolonged fast.

OBJECTIVE: To determine the changes in the plasma level of acylation stimulating protein (ASP) during a one month total fast in female subjects with marked obesity. DESIGN: Patients with marked obesity underwent a month total fast, before, during (2 weeks), and at the end of which, a variety of relevant metabolic parameters were measured. SETTING: A metabolic unit of a teaching hospital. SUBJECTS: 10 women with marked obesity were studied and the results compared with those in 16 age-matched controls. MAIN OUTCOME MEASURES: Plasma ASP, lipoprotein lipids, apoB, free fatty acid, and ketone levels. RESULTS: At baseline, fasting levels of ASP in the obese group were double that in control subjects (116 +/- 26 vs 53 +/- 30 nM P < 0.001). During the fast, ASP levels dropped progressively and were within the normal range at the end of the study (63 +/- 16 vs 53 +/- 30 nM pNS). In addition, there was a strong correlation between the plasma ASP at baseline before beginning the fast and the 4 week drop in ASP. That is, those subjects who had the highest starting ASP also had the largest 4 week drop in ASP (r2 = 0.644, P < 0.005). Of interest, as plasma ASP levels dropped, plasma free fatty acid and ketone levels rose and when all timepoints were considered, there was a significant inverse relation between plasma ASP and plasma free fatty acid (r2 = 0.295, P < 0.0002). CONCLUSIONS: The pattern of responses during the fast is that of increasing mobilization of fatty acids from adipose tissue coincident with decreased activity of the pathway responsible for the storage of adipocyte triglyceride mass. The data are consistent, therefore, with the role proposed for ASP as a major determinant of the rate of triglyceride synthesis in human adipocytes and thus a potentially important factor in the pathophysiology of obesity.

Adult↗
PubMed1995

Role of the neutral lipid accessible pool in the regulation of secretion of apoB-100 lipoprotein particles by HepG2 cells.

The rate of secretion of apoB-100-containing lipoprotein particles by HepG2 cells is determined to an important extent by post-translational mechanisms, the mass of neutral lipids clearly playing a role in this process. Our previous data indicated that cholesteryl ester might influence the proportion of newly synthesized apoB-100 molecules that are incorporated into nascent lipoproteins rather than being catabolized intracellularly shortly after they have been synthesized. The present studies, therefore, were designed: 1) to examine in more detail the relationship between the mass of triglyceride and cholesteryl ester in HepG2 cells and the rate of apoB-100 secretion, and 2) to determine whether cholesteryl ester molecules that have been synthesized and stored within these cells must undergo hydrolysis and re-esterification before being secreted with newly synthesized apoB-100 molecules. Changes in apoB-100 secretion in HepG2 cells were assessed in response to changes in intracellular triglyceride and/or cholesteryl ester pool size. This was accomplished through lipid loading of the cells by incubating them overnight with exogenously supplied very low density lipoprotein (VLDL) (with lipoprotein lipase, LPL), low density lipoprotein (LDL), oleate, or LDL + oleate. The medium was changed to fresh serum-free medium and apoB-100 secretion was shown to increase over at least 8 h. After overnight incubation with VLDL, intracellular triglyceride mass increased 6-fold, while intracellular cholesteryl ester mass increased 2-fold. Medium apoB-100 increased up to 3-fold, while apoB-100 mRNA increased by only 12%. Both heparin (10 IU/ml) and lactoferrin (20 microM) independently blocked the VLDL-mediated increases in intracellular cholesteryl ester mass (-56% and -46%) without decreasing triglyceride mass. ApoB-100 secretion was also reduced by 53% and 72%, respectively. Incubation of HepG2 cells with LDL increased intracellular cholesteryl ester mass but triglyceride mass remained unchanged. In this instance, apoB-100 secretion increased 2-fold but there was no change in apoB-100 mRNA. Overall, there was little relationship between the mass of intracellular triglyceride and the rate of apoB-100 secretion (r2 = 0.034, NS) whereas there was a strong correlation between the intracellular mass of cholesteryl ester and apoB-100 secretion (r2 = 0.67, P < 0.0005). To examine the process of cholesteryl ester secretion, intracellular triglyceride and cholesteryl ester mass were increased after incubation with LDL + oleate. The medium was then changed to fresh serum-free medium containing an acyl-CoA:cholesterol acyltransferase (ACAT) inhibitor (Sandoz compound 58-035). Although de novo cholesteryl ester synthesis was inhibited up to 89%, cholesteryl ester mass secretion remained constant with up to 15% of total cholesteryl ester mass secreted over the 8-h period. ApoB-100 secretion also remained elevated above control, with 92% of the cholesteryl ester secreted associated with apoB-100 particles (27% with d < 1.006 g/mL particles and 65% with d 1.006-1.063 g/mL particles). Therefore, not only newly synthesized cholesteryl ester but also stored cholesteryl ester can associate with newly synthesized apoB-100 molecules and can be secreted without the necessity of an intracellular hydrolysis/re-esterification step.

Apolipoprotein B-100↗
PubMed1994

Adipsin/acylation stimulating protein system in human adipocytes: regulation of triacylglycerol synthesis.

Through their capacity to store fatty acids as triacylglycerol molecules, adipocytes serve a vital physiologic role. This study presents further evidence that this process can be modulated in human adipocytes by the adipsin/acylation stimulating protein (ASP) pathway and suggests a novel function for the product of this system--ASP. The data demonstrate the following: (1) ASP stimulates triacylglycerol synthesis within adipocytes, and this occurs to a greater extent in differentiating than undifferentiated cells (242% +/- 32% vs 168% +/- 11%, p < 0.01, respectively, at an ASP concentration of 88 ng/mL; (2) ASP does not affect the Km for triacylglycerol synthesis but does substantially increase Vmax; (3) when ASP is generated in vitro through incubation of its precursor proteins under appropriate conditions, triacylglycerol synthesis increases to the same extent as when plasma-purified ASP is added to the medium; (4) human adipocytes contain mRNA for the specific serine protease adipsin and the two precursor proteins C3 and factor B required to interact for the production of ASP; and (5) the extent to which cultured differentiating adipocytes produce ASP is proportional to the degree to which they have accumulated triacylglycerol mass during differentiation (r2 = 0.7523, p < 0.0005). These findings provide the first evidence for the existence of the adipsin/ASP pathway in human adipocytes, and this may markedly enhance our understanding of the processes which regulate triacylglycerol clearance from plasma.

Acylation↗
PubMed1994

Effects of albumin on lipid synthesis, apo B-100 secretion, and LDL catabolism in HepG2 cells.

This study examines the effects of extracellular albumin on hepatic apo B-100 metabolism. To do so, a transformed human liver cell line, HepG2, was used as a hepatocyte model and the concentration of albumin in the medium was varied between 0 and 5 g%. Apo B-100 and apo A1 concentrations in the medium were determined by specific enzyme-linked immunoassay (ELISA) and intracellular synthesis of cholesterol ester and triglyceride were determined by addition of appropriate radiolabels to the medium. The data demonstrate that the reduction of extracellular albumin concentration resulted in increased apo B-100 concentration in the medium. Apo A1 secretion, however, was unaffected. While the differences in apo B-100 concentration in the medium were statistically significant (33% +/- 7%, P < 0.0025, 0 g% albumin compared to 5 g% albumin in the medium), the absolute magnitude of the effect under these conditions was relatively modest. Nevertheless, the changes were consistent and evident over incubation periods as long as 8 days. Of interest, although triglyceride synthesis was unaffected, cholesterol ester synthesis changed such that as albumin concentration decreased, synthesis of cholesterol ester increased paralleling the changes in apo B-100 (170% +/- 9%, P < 0.005). These findings were extended by studying interventions which altered cholesterol ester synthesis. Addition of the compound 58-035 (5 micrograms/ml, a specific inhibitor of acylcholesterol acyltransferase activity) resulted in substantial inhibition of cholesterol ester synthesis (39% to 66%, P < 0.025 and P < 0.005, respectively) and apo B-100 concentrations in the medium which decreased by 20% to 28%, P < 0.025. Triglyceride synthesis, in contrast, increased significantly by 32% P < 0.025. Therefore, addition of 58-035 confirmed the previous findings of a parallel relation between cholesterol ester synthesis and apo B-100 concentration in the medium. Nonetheless, albumin still had an additional inhibitory effect on cholesterol ester and apo B-100 secretion. Of interest, when chylomicron remnants (25 micrograms/ml cholesterol), which cause apo B-100 secretion to increase by more than threefold, were added to the medium, albumin now had a more pronounced absolute effect on apo B-100 secretion with a 48% inhibition observed as albumin was increased from 0 to 5 g% in the medium (P < 0.0125). The effect of extracellular albumin on the low density lipoprotein (LDL) pathway was also examined. No differences in non-specific cell association component were detected.(ABSTRACT TRUNCATED AT 400 WORDS)

Amides↗
PubMed1994

Increase in intracellular triglyceride synthesis induced by gemfibrozil.

Gemfibrozil is an effective hypotriglyceridemic agent. Its mechanism of action has not been determined with certainty, but it is generally thought to act by virtue of an effect on lipoprotein lipase. However, this study was designed to test the hypothesis that gemfibrozil directly affects intracellular triglyceride synthesis in peripheral cells, and indeed, when added to the medium, gemfibrozil markedly stimulated triglyceride synthesis in cultured human skin fibroblasts. The effect was concentration-dependent, with a maximum effect at a gemfibrozil concentration of 40 micrograms/mL. A marked increase in triglyceride synthesis was observed when either labeled glucose or labeled fatty acid were added to the medium, indicating that the stimulation was substrate-coordinated. The effect was sustained for hours, with an onset of action within minutes. Lineweaver-Burk plots were constructed, and these revealed no apparent change in Km for either oleate or glucose (8.18 +/- 2.97 v 6.40 +/- 2.10 mumol/L for oleate and 0.113 +/- 0.004 v 0.107 +/- 0.051 mmol/L for glucose, control v gemfibrozil, P = NS). By contrast, there was a significant increase in the maximum triglyceride synthetic rate (Vmax) for both oleate and glucose (14.57 +/- 0.01 v 32.66 +/- 1.85 nmol/mg/4 h for oleate and 67.72 +/- 1.22 v 156.77 +/- 17.61 nmol/mg/4 h for glucose, control v gemfibrozil, P < .025). Given the rapid onset in action, these data suggest that gemfibrozil altered the active state of a limiting step in the triglyceride synthetic pathway. The effect of gemfibrozil on triglyceride synthesis was also determined on differentiated human preadipocytes and HepG2 cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Adipocytes↗
PubMed1994

Role of cyclic nucleotides in the regulation of endothelin-1 production by human endothelial cells.

The regulation of endothelin-1 (ET-1) production by endothelial cells is likely of crucial physiological importance in the maintenance of vascular homeostasis. The aim of the present study was to explore the possible role of cyclic nucleotides in the control of ET-1 production in human umbilical vein endothelial cells (HUVEC). ET-1 release was determined by measuring levels of immunoreactive ET-1 in HUVEC-conditioned media after 6-h incubations. In the presence of 10% fetal calf serum (FCS) there was a threefold increase in ET-1 release compared with serum-free conditions (1.96 +/- 0.17 vs. 0.56 +/- 0.06 pg/micrograms protein), respectively. Inhibition of protein kinase (PK) C using staurosporine (10 nM) reduced basal ET-1 release by approximately 50% and completely prevented the response to FCS. In contrast, the addition of other PK inhibitors had little effect on basal or serum-stimulated ET-1 release at the concentrations used. N6,2'-O-dibutyryladenosine 3',5'-cyclic monophosphate (DBcAMP) produced significant alterations in ET-1 release depending on the basal level of production. Under serum-free conditions of low basal ET-1 production, DBcAMP increased ET-1 release by 68 +/- 22% but only at the highest concentration studied (1 mM). The dose-response relationship for DBcAMP was potentiated by KT-5720 (0.1 microM), an inhibitor of PKA, with a significant shift to 10-fold lower concentrations, whereas it was blocked by KT-5823 (4 microM), which can inhibit PKG.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

The adipsin-ASP pathway and regulation of adipocyte function.

The adipsin-ASP pathway provides a mechanism by which the adipocyte is able to regulate its rate of de novo triglyceride synthesis and reesterification. The adipocyte can synthesize and secrete the three proteins necessary for the formation of the effector molecule, acylation stimulating protein (ASP). ASP increases membrane transport of glucose and the activity of diacylglycerol acyltransferase and by virtue of both of these effects markedly increases the rate of triglyceride synthesis. Awareness of the pathway will allow, we believe, a new understanding of the regulation of triglyceride removal from plasma. Accordingly, the concept of microenvironmental metabolic regulation of triglyceride hydrolysis at the endothelial surface and triglyceride synthesis within the adipocyte will be presented. In addition, the pathogenetic sequence by which dysfunction of this pathway can lead to dyslipoproteinaemia will be reviewed. Emphasis, however, will be placed on the role this pathway may play in the pathogenesis of obesity and the adaptation to negative caloric balance in the obese.

Adipocytes↗
PubMed1993

The role of amino acids in ApoB100 synthesis and catabolism in human HepG2 cells.

The object of this study was to examine the effects of extracellular amino acids on hepatic apoB100 metabolism using a transformed human liver cell line (HepG2) as a hepatocyte model. The data demonstrate that reduced extracellular amino acids resulted in marked increases in apoB100 concentrations in the medium (87 +/- 10% increase; p < 0.002). These data contrast with the effects of amino acids on secretion of total hepatic protein into the medium, which increased by 541 +/- 83% at 5.5 mg/ml amino acids compared to the low (0.5 mg/ml) amino acid medium (p < 0.01), pointing to a positive relationship between the concentration of amino acids in the medium and overall secretion of protein by the HepG2 cells. No significant effect of medium amino acid concentration on intracellular synthesis of cholesterol ester, cholesterol, triglyceride, or apoA1 secretion was evident. Amino acids also affected the activity of the low density lipoprotein pathway: at lower medium amino acid concentration, specific cell-associated radioactivity and cellular degradation were increased (117 +/- 24 and 233 +/- 13% at 0.5 versus 5.5 mg/ml amino acids, respectively). Pulse-chase analysis demonstrated that there was no difference in secretion efficiency of apoB (36 +/- 6 versus 46 +/- 8% in low versus high amino acid medium, respectively), but that the initial amount of [3H]apoB synthesized was greater in the low amino acid medium (10.5 +/- 3.8 versus 5.7 +/- 1.7 x 10(3) dpm [3H]apoB/mg of cell protein; p < 0.05). In contrast, the initial amount of [3H]albumin synthesized was much greater in the high amino acid medium (26.6 +/- 5.0 versus 54.6 +/- 19.0 x 10(3) dpm [3H]albumin/mg of cell protein in low versus high amino acid medium, respectively; p < 0.05). Slot blot analysis of apoB mRNA was 87 +/- 22% higher in lower amino acid medium as compared to the high amino acid medium (p < 0.01). These results demonstrate that amino acids have a profound negative regulatory effect on apoB synthesis and secretion and may shed light on the pathogenesis of some clinical dyslipidemias such as the increased plasma apoB levels in patients treated with continuous ambulatory peritoneal dialysis.

Albumins↗
PubMed1993

Coordinate regulation of triacylglycerol synthesis and glucose transport by acylation-stimulating protein.

Acylation-stimulating protein (ASP) is the most potent recognized stimulant of triacylglycerol synthesis in human adipocytes. However, its mechanism(s) of action have not yet been elucidated in detail. The present study examines the effects of ASP on membrane transport of glucose and fatty acids in cultured human skin fibroblasts. The data demonstrate that ASP stimulated carrier-mediated glucose transport in a time- and concentration-dependent manner, an effect that was greater than and independent of that observed with insulin. ASP increased the Vmax for glucose transport with no change in the transport Km, suggesting that ASP might result in increased sugar transporters in the plasma membrane. This finding was supported by quantitative Western blot analyses using a monoclonal antibody (G3) that demonstrated an increase in the plasma membrane content of the glucose transporter (Glut 1) with a concomitant decrease in the glucose transporter content of an intracellular membrane fraction. By contrast, ASP had no effect on specific membrane transport of fatty acids. Maximal effects of ASP on triacylglycerol synthesis were demonstrated at saturating levels of both glucose and oleate. Comparable stimulation by ASP in the absence of glucose (with or without pyruvate) was also demonstrated, although the absolute rates of triacylglycerol synthesis were substantially lower. Finally, it was shown that ASP increased the apparent Vmax for triglyceride synthesis without changing its Km. Since ASP will act in the absence of extracellular glucose, ASP must have additional actions, independent of its effect on specific membrane transport of glucose by which it accelerates intracellular triglyceride synthesis.(ABSTRACT TRUNCATED AT 250 WORDS)

Biological Transport↗
PubMed1993

Substrate delivery as a determinant of hepatic apoB secretion.

The evidence that apoB particles secreted by the liver can differ in number and composition has been reviewed. No evidence has yet emerged that changes in apoB100 itself affect the rate of its secretion from the liver. The metabolic role of apoB appears to be the prevention of lipid accumulation within the liver cell: when delivery of lipid to the liver increases, apoB secretion will increase pari passu. This reality in no way detracts from the critical role played by the LDL receptor in determining the number of LDL particles in plasma, nor does it diminish the potential importance of intracellular processes such as 7 alpha-hydroxylase activity to also mediate LDL receptor activity. However, it should be obvious that variation in catabolism by itself cannot explain all that has been observed in physiological and pathological studies. On the contrary, the whole process must be taken into account--the rate at which apoB particles are added to the circulation, the rate at which they are converted to LDL, and the rate at which they are irreversibly removed from plasma--if we are to understand and appreciate this most peculiar and most important of transport systems.

Animals↗
PubMed1993

The adipsin-acylation stimulating protein system and regulation of intracellular triglyceride synthesis.

We have previously characterized an activity from human plasma that markedly stimulates triglyceride synthesis in cultured human skin fibroblasts and human adipocytes. Based on its in vitro activity we named the active component acylation stimulating protein (ASP). The molecular identity of the active serum component has now been determined. NH2-terminal sequence analysis, ion spray ionization mass spectroscopy, and amino acid composition analysis all indicate that the active purified protein is a fragment of the third component of plasma complement, C3a-desArg. As well, reconstitution experiments with complement factors B, D, and complement C3, the components necessary to generate C3a, have confirmed the identity of ASP as C3a. ASP appears to be the final effector molecule generated by a novel regulatory system that modulates the rate of triglyceride synthesis in adipocytes.

Amino Acid Sequence↗
PubMed1993

Regional differences in triacylglycerol synthesis in adipose tissue and in cultured preadipocytes.

The initial suspicion that obesity increases coronary risk has been much sharpened with the demonstration that risk is more tightly linked to abdominal than to peripheral obesity, and tighter yet again when the mass of omental adipose tissue is taken into account. These data suggest that important metabolic differences might exist between adipocytes from different regions, and indeed, it has long been appreciated that triacylglycerol hydrolysis can be stimulated to a greater extent in omental than in subcutaneous adipocytes. The present study focuses on triacylglycerol synthesis in human subcutaneous and omental adipocytes, a process which, by contrast, has received relatively little attention. Experiments were done on adipose tissue removed at laparotomy and on cultured preadipocytes. With the former, triacylglycerol synthesis was measured in the presence and absence of oleate added to the medium using radiolabeled glucose and oleate as tracers. The results demonstrate that under all conditions examined triacylglycerol synthesis in subcutaneous adipose tissue exceeded that in deep omental adipose tissue. To study the cells in more detail, preadipocytes were cultured and triacylglycerol synthesis was examined again under basal conditions and with stimulation with insulin and acylation stimulating protein (ASP). Under basal conditions, particularly when oleate was added to the medium, clear differences were present such that triacylglycerol synthesis was substantially greater in subcutaneous preadipocytes than in omentally derived preadipocytes. These differences were more pronounced when the cells were stimulated with either insulin or acylation stimulating protein. Overall, triacylglycerol synthetic capacity in subcutaneous tissue exceeded that in omental tissue. As a consequence, omental tissue as compared to subcutaneous adipose tissue would have a limited capacity to prevent fatty acids from reaching the liver and stimulating hepatic lipoprotein synthesis.

Adipose Tissue↗
PubMed1992

Pathogenesis of carbohydrate-induced hypertriglyceridemia using HepG2 cells as a model system.

This study compares the effects of glucose and fatty acid on hepatic lipid synthesis and apolipoprotein (apo) B secretion. To do so, varying concentrations of either glucose or oleic acid were added to the medium in which HepG2 cells were being incubated. Intracellular triacylglycerol and cholesteryl ester synthesis and secretion were measured by addition of radioisotopic tracers and by determination of mass, whereas apo B concentration in the medium was measured by a specific enzyme-linked immunosorbent assay. The data indicate that increasing concentrations of glucose in the medium resulted in increased synthesis of triacylglycerol within the cell and increased secretion of triacylglycerol into the medium. Apo B secretion into the medium, however, did not change, and intracellular synthesis and secretion of cholesteryl ester did not change as well. By contrast, addition of oleic acid to the medium resulted in increased synthesis and secretion of both cholesteryl ester and triacylglycerol, and this was associated with increased secretion of apo B into the medium. Thus, a carbohydrate load resulted in secretion of normal numbers of triacylglycerol-enriched apo B particles by this hepatocyte cell line, whereas a fatty acid load led to the secretion of increased numbers of apo B particles, which were essentially normal in composition.

Apolipoproteins B↗
PubMed1991

Effect of acylation stimulating protein on the triacylglycerol synthetic pathway of human adipose tissue.

Acylation stimulating protein (ASP) is a 14 kDa plasma protein which causes in vitro triacylglycerol synthesis in human adipocytes and fibroblasts to increase substantially. ASP was found to stimulate human adipose tissue microsomal glycerophosphate acyltransferase and diacylglycerol acyltransferase activities by 23% and 90%, respectively. However, phosphatidate phosphohydrolase activity showed no increase in activity, nor did microsomal acyl-CoA synthetase activity. Moreover, ASP did not decrease the apparent Km of diacylglycerol acyltransferase (DGAT), but rather increased its apparent Vmax suggesting direct interaction of ASP with DGAT.

Acyltransferases↗
PubMed1991

Effect of moderate hypertriglyceridemia on the relation of plasma total and LDL apo B levels.

The risk of premature coronary artery disease is related to an important degree to the number of particles of low density lipoproteins (LDL) in plasma, an estimate given by measurement of LDL apo B. In clinical practise, though it is total, not LDL apo B, which is measured. The purpose of the present study therefore was to compare plasma total and LDL apo B in the presence and absence of moderate hypertriglyceridemia. The results demonstrate that within the range of plasma triglyceride levels examined, i.e., values of triglyceride up to 500 mg/dl, there is close correspondence between total and LDL apo B, with the latter more than 90% of the former. VLDL composition was also examined and two patterns found in hypertriglyceridemic patients: those with normal apo B had markedly lipid enriched VLDL while those with elevated apo B had VLDL which was normal in composition except for a moderate increase in triglyceride content. Thus total apo B within the circumstances studied reflects principally LDL apo B. Moreover measurement of apo B allows distinction between two different forms of hypertriglyceridemia, only one of which - that with an increased LDL particle number - has previous work shown to be associated with increased coronary risk. Total apo B, therefore, provides additional information not available from conventional plasma and lipoprotein lipids which allows more precise physiologic classification and may lead to more rational choice of pharmacologic therapy in normolipidemic and hypertriglyceridemic patients.

Apolipoproteins B↗
PubMed1990

Modulation of chylomicron remnant metabolism by an hepatic hydroxymethylglutaryl coenzyme A reductase inhibitor.

This study was designed to test the hypothesis that in patients with elevated plasma low-density lipoprotein (LDL) apolipoprotein-apoB, chylomicron remnant clearance can be modulated by therapy with a hepatic hydroxymethyl glutaryl coenzyme A reductase inhibitor. Accordingly, chylomicron triglyceride and remnant clearance were determined following a vitamin-A fat load in 12 such patients, before and after therapy with Lovastatin (Merck, Sharp & Dohme, Rahway, NJ). Such therapy had no significant overall effect on plasma triglyceride clearance, although there was a trend to lower levels of Sf greater than 400 triglycerides at the later time points. By contrast, retinol clearance in plasma and Sf greater than 400 lipoproteins was markedly increased (30% and 40%, respectively). The data indicate, therefore, that following therapy with Lovastatin in this group of patients, chylomicron plasma remnant clearance was significantly enhanced. The exact mechanisms responsible remain to be explicated.

Apolipoproteins B↗
PubMed1990

Endothelin release is inhibited by coculture of endothelial cells with cells of vascular media.

Endothelin is a potent vasoconstrictor peptide and a smooth muscle mitogen produced in large amounts by endothelial cells in culture. To determine whether other cellular elements of the vessel wall modify the release or clearance of endothelin, we studied the effect of coculture of endothelial cells with vascular smooth muscle cells or fibroblasts on endothelin release. Endothelial cells were grown to confluence on microcarrier beads and transferred to dishes containing confluent cultures of smooth muscle cells or fibroblasts or control media only. In parallel experiments, endothelial cells on microcarrier beads were incubated in media conditioned by 48-h exposure to smooth muscle cells or fibroblasts. Endothelin concentration was determined by radioimmunoassay (rabbit anti-endothelin-1 serum). Endothelial cells alone released large amounts of endothelin: 169 +/- 60 and 982 +/- 237 pg/10(6) endothelial cells at 4 and 24 h, respectively. Endothelin accumulation was markedly reduced in coculture with smooth muscle cells or fibroblasts by 81 +/- 10 and 49 +/- 5% (P less than 0.05), respectively, at 24 h. This difference could not be explained by smooth muscle cell binding or degradation of endothelin. Furthermore, smooth muscle cell- or fibroblast-conditioned media significantly reduced endothelin release, and twofold concentration of smooth muscle cell-conditioned media fully reproduced the inhibition of endothelin release found in coculture, confirming the presence of a transferable inhibitor. Therefore, we propose that endothelin secretion from endothelial cells may be regulated by an inhibitory factor produced by the vascular media. This mechanism might limit the production of endothelin in intact vessels and thereby protect against excessive vasoconstriction or proliferation of vascular target cells.

Animals↗
PubMed1989

The effect of ASP on the adipocyte of the morbidly obese.

The control of triglyceride synthesis within the adipocyte is not fully understood. Insulin is considered to be the most potent stimulant of triglyceride synthesis. In this paper, we report on the effect of a small (14000 Da), basic (pI 9.0) protein isolated from human serum. This protein has been called acylation stimulating protein (ASP). It is a potent stimulant of triglyceride synthesis in adipocytes from both normal weight and morbidly obese subjects. Its stimulatory effect on adipocytes is both rapid, occurring between 15-30 min after the start of incubation, and prolonged, lasting for up to 3 hr. Compared to insulin, it is sixfold more potent in its effect on triglyceride synthesis. As well as acting on isolated cells, ASP also has a fourfold stimulatory effect on triglyceride synthesis in human adipose microsomes at a concentration of 25 micrograms/ml. This study indicates that ASP is a potent stimulant of triglyceride synthesis and therefore may play a role in the pathogenesis of morbid obesity.

Adipose Tissue↗