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Biomedical subjects

K Cianflone

Publications and source records attributed to K Cianflone.

At least 37 records · Page 2Linked to original sources
PubMed1999

Mechanisms involved in the regulation of free fatty acid release from isolated human fat cells by acylation-stimulating protein and insulin.

The effects of acylation-stimulating protein (ASP) and insulin on free fatty acid (FFA) release from isolated human fat cells and the signal transduction pathways to induce these effects were studied. ASP and insulin inhibited basal and norepinephrine-induced FFA release by stimulating fractional FFA re-esterification (both to the same extent) and by inhibiting FFA produced during lipolysis (ASP to a lesser extent than insulin). Protein kinase C inhibition influenced none of the effects of ASP or insulin. Phosphatidylinositol 3-kinase inhibition counteracted the effects of insulin but not of ASP. Phosphodiesterase 3 (PDE3) activity was stimulated by ASP and insulin, whereas PDE4 activity was slightly increased by ASP only. Selective PDE3 inhibition reversed the effects of both ASP and insulin on fractional FFA re-esterification and lipolysis. Selective PDE4 inhibition slightly counteracted the ASP but not the effect of insulin on fractional FFA re-esterification and did not prevent the action of ASP or insulin on lipolysis. Thus, ASP and insulin play a major role in regulating FFA release from fat cells as follows: insulin by stimulating fractional FFA re-esterification and inhibiting lipolysis and ASP mainly by stimulating fractional FFA re-esterification. For both ASP and insulin these effects on FFA release are mediated by PDE3, and for ASP PDE4 might also be involved. The signaling pathway preceding PDE is not known for ASP but involves phosphatidylinositol 3-kinase for insulin.

3',5'-Cyclic-AMP Phosphodiesterases↗
PubMed1999

Acylation stimulating protein (ASP), an adipocyte autocrine: new directions.

Acylation stimulating protein (ASP) is an adipocyte-derived protein which has potent anabolic effects on human adipose tissue for both glucose and free fatty acid (FFA) storage. Our hypothesis is that: (i) ASP is produced by adipocytes in specific response to stimuli that initiate efficient fat storage; (ii) ASP interacts with a specific adipocyte receptor triggering an intracellular signalling pathway which activates triglyceride synthesis and fat storage; and (iii) that absence (ASP knockout mouse) or excess (in normal or obese mice) of ASP will result in physiological changes of plasma fat clearance and adipose tissue metabolism. The present review focuses on advances in ASP within the last 2 years with particular emphasis on these three aspects of ASP.

Adipocytes↗
PubMed1999

Lipids and vascular disease: what we do and do not know.

The demonstration that lowering the level of low-density lipoproteins (LDL) in plasma substantially reduces the incidence of death and symptomatic coronary disease may be the most important advance ever in cardiovascular therapeutics. Indeed it has been perceived as so dramatic that it could give rise to the view that the only problem now is application. That would be unfortunate because critical questions remain unanswered. Amongst these are whether all patients with coronary disease will benefit from LDL lowering and what should be the target of such therapy. Moreover, it remains uncertain whether other approaches such as lowering triglycerides should also be strongly supported. More fundamentally, inadequacies in what has become the traditional approach to characterize risk are becoming increasingly evident. This paper will outline issues that remain unresolved and suggest clinical approaches to deal with them.

Coronary Disease↗
PubMed1999

Regulation of Plasma fatty acid metabolism.

Although adipose tissue serves a crucial function in energy storage, excess adipose tissue--that is, obesity--is often associated with diabetes and cardiovascular disease. A common thread in the weave of complications is increased plasma concentrations of fatty acids. In the present review, we have focused on two specific points that relate to obesity: (i) What are the metabolic consequences of increased free fatty acid concentrations? and (ii) What are the physiological factors that are involved in the regulation of fatty acid uptake or release from adipose tissue? We have tried to emphasize new factors that act as hormones on adipose tissue and in so doing regulate the net concentration of circulating free fatty acids.

Body Constitution↗
PubMed1999

Plasma acylation stimulating protein, adipsin and lipids in non-obese and obese populations.

BACKGROUND: Acylation stimulating protein (ASP) is a potent stimulator of TG synthesis in human adipocytes. DESIGN: In the present study, we have analysed plasma ASP and adipsin levels and their relationships to plasma lipids in non-obese and obese groups. RESULTS: The results show that the frequency distribution of ASP is skewed but that of adipsin is normal in both groups. In the non-obese population, the mean levels of plasma ASP and adipsin were 20.2 nmol L-1 (median) and 66.6 +/- 19 nmol L-1 (mean) respectively. No difference was observed between men and women for each of the parameters. In the obese population, the median plasma ASP was increased by 246% (69.9 nmol L-1) and adipsin by 31% (87.0 +/- 22.7 nmol L-1) above that of the control group. Although the levels for men and women were not statistically different for adipsin, the median ASP plasma concentration was 1.9-fold higher in obese women than in obese men (71.8 nmol L-1 vs. 37.6 nmol L-1, P < 0.05). Best subset regression analysis provided a model with variables that best predict plasma ASP [r2 = 0.160, P < 0.008 for body mass index (BMI), P < 0.05 for triacylglycerol (TG), P < 0.03 for free fatty acid (FFA)] and plasma adipsin (r2 = 0.057, P < 0.017 for BMI) in a non-obese population. In obese subjects, the model was different for plasma ASP (P = NS for any of the variables) and plasma adipsin (r2 = 0.356, P < 0.008 for FFA, P < 0.0002 for BMI, P < 0.02 for age). There was no correlation between ASP and adipsin in either the non-obese or the obese group. CONCLUSION: The present data suggest involvement of the ASP/adipsin pathway in the pathogenesis of obesity.

Adult↗
PubMed1999

Regional specificity of ASP binding in human adipose tissue.

Obesity, in particular omental (OM) adiposity, is associated with diabetes and cardiovascular disease. Thus site-specific regulation of fat storage is important to understand. Acylation-stimulating protein (ASP) is a potent stimulator of glucose transport and triglyceride synthesis in adipocytes. In the present study, we characterized receptor binding of 125I-labeled ASP to human adipocyte plasma membranes from paired OM and subcutaneous (SC) sites in normal (N) and obese (O) male (M) and female (F) subjects (n = 24). Overall, specific binding of 125I-ASP was in the order of SC > OM and O > N (in SC tissue, particularly in F). Receptor affinity of 125I-ASP was higher [lower dissociation constant (Kd)] in SC than in OM (63.6 +/- 16.2 vs. 160.7 +/- 38.6 nM, P < 0.02), especially in F (37.0 +/- 11.1 F-N and 26.3 +/- 6.7 nM F-O) and lower (higher Kd) in male OM (291.8 +/- 116.8 M-N and 149.4 +/- 56.4 M-O). The greater binding and higher affinity of 125I-ASP binding to SC suggests that ASP may be an important factor in maintaining regional adipose tissue mass. Conversely, lower binding and receptor affinity in male OM adipose tissue may contribute to the fatty acid imbalance and metabolic complications associated with this syndrome, by reducing the efficiency of adipose fatty acid trapping by the ASP pathway.

Adipose Tissue↗
PubMed1999

Enhanced triglyceride clearance with intraperitoneal human acylation stimulating protein in C57BL/6 mice.

Acylation stimulating protein (ASP), a novel adipocyte-derived autocrine protein, stimulates triglyceride synthesis and glucose transport in vitro in human and murine adipocytes. In vitro, chylomicrons increase ASP and precursor complement C3 production in adipocytes. Furthermore, in vivo, ASP production from human adipose tissue correlates positively with triglyceride clearance postprandially. The aim of the present study was to determine if intraperitoneally injected ASP accelerated triglyceride clearance in vivo after a fat load in C57Bl/6 mice. ASP increased the triglyceride clearance with a reduction of the triglyceride area under the curve over 6 h (AUC(0-6)) from 102.6 +/- 30.0 to 61.0 +/- 14.5 mg. dl(-1). h(-1) (P < 0.05), especially in the latter postprandial period (AUC(3-6); 56.2 +/- 18.0 vs. 24.9 +/- 8.9 mg. dl(-1). h(-1), P < 0.025). ASP also reduced plasma glucose both in the mice with accelerated plasma triglyceride clearance and in those with relatively delayed triglyceride clearance (P < 0.025). Therefore, ASP alters postprandial triglyceride and glucose metabolism.

Animals↗
PubMed1999

Role of cholesterol ester mass in regulation of secretion of ApoB100 lipoprotein particles by hamster hepatocytes and effects of statins on that relationship.

Our understanding of the factors that regulate the secretion of apoB100 lipoproteins remains incomplete with considerable debate as to the role, if any, for cholesterol ester in this process. This study examines this issue in primary cultures of hamster hepatocytes, a species in which both cholesterol and apoB100 metabolism are very similar to man. Addition of oleate to medium increased the mass of triglyceride and cholesterol ester within the hepatocyte and also increased the secretion of triglycerides, cholesterol ester, and apoB100 into the medium. Next, the responses of hamster hepatocytes to addition of either an HMG-CoA reductase inhibitor (lovastatin) or an acyl-CoA cholesterol acyltransferase inhibitor (58-035) to the medium, with or without added oleate, were determined. Effects of either agent were only evident in the oleate-supplemented medium in which cholesterol ester mass had been increased above basal. If oleate was not added to the medium, neither agent reduced apoB100 secretion; equally important, over the 24-hour incubation, neither agent, at the concentration used, produced any detectable change in intracellular cholesterol ester mass. However, in contrast to the estimates of mass, which were unchanged, under the same conditions radioisotopic estimates of cholesterol ester synthesis were markedly reduced. Any conclusion as to the relation of cholesterol ester mass to apoB100 secretion would therefore depend on which of the 2 methods was used. Overall, the data indicate a close correlation between the mass of cholesterol ester within the hepatocyte and apoB100 secretion from it and they go far to explain previous apparently contradictory data as to this relation. More importantly, though, taken with other available data, they indicate that the primary response of the liver to increased delivery of lipid is increased secretion rather than decreased uptake. These results point, therefore, to a hierarchy of hepatic responses to increased flux of fatty acids and increased synthesis of cholesterol that in turn suggests a more dynamic model of cholesterol homeostasis in the liver than has been appreciated in the past.

Animals↗
PubMed1999

Differential binding of triglyceride-rich lipoproteins to lipoprotein lipase.

In comparison to very low density lipoprotein (VLDL), chylomicrons are cleared quickly from plasma. However, small changes in fasting plasma VLDL concentration substantially delay postprandial chylomicron triglyceride clearance. We hypothesized that differential binding to lipoprotein lipase (LPL), the first step in the lipolytic pathway, might explain these otherwise paradoxical relationships. Competition binding assays of different lipoproteins were performed in a solid phase assay with purified bovine LPL at 4 degrees C. The results showed that chylomicrons, VLDL, and low density lipoprotein (LDL) were able to inhibit specific binding of (125)I-labeled VLDL to the same extent (85.1% +/- 13.1, 100% +/- 6.8, 90.7% +/- 23.2% inhibition, P = NS), but with markedly different efficiencies. The rank order of inhibition (K(i)) was chylomicrons (0.27 +/- 0.02 nm apoB) > VLDL (12.6 +/- 3.11 nm apoB) > LDL (34.8 +/- 11.1 nm apoB). By contrast, neither triglyceride (TG) liposomes, high density lipoprotein (HDL), nor LDL from patients with familial hypercholesterolemia were efficient at displacing the specific binding of (125)I-labeled VLDL to LPL (30%, 39%, and no displacement, respectively). Importantly, smaller hydrolyzed chylomicrons had less affinity than the larger chylomicrons (K(i) = 2.34 +/- 0.85 nm vs. 0.27 +/- 0.02 nm apoB respectively, P < 0.01). This was also true for hydrolyzed VLDL, although to a lesser extent. Chylomicrons from patients with LPL deficiency and VLDL from hypertriglyceridemic subjects were also studied. Taken together, our results indicate an inverse linear relationship between chylomicron size and K(i) whereas none was present for VLDL. We hypothesize that the differences in binding affinity demonstrated in vitro when considered with the differences in particle number observed in vivo may largely explain the paradoxes we set out to study.

Animals↗
PubMed1999

Mice lacking acylation stimulating protein (ASP) have delayed postprandial triglyceride clearance.

Acylation stimulating protein (ASP) is a 76 amino acid fragment of the third component of complement (C3) which is generated by the interaction of adipsin and factor B with C3. In vitro studies have shown that ASP can markedly increase triglyceride synthesis in adipocytes. To test the ASP pathway in vivo, C3-deficient mice, and therefore ASP-deficient mice, were generated and oral fat loads were conducted in wild-type (C3+/+) and mutant (C3-/-) animals. The principal results were: 1) postprandial triglyceride clearance was significantly delayed in mutant compared to wild-type mice; 2) this difference was more pronounced in males compared to females; 3) in both males and females, the differences were more pronounced in the second half of the postprandial period; 4) fasting and postprandial free fatty acid (FFA) were higher in C3(-/-) than in C3(+/+) males; and 5) intraperitoneal administration of ASP accelerated triglyceride clearance in C3(-/-) males. The data are consistent therefore, with the hypothesis that the ASP pathway is an important physiologic determinant of normal postprandial triglyceride clearance.

Animals↗
PubMed1998

Chylomicron-specific enhancement of acylation stimulating protein and precursor protein C3 production in differentiated human adipocytes.

Acylation stimulating protein (ASP) is a potent stimulator of adipocyte triacylglycerol storage. In vivo studies have shown that ASP production by adipocytes increases locally after a fat meal. Initial in vitro studies demonstrated increased production of ASP in the presence of chylomicrons (CHYLO). The present aim was to define the CHYLO component responsible. None of the apoproteins tested (AI, AII, AIV, CI, CII, CIII, and E) were capable of stimulating C3 (the precursor protein) or ASP production. Rather, the active component is a nonlipid, loosely associated, trypsin-sensitive molecule. High pressure liquid chromatography fractionation of the CHYLO infranate proteins identified the critical protein as transthyretin (TTR), which binds retinol-binding protein and complexes thyroxine and retinol. Addition of TTR alone, with lipid emulsion, or with respun CHYLO to human differentiated adipocytes had little effect on C3 and ASP production. By contrast, when transthyretin was added to CHYLO, C3 and ASP production were substantially enhanced up to 75- and 7. 5-fold respectively, compared with the effect of native CHYLO alone. Finally, a polyclonal antibody against TTR could inhibit stimulation of C3 and ASP production by CHYLO (by 98 and 100%, respectively) and by CHYLO infranate proteins (by 99 and 94%, respectively). We hypothesize that TTR mediates the transfer of the active components from CHYLO to adipocytes, which then stimulates increased C3 and ASP production. Thus the CHYLO provides the physiologic trigger of the ASP pathway.

Adipocytes↗
PubMed1998

Divergent responses of the liver to increased delivery of glucose or fatty acids: implications for the pathogenesis of type IV hyperlipoproteinemia.

This study examines the patterns of response of primary cultures of hamster hepatocytes to increased delivery of glucose or oleate. Increased glucose in the medium produced: (1) increased triglyceride in the cells and the medium; (2) no change in cholesterol ester in the cells or the medium; (3) no change in apo B100 secreted into the medium; (4) more apo B100 particles within the VLDL range with an increase in the VLDL triglyceride to apo B100 ratio. By contrast, increased oleate in the medium resulted in: (1) increased triglyceride in the cells and the medium; (2) increased cholesterol ester in the cells and the medium; and (3) increased apo B100 secreted into the medium. Important differences in the intracellular metabolism of triglyceride and cholesterol ester were also documented. Under all circumstances, there was substantially more radiolabelled triglyceride (overall eight times more) in the cell than in the medium, indicating that up to 90% of the newly synthesized triglyceride enters the cellular pool rather than being secreted with apo B100. By contrast, almost half of the newly synthesized cholesterol ester molecules were secreted with apo B100, pointing to an equal probability of entering the cell storage pool as opposed to being secreted. The data establish therefore two patterns of response of the liver to increased triglyceride synthesis depending on whether the substrate drive is glucose or oleate.

Animals↗
PubMed1998

Plasma acylation stimulating protein (ASP) as a predictor of impaired cellular biological response to ASP in patients with hyperapoB.

BACKGROUND: The objective of this study was to examine specific membrane binding of [125I]-acylation stimulating protein (ASP) in cultured human skin fibroblasts obtained from normal subjects and patients with hyperapoB. ASP is a small basic protein isolated from human plasma that stimulates triglyceride synthesis (TGS) and glucose transport (GT) in human skin fibroblasts and adipocytes. DESIGN: In the present study, three groups were studied: normal (NASP-NB) subjects, hyperapoB subjects with normal plasma ASP (NASP-HB) and hyperapoB subjects with high plasma ASP (HASP-HB). RESULTS: ASP-induced TGS in fibroblasts from HASP-HB subjects was significantly less than in the two control groups with normal plasma ASP (NASP-NB and NASP-HB). Similarly, ASP stimulation of GT was less in HASP-HB fibroblasts than in the NASP-HB fibroblasts or the NASP-NB subjects. Insulin-induced TGS was similar in all three groups as was insulin-stimulated GT. As well, protein kinase C-mediated stimulation was equivalent among the three groups both for GT and for TGS. There was no significant difference in the binding affinity (Kd) of [125I]-ASP to intact cells in any group. By contrast, binding of [125I]-ASP revealed a significantly lower Bmax of the HASP-HB cell lines than the NASP-NB cells and the NASP-HB cells. CONCLUSION: A decrease in the ASP cell-surface receptor concentration is responsible for decreased ASP stimulation of TGS, and GT and may contribute to the inefficient postprandial triglyceride (TG) clearance in HASP-HB subjects.

Adult↗
PubMed1998

Coordinated release of acylation stimulating protein (ASP) and triacylglycerol clearance by human adipose tissue in vivo in the postprandial period.

The objective of this study was to determine whether Acylation Stimulating Protein (ASP) is generated in vivo by human adipose tissue during the postprandial period. After a fat meal, samples from 12 subjects were obtained (up to 6 h) from an arterialized hand vein and an anterior abdominal wall vein that drains adipose tissue. Veno-arterial (V-A) gradients across the subcutaneous adipose tissue bed were calculated. The data demonstrate that ASP is produced in vivo (positive V-A gradient) With maximal production at 3-5 h postprandially. The plasma triacylglycerol (TAG) clearance was evidenced by a negative V-A gradient. It increased substantially after 3 h and remained prominent until the final time point. There was, therefore, a close temporal coordination between ASP generation and TAG clearance. In contrast, plasma insulin and non-esterified fatty acid (NEFA) had an early (1-2 h) postprandial change. Fatty acid incorporation into adipose tissue (FIAT) was calculated from V-A glycerol and non-esterified fatty acid (NEFA) differences postprandially. FIAT was negative during the first hour, implying net fat mobilization. FIAT then became increasingly positive, implying net fat deposition, and overall followed the same time course as ASP and TAG clearance. There was a direct positive correlation between total ASP production and total FIAT (r = 0.566, P < 0.05). These data demonstrate that ASP is generated in vivo by human adipocytes and that this process is accentuated postprandially, supporting the concept that ASP plays an important role in clearance of TAG from plasma and fatty acid storage in adipose tissue.

Adipose Tissue↗
PubMed1997

Characterization and modulation of LP(a) in human hepatoma HEPG2 cells.

HepG2 cells have been widely used to study factors which affect the secretion of apoB100 lipoprotein particles. The objectives of this study were to determine if Lp(a) particles were present in conditioned medium from HepG2 cells and if so, was this accumulation affected by factors which alter apoB100 lipoprotein metabolism. The data demonstrate that Lp(a) accumulated in the medium in a time dependent manner over a 48 h incubation period. Ultracentrifugation fractionation and Western blot analysis demonstrated that lipoprotein particles containing apo(a) in complex with apoB100 were present at a density consistent with human plasma Lp(a). Incubation of the HepG2 cells with LDL or VLDL caused increases in Lp(a) accumulation in the medium (+33% +/- 14%, P NS and 56% +/- 21%, P < 0.05, respectively). In contrast, apo(a) mRNA decreased (-17% +/- 3%, P < 0.01 for both LDL and VLDL incubation). Increasing concentrations of amino acids in the medium resulted in progressively less Lp(a) and apoB100 in the medium, the effect being greater on apoB100. ApoB100 mRNA levels decreased with incubation of HepG2 cells with amino acids (-22% +/- 10%, P < 0.06) whereas apo(a) mRNA levels increased significantly (+47% +/- 14%, P < 0.005). Taken together, our data show that HepG2 cells express mRNA for apo(a), and accumulate Lp(a) in the medium. The close correlation of medium Lp(a) levels with medium apoB100 levels, and not with apo(a) mRNA levels, suggests that medium Lp(a) accumulation may be a function of lipoprotein synthesis and secretion and is consistent with extracellular assembly of Lp(a) lipoprotein particles.

Amino Acids↗
PubMed1997

Acylation-stimulating protein (ASP) regulates glucose transport in the rat L6 muscle cell line.

Acylation-stimulating protein (ASP), a human plasma protein, is a potent stimulator of triglyceride synthesis and glucose transport in both human adipocytes and fibroblasts. The purpose of the present in vitro study was to examine the effect of ASP on glucose transport in muscle cells. ASP stimulated 2-deoxy-glucose transport (2-DG) in differentiated rat L6 myotubes in a time (30 min to 24 h) and concentration dependent manner (97% increase). The magnitude of the ASP effect on glucose transport was comparable to the time- and concentration-dependent effects seen with insulin (125% increase), but was additive to insulin, pointing to involvement of differential signalling pathways. ASP stimulation was dependent on cell differentiation in that glucose transport increased by only 12% in myoblasts, comparable to the effect of insulin in myoblasts (15% increase) demonstrating selective responsiveness of the differentiated myotubes to ASP and insulin. The mechanism for the ASP induced increase in glucose transport was also examined. ASP increased the Vmax for 2-DG transport by 183% (4.02 vs. 1.42 nmol/mg cell protein/30 s; ASP vs. Control, respectively). This could be explained by an increased translocation of glucose transporters (GLUT 1, GLUT 4 and GLUT 3) to the plasma membrane surface as demonstrated by Western analysis (+43% P < 0.05, +30% P < 0.05, and +49% P < 0.05, respectively). The effects of ASP were equal to those of insulin (+47%, +26% and +53% for GLUT 1, GLUT 4 and GLUT 3, respectively) and in all cases were paralleled by comparable glucose transport increases under the same incubation conditions. After long-term stimulation (24 h), Western analysis indicated that ASP had a permissive effect on insulin stimulated increases in total GLUT3 and GLUT4 cellular transporter content. These results suggest that muscle is also responsive to ASP and that ASP may play a role in glucose metabolism in both muscle and adipose tissue.

Animals↗