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Biomedical subjects

E L Bierman

Publications and source records attributed to E L Bierman.

At least 37 records · Page 2Linked to original sources
PubMed1991

Role of the protein kinase C signaling pathway in high-density lipoprotein receptor-mediated efflux of intracellular cholesterol.

These studies provide evidence that binding of HDL3 to the HDL receptor stimulates translocation and efflux of intracellular cholesterol through mechanisms involving the activation of protein kinase C. This conclusion is supported by data demonstrating that HDL is able to increase cell diacylglycerol levels and activate protein kinase C. Sphingosine, a protein kinase C inhibitor, was able to inhibit HDL3-mediated cholesterol translocation and efflux, further suggesting a role for protein kinase C in HDL receptor-dependent cholesterol efflux. Inhibition of HDL-mediated diacylglycerol formation by pertussis toxin suggests the possible involvement of a G protein-activated phospholipase. Further studies are needed to understand how activation of protein kinase C promotes cholesterol translocation and to identify the target proteins for protein kinase C phosphorylation.

Biological Transport, Active↗
PubMed1990

Nonenzymatic glycosylation of HDL resulting in inhibition of high-affinity binding to cultured human fibroblasts.

Nonenzymatic glycosylation of plasma proteins may contribute to the excess risk of developing atherosclerosis in patients with diabetes mellitus. Because high-density lipoprotein (HDL) is believed to protect against atherosclerosis and is glycosylated at increased levels in diabetic individuals, the effects of nonenzymatic glycosylation of HDL3 on binding of HDL3 to cultured fibroblasts and to the candidate HDL-receptor protein were examined. HDL3 was glycosylated in vitro with glucose alone or in combination with sodium cyanoborohydride. With this catalyst, up to 40-50% of the lysine residues could be glycosylated, resulting in a progressive drop to nearly 60% in high-affinity binding to cultured fibroblasts at 4 degrees C. Binding to the 110,000-Mr candidate HDL-receptor protein was reduced by almost 75%. At levels of HDL glycosylation equivalent to the 3-5% observed in diabetes, high-affinity binding to fibroblasts at 4 degrees C was diminished by up to 15-20%. Binding kinetic studies paradoxically suggested that glycosylated HDL3 binds with higher affinity to a reduced number of binding sites. The findings in this study suggest that nonenzymatically glycosylated HDL may be functionally abnormal and might contribute to the development of atherosclerosis in patients with diabetes mellitus.

Animals↗
PubMed1990

The relationship between sex hormones and high-density lipoprotein cholesterol levels in healthy adult men.

The objective of this study was to clarify the complex and uncertain relationship between endogenous sex hormones and high-density lipoprotein (HDL) cholesterol levels in healthy men. Fifty-five healthy adult men were consecutively recruited from an ongoing cross-sectional study of cardiovascular disease risk factors from a lipid research clinic at the University of Washington, Seattle. Subjects receiving medication were excluded. Multiple linear regression analysis identified several factors that correlated highly significantly with HDL cholesterol levels, including alcohol intake; frequency of strenuous exercise; age; levels of total cholesterol, low-density lipoprotein cholesterol, and triglyceride; and carbohydrate intake. Nearly 80% of the heterogeneity in HDL cholesterol levels could be accounted for by these factors. Despite finding significant correlations with factors known to influence HDL cholesterol levels, no correlation with estradiol level, testosterone level, or the ratio of estradiol to testosterone levels was apparent. In conclusion, endogenous sex hormones do not appear to influence HDL cholesterol levels in healthy adult men. Alternatively, a large proportion of the heterogeneity in HDL levels in this group of men can be accounted for by environmental factors. The disparity between this conclusion and others may be partially due to differences in accounting for these confounding variables.

Adult↗
PubMed1989

Intracellular transport of cholesterol in type C Niemann-Pick fibroblasts.

The purpose of this study was to determine the capacity of Niemann-Pick type C (NPC) fibroblasts to transport cholesterol from the cell surface to intracellular membranes. This is relevant in light of the observations that NPC cells display a sluggish metabolism of LDL-derived cholesterol, a phenomenon which could be explained by a defective intracellular transport of cholesterol. Treatment of NPC cells for 4 h with 0.1 mg/ml of LDL failed to increase the incorporation of [14C]oleic acid into cholesterol [14C]oleate, an observation consistent with previous reports on this cell type (Pentchev et al. (1985) Proc. Natl. Acad. Sci. USA 82, 8247). Normal fibroblasts, however, displayed the classical upregulation (6-fold over control) of the endogenous esterification reaction in response to LDL exposure. Incubation of normal or NPC fibroblasts with sphingomyelinase (100 mU/ml; Staphylococcus aureus) led to a rapid and marked increase (9- and 10-fold for normal and NPC fibroblasts, respectively, after 4 h) in the esterification of plasma-membrane-derived [3H]cholesterol suggesting that sphingomyelin degradation forced a net transfer of cholesterol from the cell surface to the endoplasmic reticulum. The similar response in normal and mutant fibroblasts to the degradation of sphingomyelin suggests that plasma membrane cholesterol can be transported into the substrate pool of ACAT to about the same extent in these two cell types. Degradation of cell sphingomyelin in NPC fibroblasts also resulted in the movement of 20-25% of the cellular cholesterol from a cholesterol oxidase susceptible pool into oxidase-resistant pools, implying that a substantial amount of plasma membrane cholesterol was internalized after sphingomyelin degradation. This cholesterol internalization was not accompanied by an increased rate of membrane internalization, as measured by [3H]sucrose uptake. Although NPC cells showed a relative accumulation of unesterified cholesterol and a sluggish esterification of LDL-derived cholesterol when exposed to LDL, these cells responded like normal fibroblasts with regard to their capacity to transport cholesterol from the cell surface into intracellular sites in response to sphingomyelin degradation. It therefore appears that NPC cells, in contrast to the impaired intracellular movement of lipoprotein-derived cholesterol, do not display a general impairment of cholesterol transport between the cell surface and the intracellular regulatory pool of cholesterol.

Cell Line↗
PubMed1989

Downregulation of high-density lipoprotein receptor in human fibroblasts by insulin and IGF-I.

High-density lipoprotein (HDL3) particles bind to a cell surface receptor, thereby promoting the efflux of cholesterol from extrahepatic nonsteroidogenic cells. This receptor appears to be upregulated by increased cell cholesterol content and also may be responsive to the growth state of cells. Because insulin can be mitogenic, the effect of insulin on HDL-receptor function was tested. HDL-receptor activity of cholesterol-loaded fibroblasts was inhibited by insulin treatment. Insulin decreased HDL binding in a log-dose fashion (-25% at 67 nM insulin) in association with increases in [3H]thymidine incorporation into DNA. HDL-mediated cholesterol efflux from cholesterol-loaded cells was diminished by insulin treatment of cells in parallel with decreased HDL binding. Insulin induced reciprocal changes in HDL- and low-density lipoprotein (LDL)-receptor activity. In cells in which these receptors were upregulated by varying cell cholesterol content, insulin increased LDL binding (+88%) and decreased HDL binding (-24%). Insulin-like growth factor I (IGF-I, 100 ng/ml) also significantly decreased HDL binding and HDL-mediated cholesterol efflux to a comparable degree. Pooled human serum similarly induced a reduction in HDL binding to its receptor. These results are consistent with the hypothesis that growth factors in general, and insulin and IGF-I in particular, decrease HDL-receptor activity, possibly to promote retention of cholesterol needed for new membrane synthesis during cell proliferation. Such a mechanism could be partly responsible for accumulation of cholesteryl esters in arterial wall cells during atherogenesis in diabetes mellitus.

Carrier Proteins↗
PubMed1989

Elevated plasma glucose and lowered triglyceride levels from omega-3 fatty acid supplementation in type II diabetes.

We studied the effect of omega-3 fatty acids (omega 3FA) on glucose homeostasis and lipoprotein levels in eight type II (non-insulin-dependent)-diabetic subjects ingesting 8 g/day omega 3FA for 8 wk as marine-lipid concentrate capsules. After omega 3FA supplementation, fasting plasma glucose levels increased 22% (P = .005) and meal-stimulated glucose increased 35% (P = .036). The percentage of glucose elevation correlated with percentage ideal body weight (r = .73, P = .04). No significant changes were seen in fasting or meal-stimulated plasma insulin, glucose disposal, or insulin-to-glucagon ratios. Very-low-density lipoprotein cholesterol and triglyceride (TG) levels showed consistent reductions of 56% (P less than .001) and 42% (P less than .001), respectively, after omega 3FA supplementation. Total cholesterol levels decreased 7% (P less than .05) without alteration in low- or high-density lipoprotein cholesterol. Thus, omega 3FA supplementation at a dose of 8 g/day significantly improves plasma TG levels but increases fasting and meal-stimulated glucose concentrations in the type II diabetic patient not treated with insulin or sulfonylurea agents. Marine-lipid concentrate capsules supplying large amounts of omega 3FAs should be used cautiously in the type II diabetic patient.

Aged↗
PubMed1989

High density lipoprotein stimulates sterol translocation between intracellular and plasma membrane pools in human monocyte-derived macrophages.

Binding of high density lipoprotein (HDL) to its receptor on cultured fibroblasts and aortic endothelial cells was previously shown to facilitate sterol efflux by initiation of translocation of intracellular sterol to the plasma membrane. After cholesterol-loaded human monocyte-derived macrophages were incubated with either [3H]mevalonolactone or lipoprotein-associated [3H]cholesteryl ester to radiolabel intracellular pools of sterol, incubation with HDL3 led to stimulation of 3H-labeled sterol translocation from intracellular sites to the cell surface which preceeded maximum 3H-labeled sterol efflux. A similar pattern was demonstrated for macrophages that were preloaded with cholesterol derived from either low density lipoprotein (LDL), acetyl-LDL, or phospholipase C-modified LDL. However, in macrophages that were not loaded with cholesterol, HDL3 stimulated net movement of 3H-labeled sterol from the plasma membrane into intracellular compartments, the opposite direction from that seen for cholesterol-loaded cells. A similar influx pattern was found in nonloaded macrophages and fibroblasts that were labeled with trace amounts of exogenous [3H]cholesterol. Cholesterol translocation from intracellular pools to the cell surface of cholesterol-loaded macrophages appeared to be stimulated by receptor binding of HDL, since chemical modification of HDL with tetranitromethane (TNM), which abolishes its receptor binding, reduced its ability to stimulate 3H-labeled sterol translocation and efflux. In nonloaded cells, however, the ability of HDL3 to stimulate sterol efflux and movement of sterol from the plasma membrane into intracellular pools was unaffected by TNM modification. Thus, binding of HDL to its receptor on cholesterol-loaded macrophages appears to promote translocation of intracellular cholesterol to the plasma membrane followed by cholesterol efflux into the medium. However, in nonloaded macrophages, HDL stimulates sterol movement from the plasma membrane into intracellular pools by a receptor-independent process.

Body Fluids↗
PubMed1988

Up-regulation of high density lipoprotein receptor activity by gamma-interferon associated with inhibition of cell proliferation.

High density lipoprotein (HDL) binds to cell surface receptors and promotes selective removal of excess cholesterol from intracellular pools. The activity of this receptor is up-regulated when cells become loaded with cholesterol, but the relative degree of up-regulation depends on the growth state of the cells. The current study demonstrates that treatment of proliferating fibroblasts with gamma-interferon (IFN) increases the activity of the HDL receptor in association with a decrease in the rate of cell proliferation. Addition of IFN during the growth phase reduced the number of cells but had little effect on total cell protein, indicating that IFN inhibited cell proliferation but produced larger cells. IFN treatment increased the number of high affinity receptors for HDL on the surface of cholesterol-loaded fibroblasts, whether receptor binding was expressed per cell or per unit of cell surface area, cell protein, or cell cholesterol. IFN treatment also appeared to increase the amount of 110-kDa HDL binding protein in fibroblast membranes that has been postulated to represent the HDL receptor molecule. The IFN-induced increase in HDL receptor activity was associated with an enhanced ability of HDL3 to remove cholesterol from intracellular pools. These results are consistent with the hypothesis that inhibition of cell proliferation increases HDL receptor-mediated transport of excess cholesterol from cells, possibly to rid cells of cholesterol that accumulates in response to a reduced rate of membrane synthesis.

Carrier Proteins↗
PubMed1988

Modification of low density lipoprotein by lipoprotein lipase or hepatic lipase induces enhanced uptake and cholesterol accumulation in cells.

Incubation of low density lipoprotein(s) (LDL) with either lipoprotein lipase or hepatic lipase led to modification of the core lipid composition of LDL. Both lipases modified LDL by substantially reducing core triglyceride content without producing marked differences in size, charge, or lipid peroxide content in comparison to native LDL. The triglyceride-depleted forms of LDL that result from treatment with these two enzymes were degraded at approximately twice the rate of native LDL by human monocyte-derived macrophages (HMDM). Lipase-modified LDL degradation was inhibited by chloroquine, suggesting lysosomal involvement in LDL cellular processing. The increased degradation by macrophages of the LDL modified by these lipases was accompanied by enhanced cholesterol esterification rates, as well as by an increase in cellular free and esterified cholesterol content. In a patient with hepatic triglyceride lipase deficiency, degradation of the triglyceride-rich LDL by HMDM was approximately half that of normal LDL. Following in vitro incubation of LDL from this patient with either lipoprotein or hepatic lipase, lipoprotein degradation increased to normal. Several lines of evidence indicate that LDL modified by both lipases were taken up by the LDL receptor and not by the scavenger receptor. 1) The degradation of lipase-modified LDL in nonphagocytic cells (human skin fibroblast and arterial smooth muscle cells) as well as in phagocytic cells (HMDM, J-774, HL-60, and U-937 cell lines) could be dissociated from that of acetylated LDL and was always higher than that of native LDL. A similar pattern was found for cellular cholesterol esterification and cholesterol mass. 2) LDL receptor-negative fibroblasts did not degrade lipase-modified LDL. 3) A monoclonal antibody to the LDL receptor inhibited macrophage degradation of the lipase-modified LDL. 4) Excess amounts of unlabeled LDL competed substantially with 125I-labeled lipase-modified LDL for degradation by both macrophages and fibroblasts. Thus, lipase-modified LDL can cause significant cholesterol accumulation in macrophages even though it is taken up by LDL and not by the scavenger receptor. This effect could possibly be related to the reduced triglyceride content in the core of LDL, which may alter presentation of the LDL receptor-binding domain of apolipoprotein B on the particle surface, thereby leading to increased recognition and cellular uptake via the LDL receptor pathway.

Antibodies, Monoclonal↗
PubMed1988

Stimulation of receptor-dependent and receptor-independent pathways of low-density lipoprotein degradation in arterial smooth muscle cells by platelet-derived growth factor.

Platelet-derived growth factor (PDGF), a powerful mitogen released by platelets, promoted the degradation of low-density lipoprotein (LDL) by cultured primate arterial smooth muscle cells and human skin fibroblasts by stimulating both receptor-mediated and LDL-receptor-independent uptake of LDL. Stimulation of LDL-receptor-independent LDL uptake and degradation by PDGF was demonstrated in three ways. First, the small amount of LDL that was degraded by LDL-receptor-negative skin fibroblasts was stimulated by PDGF. Second, PDGF led to increased degradation of LDL that had been reductively methylated to prevent its binding to LDL receptors. Third, 125I-labeled LDL degradation was stimulated by PDGF in the presence of high concentrations of unlabeled LDL, i.e., conditions under which the contribution of the LDL receptor to cellular uptake and degradation is reduced. These observations suggest that mitogens, as typified by PDGF, can facilitate the cellular delivery of LDL cholesterol by both LDL-receptor-mediated and non-LDL-receptor-mediated mechanisms to provide exogenous cholesterol for use during cell replication.

Animals↗
PubMed1988

Depletion of plasma-membrane sphingomyelin rapidly alters the distribution of cholesterol between plasma membranes and intracellular cholesterol pools in cultured fibroblasts.

This study examines the relationship between cellular sphingomyelin content and the distribution of unesterified cholesterol between the plasma-membrane pool and the putative intracellular regulatory pool. The sphingomyelin content of cultured human skin fibroblasts was reduced by treatment of intact cells with extracellularly added neutral sphingomyelinase, and subsequent changes in the activities of cholesterol-metabolizing enzymes were determined. Exposure of fibroblasts to 0.1 unit of sphingomyelinase/ml for 60 min led to the depletion of more than 90% of the cellular sphingomyelin, as determined from total lipid extracts. In a time-course study, it was found that within 10 min of the addition of sphingomyelinase to cells, a dramatic increase in acyl-CoA:cholesterol acyltransferase activity could be observed, whether measured from the appearance of plasma membrane-derived [3H]cholesterol or exogenously added [14C]oleic acid, in cellular cholesteryl esters. In addition, the cholesteryl ester mass was significantly higher in sphingomyelin-depleted fibroblasts at 3 h after exposure to sphingomyelinase compared with that in untreated fibroblasts [7.1 +/- 0.4 nmol of cholesterol/mg equivalents of esterified cholesterol compared with 4.2 +/- 0.1 nmol of cholesterol/mg equivalents of cholesteryl ester in control cells (P less than 0.05)]. The sphingomyelin-depleted cells also showed a reduction in the rate of endogenous synthesis of cholesterol, as measured by incorporation of sodium [14C]acetate into [14C]cholesterol. These results are consistent with a rapid movement of cholesterol from sphingomyelin-depleted plasma membranes to the putative intracellular regulatory pool of cholesterol. This mass movement of cholesterol away from the plasma membranes presumably resulted from a decreased capacity of the plasma membranes to solubilize cholesterol, since sphingomyelin-depleted cells also had a decreased capacity to incorporate nanomolar amounts of [3H]cholesterol from the extracellular medium, as compared with control cells. These findings confirm previous assumptions that the membrane sphingomyelin content is an important determinant of the overall distribution of cholesterol within intact cells.

Acetates↗
PubMed1988

Fatty acid specificity of the lysosomal acid cholesterol esterase in intact human arterial smooth muscle cells.

The fatty-acid specificity of the lysosomal cholesterol esterase was examined in cultured human arterial smooth muscle cells. The lysosomal compartment of cultured cells was enriched with cholesteryl esters by incubation of cells with 0.2 mg/ml low-density lipoprotein and 50 microM chloroquine for 24 h. The hydrolysis of cholesteryl esters was subsequently induced by incubating cells in a medium containing 5% lipoprotein-deficient serum without chloroquine. Cellular cholesteryl ester mass was markedly reduced after 23 h in the lipoprotein-deficient serum. Fatty-acid analysis of cholesteryl esters in cells before and after the 23 h incubation with lipoprotein-deficient serum revealed that polyunsaturated cholesteryl esters (linoleate and arachidonate) were preferentially hydrolyzed compared to cholesteryl oleate or saturated cholesteryl esters. An increase in the ratio of cholesteryl oleate to cholesteryl linoleate was observed even when the cellular activity of acyl-CoA:cholesterol acyltransferase was inhibited with Sandoz Compound 58-035. We conclude that, in human arterial smooth muscle cells, the lysosomal acid cholesterol esterase preferentially hydrolyzes polyunsaturated cholesteryl esters.

Arteries↗
PubMed1988

The effects of omega 3 and omega 6 fatty acid-enriched diets on plasma lipoproteins and apoproteins in familial combined hyperlipidemia.

There is abundant evidence that dietary omega 3 fatty acids effect a favorable change in lipoprotein profiles of normolipidemic individuals. However, there is relatively little information available on the lipoprotein responses of hyperlipidemic individuals at risk for premature coronary artery disease. We studied a group of subjects with familial combined hyperlipidemia (FCHL), as well as a group of normal controls, on three rigidly controlled diets differing primarily in their fatty acid composition. The normal subjects demonstrated significant reductions in total cholesterol, low density lipoprotein (LDL) cholesterol, and total apolipoprotein B (apo B) levels on both an omega 3 (salmon) and omega 6 (safflower) fatty acid-enriched diet when these were compared with a basal diet high in saturated fat. The primary difference in response to the polyunsaturated diets was the potent triglyceride-lowering effect of the salmon diet. The FCHL subjects demonstrated a response to the safflower diet similar to that observed in normals and also manifested a marked triglyceride lowering with the salmon diet. However, total cholesterol and total apo B levels were not lowered by the salmon diet, and LDL cholesterol and apo B levels exhibited an upward trend. Thus, individuals with FCHL, a common disorder associated with premature coronary artery disease, do not appear to have a favorable lipoprotein response to diets enriched in omega 3 fatty acids.

Apolipoproteins↗
PubMed1987

Movement of plasma-membrane sterols to the endoplasmic reticulum in cultured cells.

The spontaneous turnover of plasma-membrane sterols, as measured by their transfer to the endoplasmic reticulum, was measured in quiescent cultured human skin fibroblasts and monkey arterial smooth-muscle cells. The plasma-membrane sterol pool was pulse-labelled with trace amounts of either [3H]desmosterol or [3H]cholesterol. We then measured the enzymic conversion of [3H]desmosterol into [3H]cholesterol and of [3H]cholesterol into [3H]cholesteryl esters in intact cells. Depending on the probe used, markedly different transfer or conversion rates were found in these cells. In quiescent human skin fibroblasts, incubated in a serum-free medium, about 1.1% of the plasma-membrane [3H]desmosterol was converted into [3H]cholesterol/h, whereas in monkey arterial smooth-muscle cells the corresponding rate was 0.4%. Under similar experimental conditions, these cells esterified less than 0.02% (fibroblasts) and 0.12% (smooth-muscle cells) of the plasma-membrane [3H]cholesterol/h. The movement of sterols from the plasma membrane to the endoplasmic reticulum, as measured by the conversion of [3H]desmosterol into [3H]cholesterol was not blocked by colchicine, but was markedly enhanced by 3% (w/v) dimethyl sulphoxide. In all, these results indicate that plasma-membrane sterols of cultured cells are continuously transferred to the interior of the cell at a rate substantially higher than previously appreciated. This turnover of plasma-membrane sterol molecules took place even when there was no mass transfer of sterols into the cells.

Animals↗
PubMed1987

Binding of high density lipoproteins to cell receptors promotes translocation of cholesterol from intracellular membranes to the cell surface.

Cultured cells have on their cell surface a specific high-affinity binding site (receptor) for high density lipoproteins (HDL) which appears to promote cholesterol efflux. In this study we characterized the cellular mechanisms involved in HDL receptor-mediated transport of cholesterol from cultured human fibroblasts and bovine aortic endothelial cells. HDL3, chemically modified by tetranitromethane (TNM-HDL3), is not recognized by this receptor and was used as a control for efflux not mediated by HDL receptor binding. HDL3 and TNM-HDL3 were found to be equally effective in causing efflux of plasma membrane cholesterol radiolabeled with [3H]cholesterol. However, HDL3 was much more effective than TNM-HDL3 in causing efflux of [3H]cholesterol associated with intracellular membranes. By measuring movement of endogenously synthesized [3H]cholesterol to the plasma membrane, and into the medium, we found that HDL3 induced a rapid movement of [3H]cholesterol from a preplasma membrane compartment to the plasma membrane that preceded [3H]cholesterol efflux. This effect was not observed with TNM-HDL3. Thus, receptor binding of HDL3 appears to facilitate removal of cellular cholesterol from specific intracellular pools by initiation of translocation of intracellular cholesterol to the plasma membrane.

Animals↗