Induced Triglyceride Accumulation Research Articles

VLDL-induced triglyceride accumulation in human macrophages is mediated by modulation of LPL lipolytic activity in the absence of change in LPL mass

Mixed dyslipidemia of phenotype IIB is characterized by elevated levels of very low density lipoprotein (VLDL)-1 and VLDL-2 subfractions and of low density lipoprotein (LDL), which are associated with premature formation of atherosclerotic plaques, characterized by the presence of lipid-rich macrophage foam cells. Lipoprotein lipase (LPL) is a key factor in mediating macrophage lipid accumulation and foam-cell formation from native VLDL particles. The action of macrophage-derived LPL in the induction of intracellular lipid accumulation from triglyceride-rich lipoprotein (TRL) subfractions (VLDL-1, VLDL-2) is, however, indeterminate, as is the potential role of VLDL-1 and VLDL-2 in modulating macrophage LPL expression. We evaluated the role of LPL in the interaction of type IIB VLDL-1 and VLDL-2 with human macrophages. Both VLDL-1 and VLDL-2 subfractions induced significant accumulation of triglyceride (9.8-fold, P<0.0001, and 4.8-fold, P<0.0001, respectively) and of free cholesterol content (1.4-fold, P<0.001, and 1.2-fold, P=0.02, respectively). Specific inhibition (90%) of the lipolytic activity of endogenous LPL by tetrahydrolipstatin (THL) in the presence of VLDL-1 or VLDL-2 resulted in marked reduction in cellular loading of both triglycerides (−89%, P=0.008, and −89%, P=0.015, respectively) and free cholesterol (−76%, P=0.02, and −55%, P=0.06 respectively). Furthermore, VLDL-1 and VLDL-2 induced marked increase in macrophage-derived LPL enzyme activity (+81%, P=0.002, and +45%, P=0.02), but did not modulate macrophage-derived LPL mRNA and protein expression; consequently, LPL specific activity was significantly increased from 1.6 mU/μg at baseline to 4.1 mU/μg ( P=0.01) and 3.1 mU/μg ( P=0.05), in the presence of VLDL-1 and VLDL-2, respectively. We conclude that type IIB VLDL-1 and VLDL-2 induce triglyceride accumulation in human monocyte-macrophages primarily via the lipolytic action of LPL, which may involve stabilization and activation of the macrophage-secreted enzyme, rather than via modulation of enzyme production.

β-very low density lipoprotein induces triglyceride accumulation through receptor mediated endocytotic pathway in 3T3-L1 adipocytes

To elucidate the mechanism of triglyceride (TG) accumulation in adipocytes induced by TG-rich lipoproteins, we examined the effect of β-very low density lipoprotein ( β-VLDL) on TG accumulation in 3T3-L1 adipocytes. β-VLDL did not induce TG accumulation in 3T3-L1 preadipocytes but in 3T3-L1 adipocytes. TG accumulation was significantly inhibited by cytochalasin B, an inhibitor of receptor mediated endocytosis. In contrast, cytochalasin B did not inhibit free fatty acid induced TG accumulation in adipocytes. The binding of [ 125I] β-VLDL to preadipocytes was inhibited completely by both β-VLDL and LDL. In sharp contrast, the binding of [ 125I] β-VLDL to adipocytes was inhibited completely by β-VLDL, but partially by LDL. The VLDL receptor mRNA was only expressed in adipocytes. These results suggest that β-VLDL induced TG accumulation in adipocytes may be mediated through the VLDL receptor pathway.

Evidence that cholesteryl ester and triglyceride accumulation in J774 macrophages induced by very low density lipoprotein subfractions occurs by different mechanisms

The present investigations have examined the mechanism(s) whereby Sf 60-400 very low density lipoproteins (VLDL) from Type IV hypertriglyceridemic subjects cause cholesteryl ester and triglyceride accumulation in J774 macrophages. Both apolipoprotein (apo) E-poor and apoE-rich Type IV VLDL subfractions, isolated by heparin-Sepharose chromatography, were capable of enhancing cellular cholesterol and triglyceride content. The apoE-rich fraction was significantly more effective at inducing cholesterol esterification (P < 0.05) and accumulation of esterified cholesterol (P < 0.05), whereas both subfractions caused similar increases in cellular triglyceride content. Thus, the amount of VLDL-associated apoE determined the extent to which Type IV VLDL loaded J774 cells with cholesterol but not triglyceride. Two VLDL subfractions, Sf 60-400 and Sf 20-60, isolated from Type III subjects homozygous for apoE2, caused little or no effect on cellular esterified cholesterol content, whereas both fractions induced the same degree of cellular triglyceride accumulation as Type IV VLDL. Type IV VLDL-induced cholesteryl ester accumulation was blocked by an anti-apoE monoclonal antibody, known to block the binding of apoE to the LDL receptor; however, the increase in cellular triglyceride was unaffected. Therefore, VLDL-induced triglyceride accumulation in this cell line can occur without apoE-mediated uptake of intact VLDL particles. The addition of heparin to J774 cells resulted in a 6-fold increase in lipoprotein lipase (LPL) activity in the media, and significantly enhanced the ability of Type IV VLDL to induce cellular triglyceride accumulation (P < 0.01), but significantly decreased cellular cholesteryl ester content (P < 0.025). Finally, Sf 60-400 VLDL from two subjects homozygous for apoC-II deficiency failed to increase cellular lipid content. However, the addition of exogenous apoC-II to C-II-deficient VLDL resulted in significant increases of both triglyceride and esterified cholesterol in J774 cells. In the presence of apoC-II, the anti-apoE monoclonal antibody blocked the cellular cholesteryl ester increase induced by C-II-deficient VLDL, but had no effect on the increase in cellular triglyceride. Collectively, these experiments demonstrate that extracellular lipolysis of Sf 60-400 VLDL by LPL is required for cholesteryl ester and triglyceride accumulation in J774 macrophages. After interaction with cellular LPL, VLDL triglycerides are hydrolyzed. The resulting free fatty acids are readily taken up by the macrophage, and re-esterified into triglyceride. Lipolysis proceeds until apoE epitopes are exposed, allowing the triglyceride-depleted remnant, containing all the cholesteryl ester, to be taken up via an apoE-mediated process.

Regulation of lipoprotein lipase secretion in murine macrophages during foam cell formation in vitro. Effect of triglyceride-rich lipoproteins.

Triglyceride rich-lipoproteins induce triglyceride accumulation in macrophages, leading to foam cell formation. The correlation between cell triglyceride accumulation and lipoprotein lipase (LPL) secretion in murine macrophages and the role that LPL plays in the accumulation process were examined. LPL secretion is defined as the extracellular LPL activity that accumulates during a 4-hour incubation of treated and untreated cells in a bovine serum albumin-containing RPMI-1640 medium. LPL secretion was suppressed (up to 70%) in a dose- and time-dependent manner when J774.1 cells were incubated with chylomicrons, very low density lipoproteins, and intermediate density lipoproteins but not with low or high density lipoproteins from normolipidemic and hypertriglyceridemic subjects. Oleic acid both suppressed LPL secretion and invoked triglyceride accumulation. Suppression of LPL secretion preceded gross triglyceride accumulation, was reversible, and was not the result of a reduction in LPL mRNA. P388D1 cells neither secreted LPL nor accumulated triglyceride. Inhibition of LPL secretion by tunicamycin in both peritoneal macrophages and J774.1 cells prevented a hypertriglyceridemic very low density lipoprotein-induced triglyceride accumulation, an effect that was counteracted by addition of exogenous LPL. The results suggest that 1) extracellular hydrolysis of lipoprotein triglyceride is a major factor in inducing foam cell formation and 2) LPL secretion may be regulated by cell energy needs, and when these needs are exceeded, LPL secretion is suppressed.

Sera from preeclamptic women increase the content of triglycerides and reduce the release of prostacyclin in cultured endothelial cells

The causes of the “endothelial dysfunction” accompanying preeclampsia are unknown. Women with preeclampsia have a marked hyperlipidemia which reflects altered lipid metabolism. We asked if the hyperlipidemic sera of preeclamptic women could cause altered endothelial cell properties. Cultured endothelial cells were incubated with sera from women with preeclampsia (PE) or normal pregnancies as controls. Fifty PE-sera were tested and in 45 cases the endothelial cells acquired a large number of sudanophilic granules which by electron microscopy had lipid appearance. In 31 incubations with 31 individual control sera cellular lipid granules were observed in 4 cases. The cellular triglyceride content was increased to 153±30 compared to 48±10 μg/mg cell protein in the control cells. Furthermore, the endothelial release of prostacyclin, measured as 6-keto PGF1α, was 8.8±0.6 ng/mg cell protein in cells incubated with PE-sera as compared to 40.3±6.4 ng/mg in the control cells. Conclusion: The hyperlipidemic sera from preeclamptic women induced triglyceride accumulation in cultured endothelial cells. This was accompanied by altered endothelial function as demonstrated by reduced prostacyclin release.

Very low density lipoprotein stimulation of triglyceride accumulation in rat preadipocyte cultures

Exposure of cultured rat epididymal preadipocytes to human very low density lipoproteins (VLDL) resulted in the rapid accumulation of large amounts of cellular triglyceride which was accompanied by the appearance of numerous large cellular lipid inclusions. Addition of heparin produced a two-fold stimulation of lipoprotein induced triglyceride accumulation. Supplementation of the growth medium with either low density lipoprotein, oleic acid or artificial triglyceride emulsion did not produce cellular triglyceride levels equivalent to that obtained with VLDL. Fibroblastic cells from rat skin and lung did not accumulate triglycerides when exposed to VLDL and heparin.