Lipoprotein-X stimulates monocyte chemoattractant protein-1 expression in mesangial cells via nuclear factor-kappa B.

BACKGROUND: Lipoprotein-X (Lp-X) is an abnormal lipoprotein found in the plasma of patients with familial lecithin:cholesterol acyltransferase (LCAT) deficiency. The majority of patients with this disorder develop progressive glomerulosclerosis. One key event in the pathogenesis of glomerulosclerosis is the infiltration of monocytes into affected glomeruli. Mesangial cells can synthesize and secrete monocyte chemoattractant protein-1 (MCP-1), an important chemoattractant for monocytes. The objective of the present study was to examine the effect of Lp-X on MCP-1 expression in mesangial cells leading to an enhanced monocyte chemotaxis and to elucidate the mechanisms involved in this process. METHODS: Lp-X was isolated from the plasma of a patient with familial LCAT deficiency. After rat mesangial cells were incubated with Lp-X for four or six hours, the expression of MCP-1 mRNA was determined by nuclease protection assay, and MCP-1 protein was measured by Western immunoblotting analysis. Monocyte chemotaxis was determined by using a Micro Chemotaxis Chamber. RESULTS: Lp-X (50 to 100 nmol/mL) stimulated mesangial cell MCP-1 mRNA expression (137 to 220%) and MCP-1 protein levels (233 to 375%). Conditioned media collected from Lp-X-treated mesangial cells stimulated human acute monocytic leukemia (THP-1) monocyte chemotaxis (165 to 200%). The increase in MCP-1 expression in mesangial cells was associated with an elevation of intracellular diacylglycerol levels, and activation of protein kinase C (PKC) as well as nuclear factor-kappa B (NF-kappa B). CONCLUSION: These results suggest that Lp-X participates in the pathogenesis of glomerulosclerosis and subsequent renal failure in familial LCAT deficient patients by stimulating monocyte infiltration via a mechanism involving mesangial MCP-1 expression.

Magnesium tanshinoate B (MTB) inhibits low density lipoprotein oxidation.

Danshen, a Chinese herbal medicine has been widely used for the treatment of cardiovascular diseases. Magnesium tanshinoate B (MTB) is an active compound purified from Danshen. The objective of this study was to investigate the effect of MTB on the susceptibility of low density lipoproteins (LDL) to oxidative modification as well as on the accumulation of lipids in THP-1 derived macrophages. Aliquots of LDL were incubated with copper sulfate in the absence or presence of MTB. The degrees of oxidative modification of LDL were assessed by examining the relative gel electrophoretic mobility, by measuring the amount of thiobarbituric acid reactive substances (TBARS), and by continuous monitoring of the formation of conjugated dienes upon the increase in absorbency at 234 nm. MTB at concentrations of 1-10 microM significantly inhibited oxidative modification of LDL. Such inhibitory effect resulted in a decrease in the uptake of LDL by THP-1 derived macrophages. Taken together, these results clearly demonstrate that MTB inhibits oxidative modification of LDL and hence prevents the uptake of LDL by cultured macrophages. Such effect may be therapeutically relevant in protecting cells from lipid peroxidation in vascular disorders.

Homocysteine stimulates the expression of monocyte chemoattractant protein-1 in endothelial cells leading to enhanced monocyte chemotaxis.

Hyperhomocysteinemia has been identified as an independent risk factor for atherosclerosis. The infiltration of monocytes into the arterial wall is one of the key events during atherogenesis. Monocyte chemoattractant protein-1 (MCP-1) is a potent chemokine that stimulates the migration of monocytes into the intima of the arterial wall. The mechanism by which increased monocyte infiltration occurs in atherosclerotic lesions in patients with hyperhomocysteinemia has not been delineated. The objective of the present study was to investigate the effect of homocysteine on MCP-1 production in endothelial cells. Cells were incubated with homocysteine. The secretion of MCP-1 protein was significantly increased (195% as compared to the control) in cells treated with pathological concentrations of homocysteine. Such effect was accompanied by an increased expression of MCP-1 mRNA (176% as compared to the control) in endothelial cells which resulted in enhanced monocyte chemotaxis. The p38 MAP kinase as well as other members of the p38 MAP kinase pathway, including MKK3, MKK6, ATF-2 and Elk-1, were activated in homocysteine-treated cells. Homocysteine-induced MCP-1 expression and subsequent monocyte chemotaxis were blocked by a p38 MAP kinase inhibitor (SB203580) suggesting that the p38 MAP kinase pathway might be involved in homocysteine-induced MCP-1 expression in endothelial cells. In contrast, staurosporine, a protein kinase C inhibitor, had no effect on homocysteine-induced MCP-1 expression. In conclusion, our results indicate that homocysteine stimulates MCP-1 expression in endothelial cells leading to enhanced monocyte chemotaxis.

Very low-density lipoprotein stimulates the expression of monocyte chemoattractant protein-1 in mesangial cells.

BACKGROUND: Elevated plasma levels of very low-density lipoprotein (VLDL) are associated with an increased risk for focal glomerulosclerosis, which is analogous to atherosclerosis. One feature of focal glomerulosclerosis is the presence of foam cells derived from the infiltration of circulating monocytes. Mesangial cells are able to express monocyte chemoattractant protein-1 (MCP-1). In this study, the ability of VLDL to stimulate MCP-1 expression in mesangial cells and consequent monocyte adhesion was investigated. METHODS: For adhesion studies, mesangial cells isolated from Sprague-Dawley rats were treated with VLDL for six hours, followed by a one-hour incubation with Tamm-Horsfall protein-1 (THP-1) cells. Mesangial MCP-1 mRNA levels were determined by reverse transcription-polymerase chain reaction. MCP-1 protein was determined by solid-phase immunoassay. RESULTS: VLDL (100 to 300 microg/mL) significantly enhanced the expression and secretion of MCP-1 (54 to 285 ng/well) in mesangial cells. Such an effect was accompanied by the increased adhesion of monocytes to mesangial cells and later the formation of foam cells from monocytes after ingesting excessive amounts of VLDL lipids. VLDL-induced MCP-1 expression and monocyte adhesion were blocked by a protein kinase C inhibitor (staurosporine), as well as a calcium channel blocker (diltiazem). CONCLUSIONS: Our results demonstrate that elevated levels of VLDL, through the action of MCP-1, may contribute to the infiltration of monocytes into the mesangium and subsequent foam cell formation. Hence, VLDLs may play a role in the pathogenesis of focal glomerulosclerosis. One of the mechanisms of such effect may be mediated through the calcium-dependent protein kinase C pathway.

Homocysteine stimulates the production and secretion of cholesterol in hepatic cells.

Homocysteinemia and hypercholesterolemia are important risk factors associated with the occurrence of arteriosclerotic vascular diseases. A positive correlation between plasma levels of homocysteine and cholesterol was found in homocysteinemic patients as well as in experimental animals. In the present study, the effect of homocysteine on the production and secretion of cholesterol in human hepatoma cell line HepG2 cells was investigated. When cells were incubated with 4 mM homocysteine, the amounts of total cholesterol produced as well as the cholesterol secreted by these cells were significantly increased (from 32 +/- 5 to 74 +/- 5 nmol/mg cellular protein). Further biochemical analyses revealed that the increase in cholesterol was resulted from an enhancement in the production and secretion of the unesterified cholesterol with no concomitant change in the level of cholesteryl esters. The activity of intracellular 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase was markedly elevated by 131% and 190% after cells were incubated with homocysteine for 24 and 48 h. Homocysteine also stimulated the secretion of apo B100 by HepG2 cells (from 0.84 +/- 0.11 to 1.37 +/- 0.12 micrograms apolipoprotein B/mg cellular protein). Our results demonstrate that homocysteine stimulates the production and secretion of cholesterol and apolipoprotein B100 in HepG2 cells. The increase in the production of cholesterol induced by homocysteine may contribute to the pathogenesis of arteriosclerosis.

Role of lipoprotein-X in foam cell formation and rat mesangial cell proliferation.

1. In the present study, the effect of reconstituted lipoprotein-X (rLp-X) on lipid accumulation and foam cell formation in rat peritoneal macrophages was evaluated. Furthermore, the combined effect of rLp-X and macrophages on mesangial cell proliferation was examined. 2. Incubation of macrophages with rLp-X (177 and 387 nmol unesterified cholesterol (FC)/mL) resulted in an increase of cellular cholesterol (162%) and cholesteryl esters (223 to 245%) relative to control. 3. Oil Red O staining of macrophages treated with rLp-X revealed the presence of foam cells. 4. In conclusion, rLp-X had no effect on the proliferation of mesangial cells incubated in macrophage-conditioned medium.

Uptake and metabolism of lipoprotein-X in mesangial cells.

Progressive glomerulosclerosis is a major complication in patients with familial lecithin:cholesterol acyltransferase (LCAT) deficiency. The lack of LCAT activity results in the accumulation of an abnormal lipoprotein, lipoprotein-X (Lp-X), in the plasma of these patients. Lipoprotein-X contains high levels of unesterified cholesterol and phosphatidylcholine. Lp-X may play a role in the accumulation of lipids in the kidney, which in turn may lead to glomerulosclerosis. The objective of this study is to examine the uptake and metabolism of Lp-X by rat mesangial cells. Our results suggest that Lp-X is taken up by mesangial cells and that the lipids in Lp-X are metabolized. Lysosomes containing unesterified cholesterol and phosphatidylcholine, in a molar ratio similar to Lp-X, were synthesized to investigate the roles individual apolipoproteins (apo CI, II, III and E) play in the uptake of Lp-X. Both apo CI and CIII inhibited its uptake while apo CII (1.5 fold) and E (4 fold) stimulated the uptake of Lp-X. Very low density lipoprotein (VLDL) and low density lipoprotein (LDL) inhibited Lp-X uptake by mesangial cells. However, at higher concentrations of high density lipoprotein (HDL), the uptake of Lp-X was stimulated. Proteoglycans have an important role in regulating the uptake of Lp-X, while cytoskeleton-dependent phagocytosis and the scavenger receptor do not appear to be involved.