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.

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.

Effect of lipoprotein-X on lipid metabolism in rat kidney.

Lipoprotein-X (Lp-X) is found in the plasma of patients with familial lecithin: cholesterol acyltransferase (LCAT) deficiency syndromes. The majority of the patients with this disorder develop progressive glomerulosclerosis. In this study, the effect of Lp-X on lipid metabolism in perfused rat kidney was investigated. Lp-X was isolated from plasma of patients with familial LCAT deficiency by sequential ultracentrifugation and gel filtration column chromatography. Rat kidneys were perfused for 1-2 h with Krebs-Henseleit buffer containing 20 microM [1-(14)C]acetate or 20 microM [Me-3H]choline. In the presence of Lp-X, no significant difference in the incorporation of radioactivity into triglycerides, cholesterol, phosphocholine, CDP-choline and sphingomyelin was observed. However, incorporation of radioactivity into cholesteryl esters and phosphatidylcholine was significantly elevated in Lp-X perfused kidneys. The contents of cholesterol, cholesteryl esters and phosphatidylcholine were also significantly increased in Lp-X perfused kidneys. The increase in lipid content in the Lp-X perfused kidney is attributed to the direct deposition of Lp-X lipids into the organ. The increase in the labelling of cholesteryl esters was attributed to the increase of available substrate (cholesterol) for the acyl-CoA:cholesterol acyltransferase (ACAT) reaction. The increase in phosphatidylcholine labelling was caused by a reduced turnover of the newly synthesized labelled phosphatidylcholine during Lp-X perfusion.

Lipoprotein-X fails to inhibit hydroxymethylglutaryl coenzyme A reductase in HepG2 cells.

Abnormalities of lipid composition and metabolism are frequently observed in patients with cholestatic liver disease. Both elevated low-density lipoprotein (LDL) levels and the appearance of lipoprotein-X (LP-X) in plasma underlie the high incidence of hypercholesterolemia in this population. We tested the hypothesis that the hypercholesterolemia of cholestasis may reflect a failure of normal feedback regulation of hepatic cholesterogenesis by determining the influence of LP-X on the rate-limiting enzyme of cholesterol synthesis, hydroxymethylglutaryl coenzyme A (HMG CoA) reductase. Cultured human hepatoma (HepG2) cells were incubated in purified lipoprotein for 24 hours, harvested, and then assayed for HMG CoA reductase activity and mass. LDL isolated from either normal controls or patients with cholestasis decreased reductase activity in a dose-dependent fashion (2 to 30 micrograms cholesterol/mL media) to a level approximately 50% of that measured in cells incubated in lipid-deficient serum. LP-X failed to downregulate enzyme activity compared with LDL, with little change in reductase activity at cholesterol concentrations (30 micrograms/mL media) that produced maximal reductase inhibition by LDL. Three distinct LP-X subspecies were purified from the plasma of a patient with primary biliary cirrhosis (PBC) and tested in an analogous manner. All LP-X subspecies were similar in their inability to decrease reductase activity as compared with LDL. HMG CoA reductase mass was increased approximately twofold in cells incubated with LP-X, as estimated by Western blot analysis. These results suggest that LP-X may contribute to hypercholesterolemia in the cholestatic patient by not effectively downregulating hepatic cholesterol synthesis.

Interaction of various lipoproteins from normal and dyslipoproteinemic plasma with mouse peritoneal macrophages.

In this study we tried to elucidate the atherogenicity of various plasma lipoproteins with respect to their capability of foam cell formation. Mouse peritoneal macrophages (MPM) were incubated with increasing amounts of lipoproteins and the incorporation of 14C oleate into the cholesteryl ester fraction was followed. The results may be summarized as follows: freshly isolated Lp(a) behaves very similar to normal LDL causing no or little increase in CE formation in MPM. Lp(a) treated with dextran sulfate as well as with antibodies to Apo-a, strongly interact with scavenger receptors causing massive accumulation of CE in MPM. The abnormal lipoproteins from patients suffering from liver disease, LP-X, HDL-E cause no increase in CE formation of MPM. They behave very similar to artificial PL/FC liposomes. If on the other hand these abnormal lipoproteins are mixed with Ac-LDL, a synergistic effect was observed causing an approx. 30 per cent increase in CE-formation as compared to Ac-LDL alone. This was caused by a net transfer of FC from abnormal lipoproteins to Ac-LDL alone. This was caused by a net transfer of FC from abnormal lipoproteins to Ac-LDL. It is concluded that the lipoproteins studied in this report by itself exert no atherogenic function in MPM. They may, however, aggravate the atherogenicity of other processes known to be involved in the development of vascular diseases.

Cholesterol esterification in mouse peritoneal macrophages in the presence of pathological human plasma lipoproteins.

Mouse peritoneal macrophages were incubated with abnormal lipoproteins (LP-X, HDL-E, VLDL-p, IDl-p and LDL-p) from a patient with secondary deficiency in phosphatidylcholine-sterol acyltransferase, or with phosphatidylcholine/cholesterol liposomes, and the stimulation of cholesteryl ester formation was studied. Acetylated low density lipoproteins served as a control. It was found that macrophages incubated with LP-X, the other pathological lipoproteins or with liposomes did not show an enhanced cholesterol esterification. Also HDL-E had no effect despite of its high apoE content and the fact that apoE has been postulated to be the agonist in beta-VLDL binding to macrophages.