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Regulation of apolipoprotein B secretion from hepatocytes: Comparison between normal and WHHL rabbit hepatocytes
WS5-K-3-07 REGULATION OF APOLIPOPROTEIN B SECRETION FROM HEPATOCYTES:COMPARISON BETWEEN NORMAL AND WHHL RABBIT HEPATOCYTES. M.Tanaka, H.Otani, M.Yokode,and T.Kita, Dept.of Geriatric Medicine, Kyoto University, Kyoto, Japan We investigated the mechanism by which cellular cholesteryl ester contents regulate apolipoprotein B (apoB) secretion in rabbit hepatocyte culture system. When normal rabbit hepatocytes were incubated with increasing concentrations of LDL, there was a dose dependent increase in apoB secretion. On the other hand, at a low concentration of LDL, there was no increase in apoB secretion in WHHL rabbit hepatocytes. However, at a high concentration of LDL, a significant increase in apoB secretion could be observed in WHHL rabbit hepatocytes. When pravastatin, an HMG-CoA reductase inhibitor, was added to the culture medium, apoB secretion was decreased significantly both in normal and WHHL rabbit hepatocytes. There was a significant correlation between cellular cholesteryl ester contents and apoB secretion. Northern hybridization and RNase protection assay indicated that neither LDL nor pravastatin caused a significant change in cellular mRNA level for apoB. It was demonstrated by pulsechase experiments that decrease in cellular cholesteryl ester contents by pravastatin accerelated intracellular degradation of apoB, whereas increase in cellular cholesteryl ester contents by the addition of LDL slowed intracellular degradation rate. We conclude that cellular cholesteryl ester contents change intracellular degradation rate of apoB, and thereby regulate apoB secretion both in normal and WHHL rabbit hepatocytes.
WS5-K-3-08 HMG-CoA REDUCTASE INHIBITOR AND CHOLESTYRAMINE INDUCE HEPATIC LDL RECEPTOR AND ARE EFFECTIVE FOR LOWERING PLASMA CHOLESTEROL IN HOMOZYGOUSWATANABE HERITABLE HYPERLIPIDEMIC RABBITS K. Hayashi, J. Kurokawa, Y. Kuga, H. Kurushima I st Dept. of Internal Medicine, Hiroshima Univ., School of Medicine, Hiroshima,Japan Increased plasma cholesterol levels are causually related to an increased risk of coronary artery disease. Several kinds of intervention studies involves dietary or drug use have provided substantial evidence that lowering of total and LDL-cholesterol levels reduces the incidence of coronary heart disease and the progression of coronary atheroscterosis. Familialhypercholesterolemia(FH), a genetic disease characterized by LDL receptor deficiency and increased levels of total cholesterol and LDL-cholesterol in the blood, results in s marked increased incidence of atherusclerusis and coronary artery disease. Even in FH, the cholesterol lowering therapy with drugs and LDL-apheresis reduces the incidence of coronary heart disease and the progression of coronary atherosclerosis. In homozygous Wstanabe heritable hyperlipidemic (WHHL) rabbits, the amount of LDL bound to LDL receptor is less than 5% of that in normal rabbits. This is due to s mutant receptor for LDL that is not transported to the cell surface at a normal rate and WHHL rabbits show development of severe atherosclerosis. Therefore, homozygous and heterozygous WHHL rabbits are excelent animal models for human FH and are useful to evaluate effect of drugs for lowering plasma cholesterol, especially LDL-cholesterol. Among drugs that are highly effective for lowering plasma cholesterol, inhibitors of HMG-CoA reductese exert a strong effect on both familial and non-famillai forms of hypercholesterolemia. The main mechanism for this drug to lower plasma LDLcholesterol is reported to be the induction of hepatic LDL receptor caused by hepatic cholesterol depletion. In this study, we examined whether plasma cholesterol levels were reduced in homozygous WHHL rabbits which pocessed mutant LDL receptor by HMG-CoA reductaes inhibitor alone and in combination with cholestyramine. Fluvastatin sodium, an inhibitor of HMG-CoA reductase, was administered (50mg/kg) or in combination with 2% cholestyramine in the diet to homozygous WHHL rabbits for 4 weeks. LDL-cholesterol levels were reduced by 32% and 46% with fluvastelJn alone and the combination treatment, respectively. Hepatic LDL receptor activity was increased by 2.0- and 2.5-fold with fluvastatin alone and the combination treatment, respectively, mRNA for the LDL receptor in the liver was also increased by 2.0- and 2.0-fold with fluvastatin alone and the combination treatment, respectively. Hepatic cholesterol 7a-hydroxylase activity was not affected with fluvastatin alone and was increased by 5-fold with the combination treatment. These results suggest that the increase in hepatic LDL receptor activity by the treatment of fluvastaUn alone or in combination with cholestyramine plays a role in reduction of plasma cholesterol levels in homozygous WHHL rabbits, and that further reduction of plasma cholesterol levels obtained by the combination therapy may be attributable to the decrease in hepatic cholesterol pool used for secretion as VLDL particle by excessive coversion of cholesterol into bile acid through up-regulated hepatic cholesterol 7a -hydroxylase. Synthesized cholesterol is known to stabilize apoB in hepatocytes. Therefore, the hepatic cholesterol depletion by HMG-CoA reductase inhibitor may affect stability of hepatic apoB and impair assembly and secretion of VLDL particles from liver. In addition to the induction and increase in activity of hepatic LDL receptor, the decrease in VLDL secretion from liver may also contribute to the plasma cholesterol lowering effect of HMG-CoA reductase inhibitors in homozygous WHHL rabbits.
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WS5-K-3-08 HMG-CoA REDUCTASE INHIBITOR AND CHOLESTYRAMINE INDUCE HEPATIC LDL RECEPTOR AND ARE EFFECTIVE FOR LOWERING PLASMA CHOLESTEROL IN HOMOZYGOUSWATANABE HERITABLE HYPERLIPIDEMIC RABBITS K. Hayashi, J. Kurokawa, Y. Kuga, H. Kurushima I st Dept. of Internal Medicine, Hiroshima Univ., School of Medicine, Hiroshima,Japan Increased plasma cholesterol levels are causually related to an increased risk of coronary artery disease. Several kinds of intervention studies involves dietary or drug use have provided substantial evidence that lowering of total and LDL-cholesterol levels reduces the incidence of coronary heart disease and the progression of coronary atheroscterosis. Familialhypercholesterolemia(FH), a genetic disease characterized by LDL receptor deficiency and increased levels of total cholesterol and LDL-cholesterol in the blood, results in s marked increased incidence of atherusclerusis and coronary artery disease. Even in FH, the cholesterol lowering therapy with drugs and LDL-apheresis reduces the incidence of coronary heart disease and the progression of coronary atherosclerosis. In homozygous Wstanabe heritable hyperlipidemic (WHHL) rabbits, the amount of LDL bound to LDL receptor is less than 5% of that in normal rabbits. This is due to s mutant receptor for LDL that is not transported to the cell surface at a normal rate and WHHL rabbits show development of severe atherosclerosis. Therefore, homozygous and heterozygous WHHL rabbits are excelent animal models for human FH and are useful to evaluate effect of drugs for lowering plasma cholesterol, especially LDL-cholesterol. Among drugs that are highly effective for lowering plasma cholesterol, inhibitors of HMG-CoA reductese exert a strong effect on both familial and non-famillai forms of hypercholesterolemia. The main mechanism for this drug to lower plasma LDLcholesterol is reported to be the induction of hepatic LDL receptor caused by hepatic cholesterol depletion. In this study, we examined whether plasma cholesterol levels were reduced in homozygous WHHL rabbits which pocessed mutant LDL receptor by HMG-CoA reductaes inhibitor alone and in combination with cholestyramine. Fluvastatin sodium, an inhibitor of HMG-CoA reductase, was administered (50mg/kg) or in combination with 2% cholestyramine in the diet to homozygous WHHL rabbits for 4 weeks. LDL-cholesterol levels were reduced by 32% and 46% with fluvastelJn alone and the combination treatment, respectively. Hepatic LDL receptor activity was increased by 2.0- and 2.5-fold with fluvastatin alone and the combination treatment, respectively, mRNA for the LDL receptor in the liver was also increased by 2.0- and 2.0-fold with fluvastatin alone and the combination treatment, respectively. Hepatic cholesterol 7a-hydroxylase activity was not affected with fluvastatin alone and was increased by 5-fold with the combination treatment. These results suggest that the increase in hepatic LDL receptor activity by the treatment of fluvastaUn alone or in combination with cholestyramine plays a role in reduction of plasma cholesterol levels in homozygous WHHL rabbits, and that further reduction of plasma cholesterol levels obtained by the combination therapy may be attributable to the decrease in hepatic cholesterol pool used for secretion as VLDL particle by excessive coversion of cholesterol into bile acid through up-regulated hepatic cholesterol 7a -hydroxylase. Synthesized cholesterol is known to stabilize apoB in hepatocytes. Therefore, the hepatic cholesterol depletion by HMG-CoA reductase inhibitor may affect stability of hepatic apoB and impair assembly and secretion of VLDL particles from liver. In addition to the induction and increase in activity of hepatic LDL receptor, the decrease in VLDL secretion from liver may also contribute to the plasma cholesterol lowering effect of HMG-CoA reductase inhibitors in homozygous WHHL rabbits.
362