Studies of sterol 12alpha-hydroxylase (CYP8B1) : distribution and role for cholesterol homeostasis and gallstone formation

Sammanfattning: Bile acids are essential physiological molecules which carry out multiple functions such as facilitating the absorption of dietary lipids and regulating the metabolism of cholesterol and bile acids. The formation of bile represents the major route for elimination of excess cholesterol, either by conversion of cholesterol into bile acids or excretion of intact cholesterol in mixed micelles. The primary bile acids consist of cholic acid (CA), chenodoxycholic acid (CDCA), and in the mouse beta-muricholic acid (beta-MCA). Synthesis of CA requires sterol 12alpha-hydroxylase (CYP8B1), a membrane-bound enzyme in the endoplasmic reticulum in the hepatocytes. Cyp8b 1 -/- mice are unable to produce CA, have reduced cholesterol absorption, and a compensatory increased bile acid synthesis. The present studies aimed to investigate the cellular distribution of CYP8B1, and evaluate its role for the bile acid synthesis and cholesterol homeostasis. Specific antibodies against selected peptides of CYP8B1 were used for the immunohistochemical studies. The results demonstrated a species difference concerning the distribution of CYP8B1, as an even distribution was found throughout the liver lobuli in humans, whereas a zonal expression was seen in mice, with an increasing gradient from the portal areas towards the central veins. Such a pattern was also observed for the histochemical distribution of mouse Cyp7a1. Cyp8b1-/- mice were found to accumulate much less hepatic cholesterol than Cyp8b1+/+ mice when fed cholesterol or CA, mainly due to a decreased cholesterol absorption and increased bile acid synthesis through upregulation of the Cyp7a1 expression. The high rate of cholesterol synthesis in Cyp8b1-/- mice was associated with increased levels of SREBP2 mRNA and the corresponding precursor protein. Cholesterol-fed Cyp8b1-/- mice generated higher plasma HDL-cholesterol. After feeding a diet with cholesterol plus CA, cholesterol crystals were found in the bile of Cyp8b1 +/+ but not of Cyp8b1 -/- animals. Insulin-independent diabetes was induced by alloxan in both Cyp8b1-/- and +/+ mice fed cholesterol for 9-10 weeks. Alloxan/cholesterol treated Cyp8b1+/+ mice displayed significantly increased levels of serum cholesterol, and their bile was supersaturated with cholesterol, containing plenty of cholesterol crystals. In addition they had elevated levels of hepatic Abcg5/g8 mRNA. These effects were all absent in the Cyp8bl -/- mice or in +/+ animals treated only with alloxan. Our results indicate a crucial role of Cyp8b1/CA for cholesterol homeostasis in the mouse, suggesting that inhibition of Cyp8b1 may have beneficial effects such as preventing liver cholesterol accumulation and gallstone formation. In addition, such inhibition might also favorably affect the plasma lipid levels under diabetic condition, which may confer anti-atherogenic effects.