Cholesterol 7α-hydroxylase activity and bile acid synthesis in hepatocytes of unwearied and weaned pigs in monolayer culture

Kwekkeboom, Jaap; Princen, H.M.G.; Voorthuizen, E.M. van; Kempen, H.J.M.

doi:10.1016/0005-2760(90)90169-x

Cited by 18 publications

(16 citation statements)

References 49 publications

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“…The existence of alternative pathways in the formation of bile acids in cultured hepatocytes from the rat and pig has been suggested previously by us as an explanation for the discrepancy between cholesterol 7a-hydroxylase activity and mass synthesis of bile acids [21,32]. Based on the latter observations and on the magnitude of inhibition of bile acid synthesis by CsA, we suggest that formation of bile acids via this alternative pathway may contribute substantially to total bile acid synthesis, at least in cultured rat hepatocytes.…”

Section: Discussionmentioning

confidence: 99%

Cyclosporin A blocks bile acid synthesis in cultured hepatocytes by specific inhibition of chenodeoxycholic acid synthesis

Princen¹,

Meijer

Wolthers

et al. 1991

Biochemical Journal

102

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Bile acid synthesis, determined by conversion of [4-14C]cholesterol into bile acids in rat and human hepatocytes and by measurement of mass production of bile acids in rat hepatocytes, was dose-dependently decreased by cyclosporin A, with 52 % (rat) and 45 % (human) inhibition at 10/,M. The decreased bile acid production in rat hepatocytes was due only to a fall in the synthesis of,-muricholic and chenodeoxycholic acids (-64 % at 10 4M-cyclosporin A), with no change in the formation of cholic acid. In isolated rat liver mitochondria, 26-hydroxylation of cholesterol was potently inhibited by the drug (concn. giving half-maximal inhibition = 4/tM). These results suggest that cyclosporin A blocks the alternative pathway in bile acid synthesis, which leads preferentially to the formation of chenodeoxycholic acid.

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Section: Discussionmentioning

confidence: 99%

Cyclosporin A blocks bile acid synthesis in cultured hepatocytes by specific inhibition of chenodeoxycholic acid synthesis

Princen¹,

Meijer

Wolthers

et al. 1991

Biochemical Journal

102

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“…Materials used for isolation and culturing of pig hepatocytes and determination of cholesterol 7a-hydroxylase activity were obtained from sources described previously (16,18). Hexafluoro-2-propano1, trifluoroacetic anhydride and deoxycholic acid were obtained from Serva (Heidelberg, West Germany); pronase from Calbiochem (Behring Diagnostics, La Jolla, CA) and mithramycin from Pfizer (Rotterdam, The Netherlands).…”

Section: Methodsmentioning

confidence: 99%

“…Acids by Cultured Pig Hepatocytes. Pig hepatocytes in monolayer culture synthesize mainly hyocholic and chenodeoxycholic acids (>85% of total bile acid synthesis) and furthermore minor amounts of hyodeoxycholic and murocholic acids but not cholic acid (16). Thus, addition of taurocholic acid to the culture medium does not interfere with determination of total mass production of bile acids by these cells.…”

Section: Effect Of Taurochlic Acid On Mass Production Of Bilementioning

confidence: 98%

Bile acids exert negative feedback control on bile acid synthesis in cultured pig hepatocytes by suppression of cholesterol 7α-hydroxylase activity

Kwekkeboom¹,

Princen²,

Voorthuizen³

et al. 1990

Hepatology

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Feedback regulation of bile acid synthesis by its end products was studied in cultured hepatocytes of young weaned pigs. We previously showed that conversion of exogenous [14C] cholesterol into bile acids was suppressed by addition of bile acids to the culture medium. In the present study, the effects of bile acids on bile acid mass production and cholesterol 7 alpha-hydroxylase activity were examined. Mass production of bile acids was strongly inhibited by addition of taurocholic acid (50 and 100 mumol/L) to the culture medium. The inhibitory action was exerted specifically on activity of cholesterol 7 alpha-hydroxylase because conversion of [14C] 7 alpha-hydroxycholesterol to bile acids by pig hepatocytes was not affected. Suppression of cholesterol 7 alpha-hydroxylase activity after incubation of the hepatocytes with taurocholic acid was concentration- and time-dependent. Maximum suppression (-80%) was achieved after a 20 to 30 hr incubation of hepatocytes with 100 mumol/L of this bile acid. Decline of enzyme activity caused by 100 mumol/L taurocholic acid followed first-order kinetics with a half-life of 10 hr. Taurocholic acid had no direct effect on cholesterol 7 alpha-hydroxylase activity in homogenates of hepatocytes as assessed by addition of the bile acid to the assay mixture. The effects of several other bile acids in a concentration of 100 mumol/L on cholesterol 7 alpha-hydroxylase activity were examined in 48 hr incubations. Glycochenodeoxycholic and glycohyodeoxycholic acids, which are the major bile acids in pig bile, their unconjugated forms and also deoxycholic and cholic acid pronouncedly inhibited activity of the enzyme.(ABSTRACT TRUNCATED AT 250 WORDS)

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“…This correlation was particularly marked between uptake of cholesterol-rich lipoproteins (LDL, HDL) and synthesis of bile salts from lipoproteic cholesterol (Junker & Davis, 1989;Whiting et al, 1989;Kwekkeboom et al, 1990). Incorporation of lipoproteic cholesterol implies binding of bile salts to lipoproteins (Kramer et al, 1979;Chitranukroh & Billing, 1983;Salvioli et al, 1985;Ceryak et al, 1993).…”

Section: Discussionmentioning

confidence: 99%

Effect of crilvastatin, a new cholesterol lowering agent, on unesterified LDL‐cholesterol metabolism into bile salts by rat isolated hepatocytes

Clerc

Sbarra

Diaconescu

et al. 1995

British J Pharmacology

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The aim of these experiments was to determine the effect of crilvastatin, a new cholesterol lowering agent, on the metabolism of unesterified low density lipoprotein (LDL)‐cholesterol by rat freshly isolated hepatocytes. This preclinical model was developed as an alternative to in vivo experiments, to mimic the metabolic effects of a molecule on its target cells and to define optimal conditions for future experimentation on human hepatocytes. Cells were obtained from normolipidaemic or hypercholesterolaemic rats, hypercholesterolaemia was nutritionally induced. Incubations were performed in a medium containing 600 μm taurocholate and 50 μm or 300 μm crilvastatin. This molecule was shown in vitro to be carried by physiological transporters, i.e., albumin‐bile salt micellar associations and LDL. Crilvastatin induced a significance increase in the synthesis and secretion by hepatocytes of bile salts resulting from the metabolism of unesterified LDL‐cholesterol in both normolipidaemic and hypercholesterolaemic rats. Stimulation involved non‐conjugated as well as tauro‐ and glyco‐conjugated bile salts. These findings corroborate preliminary studies showing in vivo that crilvastatin enhances the secretion of bile acids by stimulating the uptake and incorporation of LDL‐cholesterol by the liver.

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Cholesterol 7α-hydroxylase activity and bile acid synthesis in hepatocytes of unwearied and weaned pigs in monolayer culture

Cited by 18 publications

References 49 publications

Cyclosporin A blocks bile acid synthesis in cultured hepatocytes by specific inhibition of chenodeoxycholic acid synthesis

Cyclosporin A blocks bile acid synthesis in cultured hepatocytes by specific inhibition of chenodeoxycholic acid synthesis

Bile acids exert negative feedback control on bile acid synthesis in cultured pig hepatocytes by suppression of cholesterol 7α-hydroxylase activity

Effect of crilvastatin, a new cholesterol lowering agent, on unesterified LDL‐cholesterol metabolism into bile salts by rat isolated hepatocytes

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