Genetic and dietary interactions in the regulation of HMG-CoA reductase gene expression.
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Inbred strains of mice exhibit large genetic variations in hepatic 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase activity. A tissue-specific genetic variation between the strains BALB/c and C57BL/6, resulting in about 5-fold higher levels in hepatic reductase activity in strain C57BL/6, was examined in detail. The activity difference between these two strains could be explained entirely by differences in hepatic reductase mRNA levels. In genetic crosses, the variation segregated as a single major Mendelian element. Surprisingly, the mode of inheritance was recessive since F1 mice exhibited the BALB/c levels of enzyme activity. Despite the fact that the rates of hepatic sterol synthesis also differed between the strains by a factor of about five, the altered hepatic reductase expression did not significantly influence plasma lipoprotein levels. The response to a high cholesterol, high fat diet between the strains was remarkably different. Thus, in BALB/c mice, both hepatic reductase activity and mRNA levels were affected only slightly, if at all, by cholesterol feeding, while in strain C57BL/6 mice both were reduced more than 10-fold by cholesterol feeding. Several lines of evidence, including analysis of cis-acting regulatory elements, the nonadditive mode of inheritance, and genetic studies of the HMG-CoA reductase gene locus on mouse chromosome 13, support the possibility that the variation in reductase expression is not due to a mutation of the structural gene but, rather, is determined by a trans-acting factor controlling reductase mRNA levels. The variation provides a striking example, at the molecular level, of the importance of dietary-genetic interactions in the control of cholesterol metabolism. (+info)
Use of the SeHCAT test in the investigation of diarrhoea.
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The SeHCAT test was used to investigate possible bile acid malabsorption in 166 patients presenting to a district general hospital with chronic diarrhoea of uncertain cause. Eighty-four (51%) patients had impaired SeHCAT retention. These included 23 of 28 patients with a possible type I abnormality (terminal ileal resection or disease, previous pelvic radiotherapy), 20 of 74 with a possible type II abnormality (idiopathic diarrhoea), 32 of 45 with a possible type III abnormality (post-cholecystectomy, post-vagotomy), and 9 of 19 with diarrhoea associated with diabetes. Patients with severe bile acid malabsorption demonstrated a good response to cholestyramine whereas the response in patients with a mildly abnormal SeHCAT retention was variable. Bile acid malabsorption is an important cause of diarrhoea in patients presenting with unexplained chronic diarrhoea. (+info)
Differing effects of cholesterol and taurocholate on steady state hepatic HMG-CoA reductase and cholesterol 7 alpha-hydroxylase activities and mRNA levels in the rat.
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We investigated the effects of cholesterol, cholestyramine, and taurocholate feeding on steady state specific activities and mRNA levels of hepatic 3-hydroxy-3-methylglutaryl (HMG)-CoA reductase and cholesterol 7 alpha-hydroxylase in the rat. Interruption of the enterohepatic circulation of bile acids (cholestyramine feeding) increased total HMG-CoA reductase activity 5-fold. Cholesterol and taurocholate administration suppressed total microsomal HMG-CoA reductase activities 87% and 65%, respectively. HMG-CoA reductase mRNA levels increased 3-fold with cholestyramine, did not decrease significantly with cholesterol feeding, but were markedly decreased after taurocholate treatment. Cholesterol 7 alpha-hydroxylase activity increased 4-fold with cholestyramine and 29% during cholesterol feeding, but decreased 64% with taurocholate. Cholesterol 7 alpha-hydroxylase mRNA levels rose 150% and 50% with cholestyramine and cholesterol feeding, respectively, but decreased 73% with taurocholate. The administration of cholesterol together with taurocholate prevented the decline in cholesterol 7 alpha-hydroxylase mRNA levels, but inhibition of enzyme activity persisted (-76%). Hepatic microsomal cholesterol concentrations increased 2-fold with cholesterol feeding but did not change with taurocholate or cholestyramine treatment. These results demonstrate that mRNA levels of HMG-CoA reductase are controlled by the hepatic taurocholate flux, whereas mRNA levels of cholesterol 7 alpha-hydroxylase are controlled by the cholesterol substrate supply. These end products, cholesterol and bile acids, exert post-transcriptional regulation on HMG-CoA reductase and cholesterol 7 alpha-hydroxylase, respectively. (+info)
12 alpha-hydroxylase activity in human liver and its relation to cholesterol 7 alpha-hydroxylase activity.
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Interruption of the enterohepatic circulation by cholestyramine causes a several-fold increase in bile acid synthesis, reflected in a stimulation of cholesterol 7 alpha-hydroxylase activity; the synthesis of cholic acid being stimulated to a greater extent than chenodeoxycholic acid. It is not known if this preferential increase in cholic acid is due to an increase of the 12 alpha-hydroxylase activity. The present study aimed at investigating the 12 alpha-hydroxylase activity and its relation to cholesterol 7 alpha-hydroxylase activity in liver microsomes of patients with different levels of cholesterol 7 alpha-hydroxylase activity. Liver biopsies were obtained from four gallstone-free patients, and seven untreated and two cholestyramine-treated gallstone patients undergoing cholecystectomy, and four patients with Crohn's disease undergoing intestinal resection. The combined group of cholestyramine-treated and ileum-resected patients had four times higher cholesterol 7 alpha-hydroxylase activity and two times higher 12 alpha-hydroxylase activity than the other patients. A positive correlation was obtained between cholesterol 7 alpha-hydroxylase activity and 12 alpha-hydroxylase activity (r = +0.69; n = 16). These results indicate that the increased ratio between the synthesis of cholic acid and chenodeoxycholic acid during cholestyramine treatment is due to a compensatory increase of the 12 alpha-hydroxylase activity. (+info)
Effects of pravastatin sodium alone and in combination with cholestyramine on hepatic, intestinal and adrenal low density lipoprotein receptors in homozygous Watanabe heritable hyperlipidemic rabbits.
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Pravastatin sodium (pravastatin), a tissue-selective inhibitor of 3-hydroxy-3-methylglutaryl coenzyme A reductase, was administered alone (50 mg/kg) or in combination with cholestyramine, a bile acid sequestrant resin, at the level of 2% in the diet to homozygous Watanabe heritable hyperlipidemic (WHHL) rabbits for 4 weeks. The low density lipoprotein (LDL)-cholesterol levels were reduced by 29% and 56% with pravastatin alone and the combination treatment, respectively. Hepatic LDL receptor activity was increased by 11.2- and 13.9-fold with pravastatin alone and the combination treatment, respectively. The LDL receptor activity in the untreated homozygous WHHL rabbits was only 2.5% of that in the normal rabbits. mRNA for the LDL receptor in the liver was also increased by 2.1- and 3.4-fold with pravastatin alone and the combination treatment, respectively. On the other hand, mRNA for the LDL receptor in the adrenal gland was not affected by pravastatin and the combination treatment, whereas the mRNA in the intestine was increased in both groups. These results suggest the following: 1) the induction of hepatic LDL receptor activity by the treatment of pravastatin alone or in combination with cholestyramine is the main cause of the reduction of serum cholesterol levels by these treatments even in LDL receptor-deficient animals. 2) The induction of the mRNA for the LDL receptor in the liver and intestine, but not that in the adrenal gland, might be a reflection of the tissue-selective inhibition of cholesterol synthesis by pravastatin. (+info)
Hormonal regulation of cholesterol 7 alpha-hydroxylase mRNA levels and transcriptional activity in primary rat hepatocyte cultures.
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In primary cultures of adult rat hepatocytes the level of cholesterol 7 alpha-hydroxylase steady-state mRNA markedly decreased by 72 h. However, the addition of L-thyroxine (T4) and dexamethasone synergistically returned cholesterol 7 alpha-hydroxylase steady-state mRNA levels near to that of cholestyramine-fed animals. The maximal responses to T4 and dexamethasone in serum-free medium were at 1.0 and 0.1 microM, respectively. The addition of T4 in combination with dexamethasone resulted in an 11-fold increase in transcriptional activity of the cholesterol 7 alpha-hydroxylase gene as compared to no addition controls. The specific activities of cholesterol 7 alpha-hydroxylase in microsomes prepared from cultures treated with dexamethasone and T4 were 1.56 +/- 1.17 nmol/h/mg protein which is similar to that of intact liver (1.70 +/- 0.062 nmol/h/mg protein), but lower than cholestyramine-fed animals. Cholesterol 7 alpha-hydroxylase activity was not detectable (less than 0.020 nmol/h/mg protein) at 72 h in cultures without the addition of both dexamethasone and T4. In the presence of optimal concentrations of dexamethasone and T4, glucagon (0.2 microM), or dibutyryl cAMP (50 microM) decreased (90%) cholesterol 7 alpha-hydroxylase mRNA within 6 h. Transcriptional activity decreased (62%) in 6 h following the addition of glucagon (0.2 microM) to the culture medium. The results reported in this paper suggest an important role for multiple hormones in the regulation of cholesterol 7 alpha-hydroxylase in the liver. (+info)
Differential expression of cholangiocyte and ileal bile acid transporters following bile acid supplementation and depletion.
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AIM: We have previously demonstrated that cholangiocytes, the epithelial cells lining intrahepatic bile ducts, encode two functional bile acid transporters via alternative splicing of a single gene to facilitate bile acid vectorial transport. Cholangiocytes possess ASBT, an apical sodium-dependent bile acid transporter to take up bile acids, and t-ASBT, a basolateral alternatively spliced and truncated form of ASBT to efflux bile acids. Though hepatocyte and ileal bile acid transporters are in part regulated by the flux of bile acids, the effect of alterations in bile acid flux on the expression of t-ASBT in terminal ileocytes remains unclear. Thus, we tested the hypothesis that expression of ASBT and t-ASBT in cholangiocytes and ileocytes was regulated by bile acid flux. METHODS: Expression of ASBT and t-ASBT message and protein in cholangiocytes and ileocytes isolated from pair-fed rats given control (C) and 1% taurocholate (TCA) or 5% cholestyramine (CY) enriched diets, were assessed by both quantitative RNase protection assays and quantitative immunoblotting. The data obtained from each of the control groups were pooled to reflect the changes observed following TCA and CY treatments with respect to the control diets. Cholangiocyte taurocholate uptake was determined using a novel microperfusion technique on intrahepatic bile duct units (IBDUs) derived from C, TCA and CY fed rats. RESULTS: In cholangiocytes, both ASBT and t-ASBT message RNA and protein were significantly decreased in response to TCA feeding compared to C diet. In contrast, message and protein of both bile acid transporters significantly increased following CY feeding compared to C diet. In the ileum, TCA feeding significantly up-regulated both ASBT and t-ASBT message and protein compared to C diet, while CY feeding significantly down-regulated message and protein of both bile acid transporters compared to C diet. As anticipated from alterations in cholangiocyte ASBT expression, the uptake of taurocholate in microperfused IBDUs derived from rats on TCA diet decreased 2.7-fold, whereas it increased 1.7-fold in those on CY diet compared to C diet fed groups. CONCLUSION: These data demonstrate that expression of ASBT and t-ASBT in cholangiocytes is regulated by a negative feedback loop while the expression of these transporters in terminal ileum is modified via positive feedback. Thus, while transcriptional regulatory mechanisms in response to alterations in bile acid pool size are operative in both cholangiocytes and ileocytes, each cell type responds differently to bile acid supplementation and depletion. (+info)
Stanol esters decrease plasma cholesterol independently of intestinal ABC sterol transporters and Niemann-Pick C1-like 1 protein gene expression.
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Possible mechanisms for the cholesterol-lowering effects of plant stanol esters were addressed by feeding hamsters diets containing stanol esters, cholesterol, or cholestyramine/lovastatin. ABCA1, ATP binding cassette G1 (ABCG1), ABCG5, ABCG8, and Niemann-Pick C1-like 1 (NPC1L1) mRNA levels were then estimated in duodenum, jejunum, and ileum. Plasma cholesterol was decreased by 36% and 94% in animals fed stanol esters and cholestyramine/lovastatin, respectively. Cholesterol feeding increased plasma cholesterol by 2.5-fold. Plasma plant sterols were unchanged by stanol ester feeding but became undetectable by feeding cholestyramine/lovastatin. Cholesterol and stanols accumulated in enterocytes of animals fed cholesterol and stanol esters, respectively. ABCG5 and ABCG8 mRNA levels were decreased by stanol esters and cholestyramine/lovastatin. Cholesterol feeding markedly increased ABCA1 and ABCG1 expression and modestly increased ABCG5/ABCG8. NPC1L1 mRNA was not significantly altered by any of the diets. ABCG1, ABCG5, ABCG8, and NPC1L1 mRNAs were highest in cells of the upper villus, whereas ABCA1 mRNA was highest in cells of the lower villus. The results suggest that cholesterol lowering effect of stanol esters is unrelated to changes in mRNA levels of intestinal ABC sterol transporters or NPC1L1. Cholesterol flux regulates ABC expression but not NPC1L1. The different localization of ABCA1 suggests a different function for this protein than for ABCG1, ABCG5, ABCG8, and NPC1L1. (+info)