7a-Hydroxylase activity

Matheson, H. B., Colon, I. S., and Story, J. A. (1995). Cholesterol 7a-hydroxylase activity is increased by dietary modification with psyllium hydrocolloid, pectin, cholesterol and cholestyramine in rats. /. Nutr. 125,454-M58. 

The properties of 7a-hydroxylase from pigeon liver microsomes305 and from rat liver306,307 have been further described, and new assay methods are available.308,309 Free cholesterol, rather than a cholesteryl ester, was the preferred substrate for the enzyme from rat liver microsomes,310 and the substrate pool for the hydroxylase was about one third of the total amount of cholesterol present in the microsomal preparation.309 Cholesterol 7a-hydroxylase activity is more sensitive to thyroid function than are the activities of the enzymes responsible for cholesterol synthesis,311 and (22f )-22-aminocholesterol, although having no effect on serum or liver cholesterol levels in rats, drastically reduced 7a-hydroxylase activity.312... 

Patients with familial hypercholesterolaemia exhibit lower levels of plasma cholesterol after an operation for portacaval anastomosis, and it has now been shown in rats that such an operation causes an increase in HMG-CoA reductase and cholesterol 7a -hydroxylase activities. Many transplantable human and rodent hepatomas do not control the rate of sterol biosynthesis and HMG-CoA reductase levels in response to dietary cholesterol as normal liver cells do. However, certain hepatoma cells have now been found that, although lacking feedback regulation of choles-terologenesis in vivo, retain their regulatory ability in vitro It thus appears that malignant transformation is not necessarily linked to the loss of regulation by the cell of HMG-CoA reductase activity or sterol synthesis. 

Cholesterol 7a-hydroxyIase has been partially purified from rat and rabbit liver (H5). The enzyme is located in the smooth endoplasmic reticulum and is dependent on cytochrome F-450 and NADPH-cytochrome P-450 reductase for activity (H5). The particular cytochrome P-450 associated with microsomal cholesterol 7a-hydroxylase activity constitutes a small fraction of total liver cytochrome P-450 and, in the rabbit, it appears to be a subfraction of cytochrome P-450lm4 (B28). Measurement of the activity of this enzyme by isotope incorporation is complicated by dilution of added cholesterol by endogenous microsomal cholesterol. A method has now been developed to remove cholesterol fit>m microsomes, so that the mass of 7a-hydroxycholesterol formed during enzyme assay can be accurately calculated (S25). Using this assay, cholic acid feeding was shown to suppress the activity of cholesterol 7a-hydroxylase in rat liver, whereas cholesterol feeding did not (S25). 

A short-term regulation mechanism for cholesterol 7a-hydroxylase activity has been investigated recently in rat liver. The enzyme appears to exist in two forms, which are interconverted by cytosolic fiictors (K12). These foctors may correspond to a protein kinase and a phosphatase, which have been proposed to regulate cholesterol 7a-hydroxylase activity by a phosphorylation (active form)-dephosphorylation (inactive form) mechanism (S9). Another enzyme utilizing cholesterol as substrate, acyl-CoA cholesterol O-acyltransferase (EC 2.3.1.26), may also be regulated in this way, while the biosynthetic enzyme, HMC-CoA reductase, is inhibited in the phosphory-lated form (SIO). Thus, short-term regulation of the concentration of un-esterified cholesterol in the liver may be achieved by coordinate control of these three key enzymes in cholesterol metabolism by reversible phosphorylation (SIO). 

Other clinical signs consist of progressive neurologic dysfunction, cataracts, and premature atherosclerosis (SI). The disease is inherited as an autosomal recessive trait, but is usually only detected in adults when cholesterol and cholestanol have accumulated over many years (S2). Biochemical features of the disease include striking elevations in tissue levels of cholesterol and cholestanol and the presence of unusual bile acids, termed bile alcohols, in bile. These bile alcohols are mainly 5 -cholestane-3a,7a,12a,24S, 25-pentol, Sp-diolestane-3a,7a,12a,23 ,25-pentol and 5P-du)lestane-3a,7a,12a,25-tetrol (S2). As chenodeoxycholic acid is deficient in the bile of patients with CTX, it was postulated that early bile salt precursors are diverted into the cholic acid pathway and 12a-hydroxy bile alcohols with an intact side chain accumulate because of impaired cleavage of the cholesterol side chain and decreased bile acid production (S2). HMG-CoA reductase and cholesterol 7a-hydroxylase activity are elevated in subjects with CTX (N4, N5), so that sufficient 7a-hydroxycholesterol should be available for bile acid synthesis. 

Recent investigations into the mechanism of action of these bile acids indicate that ursodeoxycholic acid has certain advantages over chenodeoxycholic acid in the context of the overall homeostasis of cholesterol metabolism (F6). In contrast to chenodeoxycholic acid, ursodeoxycholic acid does not suppress bile acid synthesis (H7), possibly because the a-orientation of the 7-hydroxyl group of chenodeoxycholic acid is required to inhibit cholesterol 7a-hydroxylase activity. Thus, cholesterol breakdown into bile acids is not reduced by ursodeoxycholic acid. Other favorable factors are that ursodeoxycholic acid has a reduced capacity to solubilize cholesterol into micellar solution compared to chenodeoxycholic acid and intestinal cholesterol absorption is decreased by this bile acid (F6, H7). However, in gallbladder bile the relative limitation of ursodeoxycholic acid for micellar solubilization of cholesterol is compensated for by an ability to transport... 

Most studies on substrate specificity of cholesterol 7a-hydroxylase have been performed with intact microsomes. Results of such studies may be difficult to interpret since the enzyme system is embedded in a lipoprotein membrane, and may not be directly accessible to potential substrates [59]. Thus, differences in the rate of 7a-hydroxylation of various steroids could be due to differences in the rate at which the substrate reaches the active site of. the enzyme rather than to differences in the intrinsic ability of the enzyme to interact catalytically with the substrate [59], Further, occurrence of 7a-hydroxylation of a certain steroid may not reflect the substrate specificity of cholesterol 7a-hydroxylase activity since different species of cytochrome P-450 are present in the microsomes. 

The possibility that the biosynthesis of bile acids is regulated by a negative feedback mechanism was supported by early experiments by Thompson and Vars [206] and Eriksson [207], who showed that the rate of bile acid synthesis in rats increased about 10-fold when a bile fistula is made. Bergstrom and Danielsson demonstrated that duodenal infusion of taurochenodeoxycholic acid in bile fistula rats restored the increased synthesis to a normal rate [208]. Danielsson et al. [44] showed that the cholesterol 7a-hydroxylase activity increased in parallel with the bile acid synthesis after cannulation of the bile duct in rats. In a subsequent work by Mosbach et al., it was reported that the incorporation of isotope from labelled acetate, mevalonate and cholesterol but not from labelled 7a-hydroxycholesterol into bile acids was inhibited by duodenal infusion of taurocholate to bile fistula rats [209]. The incorporation of isotope from labelled acetate, mevalonate and cholesterol but not from labelled 7a-hydroxycholesterol was stimulated in perfused livers of cholestyramine-treated rabbits [210]. It was concluded that there are essentially no rate-limiting steps beyond 7a-hydroxycholesterol in the biosynthesis of bile acids from acetate. Since both cholesterol and bile acid biosynthesis was subjected to negative feedback inhibition by bile acids, it cannot be excluded that inhibition of cholesterol biosynthesis precedes inhibition of the bile acid biosynthesis, and that the latter inhibition is secondary to the former. 

The specificity of the inhibitory effect of bile acids on bile acid formation has been studied by Danielsson [110]. Taurocholic acid, taurochenodeoxycholic acid and taurodeoxycholic acid, fed at the 1% level in the diet for 3-7 days, were found to inhibit cholesterol 7a-hydroxylase activity. Feeding taurohyodeoxycholic acid and... 

Since very little bile acids are transported via the lymph [224], the increased cholesterol 7a-hydroxylase activity observed after lymphatic drainage [225] is not due to depletion of bile acids. In view of the fact that the synthesis of cholesterol increases after lymphatic drainage, it is possible that the increased 7a-hydroxylation in this case may be due to increased supply of newly synthesized cholesterol. 

Treatment with thyroid hormone increases and thyroidectomy decreases cholesterol 7a-hydroxylase activity as well as HMG-CoA reductase activity [75,234,235]. As shown by Balasubramaniam et al. treatment with thyroid hormone increases first the cholesterol 7a-hydroxylase activity and then, after a time lag of several hours, HMG-CoA reductase [234]. Thus, it is likely that the rise in HMG-CoA reductase activity in this case is secondary to the depletion of cholesterol by cholesterol 7a-hydroxylase. 

Takeuchi et al. measured cholesterol 7a-hydroxylase and HMG-CoA reductase activity in fasted rats refed glucose [237]. The administration of glucose to these rats resulted in increased cholesterol synthesis after 1 h and increased cholesterol 7a-hydroxylase activity after 2 h. These effects were not noted in rats pretreated with an inhibitor of cholesterol biosynthesis, suggesting that the effects on the cholesterol 7a-hydroxylase were secondary to those on cholesterologenesis. It was suggested that the stimulatory effect of glucose was due to increased availability of cholesterol for the cholesterol 7a-hydroxylase. 

A dissociation between HMG-CoA reductase and the cholesterol 7a-hydroxylase has been reported in connection with feeding of cholesterol, tomatidine, sitosterol as well as in scurvy. Feeding cholesterol inhibits HMG-CoA reductase and in most [99,222,238,239] but not all [50] studies a stimulatory effect has been found on cholesterol 7a-hydroxylase. The stimulatory effect may be due to an expansion of the pool of cholesterol available for cholesterol 7a-hydroxylase. Feeding with tomatidine and sitosterol interferes with absorption of cholesterol from the intestine, and the increased HMG-CoA reductase activity is probably due to decreased inhibition by lymph cholesterol [240,241]. The cholesterol 7a-hydroxylase activity is only slightly increased or unaffected under these conditions [240,241]. 

Cholesterol 7a-hydroxylase activity is markedly reduced in scorbutic guinea pigs, probably due to decreased amount of the specific cytochrome P-450 [242]. HMG-CoA reductase is unaffected [243]. In another study, an increased incorporation of [ C]acetate into cholesterol in liver of scorbutic guinea pigs was reported [244]. 

Possible mechanisms for regulation of cholesterol 7a-hydroxylase activity... 

The most important information concerning mechanisms of regulation of cholesterol 7a-hydroxylase is summarized in the model shown in Fig. 14. In view of the importance of HMG-CoA reductase for cholesterol 7a-hydroxylase activity, some important modulators of HMG-CoA reductase activity have also been included (cf. Chapter 2). It should be pointed out that the regulation of biosynthesis of cholesterol shown in Fig. 14 is oversimplified. For a discussion of the regulation of HMG-CoA reductase, the reader is referred to Chapter 2 and a recent review by Brown and Goldstein [246]. 

Several investigations have reported the ability of some natural quinones to be therapeutically useful as hypolipidemic agents. Both experimental and clinical studies have indicated that a novel source of dietary fibre produced from rhubarb is potentially hypolipidemic. The increased excretion of bile acids and induction of cholesterol 7a-hydroxylase activity may account for... 

Kushwaha, R.S. and K.M. Born (1991). Effect of estrogen and progesterone on the hepatic cholesterol 7a-hydroxylase activity in... 

Reihner, E., I. Bjbrkhem, B. Angelin, S. Ewerth, and K. Einarsson (1989). Bile acid synthesis in humans Regulation of hepatic microsomal cholesterol 7a-hydroxylase activity. Gastroenterology 97, 1498-1505. 

K. Okuda, and T. Setoguchi (2002). Half-life of cholesterol 7a-hydroxylase activity and enzyme mass differ in animals and humans when determined by a monoclonal antibody against human cholesterol 7a-hydroxylase. J. Steroid Biochem. Mol. Biol. 81, 377-380. 

Nguyen, L.B., S. Shefer, G. Salen, J.Y. Chiang, and M. Patel (1996), Cholesterol 7a-hydroxylase activities from human and rat liver are modulated in vitro posttranslationally by phosphorylation/ dephosphorylation. Hepatology 24, 1468-1474. 

K. Wikvall (2000). Oxysterol 7a-hydroxylase activity by cholesterol 7a-hydroxylase (CYP7A). J. Biol. Chem. 275, 34046-34053. 

Sauter, G., M. Weiss, and R. Hoermann (1997). Cholesterol 7a-hydroxylase activity in hypothyroidism and hyperthyroidism in humans. Horm. Metab. Res. 29, 176-179. 

Section VI The patterns of diurnal variations in cholesterol synthesis and cholesterol 7a-hydroxylase activity have been found to be practically the... 

Einarsson K, Akerlund JE, Reihner E, Bjorkhem I (1992) 12a-hydroxylase activity in human liver and its relation to cholesterol 7a-hydroxylase activity. J Lipid Res 33 1591-1595... 

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