Chenodeoxycholate, biosynthesis

Fig. 9. The elaboration of the cholesteol nucleus in bile acid synthesis. (Cholic acid and chenodeoxycholic acid biosynthesis pathway).

For obvious reasons, inhibitors of cholesterol biosynthesis have aroused intense interest. Studies on a variety of species (mostly rats) have been made using 1-alkylimidazoles," colchicine," and chenodeoxycholic acid, the last work being particularly interesting as the metabolite affected HMG-CoA reductase but not cholesterol 7a-hydroxylase, the steps believed to be rate-limiting for the biosynthesis of sterols and of bile acids respectively. Arsenite, /8-mercaptoethanol, dithiothreitol, and ethanethiol all inhibited the biosynthesis of cholesterol from MVA in rat liver homogenates. The accumulation of 4,4-dimethyl-5a-cholest-8-en-3/3-ol together with the corresponding A -diene supported the view that... 

The exact contributions of these alternate pathways to total hepatic bile acid synthesis in normal subjects is not certain, although 26-hydroxylation is usually regarded as the major pathway. In addition, it should be pointed out that current views of hepatic cholic acid and chenodeoxycholic acid synthesis are based primarily oh studies carried out in the rat. More recent studies, which have involved the administration of labeled bile acid intermediates to patients, have suggested that bile acid biosynthesis is more complex than previously thought and that multiple pathways exist to convert cholesterol to bile acids (Vll). 

Cholic acid differs from chenodeoxycholic acid in having an extra hydroxyl group at C-12. The enzyme responsible for producing this difference, 7a-hydroxy-4-cholesten-3-one 12a-hydroxylase, thus acts at a key branch point in the biosynthesis of bile acids and might be expected to be regulated in order to control the relative amounts of cholic acid and chenodeoxycholic acid produced. Like other hydroxylation steps in bile acid biosynthesis, 12a-hydroxylation requires a specific form of cytochrome P-450, which is present in the cytochrome P-45OLM4 fraction of rabbit liver microsomes (H6). The activity of I2a-hydroxylase has been postulated to be decreased in patients with liver cirrhosis to explain the low proportion of cholic add relative to chenodeoxycholic add in the bile of these patients (V9). Conversely, the activity of this enzyme may be high in patients with cerebrotendinous xanthomatosis, as the bile of these individuals contains mostly cholic acid... 

S2). More recent studies have shown that patients with cirrhosis are able to efficiently convert 7a-hydroxy-cholesterol into cholic acid (G8, P8), suggesting that 12a-hydroxylase activity is near normal. Other evidence from in vivo studies in man with labeled preciusors suggests that 12a-hydroxylase activity is not important in the regulation of the ratio between cholic acid and chenodeoxycholic add in human bile (B21). The possibility that different pools of cholesterol are utilized for the biosynthesis of cholic acid and chenodeoxycholic acid is now being investigated. 

Salen et al. reported that liver microsomes from 2 patients with CTX had a decreased capacity to 24/8-hydroxylate 5)8-cholestane-3a,7a,12a,25-tetrol [185]. It was suggested that the basic metabolic defect is a relative deficiency of the 24j8-hy-droxylase. To explain the severe metabolic consequences of such a defect, it must be assumed that the 25-hydroxylase pathway is the major pathway in the biosynthesis of cholic acid. This hypothesis does not explain the marked reduction in the biosynthesis of chenodeoxycholic acid. In view of the very low activity of the microsomal 25-hydroxylase towards 5)3-cholestane-3a,7a-diol in human hver [41] it is evident that a 25-hydroxylase pathway cannot be of importance in the normal... 

Oftebro et al. reported that the mitochondrial fraction of a liver homogenate from a biopsy of a CTX patient was completely devoid of 26-hydroxylase activity [193]. The possibility that there had been a general inactivation of the mitochondrial fraction seems excluded since there was a significant 25-hydroxylase activity towards vitamin D,. There was a substantial accumulation of 5 -cholestane-3a,7a,12a-triol, the immediate substrate for the 26-hydroxylase in cholic acid biosynthesis. It was suggested that the accumulation of 5)3-cholestane-3a,7a,12a-triol would lead to increased exposure to the action of the microsomal 23-, 24- and 25-hydroxylases. The alternative 25-hydroxylase pathway would then be of importance for the formation of cholic acid in patients with CTX (Fig. 13). If the 25-hydroxylase pathway has an insufficient capacity, this would explain the accumulation of the different 25-hydroxylated intermediates in patients with CTX. A lack of the mitochondrial 26-hydroxylase would also lead to accumulation of intermediates in chenodeoxycholic acid biosynthesis such as 5)8-cholestane-3a,7a-diol and 7a-hy-droxy-4-cholesten-3-one. Such accumulation would lead to increased exposure to the microsomal 12a-hydroxylase which would yield a relatively higher biosynthesis of cholic acid. This would explain the marked reduction in the biosynthesis of chenodeoxycholic acid in patients with CTX. 

Results of various in vivo experiments with labelled bile acid precursors in patients with CTX have been published [185,190,195]. All these experiments show that there is a defect in the oxidation of the steroid side chain in the biosynthesis of cholic acid but are not fully conclusive with respect to the site of defect. Bjorkhem et al. administered a mixture of [ H]7a,26-dihydroxy-4-cholesten-3-one and [ " C]7a-hy-droxy-4-cholesten-3-one to a patient with CTX [195]. The ratio between and C in the cholic acid and the chenodeoxycholic acid isolated was 40 and 60 times higher, respectively, than normal. Similar results were obtained after simultaneous administration of H-labelled 5)3-cholestane-3a,7a,26-triol and 4- C-labelled 5j8-cholestane-3a,7a-diol. The results of these experiments are in consonance with the contention that the basic defect in CTX is the lack of the 26-hydroxylase, but do not per se completely exclude other defects in the oxidation of the side chain. 

The regulation of the overall biosynthesis of bile acids has been studied intensively during the last decade, and only a small fraction of all the pubhcations can be reviewed here. Cholesterol 7a-hydroxylase is the rate-limiting enzyme in the biosynthesis of both chohc acid and chenodeoxycholic acid. The publications in which a correlation has been demonstrated between bile acid biosynthesis and 7a-hydroxyl-ation of cholesterol have been reviewed by Myant and Mitropoulos [59]. In the present review, emphasis will be put on the feedback regulation of the cholesterol 7a-hydroxylase by the bile-acid flux through the hver, the relation between HMG-CoA reductase and cholesterol 7 -hydroxylase and possible mechanisms for the regulation. 

Mitropoulos et al. have measured the rate of excretion and the specific activities of cholic acid and chenodeoxycholic acid in bile fistula rats fed [ H]cholesterol and infused with [ " C]mevalonate or [ C]7a-hydroxycholesterol [255]. It was concluded that newly synthesized hepatic cholesterol was the preferred substrate for the formation of cholic acid. It could not be excluded, however, that part of the chenodeoxycholic acid had been formed from a pool of cholesterol different from that utilized in cholic acid biosynthesis. The mitochondrial pathway, starting with a 26-hydroxylation, could have accounted for a significant fraction of the chenodeo-... 

Biochemical studies with isolated rat hepatocytes have largely been concerned with transport mechanisms [15], secretion of bile acids [17-19], or biosynthesis of bile acids [20]. The capacity of cultured hepatocytes to convert tauro- or glyco-chenodeoxycholate to a- and )8-muricholates [19,21] and to produce bile salts (glycine or taurine conjugates) during the dark phase of the diurnal cycle [21] has been established. Demonstrations of other metabolic transformations by hepatocytes are included in the following sections. 

Biosynthesis The primary B. are synthesized in the liver from cholesterol by a complicated, multi-step reaction sequence. The 7o-hydroxy group is introduced first while the 12-hydroxy group is added later to a further intermediate with subsequent formation of both chenodeoxycholic acid and cholic acid. Deoxycholic acid is not synthesized in the liver but rather in the intestines by 7a-dehydroxylation of cholic acid by intestinal bacteria. 

In the early studies of the metabolism of cholesterol and other C27-steroids in the presence of mitochondrial preparations from rat and mouse liver, several metabolites were isolated that could be intermediates in the biosynthesis of chenodeoxycholic acid (11). Cholesterol, 5-cholestene-3j, 7a-diol, 7a-hydroxy-4"Cholesten-3-one, and 5/5-cholestane-3a,7a-diol were found to be converted into the corresponding 26-hydroxy derivatives by mitochondrial preparations. In addition, these preparations were shown to catalyze the 7a-hydroxylation of 5-cholestene-3/9,26-diol and the oxidation of 5-cholestene-3i, 7a-diol into 7a-hydroxy-4-cholesten-3-one. All the 26-hydroxy compounds were found to be transformed predominantly into chenodeoxy-... 

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