Sverre Skrede

Cerebrotendinous xanthomatosis: a defect in mitochondrial 26-hydroxylation required for normal biosynthesis of cholic acid.

Oxidation of side chain of 5 beta-cholestane-3 alpha,7 alpha,12 alpha-triol was studied in a patient with cerebrotendinous xanthomatosis (CTX) and in control subjects, using various subcellular fractions of liver homogenate and a method based on isotope dilution-mass spectrometry. In the control, 5 beta-cholestane-3 alpha,7 alpha,12 alpha-triol was converted into 5 beta-cholestane-3 alpha,7 alpha,12 alpha,26-tetrol and 3 alpha,7 alpha,12 alpha-trihydroxy-5 beta-cholestanoic acid by the mitochondrial fraction, and into 5 beta-cholestane-3 alpha,7 alpha,12 alpha,-25-tetrol by the microsomal fraction. In the CTX patient, liver mitochondria were completely devoid of 26-hydroxylase activity. The same mitochondrial fraction catalyzed 25-hydroxylation of vitamin D3. The microsomal fraction of liver of the subject with CTX contained more than 50-fold the normal amount of 5 beta-cholestane-3 alpha,7 alpha,12 alpha-triol. The basic metabolid defect in CTX appears to be a lack of the mitochondrial 26-hydroxylase. The excretion in the bile of 5 beta-cholestane-3 alpha,7 alpha,12 alpha,25-tetrol and 5 beta-cholestane-3 alpha,7 alpha,12 alpha,24 alpha,25-pentol observed in CTX patients may be secondary to the accumulation of the major substrate for the 26-hydroxylase, i. e., 5 beta-cholestane-3 alpha,7 alpha,12 alpha-triol, and exposure of this substrate to the normally less active microsomal 25-and 24-hydroxylases. It is concluded that the major pathway in the biosynthesis of cholic acid in human liver involves a mitochondrial C27-steroid 26-hydroxylation.

Demonstration of 26-hydroxylation of C27-steroids in human skin fibroblasts, and a deficiency of this activity in cerebrotendinous xanthomatosis.

26-Hydroxylation of 5 beta-cholestane-3 alpha, 7 alpha, 12 alpha-triol and other C27-steroids was demonstrated in cultured skin fibroblasts from healthy individuals. Activities in skin fibroblasts were approximately 5-10% of those previously found in human liver homogenates, and were inhibited by CO. The apparent Km was lowest for 5 beta-cholestane-3 alpha, 7 alpha, 12 alpha-triol (1.3 mumol/liter) and highest for 5-cholestene-3 beta, 7 alpha-diol (12 mumol/liter). The rate of 26-hydroxylation was highest with 7 alpha-hydroxy-4-cholesten-3-one. These characteristics are similar to those of hepatic mitochondrial C27-steroid 26-hydroxylase. In skin fibroblasts from three patients with cerebrotendinous xanthomatosis (CTX), 26-hydroxylation of C27-steroids proceeded at a rate of only 0.2-2.5% of healthy controls. No accumulation of endogenous 5 beta-cholestane-3 alpha, 7 alpha, 12 alpha-triol could be demonstrated in these cells, and the lowered formation of radioactive, 26-hydroxylated products could not be explained by dilution of the labeled exogenous substrate. The present results add strong evidence to the concept that the primary metabolic defect in CTX is a deficiency of C27-steroid 26-hydroxylase.

Serum selenium levels in liver diseases

A possible pathogenetic role of selenium deficiency in alcoholic cirrhosis of the liver has previously been discussed. In the present study serum selenium was analyzed in 5 groups of liver diseases. The method used for selenium determination was electrothermal atomic absorption, after thermal stabilization of selenium compounds by addition of nickel nitrate. The selenium level of a reference group of healthy Norwegian adults (n = 40) was 1.53 +/- 0.25 mumol/l. The serum concentrations of selenium in patients suffering from alcoholic cirrhosis, chronic active hepatitis and chronic persistent hepatitis were lowered to 40-80 per cent of those of the reference group. In alcoholic cirrhosis and chronic active hepatitis the decreased serum selenium concentrations were significantly correlated to decreased levels of albumin and prealbumin.

Diagnosis of Liver Diseases by Laboratory Results and Discriminant Analysis Identification of Best Combinations of Laboratory Tests

Patients with different liver diseases were studied by discriminant analysis. Groups of patients classified mainly on the basis of liver biopsy findings showed functional differences which permitted a consistent reclassification by discriminant functions using laboratory results. Optimal combinations of laboratory tests for the separation of liver diseases were defined. Different combinations were found, dependent on the subsets of liver diseases studied.

A novel pathway for biosynthesis of cholestanol with 7 alpha-hydroxylated C27-steroids as intermediates, and its importance for the accumulation of cholestanol in cerebrotendinous xanthomatosis.

A mixture of 7 alpha-3H- and 4-14C-labeled cholesterol was administered intravenously to rats. Cholestanol with 20-30% lower ratio between 3H and 14C than in cholesterol could be isolated from different organs. In a healthy human control, cholestanol isolated from feces had a 3H/14C ratio which was 28% lower than in administered cholesterol. Cholesterol and coprostanol reisolated in these experiments had the same ratio between 3H and 14C as in the precursor. A previously unknown pathway for formation of cholestanol, involving 7 alpha-hydroxylated intermediates, may explain these results. Under normal conditions, this pathway is responsible for at most 30% of the cholestanol synthesized from cholesterol. Intravenous administration of the 7 alpha-3H- and 4-14C-labeled cholesterol to a patient with cerebrotendinous xanthomatosis (CTX) resulted in formation of cholestanol which had 70-75% lower 3H/14C ratio. It is concluded that the novel pathway involving 7 alpha-hydroxylated intermediates is accelerated in patients with CTX. This acceleration may contribute essentially to the accumulation of cholestanol, which is a predominant feature of this disease. 7 alpha-Hydroxycholesterol and 7 alpha-hydroxy-4-cholesten-3-one might be intermediates in the novel pathway to cholestanol. After intravenous administration of 7 beta-3H-labeled 7 alpha-hydroxycholesterol in a patient with CTX, significant amounts of 3H were incorporated into plasma and fecal cholestanol. Only small amounts of 7 alpha-hydroxycholesterol and 7 alpha-hydroxy-4-cholesten-3-one are excreted into the intestine, and we therefore conclude that the 7 alpha-dehydroxylation step mainly occurs in the liver. In CTX, the synthesis of cholestanol may be accelerated because the concentrations of 7 alpha-hydroxylated bile acid intermediates in the liver are increased. A possible mechanism for the conversion of a minor fraction of 7 alpha-hydroxycholesterol into cholestanol is suggested.

Fast atom bombardment mass spectrometry in the diagnosis of cerebrotendinous xanthomatosis

Urine from a patient with cerebrotendinous xanthomatosis (CTX) was extracted with a Sep-Pak C18 cartridge and the extract was analysed by fast atom bombardment mass spectrometry. The spectra indicated the presence of glucuronidated bile alcohols with four to seven hydroxyl groups. The method is simple and rapid and is suggested as an aid in the diagnosis of CTX with possible application to prenatal diagnosis.

Role of the 26-hydroxylase in the biosynthesis of bile acids in the normal state and in cerebrotendinous xanthomatosis. An in vivo study.

On the basis of different in vitro studies, we have previously suggested that the basic metabolic defect in the rare inherited disease cerebrotendinous xanthomatosis (CTX) is a lack of a hepatic mitochondrial C27-steroid 26-hydroxylase, involved in the normal biosynthesis of bile acids (1980. J. Clin. Invest. 65: 1418-1430; 1981. J. Lipid Res. 22: 191-200; 22: 632-640). In the present work, this hypothesis was tested in vivo. One patient with CTX and two control subjects received intravenously a mixture of [4-14C]7 alpha-hydroxy-4-cholesten-3-one and [6 beta-3H]7 alpha,26-dihydroxy-4-cholesten-3-one, steroids believed to be important precursors of chenodeoxycholic acid. The ratio between 14C and 3H in cholic acid and chenodeoxycholic acid isolated from bile of the CTX-patient was approximately 1/40 and 1/60 of those of the control subjects, respectively. Another patient with CTX and one control subject received a mixture of [4-14C]5 beta-cholestane-3 alpha,7 alpha-diol and [1,2-3H]5 beta-cholestane-3 alpha,7 alpha,26-triol, both possible precursors to chenodeoxycholic acid. In this case the 14C/3H ratio in cholic acid and chenodeoxycholic acid from the patient with CTX was 1/10 and 1/15, respectively, compared with that of the control subject. The most likely explanation for these findings is that very little of the 14C-precursors, i.e. without a 26-hydroxyl group, can be converted into cholic acid and chenodeoxycholic acid because of a defect of the 26-hydroxylase step. The results obtained are in accord with our previous findings in vitro. The results further underline the importance of the 26-hydroxylase pathway in the normal biosynthesis of cholic acid and chenodeoxycholic acid in man.

Effects of clofibrate on the intracellular localization of palmitoyl-CoA hydrolase and palmitoyl-L-carnitine hydrolase in rat liver

Feeding clotibrate to rats will induce a prominent increase in the number of peroxisomes [ 1 ] and of the capacity for fatty-acid oxidation [2,3]. Clofibrate feeding also results in a great increase in the hepatic content of free CoA and long-chain acyl- CoA [4--61 and an increase of the acylCoA hydrolase activity (71. We now report that clofibrate treatment of rats results in an increase of the activity of both palmitoyl- Lcarnitine hydrolase and palmitoylCoA hydrolase. Also a subcellular redistribution of the latter enzyme is found, possibly due to release from structurally changed microsomal membranes (endoplasmatic retic- ulum) to the particle-free supernatant. 2.

Lecithin: Cholesterol acyltransferase and plasma proteins in liver disease☆

Abstract Lecithin: cholesterol acyltransferase (LCAT) activity has been measured by the method of Stokke and Norum and by the method of Glomset and Wright in plasma from 53 patients with various liver diseases. The results were compared with the concentration of plasma albumin, prealbumin and α-lipoprotein and with Normotest as well as plasma cholesterol. In most cases, the method of Stokke and Norum and the method of Glomset and Wright gave the same results. Therefore, substrate deficiency probably does not contribute much to low activities obtained by the method of Stokke and Norum in liver disease. The activity of LCAT was found to parallel the levels of plasma proteins. In acute hepatitis LCAT activity followed the proteins with short half-life. In chronic liver disease the LCAT activity was also correlated with plasma albumin. LCAT activity was normal in some patients with primary biliary cirrhosis who had subnormal levels of prealbumin, but normal or high levels of α-lipoprotein. The previously reported stimulating effect on LCAT activity with inactivated substrates from patients with primary biliary cirrhosis may be caused by high concentration of α-lipoprotein, either as substrate or as LCAT cofactor. In the total material the activity of LCAT was not correlated with the concentration of free cholesterol, but was correlated with the levels of cholesteryl esters.

Serum immunoglobulins and organ non-specific antibodies in diseases of the liver.

Serum immunoglobulins and C3 levels, auto-antibodies to smooth muscle (SMA), mitochondria (MA), and nuclei (ANA), rheumatoid factors (RF), HB-antigen and HB-antibody were studied in 9 groups of liver disease. Hypergammaglobulinaemia was a prominent feature in most groups, IgG being particularly raised in active chronic hepatitis, IgM in primary biliary cirrhosis, and IgA in alcoholic liver disease, respectively. IgE was often increased in alcoholic liver disease and was frequently low in hepatic tumours, whereas IgD showed no typical pattern in any liver disorder. SMA was most frequently found in active chronic hepatitis (68%), and MA in primary biliary cirrhosis (58%), while ANA was detected in 50% of the patients with active chronic hepatitis. However, a pronounced over-lap of tissue antibodies was observed among the various groups of liver disease, particularly in active chronic hepatitis and primary biliary cirrhosis. The concurrent presence of SMA and ANA was most frequent in active chronic hepatitis. It was not excluded that antibody titres might have provided better diagnostic discrimination, since titration of antibodies was not performed. Low C3 levels in active chronic hepatitis were correlated with low levels of other liver-synthetized proteins, and no evidence was found of increased consumption by immunologic reactions.