Formation of β-hydroxyisovalerate by an α-ketoisocaproate oxygenase in human liver

Previously we demonstrated the occurrence of a soluble dioxygenase in rat liver which converts α-ketoisocaproic acid (the keto acid analog of leucine) to β-hydroxyisovaleric acid. Herein we show that human liver contains a similar soluble enzyme which converts α-ketoisocaproate to β-hydroxisovaleric acid. We suggest this enzyme functions as a “safety valve” in liver to help prevent excessive accumulation of α-ketoisocaproate.     

Formation of 5alpha-androstane-3alpha,17beta-diol by normal and hypertrophic human prostate.

The 3-keto reduction of [1,2-3H]dihydrotestosterone to 3alpha- and 3beta-androstanediols was assessed in homogenates of 40 prostates obtained either at surgery or at medicolegal autopsy from men who had died suddenly. Formation of both androstanediols was demonstrable in cytosol and in microsomes, and both NADH and NADPH were effective cofactors for the two reactions. Formation of the two steroids was not influenced by storage of the gland for up to 8 h prior to processing. When NADPH was cofactor, the formation of 3alpha- and 3beta-androstanediol was significantly higher in microsomes and cytosol from hypertrophic than from normal glands.     

Formation of bile acid glucosides by a sugar nucleotide-independent glucosyltransferase isolated from human liver microsomes.

A heat-labile protein has been detected in microsomes from human liver which catalyzes the formation of glucosides of the bile acids chenodeoxycholic, deoxycholic, and ursodeoxycholic acids. This glucosyltransferase activity has been purified about 900-fold from human liver microsomes, resulting in homogeneity as determined by sodium dodecyl sulfate gel electrophoresis. The subunit molecular weight was calculated to be about 56,000. The enzyme was separated from bile acid UDP-glucuronosyltransferase [UDP-glucuronate beta-D-glucuronosyltransferase (acceptor-unspecific), EC 2.4.1.17] during purification and does not catalyze the formation of bile acid glucuronides. The purified glucosyltransferase utilizes lipophilic alkyl beta-D-glucopyranosides as artificial donor substrates and dolichyl phosphoglucose as natural donor for the transfer of glucose to bile acids and does not exhibit bile acid conjugating activity in the presence of sugar nucleotides such as UDP-glucose. The apparent Km values estimated for various alkyl beta-D-glucopyranosides decreased with increasing alkyl chain length from 680 X 10(-6) M for hexyl beta-D-glucopyranoside to 20 X 10(-6) M as estimated for decyl and dodecyl beta-D-glucopyranoside. The results suggest that a glucoside-conjugation pathway of bile acids exists in humans. This conjugation is catalyzed by a sugar nucleotide-independent glucosyltransferase and is therefore distinct from the known mechanisms of glycoside conjugation.     

17Beta-hydroxysteroid dehydrogenase type 2 and dehydroepiandrosterone sulfotransferase in the human liver

The liver plays important roles in the clearance and metabolism of sex steroids. Its dysfunction is considered to influence the metabolic pathways of sex steroids, and to result in gynecomastia and other abnormalities of sex steroids. However, the details of its mechanism have not been well-characterized. We therefore examined the enzymes involved in the hepatic clearance and/or metabolism of sex steroids in human liver and its disorders using immunohistochemistry to determine whether there are any abnormalities of expression of these enzymes in human liver disorders. These enzymes are 17beta-hydroxysteroid dehydrogenase (17beta-HSD) type 2, an enzyme that catalyzes the biologically active estrogen, estradiol (E2), to inactive estrogen, estrone (El), and dehydroepiandrosterone sulfotransferase (DHEA-ST), which catalyzes sulfonation of dehydroepiandrosterone (DHEA) to form biologically inactive DHEA-S. A total of 162 cases including normal liver (n=31), chronic hepatitis (n=41), liver cirrhosis (n = 21), hepatocellular carcinoma (n = 47), cholangiocellular carcinoma (n = 22) and fetal liver (n = 4) were examined by immunohistochemistry. Both enzymes were expressed in the hepatocytes around portal area and central vein in normal liver. Immunopositive area for DHEA-ST was significantly larger in chronic hepatitis than in normal liver, but that of 17beta-HSD type 2 in chronic hepatitis was not different from normal liver. There were no significant differences in the immunopositive area for both enzymes between liver cirrhosis and normal liver. In hepatocellular carcinoma, immunoreactivity for both enzymes were categorized into Group A, or low positive group, and Group B, or high positive group. The latter tended to be poorly differentiated carcinoma. In cholangiocellular carcinoma, immunopositive areas of both enzymes were significantly smaller than those of normal liver. These findings indicate that the amount of expression of the enzymes involved in metabolism and/or clearance of sex steroids per hepatocyte did not decrease in liver cirrhosis. Therefore, sex steroids' abnormalities may be due to the decreased quantity of hepatocytes associated with liver cirrhosis. In hepatocellular carcinoma, some poorly differentiated cases were associated with increased expression of 17beta-HSD type 2 but its biological significance needs to be determined by further studies.     

The effects of exercise on lipogenic enzyme activity and glyceride synthesis by liver homogenates of diabetic rats

The purpose of this study was to determine if exercise ameliorates the elevated levels of triglycerides in diabetic rats and also to determine if the capacity of liver to synthesize glycerides correlates with changes in plasma triglyceride levels. Forty female rats were divided into four groups: sedentary control, sedentary diabetic, exercised control, and exercised diabetic. Diabetes was induced by intravenous injection of alloxan (40 mg/kg), and control rats were sham-dosed with physiologic saline. All rats remained sedentary in their cages for the first week after the injections. The exercised groups were exercised for seven consecutive days for 2 h/d at 20 m/min (0 grade). All rats were killed 24 hours after the last exercise bout. Blood glucose levels were significantly higher in the diabetic group than the nondiabetic counterparts, but exercise did not affect glucose levels in either the diabetic or nondiabetic groups. Exercise, however, significantly lowered plasma triglyceride and free fatty acid levels in both diabetic and nondiabetic rats. The activities of the five enzymes involved in fatty acid synthesis were all depressed in the diabetic rats compared to controls, and exercise had no effect on the activities of the enzymes. The capacity of liver to synthesize total lipids, diglycerides, or triglycerides was not different in the diabetic rats from that of nondiabetic rats, and exercise did not change that. Only phospholipid synthesis from glycerol-3-phosphate was affected by diabetes. It is concluded that exercise ameliorates the elevations in plasma triglyceride levels that result from diabetes.(ABSTRACT TRUNCATED AT 250 WORDS)     

Formation of human hepatocytes by human hematopoietic stem cells in sheep

We took advantage of the proliferative and permissive environment of the developing preimmune fetus to develop a noninjury large animal model in sheep, in which the transplantation of defined populations of human hematopoietic stem cells resulted in the establishment of human hematopoiesis and led to the formation of significant numbers of long-lasting, functional human liver cells, with some animals exhibiting levels as high as 20% of donor (human) hepatocytes 11 months after transplantation. A direct correlation was found between hepatocyte activity and phenotype of transplanted cells, cell dose administered, source of cells used on a cell-per-cell basis (bone marrow, cord blood, mobilized peripheral blood), and time after transplantation. Human hepatocytes generated in this model retained functional properties of normal hepatocytes, constituted hepatic functional units with the presence of human endothelial and biliary duct cells, and secreted human albumin that was detected in circulation. Transplanting populations of hematopoietic stem cells can efficiently generate significant numbers of functional hepatic cells in this noninjury large animal model and thus could be a means of ameliorating or curing genetic diseases in which a deficiency of liver cells or their products threatens the life of the fetus or newborn.     

Deiodination of thyroid hormone by human liver

Liver is an important site for the peripheral production of T3 by outer ring deiodination (ORD) of T4 as well as for the clearance of plasma rT3, which is produced by inner ring deiodination (IRD) of T4 in other tissues. However, little is known about the underlying enzymatic reactions, and current concepts about thyroid hormone deiodination are largely based on studies in rat tissue. Here we describe the results of detailed studies of the catalytic properties of the iodothyronine deiodinase activity of human liver. The results demonstrated a high degree of similarity with the type I deiodinase of rat liver. The enzyme activity was found in the microsomal fraction. rT3 was the preferred substrate, since its ORD was catalyzed roughly 400 times more efficiently than the ORD or IRD of T4 or the IRD of T3. The deiodination of sulfated substrates was more rapid, as demonstrated by the roughly 30-fold increase in the IRD of T3 sulfate (T3S) compared with T3. The deiodinations exhibited ping-pong-type kinetics with dithiothreitol as the cofactor. Inhibition by propylthiouracil was uncompetitive with substrate and competitive with dithiothreitol, and PTU was an equally effective inhibitor of the ORD of rT3 and the IRD of T3S (Ki, 0.10-0.16 mumol/L). Various compounds with widely different inhibitory potencies had similar effects on ORD (rT3) and IRD (T3S). These results suggest that in human liver microsomes a single enzyme catalyzes the deiodination of the outer as well as the inner ring of iodothyronines by the same catalytic mechanism and with the same substrate specificity as the type I deiodinase of rat liver.