3-7-dihydroxycholestan-26-oic-acid and Liver-Neoplasms

We have examined the ability of HepG2 human hepatoblastoma cells and 7800 C1 Morris rat hepatoma cells to convert 3 alpha, 7 alpha, 12 alpha-trihydroxy-5 beta-cholestanoic acid (THCA) and 3 alpha, 7 alpha-dihydroxy-5 beta-cholestanoic acid (DHCA) to cholic acid and chenodeoxycholic acid, respectively. Cell extracts from both these cell lines could neither form cholic acid from THCA nor from the activated form, THCA-CoA. This suggests that both cell lines are defective in two enzyme activities involved in the pathway, the microsomal THCA-CoA ligase and the peroxisomal THCA-CoA oxidase. Furthermore, we show that the subsequent enzymes are active in the conversion to bile acids, because the product of the THCA-CoA oxidase, 3 alpha, 7 alpha, 12 alpha-trihydroxy-5 beta-cholest-24-enoyl-coenzyme A (delta 24-THCA-CoA) or delta 24-THCA in the presence of THCA-CoA ligase, are converted to cholic acid by both cell lines. HepG2 cells were able to slowly form chenodeoxycholic acid and cholic acid from 5 beta-cholestane-3 alpha, 7 alpha-diol and 5 beta-cholestane-3 alpha, 7 alpha, 12 alpha-triol, respectively, in 24- and 96-h incubations. The rate of cholic acid formation was lower than the rate for chenodeoxycholic acid and there was a clear accumulation of THCA. 7800 C1 Morris cells had no ability to form cholic acid or chenodeoxycholic acid after 96 h incubation. We conclude that these two cell lines have defects in two enzyme activities involved in the peroxisomal oxidation in bile acid formation, the microsomal THCA-CoA ligase and the peroxisomal THCA-CoA oxidase.

Topics: Animals; Bile Acids and Salts; Chenodeoxycholic Acid; Cholestanols; Cholesterol 7-alpha-Hydroxylase; Cholic Acid; Cholic Acids; Clofibric Acid; Dexamethasone; Fatty Acids; Hepatoblastoma; Humans; Liver Neoplasms; Liver Neoplasms, Experimental; Male; Oxidation-Reduction; Rats; Rats, Wistar; Retinoids; Tumor Cells, Cultured

We used capillary gas chromatography/mass spectrometry to demonstrate that a cell line derived from a well differentiated human hepatoblastoma, HepG2, synthesized and secreted the following bile acids (ng/10(7) cells/h): chenodeoxycholic acid (131.4), cholic acid (3.3), 3 alpha, 7 alpha-dihydroxy-5 beta-cholestan-26-oic acid (DHCA; 4.5), and 3 alpha, 7 alpha, 12 alpha-trihydroxy-5 beta-cholestan-26-oic acid (THCA; 32.0). Deuterium from [7 beta-2H]7 alpha-hydroxycholesterol, which was added to the media, was incorporated into newly synthesized chenodeoxycholic acid, DHCA, and THCA, but not into cholic acid. Since THCA is a known precursor of cholic acid, these data suggest that HepG2 is specifically deficient in the side chain cleavage that transforms THCA into cholic acid. Greater than 90% of the bile acids synthesized and secreted by HepG2 were unconjugated. Conjugation could not be stimulated by the addition of glycine or taurine to the media. Approximately 30% of newly synthesized DHCA and THCA were sulfated. Chenodeoxycholic acid and cholic acid were not appreciably sulfated. In summary, cultured HepG2 cells synthesize bile acid, but in a pattern distinct from that of adult human liver. This cell line may be a model for studying pathways of human bile acid synthesis, conjugation, and sulfation.

Topics: Bile Acids and Salts; Carcinoma, Hepatocellular; Cell Line; Chenodeoxycholic Acid; Cholestanols; Cholic Acid; Cholic Acids; Gas Chromatography-Mass Spectrometry; Humans; Liver Neoplasms