Cytotoxic effect of 7?-hydroxy-4-cholesten-3-one on HepG2 cells: Hypothetical role of acetaldehyde-modified ?4-3-ketosteroid-5?-reductase (the 37-kd-liver protein) in the pathogenesis of alcoholic liver injury in the rat

Abstract

We recently identified ⌬ 4 -3-ketosteroid-5␤-reductase as the 37 kd liver protein which is highly susceptible to acetaldehyde modification in rats continuously fed alcohol. The 5␤-reductase is a key enzyme involved in bile acid synthesis. We report here that the ability to degrade 7␣-hydroxy-4-cholesten-3-one (HCO) was lower in the liver cytosol of alcohol-fed rats than in control animals, suggesting an inhibition of the 5␤-reductase enzyme activity by acetaldehyde modification. We also showed that HCO exhibited a time-and concentration-dependent cytotoxicity to HepG2 cells. HCO cytotoxicity was noticeable at a concentration of 2.5 g/mL. When 10 g/mL of HCO was added to confluent cell monolayers, 57% and 37% of cells remained viable after 24 and 48 hours of treatment. The decrease in cell viability was accompanied by an increased lactic dehydrogenase activity in the culture medium. DNA extracted from HCO-treated cells showed no evidence of DNA fragmentation when analyzed by agarose gel electrophoresis. Staining with propidium iodide showed no nuclear condensation in cells. Thus, cell death by HCO treatment was caused by necrosis and not by apoptosis. Various agents, including, serum proteins, hormones, bile acids, antioxidants, Ca ϩϩ -chelators, Fe ϩϩ -chelator, CYP450 inhibitor, adenylate cyclase inhibitor, protease inhibitors, and nitric oxide synthase inhibitor, did not protect against HCO cytotoxicity. We speculate that HCO concentrations may be elevated around the pericentral area in the liver after chronic alcohol ingestion, causing local cell necrosis. The release of cellular contents and protein-acetaldehyde adducts (PAAs) may activate nonparenchymal cells and provoke autoimmune reaction. Thus, the formation of the 37 kd-PAA may play an important role in the initiation of alcoholic liver injury. (HEPATOLOGY 1998;27:100-107.)Alcohol is metabolized in the liver mainly by alcohol dehydrogenase (ADH). 1 With prolonged alcohol consumption, the metabolic pathway through cytochrome P450IIE1 becomes important. 2 Both ADH and CYPIIE1 generate acetaldehyde from ethanol.

Ethanol Induces Fatty Acid Synthesis Pathways by Activation of Sterol Regulatory Element-binding Protein (SREBP)

“…Thus, one effect of acetaldehyde could be to reduce the concentration of sterols that influence SCAP activity. One enzyme that metabolizes sterols is known to be inhibited by acetaldehyde, namely ⌬ 4 -3-ketosteroid 5-␤-reductase, an enzyme participating in the bile salt synthetic pathway (37)(38)(39)(40). It is not known if other enzymes that participate in sterol metabolism aside from HMG-CoA reductase are affected by ethanol or acetaldehyde.…”

Androgen metabolism and biotransformation in nontumoral and malignant human liver tissues and cells

“…It is important to note that the 5␤-reductase enzyme can negatively regulate the intracellular concentrations of other bioactive steroid hormones, including progesterone and, to a lesser extent, mineralcorticoids and glucocorticoids [22]. The significant decrease of 5␤-reductase activity observed in alcoholic cirrhosis and HCC tissues may also result in liver injury due to the accumulation the cytotoxic intermediate metabolite 7-␣-hydroxy-4-cholesten-3-one in bile acid synthesis [23].…”

The Dose-Response Effects of Ethanol on the Human Fetal Osteoblastic Cell Line

“…This finding is significant because the metabolites of ethanol may be more toxic than the parent compound. ( 31,32 )…”