Suppression of GST-P by treatment with glutathione-doxorubicin conjugate induces potent apoptosis in rat hepatoma cells.

A conjugate of doxorubicin and glutathione via glutaraldehyde (GSH-DXR) inhibited glutathione S-transferase (GST) activity of rat hepatoma AH66 cells, and treatment of the cells with GSH-DXR induced caspase-3 activation and DNA fragmentation. After treatment of AH66 cells with 0.1 microM GSH-DXR, GST-P (placental type of rat GST isozymes) mRNA and its protein increased transiently and then decreased thereafter compared with the levels in nontreated cells. Caspase-3 activation and DNA fragmentation were induced following the suppression of GST-P expression by treatment with GSH-DXR. When the cells were treated with 100 microM ethacrynic acid (ECA), an inhibitor of GST, DNA fragmentation and caspase-3 activation were observed. In contrast, treatment of AH66 cells with a low concentration of ECA (1 microM) that showed little inhibition of GST activity induced slight, but significantly enhanced expression and activity of GST-P, and consequent prevention of DXR- and GSH-DXR-induced DNA fragmentation. Overexpression of GST-pi (placental type of human GST isozymes) by transfection of GST-pi sense cDNA into AH66 cells decreased sensitivities to DXR and GSH-DXR, and the suppression of GST-P by transfection of the antisense cDNA into the cells increased drug sensitivity. On the other hand, there was little change in drug sensitivity caused by overexpression of site-directedly mutated GST-P in which the active-site residue Tyr39 was replaced with His (W39H) or the substrate-binding site residue Cys48 was replaced with Ser (C48S) by transfection of those cDNAs into AH66 cells. These results suggested that the suppression of GST-P in AH66 cells treated with GSH-DXR must play an important role in the induction of apoptosis.

Association of polymorphism in the alcohol dehydrogenase 2 gene with alcohol-induced testicular atrophy.

BACKGROUND: Alcohol abuse can induce testicular atrophy, but it only occurs in some alcoholics. Alcohol dehydrogenase (ADH) is located principally on the Leydig cells. METHODS: To investigate whether genetic polymorphism of alcohol dehydrogenase (ADH) 2 and aldehyde dehydrogenase (ALDH) 2 was related to alcoholic testicular atrophy, we determined restriction fragment-length polymorphisms of the ADH2 and ALDH2 genes in 43 Japanese male alcoholics and 50 healthy subjects. An orchidometer was used to determine the testicular size. RESULTS: Less than 16 ml in testicular size was defined as testicular atrophy. Testicular atrophy was found in 24 (55.8%) cases out of 43 alcoholics. Digestion with MaeIII and MboII after polymerase chain reaction amplification showed that the ADH21 allele frequency was significantly higher in patients with testicular atrophy than in those without testicular atrophy (chi2 = 4.665, p = 0.031), whereas no significant association was observed between testicular atrophy and the ALDH2 gene. CONCLUSIONS: The ADH21 allele may be associated with alcoholic testicular atrophy.

Association of restriction fragment-length polymorphisms in the alcohol dehydrogenase 2 gene with alcoholic brain atrophy.

Alcohol abuse can induce brain atrophy, but it only occurs in some alcoholics. To investigate whether genetic polymorphism of alcohol-metabolizing enzymes [including alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH)] was related to alcoholic brain atrophy, we determined restriction fragment-length polymorphisms of the ADH2 and ALDH2 genes in 77 male alcoholics. Computed tomography was used to determine the severity of brain atrophy. Digestion with MaeIII and MboII after polymerase chain reaction amplification showed that the ADH2(1) gene frequency was significantly higher in patients with brain atrophy than in those without brain atrophy (chi 2 = 9.274, p < 0.01), whereas no significant association was observed between brain atrophy and the ALDH2 gene Multivariate analysis (including age, total alcohol intake, liver cirrhosis, and ADH2 genotype) showed that the ADH2(1)/ADH2(1) genotype was associated with alcoholic brain atrophy. These findings suggest that the ADH2(1) allele may be associated with alcoholic brain atrophy.