title: Identification of the insulin signaling cascade in the regulation of alpha-class glutathione S-transferase expression in primary cultured rat hepatocytes.
abstract: We reported previously that insulin elevated alpha-class glutathione S-transferase (GSTs) protein levels in primary cultured rat hepatocytes (Kim et al., 2003b). In contrast, glucagon down-regulated alpha- and pi-class GST expression, and mechanistic research implicated cAMP and protein kinase A in this process (Kim et al., 2003b). The present study examines the signaling pathways involved in the regulation of alpha-class GST in response to insulin in primary cultured rat hepatocytes. Protein levels of GSTA1/2 and GSTA3/5 and activity of GST toward 7-chloro-4-nitrobenzo-2-oxa-1,3-diazole (NBD) were increased in an insulin concentration-dependent manner. Treatment of cells with the phosphatidylinositol 3-kinase (PI3K) inhibitors wortmannin and LY294002 [2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one] or rapamycin, an inhibitor of mammalian target of rapamycin and ribosomal p70 S6 kinase (p70S6K) phosphorylation, or with an adenovirus containing green fluorescent protein and a dominant-negative and kinase-dead Akt, effectively inhibited the insulin-mediated increase in alpha-class GST expression and GST activity toward NBD. In contrast, PD98059 (2'-amino-3'-methoxyflavone), an inhibitor of mitogen-activated protein kinase kinase, SP600125 (1,9-pyrazoloanthrone), an inhibitor of c-Jun N-terminal kinase, SB203580 [4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl)1H-imadazole], an inhibitor of p38 mitogen-activated protein kinase, or bisindolylmaleimide, a broad spectrum inhibitor of protein kinase C, did not inhibit the insulin-mediated increase in alpha-class GST protein levels in hepatocytes. These results show that PI3K/Akt/p70S6K signaling is active in the insulin-mediated up-regulation of the antioxidant defense system and that low insulin levels, as encountered in diabetes, potentially increase the susceptibility of hepatocytes to xenobiotic-mediated and/or oxidative stress-mediated damage.
authors.affiliation: Institute of Environmental Health Sciences, Wayne State University, 2727 South Avenue, Room 4000, Detroit, MI 48201, USA.
authors.forename: Sang K
authors.initials: SK
authors.lastname: Kim
authors.forename: Mohamed A
authors.initials: MA
authors.lastname: Abdelmegeed
authors.forename: Raymond F
authors.initials: RF
authors.lastname: Novak
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D004851:Epoxide Hydrolases / Q000096:biosynthesis; D005982:Glutathione Transferase / Q000096:biosynthesis*; D022781:Hepatocytes / Q000201:enzymology*; D007328:Insulin / Q000494:pharmacology*; D008297:Male; D019869:Phosphatidylinositol 3-Kinases / Q000502:physiology; D011493:Protein Kinase C / Q000502:physiology; D051057:Proto-Oncogene Proteins c-akt / Q000502:physiology; D051381:Rats; D017207:Rats, Sprague-Dawley; D038762:Ribosomal Protein S6 Kinases, 70-kDa / Q000502:physiology; D015398:Signal Transduction / Q000502:physiology*
chemical_list: D007328:Insulin; D005982:Glutathione Transferase; D019869:Phosphatidylinositol 3-Kinases; D051057:Proto-Oncogene Proteins c-akt; D038762:Ribosomal Protein S6 Kinases, 70-kDa; D011493:Protein Kinase C; D004851:Epoxide Hydrolases