| Research Abstract |
1) We have established several focal adhesion kinase (FAK)-transfected HL-60 (HL-60/FAK) cells which become resistant to oxidative stress-induced apoptosis. We observed that HL-60/FAK cells proliferate much faster than vector-transfected (HL-60/Vect) cells. This observation prompted us to investigate the mechanism how HL-60/FAK cells augment cell proliferation. Since a PKC inhibitor, chelerythrine or a PI3-kinase inhibitor, LY 294002 suppressed cell proliferation effectively, both PKC and PI-3-kinase pathways are presumed to be involved in the cell proliferation. Since cyclin D3 expression was particularly prominent and PKCα, β, and η isoforms were activated and directly associated with FAK in HL-60/FAK cells. We thus assumed that FAK activates PKC and PI3-kinase-Akt pathway, which resulted in marked induction of cyclin D3 expression and CDK activity.
2) We performed cDNA microarray screening using cytokine-chemokine and apoptosis-chip to identify responsible molecules. We found that glutathione peroxidase (GPx) mRNA was decreased and lipid peroxidation was suppressed after treatment with H_2O_2 in HL-60/FAK cells. In addition, HL-60/FAK cells have higher basal ROS levels. Basal activity and mRNA expression of GSH reductase (GRe), phospholipid hydroperoxide glutathione peroxidase (PHGPx) were markedly elevated in HL-60/FAK cells. In contrast, GPx and catalase levels were decreased in HL-60/FAK cells. Thus, we suggested that FAK upregulates antioxidant enzymes and suppresses lipid peroxidation, resulting in the anti-apoptotic state for oxidative stress.
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