Suppression of Antiapoptotic Bcl-2 Expression in Human Bladder Tumor Cells Exposed to Hypoxic Environment is Regulated by Caveolin-1-dependent AMPK Activity.

Purpose In general, AMP-activated protein kinase (AMPK) has been known to inhibit cell proliferation or promote cell death, but still remains controversial. In this study, we propose the activity of AMPK which inhibits anti-apoptotic B-cell lymphoma 2 (Bcl-2) expression by relying on the hypoxia inducible factor-1 alpha (Hif1α)-induced caveolin-1 (Cav-1) expression pathway in non-invasive human bladder tumor cells (RT4). Methods In the cells exposed to 0.5% hypoxic environment, expression and phospho-activities of the related signaling enzymes were examined by Western-blot and RT-PCR. Cell proliferation was determined by CCK-8 cell counting assay. Results The expression level of Cav-1 is very low or undetectable in RT4 cells. Hypoxia decreased significantly cell growth with a marked induction of Hif-1α and Cav-1 expression, and suppressed anti-apoptotic marker Bcl-2 expression with unchanged AMPK activity. Under hypoxic conditions, HIF-1α induced acts as a transcriptional factor of Cav-1 mRNA gene expression. The cell growth and Bcl-2 expression level suppressed under hypoxia were reversed with a decrease of the induced Hif-1α and Cav-1 levels after AMPK activation with metformin (1 mM) or phenformin (0.1 µM). In addition, the expression of HIF1α and Cav-1 induced under hypoxia was not only increased in AMPK-siRNA pretreated cells, but also reversed the suppression of Bcl-2 expression. These results suggest that HIF1α and Cav-1 expressions in hypoxic environments are regulated by the basal AMPK activity, and that the inhibition of Bcl-2 expression cannot be expected even if Cav-1 expression is increased when AMPK activity is suppressed. Conclusions We suggest a new finding for the first time that AMPK activation can regulate Hif-1α induction as well as Hif-1α-induced Cav-1 expression, and the inhibitory effect of anti-apoptotic pathway on the exposure of RT4 cells to hypoxic environment is due to Cav-1-dependent AMPK activity.