Because survival and growth of human
hepatoma cells are maintained by nutrient, especially
glucose,
glucose starvation induces acute cell death. The cell death is markedly suppressed by
hypoxia, and we have reported involvement of
AMP-activated protein kinase-alpha (AMPK-alpha), Akt, and ARK5 in
hypoxia-induced tolerance. In the current study we investigated the mechanism of
hypoxia-induced tolerance in human
hepatoma cell line HepG2. ARK5 expression was induced in HepG2 cells when they were subjected to
glucose starvation, and we found that
glucose starvation transiently induced Akt and AMPK-alpha phosphorylation and that
hypoxia prolonged phosphorylation of both
protein kinases. We also found that
hypoxia-induced tolerance was partially abrogated by blocking the Akt/ARK5 system or by suppressing AMPK-alpha expression and that suppression of both completely abolished the tolerance, suggesting that AMPK-alpha activation signaling and the Akt/ARK5 system play independent essential roles in
hypoxia-induced tolerance. By using chemical compounds that specifically inhibit
kinase activity of type I-
transforming growth factor-beta (
TGF-beta) receptor, we showed an involvement of
TGF-beta in
hypoxia-induced tolerance.
TGF-beta1 mRNA expression was induced by
hypoxia in an
hypoxia-inducible factor-1alpha-independent manner, and addition of recombinant
TGF-beta suppressed cell death during
glucose starvation even under normoxic condition. AMPK-alpha, Akt, and ARK5 were activated by
TGF-beta1, and Akt and AMPK-alpha phosphorylation, which was prolonged by
hypoxia, was suppressed by an inhibitor of
type I TGF-beta receptor. Based on these findings, we propose that
hypoxia-induced
tumor cell tolerance to
glucose starvation is caused by
hypoxia-induced
TGF-beta1 through AMPK-alpha activation and the Akt/ARK5 system.