Less information is available concerning the molecular mechanisms of cell survival after
hypoxia in hepatocytes. Therefore, this study examined the effect of
hypoxia on
DNA synthesis and its related signal cascades in primary cultured chicken hepatocytes.
Hypoxia increased [3H]
thymidine incorporation, which was increased significantly after 0-24 h of hypoxic exposure. Indeed, the percentage of cell population in the S phase was increased in
hypoxia condition. However, the release of LDH indicating cellular injury was not changed under hypoxic conditions.
Hypoxia increased Ca2+ uptake and PKC translocation from the cytosol to the membrane fraction. Among the PKC
isoforms,
hypoxia stimulated the translocation of PKC alpha and epsilon.
Hypoxia also phosphorylated the p38 and p44/42
mitogen-activated protein kinases (MAPKs), which were blocked by the inhibition of PKC. On the other hand,
hypoxia increased Akt and mTOR phosphorylation, which was blocked in the absence of intra/extracellular Ca2+. The inhibition of PKC/MAPKs or PI3K/Akt pathway blocked the
hypoxia-induced [3H]
thymidine incorporation. However,
hypoxia-induced Ca2+ uptake and PKC translocation was not influenced by
LY 294002 or Akt inhibitor and
hypoxia-induced MAPKs phosphorylation was not changed by
rapamycin. In addition,
LY 294002 or Akt inhibitor has no effect on the phosphorylation of MAPKs. It suggests that there is no direct interaction between the two pathways, which cooperatively mediated cell cycle progression to
hypoxia in chicken hepatocytes.
Hypoxia also increased the level of the
cell cycle regulatory proteins [
cyclin D(1),
cyclin E,
cyclin-dependent kinase (CDK) 2, and CDK 4] and p-
RB protein but decreased the p21 and p27 expression levels, which were blocked by inhibitors of upstream signal molecules. In conclusion, short time exposure to
hypoxia increases
DNA synthesis in primary cultured chicken hepatocytes through cooperation of Ca2+/PKC,
p38 MAPK, p44/42 MAPKs, and PI3K/Akt pathways.