Molecular ordering of hypoxia-induced apoptosis: critical involvement of the mitochondrial death pathway in a FADD/caspase-8 independent manner.

Dys-regulated growth and improper angiogenesis commonly lead to areas of hypoxia in human tumors. Hypoxia is known to be associated with a worse outcome since a lack of oxygen interferes with the efficacy of chemotherapy or radiotherapy. In parallel, hypoxia-induced apoptosis may also impose a selection pressure favoring growth of more resistant tumor cells. However, the mechanisms of hypoxia-induced apoptosis and the relative contribution of intrinsic and extrinsic apoptotic pathways are not understood. Therefore, Jurkat cell lines with defined defects in the extrinsic or intrinsic signaling cascades were used to evaluate the role of either pathway for induction of apoptosis under hypoxic conditions. Jurkat cells were incubated in hypoxia and the rate of apoptosis induction was determined by Western blotting, fluorescence microscopy and flow cytometry. Hypoxia-induced apoptosis was not affected by lack of caspase-8 or FADD, whereas overexpression of Bcl-2 or expression of dominant-negative caspase-9 mutant rendered the cells resistant to hypoxia-induced apoptosis. These results suggest that hypoxia-induced apoptosis mainly relies on intrinsic, mitochondrial pathways, whereas extrinsic pathways have no significant implications in this process. Thus, in human tumors, hypoxia will mainly lead to the selection of hypoxia-resistant cells with defects in mitochondrial apoptosis signaling pathways.