Apoptosis-inducing factor determines the chemoresistance of non-small-cell lung carcinomas.

Non-small-cell lung carcinomas (NSCLCs) are resistant to the induction of apoptosis by conventional anticancer treatment. However, NSCLC cell lines are sensitive to the action of the broad protein kinase inhibitor, staurosporine (STS). In the NSCLC cell line U1810, STS induced the mitochondrial release of apoptosis-inducing factor (AIF) and cytochrome c (Cyt c) followed by activation of caspases, nuclear condensation, DNA fragmentation and finally cell death. Although preincubation of U1810 cells with the broad-spectrum caspase inhibitor z-VAD.fmk delayed the occurrence of nuclear apoptosis induced by STS, it did not impede mitochondrial alterations (such as the release of Cyt c and AIF) and cell death to occur. Moreover, the microinjection of neither Cyt c nor recombinant active caspase-3 into the cytoplasm promoted nuclear apoptosis-related changes in U1810 cells. Evaluation of the role of the caspase-independent factor AIF in STS-mediated death revealed that, upon immunodepletion of AIF, cytosols from STS-treated U1810 lost their capacity to induce nuclear condensation when incubated with isolated nuclei. In addition, microinjection of an anti-AIF antibody prevented AIF from translocating to the nuclei of STS-treated U1810 cells and reduced STS-induced cell death. Finally, although the transfection-enforced overexpression of AIF was not sufficient to induce cell death, it did enhance STS-mediated cell killing. Altogether, these results indicate that activation of caspases is not sufficient to kill U1810 cells and rather suggests an important role for the AIF-mediated mitochondrial-mediated death pathway.

Induction of apoptosis-like mitochondrial impairment triggers antioxidant and Bcl-2-dependent keratinocyte differentiation.

Terminally differentiated keratinocytes are dead enucleated squams. We showed previously that the mitochondria-dependent cell death pathway might be gradually activated as differentiation progresses. In this study, we demonstrated that protoporphyrin IX, staurosporine, and rotenone induced apoptotic-like changes in the mitochondria, and early differentiation of keratinocytes without inducing apoptosis. Kinetics studies established that differentiation-related changes, including growth arrest, flattened morphology, stratification, and keratin 10 (K10) expression, were downstream of mitochondrial depolarization and proliferation, reactive oxygen species (ROS) production, and release of cytochrome c and apoptosis-inducing factor. When these changes were prevented by overexpressing Bcl-2 or pharmacologically decreasing the ROS level, K10 upregulation was inhibited, implying that the differentiated phenotype and K10 expression require apoptotic mitochondria, ROS being the most likely differentiation-mediating factor. Our data also suggest that the same mitochondria-affecting stimuli can induce either differentiation or apoptosis, depending on the keratinocyte's competency to undergo differentiation, a competency that may be controlled by Bcl-2.