Int J Biol Sci 2018; 14(14):1974-1984. doi:10.7150/ijbs.28050
Nitric oxide prevents H2O2-induced apoptosis in SK-N-MC human neuroblastoma cells
Laboratory of Veterinary Biochemistry and Molecular Biology, College of Veterinary Medicine, Chungbuk National University, Cheongju, Chungbuk 28644, Republic of Korea
Nitric oxide (NO) is a cellular signaling molecule in many physiological and pathological processes including neuroprotector. Here we examined the antiapoptotic effect of NO in SK-N-MC cells. H2O2 treatment (10-200 μM) induced cell death in a dose-dependent manner and pretreatment of cells with 100 μM S-nitroso-N-acetylpenicillamine (SNAP), an NO donor, attenuated the occurrence of H2O2-induced cell death. DAPI staining showed H2O2-induced nuclear fragmentation and NO treatment suppressed it. NO inhibited the proteolytic activation of caspase-3 and mitochondrial cytochrome c release. Treatment of soluble guanylyl cyclase inhibitor ODQ decreased the protective effect of SNAP on H2O2-treated cells and increased caspase 3-like enzyme activity and activation, cytochrome c release, PARP cleavage, and DNA fragmentation, indicating that cGMP is a key mediator in NO-mediated antiapoptosis. The cGMP analog 8-Br-cGMP blocked H2O2-induced apoptotic cell death; reduction of caspase-3 enzyme, cytochrome c release, and caspase-8 and -9. These preventive effects of SNAP and 8-Br-cGMP were suppressed by PKG inhibitor KT5823. Levels of PKGI, PKGII, and p-VASP proteins were increased by SNAP and 8-Br-cGMP and suppressed by KT5823 treatment. These results indicate that PKG is a downstream signal mediator in the suppression of apoptosis by NO and cGMP. Akt activation was inhibited the PI3K inhibitors LY294002 and Wortmannin, resulting in the inhibition of cell viability and increase of cytochrome c release. SNAP induced phosphorylation of Akt and Bad and then increased the interactions between 14-3-3β and p-Bad. These data suggest that the NO suppresses H2O2-induced SK-N-MC cell apoptosis by suppressing apoptosis signal mediating the interaction between 14‐3‐3β and Bad phosphorylation via PKG/PI3K/Akt.
Keywords: Nitric oxide, hydrogen peroxide, PKG, PI3K, Akt
Yoo YM, Jung EM, Ahn C, Jeung EB. Nitric oxide prevents H2O2-induced apoptosis in SK-N-MC human neuroblastoma cells. Int J Biol Sci 2018; 14(14):1974-1984. doi:10.7150/ijbs.28050. Available from http://www.ijbs.com/v14p1974.htm