Purpose To investigate the effect of a novel mitochondria targeted peptide MTP-131 against H2O2-induced oxidative damage in undifferentiated retinal ganglion cells (RGC-5).
Methods Immortalized RGC-5 cells were cultured with Dulbecco’s modified Eagles’ medium (DMEM). After pretreated with MTP-131 for 1 h, sustained oxidative stress was applied by subjecting undifferentiated RGC-5 cells to 500μM H2O2 for 24 h.Cell viability was measured by lactate dehydrogenase (LDH) assay. Changes of mitochondrial membrane potential (ΔΨm) and generation of intracellular reactive oxygen species (ROS) were measured by flow cytometry and confocal microscopy, using tetramethylrhodamine methyl ester (TMRM) and 2,7-dichlorofluorescin diacetate (H2DCFDA), respectively. Annexin V and propidium iodide (PI) staining was used for assessment of apoptosis. Release of Cytochrome c was analyzed by confocal. Data were analyzed with Graphpad Prism 5.0 software and P<0.05 was considered to be statistically significant.
Results Compared with cells treated with H2O2 alone, pretreatment with MTP-131 inhibited H2O2-induced cytotoxicity and reduced LDH release in a dose-dependent manner. Mitochondrial depolarization and ROS generation were also prevented by MTP-131 pretreatment. MTP-131 could also inhibit Cytochrome c release from mitochondria to cytoplasm, and significantly reduce apoptosis in RGC-5 cells.
Conclusion Mitochondria-targeted peptide MTP-131 has a protective effect against oxidative stress induced apoptosis in undifferentiated RGC-5 cells.