miR-155-driven dysregulation of the PPARγ/PGC-1α/ERRα-Drp1 axis contributes to mitochondrial damage in hypoxic/ischemic neuronal cells

Methods

Oxygen-glucose deprivation (OGD)-treated Neuro-2a cells were employed to investigate the effects of hypoxia/ischemia on miR-155 expression. The cells transfected with miR-155 mimic and miR-155 inhibitor were used to clarify the the role of miR-155 in OGD-induced mitochondrial damage. The expression levels of PGC-1α, PPARα, PPARβ/δ, PPARγ, ERRα, and Drp1 were assessed using qRT-PCR, Western blotting, and immunofluorescence staining, and their interactions were identified by confocal colocalization and co-immunoprecipitation experiments.

Mitochondrial functions were evaluated by measuring ATP content, ROS, MMP, and mPTP opening. Chromatin immunoprecipitation (ChIP) assay was performed to detect the binding of ERRα to the Drp1 promoter. Results: miR-155 expression in OGD-treated neuronal cells exhibited a significant upregulation in a time-dependent manner.

Elevated miR-155 induced excessive mitochondrial fission and dysfunction, as evidenced by a decreased ATP content, increased ROS generation, depolarized MMP, and mPTP opening, which is accompanied by a markedly reduced expression of PPAR family members, particularly the PPARγ. Mechanistically, miR-155 weakened the interaction between PGC-1α and PPARγ by suppressing their expressions and thus downregulated ERRα expression, which prevents the binding of ERRα to the Drp1 promoter, thereby relieving the repression of the Drp1 promoter by ERRα.

These findings suggest that PGC-1α and ERRα negatively regulate Drp1 expression in neuronal cells. Notably, inhibition of miR-155 could ameliorate mitochondrial dysfunction by improving the OGD-induced dysregulation of PPARγ/PGC-1α/ERRα-Drp1 axis in Neuro-2a cells, providing new insights into the mechanisms of ischemia/hypoxia-induced mitochondrial dysfunction. Conclusion: Our findings reveal a novel mechanism by which miR-155 contributes to mitochondrial damage by driving the ischemia/hypoxia-induced dysregulation of PPARγ/PGC-1α/ERRα-Drp1 axis in neuronal cells, identifying potential new therapeutic targets for the treatment of post-ischemic mitochondrial damage.