Transfer of MSCs mitochondria facilitate the recovery of stroke

Abstract

Objective To explore MSCs mitochondria transfer mediated protection after stroke. Methods Isolation and primary culture of bone MSCs of mice. FACS was used to identify the purity of primary MSCs. Primary neuron cultures were prepared from the cerebral cortex of postnatal day 0 (P0) SD rats. OGD (oxygen-glucose deprivation) was performed on the primary neurons. OGD neurons were grouped into MCM group (MSCs conditioned mitochondria, MCM) , mdMCM group (depleted extracellular mitochondria in MCM, mdMCM) , OGD group and normal neurons which labeled as Neuron group. MitoTracker was used to track the mitochondria transfer from MSCs to OGD neurons. Detection kits was used to analyze the ATP level and viability of neurons. Membrane potential of mitochondria was analyzed by detection kit to measure the function of mitochondria. The expression of Miro1 protein was performed by Western Blot. The protection of MSCs transplant was performed by MCAO and MSCs injection. Statistical analysis was performed using analysis of variance and t-tests. Results The purity of primary cultured MSCs was higher than 99﹪. Mitochondria transfer from MSCs to OGD neurons was observed. The transfer of mitochondria protected OGD neurons by increasing the ATP level, and the mdMCM group ATP level were significant lower than MCM group. (0.634±0.023 in OGD group, 1.623±0.039 in MCM group and 0.645±0.011 in mdMCM group, F = 3413.62, P < 0.01) ; And the viability of neurons in MCM group higher than in OGD group, and the viability of neurons in mdMCM group were significant lower than MCM group[ (73.7±1.12) ﹪ in OGD group, (83.3±1.57) ﹪ in MCM group and (72.9±1.25) ﹪ in mdMCM group, F = 654.280, P < 0.01]. And the membrane potential loss of mitochondria in OGD and mdMCM groups were significantly higher than in MCM group. (70.3﹪ in OGD group, 44.7﹪ in MCM group and 67.7﹪ in mdMCM group, P < 0.05) . After intervention of Miro1, the ATP level increased in Miro1-overexpression group (2.304 vs 1.611 in control group, P = 0.034) ; the neuronal viability also increased in Miro1-overexpression group (90.4﹪ vs 81.7﹪ in control group, P = 0.040) . After MCAO, the infarct volume of mice reached 38.4﹪, while the MSCs-injected mice decreased to 14.4﹪ (P = 0.004) . Conclusion MSCs transplantation decreased the infarct volume of MCAO mice and the protection may be mediated by Miro1 protein. Key words: MSCs; Stroke; Miro1; Mitochondria transfer