Miro1 protects against brain injury after CPR in rats by enhancing the effect of BMSCs on mitochondrial homeostasis

Abstract Background Mitochondrial dyshomeostasis plays an important role in neuronal damage after cerebral ischemia-reperfusion, and Miro1 is a core protein that regulates mitochondrial homeostasis. In this study, we aimed to investigate the neuroprotective effects of bone marrow-derived mesenchymal stem cells (BMSCs) via mitochondrial homeostasis in rats after cardiac arrest (CA), and to clarify the role that the protein Miro1 plays in this protective efficacy. Methods The study compared the effects of BMSCs in which Miro1 was overexpressed BMSCs (BMSCs-mirohi), knocked down (BMSCs-mirolo), and unmodified BMSCs on mitochondrial homeostasis in hippocampal neurons to evaluate their neuroprotective effects of these cells in a rat model of global cerebral ischemia-reperfusion injury. Rats underwent CA modeling for 5 min and received cardiopulmonary resuscitation (CPR). Two hours after the restoration of spontaneous circulation, 1 mL of PBS or 1 mL containing 1 × 106 BMSCs (normal, mirohi, or mirolo) were injected via the femoral vein. The neurological function of rats was assessed based on Neurological Disability Score (NDS) values. Brain histopathological examination was conducted to evaluate brain injury by measuring oxidative stress levels and the apoptosis rate of hippocampal neurons. Immunoblotting and transmission electron microscopy were applied to detect the expression of mitophagy-related proteins in hippocampal neurons. Immunofluorescence was used to track the mitochondria in BMSCs and observe mitochondrial transfer. Additionally, the membrane potential level, oxidative stress level, and ATP content of mitochondria in hippocampal neurons were measured to assess the impact of transplanted BMSCs on mitochondrial quality in these hippocampal neurons. Results Immunofluorescence staining revealed the presence of mitochondria from MitoTracker-labeled BMSCs in rat hippocampal neurons post-CPR. Additionally, the fluorescence intensity of TOMM20 was notably increased following the transplantation of BMSCs. Through immunoblotting experiments, we identified that BMSCs amplified the post-CPR protein expression of LC3, p62, PINK1 and parkin in hippocampal neurons. The number of autophagosomes significantly increased in hippocampal neurons following BMSC transplantation, as observed through transmission electron microscopy. Flow cytometry, Hematoxylin and Eosin (HE) staining, and NDS scoring indicated that BMSCs effectively reduced reactive oxygen species accumulation in hippocampal neurons and mitochondria after CPR. Furthermore, they restored mitochondrial membrane potential and ATP levels in the hippocampus while decreasing apoptosis, ultimately contributing to the restoration of neurological function. Additionally, unlike BMSCs-mirolo, BMSCs-mirohi were able to significantly enhance the efficiency of BMSC-mediated mitochondrial transfer and enhance mitophagy. This amplification, in turn, was found to bolster the protective impact of BMSCs on hippocampal neurons during CPR, thereby contributing to the restoration of rat neurological function. Conclusions These analyses revealed that BMSC transplantation has a dual protective effect by facilitating healthy mitochondrial transfer and promoting the autophagic degradation of damaged mitochondria, effectively enhancing hippocampal neuronal mitochondrial function following CA while reducing neuronal apoptosis, restoring neuronal function, and alleviating neuropathological damage. Moreover, Miro1 can enhance the efficiency of mitochondrial transfer and promote BMSC-mediated mitophagy induction, thereby optimizing the therapeutic effect of BMSCs.