Mitochondrial Transplantation Restores Bioenergetics after Spinal Cord Injury
Authors: Gollihue Jenna , Patel Samir , Eldahan Khalid , Cox David , Rabchevsky Alexander 
Published: November 2023

Traumatic spinal cord injury (SCI) has devastating, life‐long consequences and there are currently no effective pharmacological interventions to prevent permanent loss of function. Mitochondria, the energy producing organelles found in most cells, become severely damaged after SCI and release several detrimental factors such as cytochrome c, reactive oxygen and nitrogen species (ROS/RNS), and calcium resulting in decreased energy production, lipid and protein oxidation, and cell death. Treatments designed to ameliorate mitochondrial damage and maintain bioenergetics of injured spinal cord tissues are being tested with some promising results; a principle caveat is that not all pathophysiological factors can be targeted simultaneously. Therefore, we posited that mitochondrial replacement strategies could be a multi‐mechanistic approach to provide damaged tissues with increased ATP production, greater calcium buffering capacity, decreased ROS and RNS release and increased endogenous antioxidants. We set out to test the hypothesis that exogenous mitochondria can be transplanted into the injured spinal cord and enter host cells to preserve acute bioenergetics that fosters long‐term tissue sparing and improved functional recovery. We used transgenically modified PC12 cells in which mitochondria are labeled with tGFP allowing for their visualization after transplantation. At 20–30 min after severe contusion SCI, isolated tGFP‐labeled mitochondria were microinjected into the spinal cord of adult, female Sprague‐Dawley rats. Separate injury cohorts were designated to 1) establish the dose‐dependent effects of transplantation on cellular respiration after 24 hours, 2) demonstrate mitochondrial incorporation into various cell types and 3) assess long‐term hindlimb motor function, pain sensitivity, and tissue sparing. Preliminary results show that compared to 50 and 150ug dosages, respiration is significantly restored after 100ug tGFP mitochondrial transplantation compared to injury + vehicle. Confocal imaging after 50ug dosage injections shows tGFP mitochondria localized within both microglia/macrophages and endothelial cells at 24 and 48 hours after injections, but no evidence of tGFP mitochondria present after 7 days. The 100 and 150ug dosage groups are being evaluated similarly to establish the propensity of cell‐type incorporation after 24hr, 48hr and 7 days. Hindlimb functional recovery over 6 weeks is also being assessed following 100ug mitochondrial transplantation with concurrent analyses of kinematic stepping patterns and pain withdrawal latencies. While preliminary data appear to show insignificant trends, there are no differences in over‐ground locomotion or kinematic stepping patterns compared to vehicle‐injected injured groups (blinded). Additional injury cohorts are undergoing behavioral analyses and terminal morphometric assessments of spared white and gray matter will determine whether improved bioenergetics following mitochondrial transplantation results in long‐term tissue sparing, thus supporting this novel therapeutic intervention to promote recovery after traumatic SCI.Support or Funding InformationNIH Fellowship 5T32NS077889 (JLV)NIH F31NS093904‐01A1 (JLV)R21NS096670 (AGR)Conquer Paralysis Now (AGR)SCoBIRC Chair Endowment (AGR)NIH/NINDS 2P30NS051220 (AGR)