Effects of mitochondrial transplantation on intracellular antiviral pathways, restorative actions, and immunomodulatory responses in rig-i agonist-activated lung cells.

Background & Aim Viral respiratory infections impact mitochondrial function, altering the lung's immune response and contributing to disease pathogenesis through impaired energy metabolism, increased oxidative stress, and mitochondrial dynamics disruption. Mitochondrial transfer-based therapy, particularly those derived from mesenchymal stromal cells (MSCs), emerges as a promising therapeutic strategy for restoring mitochondrial function in impaired cells. We explore the effects of MSC-derived mitochondria transplantation on lung cells activated with a retinoic acid-inducible gene-I (RIG-I) receptor agonist and their immunomodulatory impact on immune cells. RIG-I is critical in triggering antiviral responses to control viral replication in response to virus-specific RNA structures. Upon viral RNA detection, RIG-I recruits the mitochondrial antiviral signaling protein (MAVS), which activates signaling cascades and produces interferons and cytokines. Methods, Results & Conclusion Mitochondria and MAVS-enriched mitochondria were isolated from MSC. Using an in vitro model of lung epithelial cells (A549) activated with a RIG-I agonist (synthetic viral-RNA), we assessed the impact of mitochondria transplantation on cell viability (Flow cytometry,FC), ATP production (Fluorometry), mitochondria membrane potential (MMP)(FC), Oxygen Consumption Rate (OCR,Seahorse), reactive oxygen species (ROS)(FC) and activation of antiviral MAVS pathway (qRT-PCR,WB). Also, we investigate the immunomodulatory effects of transplanted lung cells on PBMC-derived immune cells. Our results reveal that activated lung cells exposed to mitochondria-isolated rich fractions demonstrated restored OCR, increased viability through mitigated apoptosis, recovered ATP levels and MMP, and reduced ROS production. In macrophages subjected to a pro-inflammatory environment, the transplantation intervention induced polarization towards an anti-inflammatory M2-like macrophage phenotype. In lymphocytes, an increase towards the T-regulatory phenotype was observed. These findings underscore the efficacy of mitochondrial-derived MSC transplantation in ameliorating mitochondrial and cellular dysfunction in viral RNA-activated lung cells. Also, upon acceptance of mitochondria by activated lung cells, these cells showed the capacity to alter the pro-inflammatory microenvironment towards an anti-inflammatory state. Mitochondria-based therapy presents a promising avenue for enhancing the treatment of viral respiratory diseases. Supported by Fondecyt N°1211376.