340. Cell Penetrating Peptide Can Improve the Efficacy of Mitochondrial Transfer
Authors: Daisuke Kami, Tomoya Kitani, Yuki Takaoka, Yu Yoshimoto, Yasuhiko Tabata, Satoshi Gojo
Journal: Molecular Therapy (2016)

Cell-based therapies for organ regeneration are currently emerging as a new promising approach to treat various diseases, including cardiovascular and neurodegenerative diseases. However, the mechanism supporting these therapeutic effects remains poorly understood. Several recent studies suggest that the intercellular transfer of organelles, including mitochondria, might contribute to these regenerative responses. Mitochondria are considered cellular "power plants" because they largely synthesize the universal energy "currency" of the cells, i.e., adenosine triphosphate (ATP). Mitochondrial dysfunction is associated with many diseases, including metabolic and neurodegenerative disorders. In vitro experiments revealed that mitochondrial DNA (mtDNA)-depleted mammalian cells recovered aerobic respiration after intercellular mitochondrial transfer from intact cells. The therapeutic potential of mitochondrial transfer was supported by an in vivo study conducted on rabbit model of myocardial infarction. Direct injection of autologous mitochondria into the ischemic heart considerably improved post-infarct cardiac functions. Mitochondria dysfunction also plays central role in the pathogenesis of ischemia reperfusion injury and subsequent cardiomyocytes death. The beneficial effect of mitochondria replenishment for damaged cells and tissues has been reported recently. Our previous studies demonstrated that the mitochondrial transfer could occur by simple co-culture of mammalian cells with isolated mitochondria. We assessed the impact of the transfer on the mitochondrial function in the recipient cells. The mechanism of mitochondrial internalization was investigated using endocytosis inhibitors. We showed that mammalian cells engulfed isolated mitochondria by macropinocytosis, which survived in heteroplasmy state and functioned to generate ATP in recipient cells. However, the internalization for isolated mitochondria remains inefficient to apply as a clinical therapeutic strategy. Here, we report the use of TAT, which is cell penetrating peptide derived of human immunodeficiency virus, dextran complexes (TAT-DEX) to enhance the cellular uptake of exogenous mitochondria and improve the protective effect of mitochondria replenishment on rat neonatal cardiomyocytes (NRCM) from IR injury in vitro. TAT-DEX modification significantly neutralized surface negative charge in mitochondria isolated from the human endometrium-derived mesenchymal cells and H9c2 rat cardiomyoblast. TAT-DEX modified mitochondria isolated from H9c2 showed a significantly higher level of cellular uptake. Compared with control mitochondria, co-incubation of TAT-DEX modified mitochondria with NRCM during IR significantly reduced IR-induced reactive oxygen species production and apoptosis in NRCM. These results indicate that surface modification of isolated mitochondria at the molecular level can improve the efficiency of cellular uptake of exogenous mitochondria, and enhanced mitochondrial internalization into cardiomyocytes may improve the potential effect of mitochondria replenishment therapy in myocardial IR injury.