Xenotransplantation of mitochondrial electron transfer enzyme, Ndi1, in myocardial reperfusion injury

• Published in: PloS one 2011-02, Vol.6 (2), p.e16288
• Main Authors: Perry, Cynthia N, Huang, Chengqun, Liu, Wayne, Magee, Najib, Carreira, Raquel Sousa, Gottlieb, Roberta A
• Format: Article
• Language: English
• Subjects: Animals; Animals, Newborn; Apoptosis; ATP; Autophagy; Baking yeast; Biology; Cardiac muscle; Cardiomyocytes; Cell death; Cell Death - genetics; Cells (Biology); Cells, Cultured; Cloning; Cytochrome; Damage; Dehydrogenases; Dendritic cells; Drosophila; Electron transfer; Electron transport; Electron Transport - genetics; Electron transport chain; Electron Transport Complex I - genetics; Electron Transport Complex I - metabolism; Electrons; Enzymes; Fusion protein; Gene Transfer Techniques; Genetic Therapy - methods; Heart; Heart attacks; Hypertension; Insects; Ischemia; Lipids; Localization; Mammalian cells; Mammals; Membranes; Microscopy; Mitochondria; Mitochondria, Heart - genetics; Mitochondria, Heart - metabolism; Mitochondrial DNA; Myocardial Reperfusion Injury - genetics; Myocardial Reperfusion Injury - metabolism; Myocardial Reperfusion Injury - therapy; Myocytes; Myocytes, Cardiac - metabolism; Myocytes, Cardiac - pathology; NAD; NADH; NADH-quinone oxidoreductase; Neonates; Nicotinamide adenine dinucleotide; Oxidation; Oxidoreductase; Oxygen; Peptide Fragments - chemistry; Peptide Fragments - genetics; Peptide Fragments - metabolism; Phosphorylation; Preservation; Proteins; Quinones; Rats; Reactive oxygen species; Recombinant Proteins - genetics; Recombinant Proteins - metabolism; Reperfusion; Reperfusion injury; Rodents; Rotenone; Saccharomyces cerevisiae; Saccharomyces cerevisiae - genetics; Saccharomyces cerevisiae Proteins - genetics; Saccharomyces cerevisiae Proteins - metabolism; Superoxides; tat Gene Products, Human Immunodeficiency Virus - chemistry; tat Gene Products, Human Immunodeficiency Virus - genetics; tat Gene Products, Human Immunodeficiency Virus - metabolism; Tissue Distribution - genetics; Toxicity; Transplantation, Heterologous; Ventricle; Xenografts; Xenotransplantation
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0016288

A significant consequence of ischemia/reperfusion (I/R) is mitochondrial respiratory dysfunction, leading to energetic deficits and cellular toxicity from reactive oxygen species (ROS). Mammalian complex I, a NADH-quinone oxidoreductase enzyme, is a multiple subunit enzyme that oxidizes NADH and pumps protons across the inner membrane. Damage to complex I leads to superoxide production which further damages complex I as well as other proteins, lipids and mtDNA. The yeast, S. cerevisiae, expresses internal rotenone insensitive NADH-quinone oxidoreductase (Ndi1); a single 56 kDa polypeptide which, like the multi-subunit mammalian complex I, serves as the entry site of electrons to the respiratory chain, but without proton pumping. Heterologous expression of Ndi1 in mammalian cells results in protein localization to the inner mitochondrial membrane which can function in parallel with endogenous complex I to oxidize NADH and pass electrons to ubiquinone. Expression of Ndi1 in HL-1 cardiomyocytes and in neonatal rat ventricular myocytes protected the cells from simulated ischemia/reperfusion (sI/R), accompanied by lower ROS production, and preservation of ATP levels and NAD+/NADH ratios. We next generated a fusion protein of Ndi1 and the 11aa protein transduction domain from HIV TAT. TAT-Ndi1 entered cardiomyocytes and localized to mitochondrial membranes. Furthermore, TAT-Ndi1 introduced into Langendorff-perfused rat hearts also localized to mitochondria. Perfusion of TAT-Ndi1 before 30 min no-flow ischemia and up to 2 hr reperfusion suppressed ROS production and preserved ATP stores. Importantly, TAT-Ndi1 infused before ischemia reduced infarct size by 62%; TAT-Ndi1 infused at the onset of reperfusion was equally cardioprotective. These results indicate that restoring NADH oxidation and electron flow at reperfusion can profoundly ameliorate reperfusion injury.