Targeted transplantation of mitochondria to hepatocytes

BACKGROUND: Mitochondrial defects in hepatocytes can result in liver dysfunction and death. Hepatocytes have cell-surface asialoglycoprotein receptors (AsGRs) which internalize AsGs within endosomes. The aim of this study was to determine whether mitochondria could be targeted to hepatocytes by AsGR-mediated endocytosis. MATERIALS AND METHODS: An AsG, AsOR, was linked to polylysine to create a conjugate, AsOR-PL, and complexed with healthy and functional mitochondria (defined by normal morphology, cytochrome c assays, and oxygen-consumption rates). Huh7 (AsGR(+)) and SK Hep1 (AsGR(−)) cells were treated with a mitochondrial toxin to form Huh7-Mito(−) and SK Hep1-Mito(−) cells, lacking detectable mitochondrial DNA. An endosomolytic peptide, LLO, was coupled to AsOR to form AsOR-LLO. A lysosomal inhibitor, amantadine, was used in mitochondria-uptake studies as a control for nonspecific endosomal release. RESULTS: Coincubation of complexed mitochondria and AsOR-LLO with Huh7-Mito(−) cells increased mitochondrial DNA to >9,700-fold over control at 7 days (P<0.001), and increased mitochondrial oxygen-consumption rates to >90% of control by 10 days. CONCLUSION: Rescue of mitochondria-damaged hepatocytes can be achieved by targeted uptake of normal mitochondria through receptor-mediated endocytosis.