Date of Completion

8-20-2015

Embargo Period

8-11-2021

Major Advisor

Dr. George Y Wu

Associate Advisor

Dr. Blanka Rogina

Associate Advisor

Dr. Gordon Carmichael

Associate Advisor

Dr. Peter Maye

Associate Advisor

Dr. Catherine Wu

Field of Study

Biomedical Science

Degree

Doctor of Philosophy

Open Access

Open Access

Abstract

Hepatocyte damage caused by genetic and acquired defects in mitochondria can result in severe liver dysfunction causing disease and death. Mammalian hepatocytes possess specific receptors on their surfaces called asialoglycoprotein receptors (AsGRs), which can recognize and bind asialoglycoproteins (AsGs). After binding, AsGs are internalized by receptor-mediated endocytosis within membrane-limited vesicles, endosomes, which ultimately fuse with lysosomes resulting in degradation. This pathway represents a natural mechanism by which substances outside cells can gain access to the interior of liver cells. In the past, small molecules have been bound to AsGs to deliver them specifically to the liver. The aim of this study was to determine whether AsGs could serve as targetable carriers for mitochondria to permit targeting specifically to hepatocytes by the AsGR pathway. We describe here, a method by which mitochondria can be coated with AsGs by a non-damaging interaction. We converted an AsG into a highly positively charged molecule, and took advantage of the fact that the surface of mitochondria is negatively charged. By mixing a positively charged AsG protein conjugate with mitochondria, the conjugate could bind to mitochondria in a strong, but non-damaging electrostatic (charge-charge) interaction forming protein–mitochondrial complexes. This results in recognition and internalization of mitochondria specifically by hepatocytes. We have also developed a method by which AsGs–mitochondria complexes can be released from endosomes. This allows escape from endosomes before lysosomal digestion and cytoplasmic delivery of healthy mitochondria to the cells. The system can result in rescue of cells rendered mitochondria-free by drug toxicity, and proliferation of both host cells and transplanted mitochondria. Currently, there is no treatment for mitochondrial dysfunction, and no means to repair or replace damaged mitochondria. Targeting healthy mitochondria to hepatocytes with defective or damaged mitochondria by the AsGR pathway would be expected to confer on those cells a selective survival advantage. As a result, those cells would be expected to proliferate, eventually replace defective cells reversing hepatotoxicity.

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