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Thesis Format

Alternative Format

Degree

Master of Science

Program

Pathology and Laboratory Medicine

Supervisor

Peng, Tianqing

Abstract

Doxorubicin (DOX) blocks the autophagic flux in cardiomyocytes by inhibiting lysosome acidification. The acidic pH of lysosomes is maintained by the V-ATPase pump. NAD+ is an essential cofactor for the maintenance of cellular homeostasis. DOX treatment significantly depletes the NAD+ levels in cardiomyocytes. This study investigated the potential of the NAD+ precursor, nicotinamide mononucleotide (NMN), in preventing DOX-induced cardiotoxicity. DOX induced cell injury and altered the lysosomal pH of H9c2 cells, an in vitro model of cardiomyocytes. These effects of DOX were attenuated by NMN. The protection conferred by NMN was offset by inhibition of V-ATPase activity with bafilomycin A. Furthermore, NMN prevented the DOX induced hyperacetylation of the V-ATPase subunit ATP6V0d1, a critical protein involved in the maintenance of V-ATPase activity. In summary, NMN protects cardiomyocytes from DOX induced toxicity by maintaining the pH of lysosomes. Thus, NMN holds potential in combating the deleterious impacts of Doxorubicin on the heart.

Summary for Lay Audience

Cancer is one of the leading causes of death worldwide. Nearly 1 in 2 Canadians develop cancer during their lifetime. The harsh reality of cancer does not end with its onset. The devastating dark side of the available treatment options like chemotherapy impacts the patient’s quality of life. Doxorubicin, the chemotherapy antibiotic, is routinely used for the treatment of multiple cancers. Its deadliest side effect is heart injury, which leads to heart failure. In fact, over 50% of the childhood cancer survivors develop heart disease in the later stages of their life. Doxorubicin impairs a cellular process known as autophagy in heart cells. Autophagy or cellular recycling is a self-eating process by which the damaged components in the cells can be degraded and recycled. This recycling process occurs in designated recycling centres or specialized compartments called lysosomes in the cell. Efficient functioning of lysosomes is rendered by their acidic nature or low internal pH. This helps the lysosomal proteins to break down the cellular debris. The low pH in lysosomes is achieved with the help of a cellular pump called V-ATPase positioned on the lysosomal membrane. V-ATPase transports protons into the lysosome and maintains optimal pH. Doxorubicin increases the pH of lysosomes and it could most likely be by damaging the V-ATPase pump, but the exact mechanism remains unknown.

NAD+ is a critical molecule in the body which is required for many essential reactions in the cell. Doxorubicin decreases the natural levels of NAD+ in the body and compromises the normal functioning of the cell. NAD+ depletion effects the lysosome function too. This study aims to identify the impact of Doxorubicin treatment on the pH of lysosomes and the V-ATPase pump in the heart cells. Nicotinamide mononucleotide (NMN) is a vitamin B3 derivative and NAD+ supplement that provides many essential benefits to the body. The role of NMN in preventing the harmful effects of Doxorubicin have been investigated in this study. The results indicate that NMN has promising potential in protecting the heart from the side effects of Doxorubicin.

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