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Quantification of Viral Genome Uptake After AAV6 RR1.2 Injection in Mice with Dilated Cardiomyopathy

Undergraduate #12
Discipline: Biological Sciences
Subcategory: Biomedical Engineering

Yarid Mera - University of Washington


The reduction of cardiac function can be caused by many different factors including myocardial infarction and genetic cardimyopathies. There are limited pharmacological treatments to improve heart performance in these conditions. However, the Heart and Muscle Mechanics (HAMM) laboratory is investigating a novel gene therapy, which can improve cardiac performance. Studies have shown that dATP (2-deoxyadenosine triosphosphate), a naturally present nucleotide used by the cell during DNA synthesis, works as a better energy substrate for muscle contraction than ATP (Regnier, 2000; Adhikari, 2004). dATP allows for an increase in contractile capacity due to faster acto-myosin cross-bridge turnover. The dATP is produced by the enzyme Ribonucleotide Reductase (RNR) during DNA synthesis and cell division. By overexpressing RNR, more dATP can be produced, allowing for contractions with more force (Korte, 2011). Previous studies have shown that increasing the expression of RNR in cardiac cells increases contraction both in animals and in cell culture (Korte, 2011; Novakowski, 2013). In the current study, we are investigating if increased expression of RNR can improve cardiac function in a genetic model of dilated cardiomyopathy caused by mutation in tropomyosin (D230N). Mice with dilated cardiomyopathy received a systemic injection of an adveno-associated virus serotype 6(AAV6) that restricts the expression of RNR to cardiac cells. Tissue samples were harvested for DNA and protein analysis to determine uptake of the virus. Through qPCR, it was shown that there was successful uptake of the viral genome into the mice genome, with an average uptake rate of 0.15 vector genome per nuclei. Uptake of the virus also occurred in other tissues, such as liver and lung tissue. However through western blotting, it was shown that viral expression only occurred in the heart. Following the success of the RNR overexpression, cardiac function will be measured to determine improvements in cardiac contractions. The success of this study could lead to the development of a new gene therapeutic treatment for those suffering from dilated cardiomyopathy.

Funder Acknowledgement(s): University of Washington GenOM Project (NIH 5R25HG007153-03) and the donation from Anne Dinning and Michael Wolf.

Faculty Advisor: Michael Regnier,

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This material is based upon work supported by the National Science Foundation (NSF) under Grant No. DUE-1930047. Any opinions, findings, interpretations, conclusions or recommendations expressed in this material are those of its authors and do not represent the views of the AAAS Board of Directors, the Council of AAAS, AAAS’ membership or the National Science Foundation.

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