Authors: Dennis Le, Zach Salyers, and Terence Ryan
Faculty Mentor: Terence Ryan
College: College of Health and Human Performance
Peripheral arterial disease (PAD) – the narrowing of blood vessels in the limb from plaque buildup – is the third leading cause of atherosclerotic cardiovascular mortality. Critical limb ischemia (CLI) – the most severe form of PAD – requires urgent care. In absence of effective revascularization, the mortality rates of CLI patients exceed 50% by 5 years with CLI conditions. This project aimed to develop and validate genetic therapy tools for potential CLI treatment through one of the underlying biological changes experienced among PAD patients. CLI muscle biopsies from Ryan Lab revealed a decreased expression of PFKFB3 (6-phosphofructo-2-kinase/2,6-bisphosphatase 3). PFKFB3 – a bifunctional enzyme regulating an allosteric glycolytic activator – has an established upregulating role in the anaerobic energy production within our cells. PFKFB3 knockdown and overexpression vectors were constructed and evaluated to ensure tissue specificity and gene manipulation. In mouse tibialis anterior (TA) muscle, the downregulating vector revealed an overall 62.9% knockdown of PFKFB3 mRNA while the upregulating vector showed a 110-fold increase (P < 0.05). Verifying the tools’ functionality and specificity confirmed our goal of manipulating the targeted cells and their genome before conducting any biological study of hindlimb ischemic mice mimicking CLI conditions with implementation of the PFKFB3 vector in the TA.