Abstract 14878: Downregulation of Gene p63 Enhances Reprogramming of Human Cardiac Fibroblasts Towards a Cardiac Fate

• Published in: Circulation (New York, N.Y.) N.Y.), 2019-11, Vol.140 (Suppl_1 Suppl 1), p.A14878-A14878
• Main Authors: Pinnamaneni, Jaya Pratap, Singh, Vivek P, Kim, Mary, Almousa, Ayman, Sanagasetti, Deepthi, Pugazenthi, Aarthi, Mathison, Megumi, Wang, Kai, So, Davis, Elsenousi, Abdussalam, Shafii, Alexis E, Yang, Jianchang, Rosengart, Todd K
• Format: Article
• Language: English
• ISSN: 0009-7322; 1524-4539
• DOI: 10.1161/circ.140.suppl_1.14878

IntroductionDirect reprogramming represents a promising new strategy in treating heart failure by allowing in situ transdifferentiation of cardiac fibroblasts into functional cardiomyocyte-like cells (iCMs). However, recent findings show that the reprogramming factors used in rodent cell transdifferentiation (e.g., Gata4, Mef2c, Tbx5 [GMT]) failed to reprogram human cardiac fibroblasts (HCFs), indicating a more complex & resistant feature in higher species.HypothesisBased on our recent success in murine cells, we hypothesized that downregulation of an antiplasticity gene p63 could overcome the resistance in HCFs & improve their reprogramming efficiency towards a cardiac fate.MethodsAdult HCFs were prepared from heart failure patients. To induce iCM reprogramming, isolated cells were treated with lentiviral-GFP vectors encoding scramble, shp63, GMT, or shp63+Hand2/Myocardin (HM) (Dose = 20 MOI). After 2 weeks, cells were assessed for iCM phenotype changes, using qRT-PCR, flow cytometry (FACS) & immunofluorescence (IF) assays. Co-IP assays were executed for mechanism analysis & for co-culture assays, HCFs were cultured with neonatal rat cardiomyocytes & maintained in defined medium for up to 7 weeks.ResultsFACS analysis showed that HCFs treated with shp63 in combination with HM yielded significant percentage of cells expressing the cardiac-specific troponin protein (cTnT) Vs control (11±1.3% Vs 0.6±0.14%; N = 5, mean±SEM, p