Dystrophin and calcium current are decreased in cardiomyocytes expressing Cre enzyme driven by αMHC but not TNT promoter

Abstract

BackgroundThe Cre/lox system is a potent technology to control gene expression in mouse tissues. However, cardiac alterations following cardiac-specific Cre enzyme expression in non-loxP-flanked genome heart have been reported. Recently, many loxP like sites have been identified in the wild-type mouse genome. Interestingly one of them is localized in the Dmd gene encoding the dystrophin protein known to be crucial for stabilization of cardiac voltage-gated ion channels Nav1.5 and Cav1.2. AimHere, we studied the potential alteration of dystrophin expression in adult alpha-myosin heavy chain (MHC)-Cre mice, which are extensively used for cardiac-specific recombination, and investigated Troponin T (TNT)-Cre mice as a potential alternative. MethodsCardiac-specific MHC-Cre and TNT-Cre mouse lines expressing Cre recombinase under the control of the cardiac-specific alpha-myosin-heavy chain, and rat cardiac troponin T2 promoter respectively were used. Western blots, quantitative RT-PCR, immunostainings, and patch-clamp experiments were performed to characterize MHC-Cre and TNT-Cre mouse hearts and cardiomyocytes. ResultsDystrophin protein level was decreased in hearts from 12-week-old MHC-Cre+ mice compared to MHC-Cre-. Reduction of dystrophin was more pronounced with age. No significant difference was observed between 8-week-old MHC-Cre+ mice and MHC-Cre-. Immunostainings performed on cardiac sections showed reduced dystrophin signal at the lateral membrane of MHC-Cre+ cardiomyocytes. Quantitative RT-PCR showed decreased mRNA levels of Dmd gene encoding dystrophin. Finally, patch-clamp experiments showed a significant decrease in calcium current (ICaL) in adult MHC-Cre+ cardiomyocytes compared to MHC-Cre-. Neither dystrophin nor ICaL was reduced in adult TNT-Cre+ mouse hearts compared to TNT-Cre-. ConclusionIn contrary to TNT-Cre+ mice, the sole expression of Cre recombinase can alter the cardiac phenotype of MHC-Cre+ mice. Thus, researchers should include the "Cre-only" condition as control condition when designing experiments with Cre mouse strains.