An Integrated Evaluation Framework for Adult Heart Regeneration.

Abstract

Myocardial infarction (MI) remains a primary global cause of mortality due to irreversible cardiomyocyte loss. While promoting endogenous regeneration offers a highly promising therapy, its validation is critically hindered by the lack of standardised, comprehensive methodologies for quantitative assessment. To address this, we developed an integrated, systematically validated experimental framework for the precise evaluation of heart regeneration after ischemic injury in adult mice. This framework standardises four key assessment dimensions: (1) serial echocardiography mapping for precise serial evaluation of left ventricular function and structural changes; (2) standardised Masson's trichrome staining with systematic transverse sampling across five sections for reproducible quantification of post-ischemic scar burden; (3) stringent multi-marker strategy (EdU, pH 3 and Aurora B) for the accurate quantification of cardiomyocyte cell-cycle reactivation; (4) total cardiomyocyte enumeration achieved by employing enzymatic digestion to derive definitive quantitative metrics, confirming successful cell replenishment. This integrated framework establishes detailed and reproducible methodological standards to significantly enhance the accuracy and reliability of heart regeneration research. It thereby provides a robust reference for the standardised assessment of heart regeneration in adult mice.