Mitochondrial Fission via DRP1-Mediated GTPase Activity Versus MFF/FIS1-Dependent Receptor Scaffolding: A Mechanistic and Functional Comparison in Hypoxia-Induced Cardiomyocyte Apoptosis

Abstract

Mitochondrial dynamics govern cardiomyocyte fate under hypoxic stress, yet the relative contributions of DRP1-driven GTPase hydrolysis and receptor-scaffolded fission through MFF/FIS1 complexes remain incompletely resolved. Using primary neonatal rat ventricular cardiomyocytes subjected to controlled hypoxia–reoxygenation, we systematically compared pharmacological DRP1 inhibition (Mdivi-1), siRNA-mediated MFF/FIS1 knockdown, and combinatorial suppression across indices of mitochondrial morphology, membrane potential (ΔΨm), cytochrome c release, and caspase-3 activation. Super-resolution STED microscopy and flow cytometric analyses revealed that MFF/FIS1 scaffolding predominantly initiates pathological fission events, whereas DRP1 GTPase activity amplifies downstream apoptotic signaling. Combinatorial inhibition achieved 74% reduction in apoptotic index compared to single-pathway targeting. These findings reframe the hierarchical architecture of the mitochondrial fission cascade and suggest dual-pathway therapeutic targeting as a superior cardioprotective strategy.