Suppression of Calpain Attenuates Coxsackievirus B3-Induced Myocarditis by Regulating RIPK1/RIPK3/MLKL Pathway

Viral myocarditis caused by CVB3 inflicts substantial myocardial damage and predisposes to heart failure. Although necroptosis contributes to disease progression, the mechanisms governing this death pathway during CVB3 infection remain unclear. We found that calpain and its inhibitor Cast were dynamically expressed during CVB3 infection, correlating with cardiac damage, inflammation, and fibrosis. Further investigation revealed that CVB3 dynamically induced cardiomyocyte necroptosis by activating the RIPK1/RIPK3/MLKL pathway, which was prominently activated in the early acute phase but potentially attenuated in later stages. Immunofluorescence analysis supported the translocation of p-MLKL from the nucleus to the plasma membrane during disease progression. Importantly, both in vivo and in vitro experiments supported that calpain inhibition ALLN attenuated VMC via the RIPK1/RIPK3/MLKL pathway, and improved cardiac function post-CVB3 infection. The calpain activator PMMA exacerbated this process. Our findings reveal that calpain inhibition attenuated VMC by modulating the RIPK1/RIPK3/MLKL pathway, uncovering a novel regulatory mechanism in VMC.