Inhibition of calpain reduces cell apoptosis by suppressing mitochondrial fission in acute viral myocarditis

Abstract

Cardiomyocyte apoptosis is critical for the development of viral myocarditis (VMC), which is one of the leading causes of cardiac sudden death in young adults. Our previous studies have demonstrated that elevated calpain activity is involved in the pathogenesis of VMC. This study aimed to further explore the underlying mechanisms. Neonatal rat cardiomyocytes (NRCMs) and transgenic mice overexpressing calpastatin were infected with coxsackievirus B3 (CVB3) to establish a VMC model. Apoptosis was detected with flow cytometry, TUNEL staining, and western blotting. Cardiac function was measured using echocardiography. Mitochondrial function was measured using ATP assays, JC-1, and MitoSOX. Mitochondrial morphology was observed using MitoTracker staining and transmission electron microscopy. Colocalization of dynamin-related protein 1 (Drp-1) in mitochondria was examined using immunofluorescence. Phosphorylation levels of Drp-1 at Ser637 site were determined using western blotting analysis. We found that CVB3 infection impaired mitochondrial function as evidenced by increased mitochondrial ROS production, decreased ATP production and mitochondrial membrane potential, induced myocardial apoptosis and damage, and decreased myocardial function. These effects of CVB3 infection were attenuated by inhibition of calpain both by PD150606 treatment and calpastatin overexpression. Furthermore, CVB3-induced mitochondrial dysfunction was associated with the accumulation of Drp-1 in the outer membrane of mitochondria and subsequent increase in mitochondrial fission. Mechanistically, calpain cleaved and activated calcineurin A, which dephosphorylated Drp-1 at Ser637 site and promoted its accumulation in the mitochondria, leading to mitochondrial fission and dysfunction. In summary, calpain inhibition attenuated CVB3-induced myocarditis by reducing mitochondrial fission, thereby inhibiting cardiomyocyte apoptosis.

Mitochondrial calpain-1 activates NLRP3 inflammasome by cleaving ATP5A1 and inducing mitochondrial ROS in CVB3-induced myocarditis

“…Inhibiting mitochondrial permeability transition pore opening through cyclosporin A treatment reduced Abcc6-dependent cardiac necrosis and calcification following CVB3 infection in mice [ 35 ]. Furthermore, curtailing excessive mitochondrial fission with Dynamin-related protein 1 inhibitor rescued myocardial injury induced by CVB3 [ 44 ]. However, the precise molecular mechanisms underlying the association between mitochondria and myocarditis development need to be investigated further.…”

Brain-targeted heptapeptide-loaded exosomes attenuated ischemia–reperfusion injury by promoting the transfer of healthy mitochondria from astrocytes to neurons

“…As a result, under pathological stress, mitochondrial dynamin-related protein-1 (Drp1) as a key mitochondrial regulatory protein predominantly interacts with fission 1 (Fis1), causing an exaggerated fission process, as evidenced by excessive mitochondrial fragmentation, production of reactive oxygen species (ROS) and loss of mitochondrial membrane potential [ 10 ]. Therefore, marked pathological fragmentation of mitochondria is produced, and mitochondrial function is significantly disrupted by decreasing adenosine triphosphate (ATP) and mitochondria membrane potential [ 11 ]. Finally, damaged astrocytic mitochondria are further released, enter adjacent neurons and induce fusion with neuronal mitochondria, inducing neuronal mitochondrial dysfunction, amplifying neuronal damage and worsening neurological outcomes [ 12 , 13 ].…”

TRPC6 mediates high glucose-induced mitochondrial fission through activation of CDK5 in cultured human podocytes

“…Calpain-1, activated by Ca 2+ influx, is a key downstream target of TRPC6 ( Zhang et al, 2014 ; Verheijden et al, 2018 ). Previous studies have shown that the activation of calpain induces mitochondrial fission by mediating Drp1 phosphorylation, resulting in myocardial apoptosis ( Shi et al, 2021 ; Zhang et al, 2021 ). Studies have identified that inhibition of either calpain or calcineurin leads to downregulated Drp1 and Fis1 levels, and upregulated Opa1 levels and MMP during exposure to oxidative stress.…”

“…Calpain-1, one of the nonlysosomal cysteine proteases consisting of an 80 kDa isoform-specific catalytic domain, has been widely demonstrated to be activated by TRPC6-dependent Ca 2+ influx in podocytes ( Verheijden et al, 2018 ). The effect of calpain-1 on mitochondrial fission has been reported in cardiac diseases ( Shi et al, 2021 ), whereas it is unclear in podocytes.…”