The DNA-dependent protein kinase catalytic subunit exacerbates endotoxemia-induced myocardial microvascular injury by disrupting the MOTS-c/JNK pathway and inducing profilin-mediated lamellipodia degradation.

The DNA-dependent protein kinase catalytic subunit (DNA-PKcs) promotes pathological mitochondrial fission during septic acute kidney injury. The mitochondrial open reading frame of the 12S rRNA type-c (MOTS-c) is a mitochondria-derived peptide that exhibits anti-inflammatory properties during cardiovascular illnesses. We explored whether endotoxemia-induced myocardial microvascular injury involved DNA-PKcs and MOTS-c dysregulation.To induce endotoxemiaendothelial cell-specificknockout mice were injected intraperitoneally with a single dose of lipopolysaccharide (10 mg/kg) and evaluated after 72 h.Lipopolysaccharide exposure increased DNA-PKcs activity in cardiac microvascular endothelial cells, while pharmacological inhibition or endothelial cell-specific genetic ablation ofreduced lipopolysaccharide-induced myocardial microvascular dysfunction. Proteomic analyses showed that endothelialablation primarily altered mitochondrial protein expression. Verification assays confirmed that DNA-PKcs drastically repressedtranscription by inducing mtDNA breaks via pathological mitochondrial fission. Inhibitingneutralized the endothelial protective effects ofablation, whereas MOTS-c supplementation enhanced endothelial barrier function and myocardial microvascular homeostasis under lipopolysaccharide stress. In molecular studies, MOTS-c downregulation disinhibited c-Jun N-terminal kinase (JNK), allowing JNK to phosphorylate profilin-S173. Inhibiting JNK or transfecting cells with a profilin phosphorylation-defective mutant improved endothelial barrier function by preventing F-actin depolymerization and lamellipodial degradation following lipopolysaccharide treatment.DNA-PKcs inactivation during endotoxemia could be a worthwhile therapeutic strategy to restore MOTS-c expression, prevent JNK-induced profilin phosphorylation, improve F-actin polymerization, and enhance lamellipodial integrity, ultimately ameliorating endothelial barrier function and reducing myocardial microvascular injury.