Calcium dysregulation disrupts mitochondrial homeostasis by interfering AMPK/Drp1 pathway to aggravate plaque progression and instability.

Rationale: Inactivation of Cys(674) (C674) of sarcoplasmic/endoplasmic reticulum calcium ATPase 2 (SERCA2) disrupts intracellular calcium (Ca(2+)) homeostasis and SERCA2 dysfunction has been implicated in the pathogenesis of atherosclerosis and aortic aneurysms. However, the precise role of SERCA2 dysfunction in aortic smooth muscle cells (SMCs) and its contribution to atherosclerosis remains unclear. Methods: We Used heterozygous SERCA2 C674S knock-in (SKI) mice to mimic the partial irreversible oxidation inactivation of C674 thiol under pathological conditions. The whole aorta and aortic root were isolated for immunohistological analysis, RNA sequencing and proteomic analysis. The primary SMCs were collected for cell culture, protein expression and immunofluorescence analysis. Results: Compared with SMCs from WT mice, SKI SMCs demonstrated abnormally activated AMPK/Drp1 pathway adenosine 5'-monophosphate-activated protein kinase (AMPK)/dynamin related protein 1 (Drp1) pathway, and mitochondrial disorders, including increased cytosolic/mitochondrial Ca²⁺ level, oxidative stress, ATP depletion, decreased mitochondrial membrane potential (Δψm), and disrupted mitochondrial dynamics. In SKI SMCs, stimulation of AMPK by metformin or 5-aminoimidazole-4-carboxamide-1-β-D-ribofuranoside (AICAR), or inhibition of Drp1 with mitochondrial division inhibitor 1 (Mdivi-1), restored mitochondrial homeostasis, mitigated excessive matrix metalloproteinase 2 and SMC apoptosis, thereby preserved SMC function. In vivo administration of metformin and Mdivi-1 both ameliorated atherosclerosis triggered by SERCA2 dysfunction and particularly enhanced plaque stability. Conclusions: SERCA2 dysfunction accelerates atherosclerotic plaques formation and increases plaque vulnerability by disrupting the AMPK/Drp1 pathway in aortic SMCs, leading to mitochondrial disorders and impairing SMCs function. Targeting of AMPK or Drp1 pharmacologically may offer promising therapeutic avenues for atherosclerosis, particularly in reducing atherosclerotic plaques vulnerability.