title: Calpain-1 Mediated Disorder of Pyrophosphate Metabolism Contributes to Vascular Calcification Induced by oxLDL.
abstract: We previously reported that oxidized low density lipoprotein (oxLDL) accelerated the calcification in aorta of rats and rat vascular smooth muscle cells (RVSMCs). However, the molecular mechanism underlying the acceleration remains poorly understood. The present study aimed to investigate the role of calpain-1, Ca2+-sensitive intracellular cysteine proteases, in the vascular calcification of rats treated with both high dose of vitamin D2 and high cholesterol diet. The results showed that calpain activity significantly increased in calcified aortic tissue of rats and RVSMCs treated with oxLDL. Specific calpain inhibitor I (CAI, 0.5mg/kg, intraperitoneal) inhibited the vascular calcification in rats with hypercholesterolemia accompanied by the increase in the level of extracellular inorganic pyrophosphate (PPi), the endogenous inhibitor of vascular calcification. In addition, CAI increased the content of adenosine triphosphate (ATP), decreased the activity, mRNA and protein expression of alkaline phosphatase (ALP) and reduced the production of superoxide anion in calcified aortic tissue. CAI also increased the activity of ATP synthase as well as protein expression of ATP5D, δ subunit of ATP synthase. In the in vitro study, suppression of calpain-1 using siRNA assay inhibited the calcium deposition, increased the levels of PPi and ATP, improved the activity of ATP synthase as well as protein expression of ATP5D in RVSMCs treated with oxLDL. Calpain-1 suppression also decreased the activity, mRNA and protein expression of ALP and reduced the mitochondrial ROS (Mito-ROS) production in RVSMCs. However, mito-TEMPO, the mitochondria-targeted ROS scavenger, reduced the calcium deposition, increased the PPi in culture medium, decreased the activity, mRNA and protein expression of ALP in RVSMCs treated with oxLDL. Taken together, the results suggested that calpain-1 activation plays critical role in vascular calcification caused by oxLDL, which might be mediated by PPi metabolism disorder. The results also implied that Mito-ROS might contribute to the PPi metabolism disorder through regulation of the activity and expression of ALP.
authors.affiliation: Key Laboratory of Cardio- and Cerebro-vascular Drug Research of Liaoning Province, Drug Research Institute, Liaoning Medical University, Jinzhou, China.
authors.forename: Futian
authors.initials: F
authors.lastname: Tang
authors.affiliation: Cerebrovascular Diseases Center of Gansu Provincial Hospital, Lanzhou, China.
authors.forename: Erqing
authors.initials: E
authors.lastname: Chan
authors.affiliation: Key Laboratory of Cardio- and Cerebro-vascular Drug Research of Liaoning Province, Drug Research Institute, Liaoning Medical University, Jinzhou, China.
authors.forename: Meili
authors.initials: M
authors.lastname: Lu
authors.affiliation: Guangzhou Institute of Sports Science, Guangzhou, China.
authors.forename: Xiaowen
authors.initials: X
authors.lastname: Zhang
authors.affiliation: Key Laboratory of Cardio- and Cerebro-vascular Drug Research of Liaoning Province, Drug Research Institute, Liaoning Medical University, Jinzhou, China.
authors.forename: Chunmei
authors.initials: C
authors.lastname: Dai
authors.affiliation: Key Laboratory of Cardio- and Cerebro-vascular Drug Research of Liaoning Province, Drug Research Institute, Liaoning Medical University, Jinzhou, China.
authors.forename: Meng
authors.initials: M
authors.lastname: Mei
authors.affiliation: Key Laboratory of Cardio- and Cerebro-vascular Drug Research of Liaoning Province, Drug Research Institute, Liaoning Medical University, Jinzhou, China.
authors.forename: Suping
authors.initials: S
authors.lastname: Zhang
authors.affiliation: Key Laboratory of Cardio- and Cerebro-vascular Drug Research of Liaoning Province, Drug Research Institute, Liaoning Medical University, Jinzhou, China.
authors.forename: Hongxin
authors.initials: H
authors.lastname: Wang
authors.affiliation: Guangzhou Vocational and Technical College, Guangzhou, China.
authors.forename: Qing
authors.initials: Q
authors.lastname: Song
mesh_terms: D000818:Animals; D001011:Aorta / Q000378:metabolism / Q000473:pathology; D002118:Calcium / Q000378:metabolism; D002154:Calpain / Q000378:metabolism*; D002460:Cell Line; D011756:Diphosphates / Q000378:metabolism*; D006023:Glycoproteins / Q000378:metabolism; D006937:Hypercholesterolemia / Q000150:complications / Q000378:metabolism* / Q000473:pathology; D008077:Lipoproteins, LDL / Q000378:metabolism*; D008297:Male; D008928:Mitochondria / Q000378:metabolism / Q000473:pathology; D009131:Muscle, Smooth, Vascular / Q000378:metabolism / Q000473:pathology; D032389:Myocytes, Smooth Muscle / Q000378:metabolism / Q000473:pathology; D006180:Proton-Translocating ATPases / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D061205:Vascular Calcification / Q000150:complications / Q000378:metabolism* / Q000473:pathology
chemical_list: D011756:Diphosphates; D006023:Glycoproteins; D008077:Lipoproteins, LDL; C039304:calpain inhibitors; C099252:oxidized low density lipoprotein; C107241:diphosphoric acid; D002154:Calpain; C000606071:Capn1 protein, rat; D006180:Proton-Translocating ATPases; D002118:Calcium