Calpain-mediated proteolytic cleavage of troponin I induced by hypoxia or metabolic inhibition in cultured neonatal cardiomyocytes

While ischemic damage to myofibrillar proteins is thought to be responsible in part for depressed cardiac function, the relation between myofilament p rotein breakdown and chronic hypoxia has not been defined. We previously ch aracterized a chemical hypoxia model of neonatal cardiomyocytes mediated by 1 mM azide that exhibits features of calpain activation (Mol Cell Biochem 178:141-149, 1998). We here show that both hypoxia and azide-mediated metab olic inhibition induced heme oxygenase-1 expression, and caused cell death associated with lipid peroxidation. While blocking calcium influx or inhibi ting calpain activity efficiently attenuated hypoxia-induced cell injury, i t failed to prevent cell injury caused by adenoviral overexpression of the tumor suppressor protein p53. Inhibitors of caspases, on the other hand, su ppressed cell injury caused by p53 overexpression. Hypoxia caused selective cleavage of troponin I (TnI), which could be suppressed by either nifedipi ne or calpeptin. Other myofilament proteins such as troponin T, myosin heav y chain, and actin appeared to remain largely intact. p53-mediated cell inj ury exhibited proteolysis of the caspase protein substrate lamin B without appreciable breakdown of TnI. We suggest that calpain-induced TnI breakdown may constitute a unique biochemical marker associated with chronically hyp oxic cardiomyocytes.