Release kinetics of intact and degraded troponin I and T after irreversible cell damage

PurposeWe characterized the release kinetics of cardiac troponin I and T in relation to lactate dehydrogenase (LDH) from cardiomyocytes before and after the transition from reversible to irreversible cell damage.MethodsCardiomyocytes were exposed to mild metabolic inhibition (1 mmol/L sodium azide) to induce a necrotic cell death process that is characterized by a reversible (0–12 h) and irreversible phase (12–30 h). At various time intervals cells and media were collected and analyzed for LDH activity, intact cTnI and cTnT, and their degradation products.ResultsDuring the first 12 h of metabolic inhibition, cell viability was unchanged with no release of intact cTnI and cTnT nor their degradation products. Between 12 and 30 h of azide treatment, cardiomyocytes showed progressive cell death accompanied by release of intact cTnI (29 kDa), intact cTnT (39 kDa), four cTnI degradation products of 26, 20, 17 and 12 kDa, and three cTnT degradation products of 37, 27 and 14 kDa. Possibly due to degradation, there is progressive loss of cTnI and cTnT protein that is obviously undetected by the antibodies used.ConclusionsMetabolic inhibition of cardiomyocytes induces a parallel release of intact cTnI and cTnT and their degradation products, starting only after onset of irreversible cardiomyocyte damage.