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


METHODS
Cardiomyocytes 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.


RESULTS
During 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.


CONCLUSIONS
Metabolic inhibition of cardiomyocytes induces a parallel release of intact cTnI and cTnT and their degradation products, starting only after onset of irreversible cardiomyocyte damage.
Hessel
M H M
MH
Department of Cardiology, C5-P, Leiden University Medical Center, Albinusdreef 2, P.O. Box 9600, 2300 RC, Leiden, The Netherlands.
Michielsen
E C H J
EC
Atsma
D E
DE
Schalij
M J
MJ
van der Valk
E J M
EJ
Bax
W H
WH
Hermens
W T
WT
van Dieijen-Visser
M P
MP
van der Laarse
A
A
D000818:Animals; D000831:Animals, Newborn; D018929:Cell Culture Techniques; D016923:Cell Death / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D003470:Culture Media / Q000032:analysis; D016895:Culture Media, Serum-Free / Q000032:analysis; D004791:Enzyme Inhibitors / Q000633:toxicity; D006352:Heart Ventricles / Q000166:cytology; D007118:Immunoassay; D007700:Kinetics; D007770:L-Lactate Dehydrogenase / Q000032:analysis / Q000378:metabolism; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology*; D009336:Necrosis / Q000139:chemically induced / Q000473:pathology; D051381:Rats; D017208:Rats, Wistar; D019810:Sodium Azide / Q000633:toxicity; D019210:Troponin I / Q000032:analysis / Q000378:metabolism*; D020107:Troponin T / Q000032:analysis / Q000378:metabolism*
D003470:Culture Media; D016895:Culture Media, Serum-Free; D004791:Enzyme Inhibitors; D019210:Troponin I; D020107:Troponin T; D019810:Sodium Azide; D007770:L-Lactate Dehydrogenase