Inhibition of tumor necrosis factor-alpha-dependent cardiomyocyte apoptosis by metallothionein.
Myocardial cell death is an important cellular event of heart failure. Tumor necrosis factor-alpha (TNF) accumulates in the failing heart and causes myocyte apoptosis, but the mechanism of this action is unclear. This study was undertaken to examine the relationship between TNF-induced cardiomyocyte apoptosis and activation of p38 mitogen-activated protein kinase (MAPK) through oxidative stress. Primary cultures of neonatal cardiomyocytes isolated from transgenic mouse hearts that overexpress metallothionein (MT) as well as cardiomyocytes isolated from wild-type mice were used. The treatment of wildtype cardiomyocytes with TNF at 10 ng/mL induced apoptosis, as detected by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling and confirmed by Annexin V-fluorescein isothiocyanate binding. The apoptotic effect of TNF was significantly inhibited in the MT-overexpressing cardiomyocytes. Corresponding to the apoptotic effect, TNF at 10 ng/mL caused rapid phosphorylation of p38 MAPK in wild-type cardiomyocytes. The activation of p38 MAPK was further confirmed by an in vivo experiment treating the mice with TNF and measuring p38 MAPK activity using an immune complex kinase assay. The activation of p38 MAPK was not observed in the MT-overexpressing cardiomyocytes either in vitro or in vivo. Importantly, TNF-induced accumulation of reactive oxygen species was dramatically reduced in the MT-overexpressing cardiomyocytes as determined by a carboxy-H(2)-DCFDA staining method. This study thus suggests that p38 MAPK activation is likely involved in TNFinduced cardiomyocyte apoptosis, which is also related to reactive oxygen species accumulation.
Klein
Jon B
JB
Department of Medicine, University of Louisville School of Medicine, Louisville, KY 40202, USA. jon.klein@louisville.edu
Wang
Guang-Wu
GW
Zhou
Zhanxiang
Z
Buridi
Abdul
A
Kang
Y James
YJ
D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D002478:Cells, Cultured; D008668:Metallothionein / Q000096:biosynthesis / Q000235:genetics / Q000502:physiology*; D051379:Mice; D008822:Mice, Transgenic; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D032383:Myocytes, Cardiac / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D017382:Reactive Oxygen Species / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000037:antagonists & inhibitors* / Q000502:physiology*; D048051:p38 Mitogen-Activated Protein Kinases
D017382:Reactive Oxygen Species; D014409:Tumor Necrosis Factor-alpha; D008668:Metallothionein; D020928:Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases