Norepinephrine induces apoptosis in neonatal rat cardiomyocytes through a reactive oxygen species-TNF alpha-caspase signaling pathway.
OBJECTIVE
Norepinephrine (NE)-induced apoptosis in cardiomyocytes is an important cause of heart failure. Previous studies revealed that reactive oxygen species (ROS) are involved in apoptosis. Tumor necrosis factor-alpha (TNF), a well-known mediator that stimulates apoptosis, is not only produced by macrophages but also by cardiomyocytes. Until now, the role of TNF and its relationship to ROS in NE-induced apoptosis of cardiomyocytes have never been investigated.


METHODS
Neonatal rat cardiomyocytes were treated with various concentrations of NE. Apoptosis of cardiomyocytes was determined using the TUNEL assay. The level of secreted TNF was measured by ELISA and TNF mRNA expression was determined by semiquantitative reverse transcriptional polymerase chain reaction. Caspase activity was measured by a fluorogenic protease assay kit. Anti-TNF antibodies, caspase inhibitors and antioxidants (N-acetyl-L-cysteine or vitamin C) were added to determine if they could inhibit the apoptotic effect of NE.


RESULTS
NE induced apoptosis of cardiomyocytes in a dose- and time-dependent manner. NE up-regulated TNF mRNA expression and increased TNF secretion and caspase-2,-3,-6, and -9 activities. A neutralizing anti-TNF antibody and caspase-2 and -3 inhibitors significantly attenuated NE-induced apoptosis. Antioxidants completely abrogated NE-induced TNF secretion, caspase activation, and apoptotic death.


CONCLUSIONS
NE induced apoptosis in neonatal rat cardiomyocytes through a ROS-TNF-caspase signaling pathway.
Fu
Yun-Ching
YC
Department of Education and Research, Taichung Veterans General Hospital, and Institute of Clinical Medicine, National Yang-Ming University, Taipei, Taiwan, ROC.
Chi
Ching-Shiang
CS
Yin
Sui-Chu
SC
Hwang
Betau
B
Chiu
Yung-Tsung
YT
Hsu
Shih-Lan
SL
D000818:Animals; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D000975:Antioxidants / Q000494:pharmacology; D017209:Apoptosis; D020169:Caspases / Q000378:metabolism; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D032383:Myocytes, Cardiac / Q000378:metabolism* / Q000473:pathology*; D009638:Norepinephrine / Q000494:pharmacology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D017382:Reactive Oxygen Species / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects*; D013997:Time Factors; D014409:Tumor Necrosis Factor-alpha / Q000276:immunology / Q000378:metabolism
D000911:Antibodies, Monoclonal; D000975:Antioxidants; D017382:Reactive Oxygen Species; D014409:Tumor Necrosis Factor-alpha; D020169:Caspases; D009638:Norepinephrine