Involvement of c-Jun NH2-terminal kinase and nitric oxide-mediated mitochondria-dependent intrinsic pathway signaling in cardiotoxin-induced muscle cell death: role of testosterone.
To test the hypothesis that c-Jun NH2-terminal kinase (JNK) and nitric oxide (NO)-mediated signaling plays an important role in muscle cell apoptosis, we examined the contribution of these molecules in muscle cell apoptosis during cardiotoxin (ctx)-induced muscle injury in mice. Compared to controls, where no apoptosis was detected, the percent of muscle cell apoptosis rose significantly (P < 0.05) at 4 h (27%) after ctx-treatment and increased further progressively up to 16 h posttreatment (80%), before it fell again at 24 h posttreatment (38%). Initiation of apoptosis was preceded by JNK activation and elevated levels of B-cell lymphoma-2 (BCL-2) in the mitochondrial fractions (BAX levels remained unaffected). Ctx treatment also resulted in the inactivation of BCL-2 through phosphorylation at serine 70, thereby perturbing the BAX/BCL-2 rheostat, and the subsequent activation of the cytochrome c-mediated death pathway. Concomitant administration of SP600125, a selective JNK inhibitor, or aminoguanidine (AG), a selected inducible nitric oxide synthase (iNOS) inhibitor, effectively diminished BCL-2 phosphorylation, suppressed cytochrome c release from mitochondria and caspase activation, and significantly prevented ctx-induced muscle cell apoptosis. In additional studies, we examined the role of testosterone in preventing such ctx-induced muscle cell apoptosis. Collectively, the present study emphasizes the role of a new signal transduction pathway involving JNK and iNOS that promotes ctx-induced myocyte apoptosis by provoking BCL-2 phosphorylation, leading to its inactivation, and subsequent activation of the intrinsic pathway signaling. Testosterone therapy has no protective effect in acute muscle injury associated with increased muscle cell death after ctx-treatment.
Sinha-Hikim
Indrani
I
Division of Endocrinology, Metabolism, and Molecular Medicine, Charles R. Drew University, Los Angeles, CA 90059, USA. insinhah@cdrewu.edu
Braga
Mellisa
M
Shen
Ruoqing
R
Sinha Hikim
Amiya P
AP
D000818:Animals; D054715:Cardiotoxins / Q000494:pharmacology*; D016923:Cell Death / Q000187:drug effects; D045744:Cell Line, Tumor; D006801:Humans; D048031:JNK Mitogen-Activated Protein Kinases / Q000502:physiology*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008928:Mitochondria / Q000502:physiology*; D018485:Muscle Fibers, Skeletal / Q000166:cytology / Q000187:drug effects* / Q000201:enzymology*; D009569:Nitric Oxide / Q000502:physiology*; D015398:Signal Transduction / Q000502:physiology*; D013739:Testosterone / Q000502:physiology*