Apoptosis in rat cardiac myocytes induced by Fas ligand: priming for Fas-mediated apoptosis with doxorubicin.
Fas/Fas ligand (FasL) is well known for its role in delivering apoptotic signals; however, it is unclear whether FasL can mediate apoptosis in cardiomyocytes. We hypothesized that apoptosis via Fas/FasL system may be augmented in damaged cardiomyocytes. To determine whether FasL mediates cardiomyocyte apoptosis, recombinant FasL (rFasL) was added to the culture of neonatal rat ventricular myocytes pretreated with and without doxorubicin. Without doxorubicin, high dose of rFasL caused an increase in TUNEL-positive cardiomyocytes and a mild decrease in MTT activities. When cardiomyocytes were pretreated with doxorubicin (0.5 microM), rFasL dramatically augmented TUNEL-positive cardiomyocytes in a concentration-dependent manner, which was accompanied with nuclear fragmentations. The rFasL also caused a concentration-dependent reduction in MTT activities in cardiomyocytes. The rFasL-induced caspase-8 activity was greatly facilitated by pretreatment of doxorubicin. TUNEL-positive nuclei with rFasL was inhibited by Fas-Fc, neutralizing agent of rFasL, and Z-IETD-FMK, caspase-8 inhibitor. Fas mRNA transcript by RT-PCR was up-regulated in cardiomyocytes with doxorubicin. We conclude that FasL can induce cardiomyocyte apoptosis particularly when cardiomyocyte becomes susceptible for Fas-mediated apoptosis.
Yamaoka
M
M
The First Department of Internal Medicine, Yamagata University School of Medicine, Iida-Nishi, Japan.
Yamaguchi
S
S
Suzuki
T
T
Okuyama
M
M
Nitobe
J
J
Nakamura
N
N
Mitsui
Y
Y
Tomoike
H
H
D000818:Animals; D017209:Apoptosis; D053181:Caspase 8; D053453:Caspase 9; D061945:Caspase Inhibitors; D002478:Cells, Cultured; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D004317:Doxorubicin / Q000494:pharmacology*; D053222:Fas Ligand Protein; D006352:Heart Ventricles / Q000166:cytology; D006801:Humans; D008562:Membrane Glycoproteins / Q000378:metabolism* / Q000494:pharmacology; D009842:Oligopeptides / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D011994:Recombinant Proteins / Q000378:metabolism / Q000494:pharmacology; D015854:Up-Regulation; D019014:fas Receptor / Q000235:genetics
D061945:Caspase Inhibitors; D015853:Cysteine Proteinase Inhibitors; C505661:FASLG protein, human; D053222:Fas Ligand Protein; C505662:Faslg protein, rat; D008562:Membrane Glycoproteins; D009842:Oligopeptides; D012333:RNA, Messenger; D011994:Recombinant Proteins; C403753:benzyloxycarbonyl-isoleucyl-glutamyl-threonyl-aspartic acid fluoromethyl ketone; D019014:fas Receptor; D004317:Doxorubicin; C505530:CASP8 protein, human; C506433:CASP9 protein, human; C505528:Casp8 protein, rat; C506435:Casp9 protein, rat; D053181:Caspase 8; D053453:Caspase 9