Overexpression of endoplasmic reticulum-resident chaperone attenuates cardiomyocyte death induced by proteasome inhibition.
OBJECTIVE
Proteasome inhibitors are a novel class of anticancer agents that induce tumour cell death via endoplasmic reticulum (ER) stress. Since ER stress is involved in the development of heart failure, we investigated the role of ER-initiated cardiomyocyte death by proteasome inhibition.


RESULTS
Rat neonatal cardiomyocytes were used in this study. Proteasome activity was assayed using proteasome peptidase substrates. Cell viability and apoptosis were measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenol tetrazolium bromide and flow cytometry, respectively. Western blot analysis, real-time polymerase chain reaction (PCR) and reverse transcriptional PCR were used to detect the expression of protein and messenger ribonucleic acid (RNA). The location of overexpressed glucose-regulated protein (GRP) 78 was observed by confocal fluorescence microscopy. Proteasome inhibition induced cardiomyocyte death and activated ER stress-induced transcriptional factor ATF6, but not XBP1 (X-box binding protein 1), without up-regulating ER chaperones. ER-initiated apoptosis signalling, including cytosine-cytosine-adenine-adenine-thymine enhancer-binding protein (C/EBP) homologous protein (CHOP), c-Jun-N-terminal kinase (JNK), and caspase-12, was activated by proteasome inhibition. Short interference RNA targeting CHOP, but not the blockage of caspase-12 or JNK pathway, attenuated cardiomyocyte death. Overexpression of GRP78 suppressed both CHOP expression and cardiomyocyte death by proteasome inhibition.


CONCLUSIONS
These findings demonstrate that proteasome inhibition induces ER-initiated cardiomyocyte death via CHOP-dependent pathways without compensatory up-regulation of ER chaperones. Supplement and/or pharmacological induction of GRP78 can attenuate cardiac damage by proteasome inhibition.
Fu
Hai Ying
HY
Department of Cardiovascular Medicine, National Cardiovascular Center, Suita, Osaka 565-8565, Japan.
Minamino
Tetsuo
T
Tsukamoto
Osamu
O
Sawada
Tamaki
T
Asai
Mitsutoshi
M
Kato
Hisakazu
H
Asano
Yoshihiro
Y
Fujita
Masashi
M
Takashima
Seiji
S
Hori
Masatsugu
M
Kitakaze
Masafumi
M
D051702:Activating Transcription Factor 6 / Q000378:metabolism; D000818:Animals; D000831:Animals, Newborn; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D053454:Caspase 12 / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D004721:Endoplasmic Reticulum / Q000187:drug effects* / Q000201:enzymology / Q000473:pathology; D006360:Heat-Shock Proteins / Q000378:metabolism; D048031:JNK Mitogen-Activated Protein Kinases / Q000378:metabolism; D007976:Leupeptins / Q000494:pharmacology; D018832:Molecular Chaperones / Q000235:genetics / Q000378:metabolism*; D032383:Myocytes, Cardiac / Q000187:drug effects* / Q000201:enzymology / Q000473:pathology; D009842:Oligopeptides / Q000494:pharmacology; D018384:Oxidative Stress / Q000187:drug effects; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D061988:Proteasome Inhibitors; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000378:metabolism; D051381:Rats; D015398:Signal Transduction / Q000187:drug effects; D051742:Transcription Factor CHOP / Q000378:metabolism; D014415:Tunicamycin / Q000494:pharmacology; D000091342:Endoplasmic Reticulum Chaperone BiP
D051702:Activating Transcription Factor 6; D000970:Antineoplastic Agents; D053454:Caspase 12; D015853:Cysteine Proteinase Inhibitors; D006360:Heat-Shock Proteins; D048031:JNK Mitogen-Activated Protein Kinases; D007976:Leupeptins; D018832:Molecular Chaperones; D009842:Oligopeptides; D046988:Proteasome Endopeptidase Complex; D061988:Proteasome Inhibitors; D034741:RNA, Small Interfering; D051742:Transcription Factor CHOP; D014415:Tunicamycin; C497113:Atf6 protein, rat; C051684:GRP78 protein, rat; D000091342:Endoplasmic Reticulum Chaperone BiP; C072553:benzyloxycarbonylleucyl-leucyl-leucine aldehyde; C078846:epoxomicin