title: Delayed cytoprotection induced by hypoxic preconditioning in cultured neonatal rat cardiomyocytes: role of GRP78.
abstract: Hypoxic preconditioning (HPC) has been well demonstrated to have potent protective effects in many cell types; however, the mechanisms responsible for this phenomenon are not fully understood. Recently, glucose-regulated protein 78 (GRP78), an inducible molecular chaperon, was indicated to be associated with ischemic preconditioning. We hypothesized that HPC protects cardiomyocytes against hypoxia by inducing GRP78 in cultured neonatal rat cardiomyocytes. HPC was induced by exposing cardiomyocytes to brief hypoxia (1% O(2), 30 min) followed by reoxygenation. GRP78 was expressed constitutively in cultured cardiomyocytes and its expression was enhanced at 12 h, peaked at 24 h (207.3+/-23.6% of the baseline), and was sustained for up to 72 h after HPC. Twenty-four hours after HPC, the myocytes were subjected to prolonged hypoxia (1% O(2), 12 h). The lactic dehydrogenase (LDH) release and malondialdehyde (MDA) content were reduced, while cell viability and superoxide dismutase (SOD) activity were increased in the preconditioned cells compared with the non-HPC cells. The GRP78 protein level was higher in cells exposed to both HPC and hypoxia than in the cells exposed to HPC alone or hypoxia alone. Heat shock protein 70 (HSP70) was induced in parallel by late HPC. Transfection of GRP78 antisense oligonucleotides blocked GRP78 expression but not HSP70, resulting in attenuated cardioprotection afforded by late HPC. Furthermore, inducing GRP78 by gene transfer protected cardiomyocytes from hypoxic injury. These findings demonstrate that the induction of GRP78 partially mediates the late HPC, suggesting that GRP78 is a novel mechanism responsible for the late cytoprotection of HPC.
authors.affiliation: Department of Physiology, Second Military Medical University, Shanghai 200433, China.
authors.forename: Yan-Xia
authors.initials: YX
authors.lastname: Pan
authors.forename: An-Jing
authors.initials: AJ
authors.lastname: Ren
authors.forename: Juan
authors.initials: J
authors.lastname: Zheng
authors.forename: Wei-Fang
authors.initials: WF
authors.lastname: Rong
authors.forename: Hong
authors.initials: H
authors.lastname: Chen
authors.forename: Xiao-Hong
authors.initials: XH
authors.lastname: Yan
authors.forename: Chang
authors.initials: C
authors.lastname: Wu
authors.forename: Wen-Jun
authors.initials: WJ
authors.lastname: Yuan
authors.forename: Li
authors.initials: L
authors.lastname: Lin
mesh_terms: D000818:Animals; D000831:Animals, Newborn; D015687:Cell Hypoxia / Q000187:drug effects; D002478:Cells, Cultured; D019610:Cytoprotection / Q000187:drug effects; D005786:Gene Expression Regulation / Q000187:drug effects; D018840:HSP70 Heat-Shock Proteins / Q000378:metabolism; D006360:Heat-Shock Proteins / Q000037:antagonists & inhibitors / Q000096:biosynthesis*; D019157:Ischemic Preconditioning, Myocardial; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008315:Malondialdehyde / Q000378:metabolism; D018832:Molecular Chaperones / Q000037:antagonists & inhibitors / Q000096:biosynthesis*; D032383:Myocytes, Cardiac / Q000378:metabolism* / Q000473:pathology; D020319:Oligodeoxyribonucleotides, Antisense / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D013482:Superoxide Dismutase / Q000378:metabolism; D013997:Time Factors; D000091342:Endoplasmic Reticulum Chaperone BiP
chemical_list: D018840:HSP70 Heat-Shock Proteins; D006360:Heat-Shock Proteins; D007770:L-Lactate Dehydrogenase; D008315:Malondialdehyde; D018832:Molecular Chaperones; D020319:Oligodeoxyribonucleotides, Antisense; D013482:Superoxide Dismutase; C051684:GRP78 protein, rat; D000091342:Endoplasmic Reticulum Chaperone BiP