title: Neuron-derived orphan receptor 1 transduces survival signals in neuronal cells in response to hypoxia-induced apoptotic insults.
abstract: OBJECT Hypoxia can induce cell death or trigger adaptive mechanisms to guarantee cell survival. Neuron-derived orphan receptor 1 (NOR-1) works as an early-response protein in response to a variety of environmental stresses. In this study, the authors evaluated the roles of NOR-1 in hypoxia-induced neuronal insults. METHODS Neuro-2a cells were exposed to oxygen/glucose deprivation (OGD). Cell viability, cell morphology, cas-pase-3 activity, DNA fragmentation, and cell apoptosis were assayed to determine the mechanisms of OGD-induced neuronal insults. RNA and protein analyses were carried out to evaluate the effects of OGD on expressions of NOR-1, cAMP response element-binding (CREB), and cellular inhibitor of apoptosis protein 2 (cIAP2) genes. Translations of these gene expressions were knocked down using RNA interference. Mice subjected to traumatic brain injury (TBI) and NOR-1 was immunodetected. RESULTS Exposure of neuro-2a cells to OGD decreased cell viability in a time-dependent manner. Additionally, OGD led to cell shrinkage, DNA fragmentation, and cell apoptosis. In parallel, treatment of neuro-2a cells with OGD time dependently increased cellular NOR-1 mRNA and protein expressions. Interestingly, administration of TBI also augmented NOR-1 levels in the impacted regions of mice. As to the mechanism, exposure to OGD increased nuclear levels of the transcription factor CREB protein. Downregulating CREB expression using RNA interference simultaneously inhibited OGD-induced NOR-1 mRNA expression. Also, levels of cIAP2 mRNA and protein in neuro-2a cells were augmented by OGD. After reducing cIAP2 translation, OGD-induced cell death was reduced. Sequentially, application of NOR-1 small interfering RNA to neuro-2a cells significantly inhibited OGD-induced cIAP2 mRNA expression and concurrently alleviated hypoxia-induced alterations in cell viability, caspase-3 activation, DNA damage, and cell apoptosis. CONCLUSIONS This study shows that NOR-1 can transduce survival signals in neuronal cells responsible for hypoxiainduced apoptotic insults through activation of a CREB/cIAP2-dependent mechanism.
authors.affiliation: Departments of 1 Neurosurgery.|Comprehensive Cancer Center.
authors.forename: Chung-Ching
authors.initials: CC
authors.lastname: Chio
authors.affiliation: Departments of 3 Neurosugery and.
authors.forename: Li
authors.initials: L
authors.lastname: Wei
authors.affiliation: Anesthesiology and.
authors.forename: Tyng Guey
authors.initials: TG
authors.lastname: Chen
authors.affiliation: Department of Neurosurgery.
authors.forename: Chien-Min
authors.initials: CM
authors.lastname: Lin
authors.affiliation: Anesthesiology, and.
authors.forename: Ja-Ping
authors.initials: JP
authors.lastname: Shieh
authors.affiliation: Neurology, Chi-Mei Medical Center, Tainan;
authors.forename: Poh-Shiow
authors.initials: PS
authors.lastname: Yeh
authors.affiliation: Comprehensive Cancer Center.|Graduate Institute of Medical Sciences, Taipei Medical University; |Brain Disease Research Center, Wan-Fang Hospital, Taipei Medical University; and.|Anesthetics and Toxicology Research Center, Taipei Medical University Hospital, Taipei, Taiwan.
authors.forename: Ruei-Ming
authors.initials: RM
authors.lastname: Chen
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D053148:Caspase 3 / Q000378:metabolism; D015687:Cell Hypoxia / Q000502:physiology*; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000502:physiology; D017362:Cyclic AMP Response Element-Binding Protein / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004195:Disease Models, Animal; D005947:Glucose / Q000172:deficiency*; D000860:Hypoxia / Q000503:physiopathology*; D008297:Male; D051379:Mice; D008813:Mice, Inbred ICR; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D009474:Neurons / Q000502:physiology*; D012333:RNA, Messenger / Q000378:metabolism; D011987:Receptors, Steroid / Q000378:metabolism*; D011988:Receptors, Thyroid Hormone / Q000378:metabolism*
chemical_list: D017362:Cyclic AMP Response Element-Binding Protein; D004268:DNA-Binding Proteins; D009419:Nerve Tissue Proteins; C490411:Nr4a3 protein, mouse; D012333:RNA, Messenger; D011987:Receptors, Steroid; D011988:Receptors, Thyroid Hormone; C505409:Casp3 protein, mouse; D053148:Caspase 3; D005947:Glucose
keywords: CREB = cAMP response element-binding; CREB/cIAP2; DMEM = Dulbecco’s modified Eagle’s medium; DMSO = dimethyl sulfoxide; ELISA = enzyme-linked immunosorbent assay; GBM = glioblastoma multiforme; NOR-1; NOR-1 = neuron-derived orphan receptor 1; OGD = oxygen/glucose-deprivation; PAGE = polyacrylamide gel electrophoresis; PBS = phosphate-buffered saline; PCNA= proliferating cell nuclear antigen; PI = propidium iodide; PMSF = phenylmethylsulfonyl fluoride; RNAi = RNA interference; RT-PCR = reverse transcription polymerase chain reaction; SDS = sodium dodecyl sulfate; TBI = traumatic brain injury; cIAP2 = cellular inhibitor of apoptosis protein 2; hypoxia; mAb = monoclonal antibody; neuronal cell; pAb = polyclonal antibody; siRNA = small interfering RNA; survival signals; traumatic brain injury