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| 瞬时受体电位阳离子通道M7在海马神经元损伤中的作用 |
| Roles of transient receptor potential melastatin 7 in hippocampal neuron injury |
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| DOI：10.12089/jca.2018.01.017 |
| 中文关键词: TRPM7 七氟醚 缺血预处理 缺血缺氧性损伤 神经元凋亡 炎症反应 |
| 英文关键词: Transient receptor potential melastatin 7 Sevoflurane Ischemic preconditioning Oxygen-glucose deprivation Neuronal apoptosis Inflammatory responses |
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| 中文摘要: |
| 目的 探讨瞬时受体电位阳离子通道 M7(TRPM7)在七氟醚预处理缓解缺血缺氧性损伤(OGD)后海马神经元损伤中的作用。方法 出生1 d的SD大鼠,提取海马神经元,将其随机分为五组：对照组(C组)、七氟醚预处理组(Sev组)、OGD组、七氟醚预处理+OGD组(SD组)和七氟醚预处理+缓激肽(TRPM7特异性激动剂)+OGD(B组)。缺糖缺氧1.5 h后复糖复氧,再正常培养24 h以制备OGD模型。C组海马神经元仅做正常培养;Sev组海马神经元行2%七氟醚预处理1 h;OGD组海马神经元仅制备OGD模型;SD组海马神经元行2%七氟醚预处理1 h,24 h后制备OGD模型; B组神经元于七氟醚预处理前15 min在培养基中加入缓激肽(TRPM7特异性激动剂,终浓度200 μmol/L),之后行2%七氟醚预处理1 h,24 h后制备OGD模型。正常培养24 h后,分别采用MTT法检测神经元相对存活指数,TUNEL凋亡染色法检测神经元凋亡率,Western blot检测TRPM7蛋白含量,实时定量PCR法检测TRPM7 mRNA表达水平,ELISA法测定神经元IL-1β和TNF-α蛋白含量。结果 OGD组海马神经元TRPM7蛋白含量及mRNA表达水平、凋亡率、IL-1β、TNF-α mRNA表达水平及上清蛋白含量明显高于C组(P<0.05),而相对存活指数明显降低于C组(P<0.05)。SD组海马神经元TRPM7蛋白含量及mRNA表达水平、凋亡率、IL-1β、TNF-α mRNA表达水平及上清蛋白含量明显低于OGD组(P<0.05),而相对存活指数明显高于OGD组(P<0.05)。B组海马神经元TRPM7蛋白含量及 mRNA表达水平、凋亡率、IL-1β、TNF-α mRNA表达水平及上清蛋白含量明显高于SD组(P<0.05),而相对存活指数明显低于SD组(P<0.05)。结论 七氟醚预处理可通过缓解神经元TRPM7过度表达,减轻缺血缺氧性损伤后海马神经元凋亡和炎症反应。 |
| 英文摘要: |
| Objective To investigate the role of transient receptor potential melastatin 7 (TRPM7) in the protective role of sevoflurane preconditioning against hippocampal neuron injury caused by oxygen glucose deprivation (OGD). Methods Hippocampal neurons were harvested from postnatal day 1 SD rats, and randomly divided into 5 groups: control group (group C), sevoflurane group (group Sev), oxygen-glucose deprivation group (group OGD), sevoflurane+OGD group (group SD) and sevoflurane+OGD+bradykinin group (group B). To build up the model of OGD, the neurons were cultured in a deoxygenated glucose-free medium and exposed to 95% N2 and 5% CO2 in an anaerobic chamber equilibrated at 37℃ for 1.5 h, followed by replacement with glucose-containing medium and return to a standard incubator for additional 24 h. The neurons in group C received no treatment. Group OGD was preconditioned with 2% sevoflurane for 1 h. The neurons in group OGD were subjected to OGD. Group SD was preconditioned with 2% sevoflurane for 1 h, followed by OGD at 24 h after the sevoflurane exposure. The neurons in group B was incubated in a medium supplemented with 200 μmol/L bradykinin (the selective agonist of TRPM7), followed sequentially by the preconditioning of 2% sevoflurane for 1 h and then OGD challenge. Twenty-four hours after re-oxygenation, The relative neuronal cell viability was determined by MTT assay, the neuronal apoptotic rate was analyzed by TUNEL assay, the protein expression of TRPM7 was detected by Western blot, the mRNA level of TRPM7 was estimated by real-time PCR, the neuronal release of IL-1β and TNF-α in the serum was measured by ELISA. Results Compared with group C, the mRNA and protein levels of TRPM7, the neuronal apoptotic rate, the mRNA and supernatant protein levels of IL-1β and TNF-α were significantly up-regulated in group OGD (P<0.05), whereas the cell viability was decreased (P<0.05). Compared with group OGD, the mRNA and protein levels of TRPM7, the neuronal apoptotic rate, the mRNA and supernatant protein levels of IL-1β and TNF-α were significantly down-regulated in group SD (P<0.05), whereas the cell viability was increased (P<0.05). Compared with group SD, the mRNA and protein levels of TRPM7, the neuronal apoptotic rate, the mRNA and supernatant protein levels of IL-1β and TNF-α were significantly p-regulated in group B (P<0.05), whereas the cell viability was decreased (P<0.05). Conclusion Sevoflurane attenuates apoptosis and inflammatory responses induced by OGD via reduction of the overexpression of TRPM7 in the hippocampal neurons. |
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