幼鼠癫痫模型海马神经元NMDA受体表达与MMP-3表达相关性
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摘要
目的探讨幼鼠癫痫模型海马神经元N-甲基-D-天冬氨酸受体(NMDAR)表达与基质金属蛋白酶-3(MMP-3)表达的相关性。方法将60只2~4周龄SPF级C57/BL6小鼠随机分为正常组、癫痫组、MMP-3抑制剂组、NMDAR抑制剂组,每组15只。采用匹罗卡品建立幼鼠癫痫模型;采用RT-PCR检测海马神经元NMDAR_(2A)、NMDAR_(2B)、MMP-3 mRNA表达;采用尼氏染色观察海马区神经元形态;采用Western blot检测海马区P-糖蛋白(P-gp)、钙调蛋白激酶Ⅱ(CAMKⅡ)蛋白表达。结果幼鼠癫痫模型海马区NMDAR_(2A/B)、MMP-3 mRNA以及P-gp、CAMKⅡ蛋白较正常组显著增加(P<0.05)。注射NMDA受体抑制剂或MMP-3抑制剂可显著降低幼鼠癫痫模型NMDAR_(2A/B)、MMP-3 mRNA及P-gp、CAMKⅡ蛋白表达,且明显改善海马区神经元结构。结论幼鼠癫痫模型海马神经元MMP-3表达与MNDAR表达可能具有相关性。
Objective To explore the relativity of N-methyl-D-aspartic acid(NMDA) receptors with Matrix metalloproteinases-3(MMP-3) expression in hippocampal neurons in juvenile mice with epilepsy.Methods Sixty juvenile mice were divided into 4 groups of 15 animals each, including control, epilepsy model, NMDA receptor(NMDAR) 2A, B inhibitor and MMP-3 inhibitor groups. The epilepsy model was established in all the groups except the control group. The expressions of NMDAR 2 A, and MMP-3 RNA in the hippocampal tissues were detected by RT-PCR in all the groups. The morphology of hippocampal neurons was observed by Nissl staining in all the groups. The expressions of P-glycoprotein(P-gp) and Calmodulin kinase Ⅱ(CAMK Ⅱ) proteins in the hippocampal tissues were detected by Western blot in all the groups.Results The levels of the expressions of NMDAR2A, B and MMP-3 mRNA P-gp and CAMK Ⅱ in the hippocampus tissues were significantly higher in the epilepsy model group than those in the control group(P<0.05). The levels of NMDAR2A, B and MMP-3 mRNA, and P-gp and CAMK Ⅱ in the hippocampal tissues were significantly lower in NMDAR2A, B inhibitor and MMP-3 inhibitors groups than those in the epilepsy model group(P<0.05). The structures in the hippocampal neurons were significantly improved in the NMDAR2A, B in hibitor and MMP-3 inhibitor groups compared with those inthe epilepsy model group.Conclusions The expression of MMP-3 may be positively related to the expression of NMDAR in the hippocampal tissues in the juvenile mice with epilepsy.
Objective To explore the relativity of N-methyl-D-aspartic acid(NMDA) receptors with Matrix metalloproteinases-3(MMP-3) expression in hippocampal neurons in juvenile mice with epilepsy.Methods Sixty juvenile mice were divided into 4 groups of 15 animals each, including control, epilepsy model, NMDA receptor(NMDAR) 2A, B inhibitor and MMP-3 inhibitor groups. The epilepsy model was established in all the groups except the control group. The expressions of NMDAR 2 A, and MMP-3 RNA in the hippocampal tissues were detected by RT-PCR in all the groups. The morphology of hippocampal neurons was observed by Nissl staining in all the groups. The expressions of P-glycoprotein(P-gp) and Calmodulin kinase Ⅱ(CAMK Ⅱ) proteins in the hippocampal tissues were detected by Western blot in all the groups.Results The levels of the expressions of NMDAR2A, B and MMP-3 mRNA P-gp and CAMK Ⅱ in the hippocampus tissues were significantly higher in the epilepsy model group than those in the control group(P<0.05). The levels of NMDAR2A, B and MMP-3 mRNA, and P-gp and CAMK Ⅱ in the hippocampal tissues were significantly lower in NMDAR2A, B inhibitor and MMP-3 inhibitors groups than those in the epilepsy model group(P<0.05). The structures in the hippocampal neurons were significantly improved in the NMDAR2A, B in hibitor and MMP-3 inhibitor groups compared with those inthe epilepsy model group.Conclusions The expression of MMP-3 may be positively related to the expression of NMDAR in the hippocampal tissues in the juvenile mice with epilepsy.
引文
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[13]邓镇,罗淼珊,赵元淑,等.Glu N2A抑制剂对癫痫持续状态大鼠脑内P-糖蛋白表达的影响[J].华中科技大学学报(医学版),2014,43(2):168-172.
[2]Miyamoto T,Stein L,Thomas R,et al.Phosphorylation of tau at Y18,but not taufyn binding,is required for tau to modulate NMDA receptor-dependent excitotoxicity in primary neuronal culture[J].Mol Neurodegener,2017,12:41-60.
[3]Addis L,Virdee JK,Vidler LR,et al.Epilepsy-associated GRIN2A mutations reduce NMDA receptor trafficking and agonist potency-molecular profiling and functional rescue[J].Sci Rep,2017,2(7):66-90.
[4]Huntley GW.Synaptic circuit remodelling by matrix metalloproteinases in health and disease[J].Nat Rev Neurosci,2012,13(11):743-757.
[5]Conant K,Wang Y,Szklarczyk A,et al.Matrix metalloproteinase-dependent shedding of intercellular adhesion molecule-5 occurs with long-term potentiation[J].Neuroscience,2010,166(2):508-521.
[6]杨展能,姜靓婧,顾仕红,等.NMDA受体在匹罗卡品癫痫小鼠海马星形胶质细胞活化中的作用[J].东南大学学报(医学版),2017,36(2):129-134.
[7]Carvajal FJ,Mattison HA,Cerpa W.Role of NMDA receptor-mediated glutamatergic signaling inchronic and acute neuropathologies[J].Neural Plast,2016,6:1-20.
[8]Yashiro K and Philpot BD.Regulation of NMDA receptor subunit expression and its implications for LTD,LTP,and metaplasticity[J].Neuropharmacology,2008,55(7):1081-1094.
[9]Di Maio R,Mastroberardino PG,Hu X,et al.Pilocarpine alters NMDA receptor expression and function in hippocampal neurons:NADPH oxidase and ERK1/2 mechanisms[J].Neurobiol Dis,2011,42(3):482-495.
[10]Kim,JY,Ko,AR,Hyun HW,et al.PDI regulates seizure activity via NMDA receptor redox in rats[J].Sci Rep,2017,7:42491.
[11]Brzdak P,Wlodarczyk J,Mozrzymas JW,et al.Matrix metalloprotease 3 activity supports hippocampal epsp-to-spike plasticity following patterned neuronal activity via the regulation of NMDAR function and calcium flux[J].Mol Neurobiol,2017,54:804-816.
[12]Daoudal G,Debanne D.Long-term plasticity of intrinsic excitability:learning rules and mechanisms[J].Learn Mem,2003,10(6):456-465.
[13]邓镇,罗淼珊,赵元淑,等.Glu N2A抑制剂对癫痫持续状态大鼠脑内P-糖蛋白表达的影响[J].华中科技大学学报(医学版),2014,43(2):168-172.