miRNA21在戊四氮致癫痫大鼠马神经元中的作用及其机制的研究
|
摘要
目的:探讨miRNA21在戊四氮致癫痫大鼠海马组织中的表达情况及其可能的作用机制。方法:24只健康雄性wister大鼠随机分为正常组和癫痫组,记录两组大鼠的Racine等级评分对癫痫大鼠模型的建立进行评价;应用生物信息学方法筛选癫痫大鼠海马中高差异性表达的并靶向Bcl-2的microRNA;应用qRT-PCR方法验证癫痫大鼠海马中microRNA21的表达情况;应用Western blot和免疫组织化学方法检测大鼠海马中Bcl-2的表达情况;应用Hoechst 33258检测大鼠海马神经元的凋亡情况。结果:癫痫组大鼠的Racine评分明显高于正常对照组(P<0.01);与正常组相比,癫痫组大鼠海马中的miRNA21表达降低(P<0.01),Bcl-2表达减少(P<0.01)。结论:在癫痫大鼠海马神经元中miRNA21的表达量下调,且miRNA21可能通过调节Bcl-2的表达含量进一步导致海马神经元凋亡,进而对癫痫大鼠产生影响。
Objective: To investigate the expression of miRNA21 in hippocampus of rats with pentylenetetrazol-induced epilepsy and its possible mechanism.Methods: Twenty-four healthy male wister rats was randomly divided into two groups, the normal group and the epilepsy group. The Racine grade scores of the two groups of rats were recorded to evaluate the establishment of a rat model of epilepsy. The differentially expressed of microRNAs targeting bcl-2 in the hippocampus of epileptic rats was screened by bioinformatics method. The expression of microRNA21 in hippocampus of epileptic rats was verified by qRT-PCR. Western blot and immunohistochemistry were used to detect the expression of Bcl-2 in rat hippocampus; Hoechst 33258 was used to detect the apoptosis of rat hippocampal neurons. Results: The Racine score of the rats in the epilepsy group was significantly higher than that in the normal control group(P<0.01). Compared with the normal group, the expression of miRNA21 in the hippocampus of the epileptic group decreased(P<0.01) and the expression of Bcl-2 decreased(P< 0.01). Conclusion: Epilepsy can up-regulate the expression of miRNA21 in rat hippocampal neurons, and it may further induce apoptosis of hippocampal neurons by down-regulating Bcl-2, which may affect epileptic rats.
Objective: To investigate the expression of miRNA21 in hippocampus of rats with pentylenetetrazol-induced epilepsy and its possible mechanism.Methods: Twenty-four healthy male wister rats was randomly divided into two groups, the normal group and the epilepsy group. The Racine grade scores of the two groups of rats were recorded to evaluate the establishment of a rat model of epilepsy. The differentially expressed of microRNAs targeting bcl-2 in the hippocampus of epileptic rats was screened by bioinformatics method. The expression of microRNA21 in hippocampus of epileptic rats was verified by qRT-PCR. Western blot and immunohistochemistry were used to detect the expression of Bcl-2 in rat hippocampus; Hoechst 33258 was used to detect the apoptosis of rat hippocampal neurons. Results: The Racine score of the rats in the epilepsy group was significantly higher than that in the normal control group(P<0.01). Compared with the normal group, the expression of miRNA21 in the hippocampus of the epileptic group decreased(P<0.01) and the expression of Bcl-2 decreased(P< 0.01). Conclusion: Epilepsy can up-regulate the expression of miRNA21 in rat hippocampal neurons, and it may further induce apoptosis of hippocampal neurons by down-regulating Bcl-2, which may affect epileptic rats.
引文
1 Begley CE,Beghi E,Beran RG,et al.ILAE commission on the burden of epilepsy,subcommission on the economic burden of epilepsy:final report 1998-2001[J].Epilepsia,2002,43(6):668-673.
2 Devinsky O.Diagnosis and treatment of temporal lobe epilepsy[J].Rev Neurol Dis,2004,1(1):2-9.
3 L?scher W,Klitgaard H,Twyman RE,et al.New avenues for anti-epileptic drug discovery and development[J].Nat Rev Drug Discov,2013,12(10):757-776.
4 Brodie MJ,Barry SJ,Bamagous GA,et al.Patterns of treatment response in newly diagnosed epilepsy[J].Neurology,2012,78(20):1548-1554.
5 Tanaka N,Toyooka S,Soh J,et al.Downregulation of microRNA-34 induces cell proliferation and invasion of human mesothelial cells[J].Oncol Rep,2013,29(6):2169-2174.
6 Mikkonen M,Soininen H,K?lvi?nen R,et al.Remodeling of neuronal circuitries in human temporal lobe epilepsy:increased expression of highly polysialylated neural cell adhesion molecule in the hippocampus and the entorhinal cortex[J].Ann Neurol,1998,44(6):923-934.
7 Vezzani A,Aronica E,Mazarati A,et al.Epilepsy and brain inflammation[J].Exp Neuro,2013,244:11-21.
8 Henshall DC.MicroRNA and epilepsy:profiling,functions and potential clinical applications[J].Curr Opin Neurol,2014,27(2):199-205.
9 Hu K,Xie YY,Zhang C,et al.MicroRNA expression profile of the hippocampus in a rat model of temporal lobe epilepsy and miR-34a-targeted neuroprotection against hippocampal neurone cell apoptosis post-status epilepticus[J].BMC Neurosci,2012,13:115.
10 金绍静,马巍,张淑红,等.miR34a在戊四氮致癫痫大鼠中通过下调Bcl-2导致神经元凋亡[J].中风与神经疾病杂志,2018,35(7):617-621.
11 Bengzon J,Mohapel P,Ekdahl CT,et al.Neuronal apoptosis after brief and prolonged seizures[J].Prog Brain Res,2002,135:111-119.
12 Kotloski R,Lynch M,Lauersdorf S,et al.Repeated brief seizures induce progressive hippocampal neuron loss and memory deficits[J].Prog Brain Res,2002,135:95-110.
13 Zhang X,Cui SS,Wallace AE,et al.Relations between brain pathology and temporal lobe epilepsy[J].J Neurosci,2002,22(14):6052-6061.
14 Liou AKF,Clark RS,Henshall DC,et al.To die or not to die for neurons in ischemia,traumatic brain injury and epilepsy:a review on the stress-activated signaling pathways and apoptotic pathways[J].Prog Neurobiol,2003,69(2):103-142.
15 Schmutzhard E,Pfausler B.Complications of the management of status epilepticus in the intensive care unit[J].Epilepsia,2011,52(Suppl 8):39-41.
16 Nobili P,Colciaghi F,Finardi A,et al.Continuous neurodegeneration and death pathway activation in neurons and glia in an experimental model of severe chronic epilepsy[J].Neurobio of Dis,2015,83:54-66.
17 Henshall DC,Chen J,Simon RP.Involvement of caspase-3-like protease in the mechanism of cell death following focally evoked limbic seizures[J].J Neurochem,2000,74(3):1215-1223.
18 ZhuXJ,Shen K,Bai Y,et al.NADPH oxidase activation is required for pentylenetetrazole kindling-induced hippocampal autophagy[J].Free Radic Biol Med,2016,94:230-242.
19 Pitk?nen A,Lukasiuk K.Mechanisms of epileptogenesis and potential treatment targets[J].Lancet Neuro,2011,10(2):173-186.
20 王艺,李智平,施忆贇.戊四氮点燃大鼠慢性癫癎模型的建立及评价[J].复旦学报(医学版),2006,33(2):206-208,228.
21 Song YS,Joo HW,Park IH,et al.Bone marrow mesenchymal stem cell-derived vascular endothelial growth factor attenuates cardiac apoptosis via regulation of cardiac miRNA-23a and miRNA-92a in a rat model of myocardial infarction[J].PLoS One,2017,12(6):e0179972.
22 Cao X,Cai ZG,Liu JY,et al.MiRNA-504 inhibits p53-dependent vascular smooth muscle cell apoptosis and may prevent aneurysm formation[J].Mol Med Rep,2017,16(3):2570-2578.
23 Eacker SM,Dawson TM,Dawson VL.Understanding microRNAs in neurodegeneration[J].Nat Rev Neurosci,2009,10(12):837-841.
24 O'Neill KL,Huang K,Zhang JJ,et al.Inactivation of prosurvival Bcl-2 proteins activates Bax/Bak through the outer mitochondrial membrane[J].Genes Dev,2016,30(8):973-988.
25 Kalkavan H,Green DR.MOMP,cell suicide as a BCL-2 family business[J].Cell Death Differ,2018,25(1):46-55.
26 CaoCL,NiuHJ,Kang SP,et al.MiRNA-21 sensitizes gastrointestinal stromal tumors (GISTs) cells to Imatinib via targeting B-cell lymphoma 2 (Bcl-2)[J].Eur Rev Med Pharmacol Sci,2016,20(17):3574-3581.
2 Devinsky O.Diagnosis and treatment of temporal lobe epilepsy[J].Rev Neurol Dis,2004,1(1):2-9.
3 L?scher W,Klitgaard H,Twyman RE,et al.New avenues for anti-epileptic drug discovery and development[J].Nat Rev Drug Discov,2013,12(10):757-776.
4 Brodie MJ,Barry SJ,Bamagous GA,et al.Patterns of treatment response in newly diagnosed epilepsy[J].Neurology,2012,78(20):1548-1554.
5 Tanaka N,Toyooka S,Soh J,et al.Downregulation of microRNA-34 induces cell proliferation and invasion of human mesothelial cells[J].Oncol Rep,2013,29(6):2169-2174.
6 Mikkonen M,Soininen H,K?lvi?nen R,et al.Remodeling of neuronal circuitries in human temporal lobe epilepsy:increased expression of highly polysialylated neural cell adhesion molecule in the hippocampus and the entorhinal cortex[J].Ann Neurol,1998,44(6):923-934.
7 Vezzani A,Aronica E,Mazarati A,et al.Epilepsy and brain inflammation[J].Exp Neuro,2013,244:11-21.
8 Henshall DC.MicroRNA and epilepsy:profiling,functions and potential clinical applications[J].Curr Opin Neurol,2014,27(2):199-205.
9 Hu K,Xie YY,Zhang C,et al.MicroRNA expression profile of the hippocampus in a rat model of temporal lobe epilepsy and miR-34a-targeted neuroprotection against hippocampal neurone cell apoptosis post-status epilepticus[J].BMC Neurosci,2012,13:115.
10 金绍静,马巍,张淑红,等.miR34a在戊四氮致癫痫大鼠中通过下调Bcl-2导致神经元凋亡[J].中风与神经疾病杂志,2018,35(7):617-621.
11 Bengzon J,Mohapel P,Ekdahl CT,et al.Neuronal apoptosis after brief and prolonged seizures[J].Prog Brain Res,2002,135:111-119.
12 Kotloski R,Lynch M,Lauersdorf S,et al.Repeated brief seizures induce progressive hippocampal neuron loss and memory deficits[J].Prog Brain Res,2002,135:95-110.
13 Zhang X,Cui SS,Wallace AE,et al.Relations between brain pathology and temporal lobe epilepsy[J].J Neurosci,2002,22(14):6052-6061.
14 Liou AKF,Clark RS,Henshall DC,et al.To die or not to die for neurons in ischemia,traumatic brain injury and epilepsy:a review on the stress-activated signaling pathways and apoptotic pathways[J].Prog Neurobiol,2003,69(2):103-142.
15 Schmutzhard E,Pfausler B.Complications of the management of status epilepticus in the intensive care unit[J].Epilepsia,2011,52(Suppl 8):39-41.
16 Nobili P,Colciaghi F,Finardi A,et al.Continuous neurodegeneration and death pathway activation in neurons and glia in an experimental model of severe chronic epilepsy[J].Neurobio of Dis,2015,83:54-66.
17 Henshall DC,Chen J,Simon RP.Involvement of caspase-3-like protease in the mechanism of cell death following focally evoked limbic seizures[J].J Neurochem,2000,74(3):1215-1223.
18 ZhuXJ,Shen K,Bai Y,et al.NADPH oxidase activation is required for pentylenetetrazole kindling-induced hippocampal autophagy[J].Free Radic Biol Med,2016,94:230-242.
19 Pitk?nen A,Lukasiuk K.Mechanisms of epileptogenesis and potential treatment targets[J].Lancet Neuro,2011,10(2):173-186.
20 王艺,李智平,施忆贇.戊四氮点燃大鼠慢性癫癎模型的建立及评价[J].复旦学报(医学版),2006,33(2):206-208,228.
21 Song YS,Joo HW,Park IH,et al.Bone marrow mesenchymal stem cell-derived vascular endothelial growth factor attenuates cardiac apoptosis via regulation of cardiac miRNA-23a and miRNA-92a in a rat model of myocardial infarction[J].PLoS One,2017,12(6):e0179972.
22 Cao X,Cai ZG,Liu JY,et al.MiRNA-504 inhibits p53-dependent vascular smooth muscle cell apoptosis and may prevent aneurysm formation[J].Mol Med Rep,2017,16(3):2570-2578.
23 Eacker SM,Dawson TM,Dawson VL.Understanding microRNAs in neurodegeneration[J].Nat Rev Neurosci,2009,10(12):837-841.
24 O'Neill KL,Huang K,Zhang JJ,et al.Inactivation of prosurvival Bcl-2 proteins activates Bax/Bak through the outer mitochondrial membrane[J].Genes Dev,2016,30(8):973-988.
25 Kalkavan H,Green DR.MOMP,cell suicide as a BCL-2 family business[J].Cell Death Differ,2018,25(1):46-55.
26 CaoCL,NiuHJ,Kang SP,et al.MiRNA-21 sensitizes gastrointestinal stromal tumors (GISTs) cells to Imatinib via targeting B-cell lymphoma 2 (Bcl-2)[J].Eur Rev Med Pharmacol Sci,2016,20(17):3574-3581.