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目的探讨选择性细胞周期素依赖性激酶抑制剂 olomoucine 对癫痫持续状态(SE)后神经元凋亡的影响。方法建立氯化锂-匹罗卡品 SE 模型,随机分为生理盐水组、SE 组和 olomoucine干预组。应用免疫荧光化学法检测神经元核心抗原(NeuN)和周期素蛋白 B1表达,通过 TUNEL 方法检测神经元凋亡情况;应用逆转录-聚合酶链反应(RT-PCR)检测皮质及海马白介素-1β(IL-1β)和肿瘤坏死因子-α(TNF-α)mRNA 表达情况。统计方法采用方差分析与 t 检验。结果 (1)生理盐水组TUNEL 示阳性神经元稀少,SE 后显著增加(P<0.05),olomoucine 干预组明显少于 SE 组(P<0.05),尤其齿状回门区与生理盐水组相比差异没有统计学意义(P>0.05);(2)与 SE 组比较,olomoucine 干预组细胞周期素 B1阳性神经元数目减少,差异具有统计学意义(P<0.05);(3)生理盐水组 IL-1β、TNF-αmRNA 表达量稀少,SE 后显著增加(P<0.05),除皮质 TNF-αmRNA 外,olomoucine 干预组较 SE组均明显降低(P<0.05)。结论 olomoucine 可通过调控细胞周期,抑制癫痫持续状态后神经元重新进入细胞周期,以及抑制活化的胶质细胞分泌炎性细胞因子,抑制癫痫持续状态后神经元凋亡而减轻脑损伤。
Objective To investigate the effect of olomoucine, a selective cyclin-dependent kinase inhibitor, on neuronal apoptosis after status epilepticus (SE). Methods Lithium-pilocarpine SE model was established and randomly divided into normal saline group, SE group and olomoucine intervention group. Immunocytochemistry was used to detect the expression of neuronal core antigen (NeuN) and cyclin B1, and neuronal apoptosis was detected by TUNEL method. The levels of interleukin-1β in cortex and hippocampus were detected by reverse transcription-polymerase chain reaction (RT-PCR) (IL-1β) and tumor necrosis factor-α (TNF-α) mRNA expression. Statistical methods using analysis of variance and t test. Results (1) TUNEL positive neurons were scarce in NS group and significantly increased in SE group (P <0.05), while olomoucine group was significantly less than that in SE group (P <0.05). In particular, (2) Compared with SE group, olomoucine intervention group decreased the number of cyclin B1-positive neurons, the difference was statistically significant (P <0.05); (3) The normal saline group IL (P <0.05). In addition to cortical TNF-αmRNA, olomoucine intervention group was significantly lower than SE group (P <0.05). Conclusion Olomoucine attenuates brain injury by regulating the cell cycle, inhibiting the re-entry of neurons into the cell cycle after the status epilepticus, inhibiting the secretion of inflammatory cytokines by activated glial cells and inhibiting the neuronal apoptosis after status epilepticus.