冷诱导RNA结合蛋白表达在颅脑损伤大鼠亚低温治疗中的作用及机制

发布时间：2018-06-30 05:15

本文选题：颅脑损伤 + 冷诱导RNA结合蛋白　；参考：《山东医药》2016年42期

【摘要】：目的探讨冷诱导RNA结合蛋白(CIRP)表达在颅脑损伤(TBI)大鼠亚低温(MH)治疗过程中的作用,并分析其机制。方法将大鼠随机分为A、B、C组,分别一次性鞘内注射0.1 m L生理盐水、空白AD5-GFP、AD5-GFP-CIRP-siRNA;每组再分为4个亚组,即假手术组(Sham组)、TBI组、MH组、TBI+MH组。TBI、TBI+MH组采用液压打击装置进行TBI造模,Sham组及MH组不予打击;TBI造模成功后,MH组、TBI+MH组行MH治疗48 h。于MH开始治疗后30 min及6、12、24、48、72 h,采用Western blot法检测下丘脑部位CIRP、Ras、Raf、ERK-1/2、pERK-1/2蛋白;MH开始治疗后96 h取大鼠脑皮层、海马、下丘脑部位脑组织,采用TUNEL法检测凋亡脑细胞、RTPCR法检测CIRP mRNA。结果 A、B、C组中,TBI、TBI+MH组皮层、海马、下丘脑部位神经细胞凋亡指数均较Sham、MH组升高(P均0.05);A、B组中,TBI+MH组各部位神经细胞凋亡指数较TBI组降低(P均0.05),且该组下丘脑部位较皮层、海马区神经细胞凋亡指数下降明显(P均0.05);在C组中,TBI+MH组各部位神经细胞凋亡指数较TBI组无差异(P均㧐0.05)。在A、B组中,MH、TBI+MH组各部位CIRP mRNA较Sham组升高(P均0.05),下丘脑部位较皮层、海马区CIRP mRNA表达升高(P均0.05);而在C组中,TBI、MH、TBI+MH组各部位CIRP mRNA较Sham组均无差异(P均㧐0.05),各部位CIRP mRNA表达无差异(P均㧐0.05)。在A、B组中,TBI组CIRP蛋白自创伤后12、24 h较创伤后30 min升高(P均0.05),随后开始下降,在48、72 h无差异(P均0.05);MH、TBI+MH组CIRP蛋白表达自MH开始后6 h升高(P均0.05),于48 h达高峰,72 h呈下降趋势;在C组中,各组CIRP蛋白表达均无差异(P均0.05)。TBI、MH、TBI+MH组Ras激活度(Ras/Raf值)均进行性升高,于24 h达到峰值,随后进行性下降;MH、TBI+MH组可导致ERK-1/2激活度(p-ERK-1/2/ERK-1/2值)峰值前移,并迅速降低其激活度,而C组中MH、TBI+MH组此作用消失。结论在MH治疗TBI过程中,CIRP因低温作用而在皮层、海马、下丘脑部位过表达,其中下丘脑部位更加明显;CIRP作用机制可能通过直接调控ERK-1/2激活,使其激活度随时间推移迅速降低,减少对应部位脑组织细胞凋亡,从而发挥神经保护作用。
[Abstract]:Objective to investigate the role of cold-induced RNA-binding protein (CIRP) expression in the treatment of mild hypothermia (MH) in rats with craniocerebral injury (TBI) and its mechanism. Methods the rats were randomly divided into two groups: group A, B, C, and intrathecal injection of 0.1 mL normal saline respectively, and the rats were divided into 4 subgroups, each group was treated with AD5-GFPN AD5-GFP-CIRP-siRNAs, and each group was divided into 4 subgroups. That is, sham-operated group (Sham group), TBI group (MH group), TBI group (TBI group), TBI group, TBI group. At 30 min after treatment and 61212 ~ (24) and 48 ~ 72 h after treatment, the rat cerebral cortex, hippocampus and hypothalamus brain tissues were detected by Western blot method with Western blot method. The apoptotic brain cells were detected by Tunel and RT PCR of apoptotic brain cells (RT PCR) by using Tunel method. The rat brain cortex, hippocampus and hypothalamus brain tissues were detected by Western blot method. The rat brain cortex, hippocampus and hypothalamus were detected by Western blot method using Western blot method. The rat brain cortex, hippocampus and hypothalamus brain tissues were detected by RT-PCR with Western blot method. Results the apoptotic index of neurons in the cortex, hippocampus and hypothalamus of TBI group was significantly higher than that in the Shambone MH group (P 0.05). The apoptotic index of neurons in TBI group was lower than that in TBI group (P 0.05), and the apoptotic index in hypothalamus was lower in TBI group than in TBI group (P 0.05), and the apoptotic index in hypothalamus was lower in TBI group than in TBI group (P 0.05). The apoptotic index of hippocampal neurons decreased significantly (all P 0.05), and there was no significant difference between TBI MH group and TBI group (P 0.05). The CIRP mRNA expression in the hypothalamus was higher than that in the cortex and hippocampus (P 0.05), but there was no significant difference in the CIRP mRNA expression between the TBI group and the Sham group (P 0.05), but there was no significant difference in the CIRP mRNA expression between the TBI group and the Sham group (P < 0.05), while the expression of CIRP mRNA in the hypothalamus was higher than that in the cortex and hippocampus (P 0.05), while in the TBI group there was no significant difference between the two groups (P < 0.05), and there was no significant difference in the expression of CIRP mRNA between the two groups (P < 0.05). The expression of CIRP protein in TBI group increased from 1224 hours after trauma to 30 min after trauma (P 0.05), and then decreased. There was no significant difference (P 0. 05) at 48 ~ 72 h. The expression of CIRP protein in TBI group increased 6 hours after the onset of MHBI (all P 0. 05), and decreased at 48 h and 72 h after trauma. In group C, there was no difference in the expression of CIRP protein in all groups (P 0.05). The Ras activation (Ras-Raf value) in TBI group increased progressively and reached its peak value at 24 h, and then decreased progressively, which resulted in the peak value of ERK-1 / 2 activation (p-ERK-1 / 2 / ERK-1 / 2). The activation degree of MHG TBI MH group was decreased rapidly, but the effect of MHX TBI MH group disappeared in C group. Conclusion CIRP is overexpressed in cortex, hippocampus and hypothalamus due to hypothermia during the treatment of TBI. The mechanism of CIRP in hypothalamus is more obvious, and the activation degree of CIRP decreases rapidly over time through direct regulation of ERK-1 / 2 activation. Reduce the apoptosis of brain cells in the corresponding areas, thus play a neuroprotective role.
【作者单位】：天津中医药大学第二附属医院;天津市环湖医院神经外科研究所;天津中医药大学研究生院;
【基金】：国家自然科学基金资助项目(81303091)
【分类号】：R651.15

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