TLR3信号通路在大鼠七氟烷预处理体外循环脑保护中作用研究

发布时间：2018-05-20 00:37

本文选题：TLR3 + 七氟烷　；参考：《大连医科大学》2014年硕士论文

【摘要】：目的：探讨TLR3信号通路在大鼠七氟烷预处理减轻体外循环脑损伤中的作用，及其可能的分子机制。方法：雄性SD大鼠64只。随机分为3组：假手术组（H组，n=8），CPB组（C组，n=24）和七氟烷预处理组（S组，n=32）。假手术组(H组)：气管插管机械通气，血管穿刺置管，不进行体外循环转流。CPB组（C组）：在右股动脉和右侧颈内静脉穿刺置管，建立无血预充大鼠体外循环模型。七氟烷预处理(S组)：2.4%七氟烷预处理1h，建立体外循环模型。C组和S组进行体外循环1h。选择麻醉后体外循环前（T0）、体外循环30分钟(T1)、体外循环1h(T2)、体外循环后1h(T3)、体外循环后2h(T4)和体外循环后3h(T5)，采集血样，记录大鼠生命体征。C组和S组在T0、T1、T3和T5时，取8只大鼠处死取脑组织样本。使用ELISA酶联免疫法检测血清S100-β、IL-6和IFNβ浓度变化；ELISA酶联免疫法检测左侧海马组织TLR3、TRIF表达情况；采用Western-Blot法检测左侧海马组织TLR3表达情况；选取右侧海马，使用TUNEL法检测神经元调亡情况。结果：三组大鼠各时间点平均动脉压（MAP）、心率（HR）、红细胞压积（Hct）、直肠温、pH值、PaCO2、PaO2，BE值、K+组间比较差异无统计学意义（P0.05）。仅体外循环期间MAP、HR和Hct呈明显降低（与T0比较，P0.05）。对比H组，C组与S组血清S100-β和IL-6均升高（P0.05）。C组与S组血清S100-β和IL-6浓度转流期间明显升高，转流结束后逐渐降低（与T0比较，P0.05）；对比C组T1~T5时，S组血清S100-β和IL-6浓度出现下降（P0.05）。 C组与S组在T1~T5时，血清IFNβ浓度比H组明显升高（P0.05）；与C组对比,除外T0时S组各时间点血清IFNβ浓度均升高（P0.05）。对比H组，，C组与S组TLR3和TRIF蛋白水平均出现明显升高（P0.05）。C组和S组T2时TLR3和TRIF蛋白水平明显升高，T3时TLR3和TRIF蛋白水平降低（与T0比较，P0.05）。对比H组T1、T3、T5时，C组与S组TLR3和TRIF蛋白水平均升高（P0.05）。 Western blot结果显示，C组和S组TLR3蛋白表达量比H组显著升高（与T0比较，P0.05）；S组和C组相比，TLR3表达量明显升高（P0.05）。 TUNEL法检测神经元调亡显示，H组海马神经元凋亡神经元数目极少。对比H组，C组及S组海马凋亡神经元明显增加（P0.05）；S组海马凋亡神经元数量比C组明显降低（P0.05）。结论： 1.2.4%七氟烷预处理对CPB脑损伤具有一定保护作用。 2.七氟烷预处理可通过影响TLR3信号通路，使其表达上调，减轻CPB炎症反应造成的脑损伤进而发挥脑保护作用。 3. TLR3信号通路参与七氟烷预处理脑保护作用，可能是通过七氟烷预先激活TLR3，使下游TRIF表达上调，抑制促炎性因子IL-6的表达，上调抗炎性因子IFNβ的表达，从而对抗炎症损害起到脑保护作用。
[Abstract]:Aim: to investigate the role of TLR3 signaling pathway in the protective effect of sevoflurane preconditioning on brain injury after cardiopulmonary bypass (CPB) and its possible molecular mechanism. Methods: 64 male SD rats were used. They were randomly divided into 3 groups: sham operation group (group H) and group C (group C) and group S (group S). Group H: mechanical ventilation of tracheal intubation, catheterization without cardiopulmonary bypass. Group C: puncture of right femoral artery and right internal jugular vein to establish rat model of cardiopulmonary bypass without blood supply. The model of cardiopulmonary bypass (CPB) was established in group C and group S for 1 hour after preconditioning with sevoflurane for 1 hour in group S: 2. 4% sevoflurane. Blood samples were collected before cardiopulmonary bypass (CPB), 30 minutes after cardiopulmonary bypass (CPB), 1 hour after cardiopulmonary bypass (CPB), 2 hours after cardiopulmonary bypass (CPB), and 3 hours after cardiopulmonary bypass (CPB). Blood samples were collected, and the vital signs of rats were recorded at T _ 0T _ 1T _ 3 and T _ 5 in group S and group C. Eight rats were killed and brain tissue samples were taken. The changes of serum S100- 尾 -IL-6 and IFN 尾 concentration were detected by ELISA enzyme-linked immunosorbent assay (ELISA). The expression of TLR3TIF in left hippocampus was detected by Elisa, TLR3 expression in left hippocampal tissue was detected by Western-Blot method, and the right hippocampus was selected. The apoptosis of neurons was detected by TUNEL method. Results: There was no significant difference in mean arterial pressure (MAPV), heart rate (HRT), HCT (HCT), rectal temperature and pH value (Paco _ 2 / Pao _ 2 ~ (2 +) be) between the three groups (P 0.05). During cardiopulmonary bypass (CPB), MAPHR and Hct decreased significantly (compared with T0, P 0.05). The levels of serum S100- 尾 and IL-6 in group C and group S were significantly higher than those in group C and group S during bypass, and gradually decreased after bypass (compared with T0, P 0.05), and serum S100- 尾 and IL-6 in group C and group S were decreased (P0.05A) when compared with that in group C and group S (P < 0.05). The serum S100- 尾 and IL-6 levels in group C and S were significantly lower than those in group C and group S (P < 0.05), and the levels of serum S100- 尾 and IL-6 in group C were significantly lower than those in group S. The concentration of serum IFN 尾 in group C and group S was significantly higher than that in group H at T1~T5, and that in group S was significantly higher than that in group C at all time points except T0, with the exception of T0, the concentration of serum IFN 尾 in group S was significantly higher than that in group H (P 0.05). Compared with group H and group S, the protein levels of TLR3 and TRIF in group C and group S were significantly increased. The protein levels of TLR3 and TRIF in group C and group S were significantly higher than those in group T 3. The protein levels of TLR3 and TRIF in group C were significantly higher than those in group S (P 0.05 compared with T0). The protein levels of TLR3 and TRIF in group C and group S were significantly higher than those in group T _ 1, T _ 3 and T _ 5 in H group (P 0.05). The results of Western blot showed that the expression of TLR3 protein in group C and group S was significantly higher than that in group H (compared with T0, the expression of TLR3 in group S and group C was significantly higher than that in group T 0. The number of apoptotic neurons in hippocampal neurons in H group was very small by TUNEL assay. Compared with group C and group S, the number of apoptotic neurons in hippocampus of group H and group S was significantly increased than that of group C, and the number of apoptotic neurons in group S was significantly lower than that in group C. Conclusion: 1.2.4% sevoflurane pretreatment had a protective effect on CPB brain injury. 2. Sevoflurane preconditioning can up-regulate the expression of TLR3 signal pathway and attenuate the brain injury caused by CPB inflammation and then play a protective role in the brain. 3. The TLR3 signaling pathway is involved in the protective effect of sevoflurane preconditioning. It may be that sevoflurane preactivates TLR3, up-regulates the expression of downstream TRIF, inhibits the expression of pro-inflammatory factor IL-6 and up-regulates the expression of anti-inflammatory factor IFN 尾. Thus, anti-inflammatory damage plays a brain protection role.
【学位授予单位】：大连医科大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R614

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