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目的:探讨心肺复苏后脑损伤中TLR4的表达及其意义。方法:野生型(WT:C3H/HeN)和突变型(TLR4-/-:C3H/HeJ)小鼠经静脉注射氯化钾诱导心搏骤停的方法建立心搏骤停/心肺复苏CA/CPR小鼠模型,分为假手术组和模型组。采用免疫组织化学法观察脑组织TLR4和NF-κB分布并测定阳性反应的平均光密度值。采用酶联免疫吸附法检测CA/CPR脑损伤后脑组织TNF-α含量,采用SDS聚丙烯酰胺凝胶电泳分析(Western blot)方法检测细胞间黏附分子ICAM-1的表达。结果:在小鼠脑组织观察到WT脑组织中TLR4蛋白有增强表达,与假手术组比较差异有统计学意义(P<0.01)。与假手术组相比,WT和TLR4-/-模型组NF-κB阳性细胞明显增加,差异有统计学意义(P<0.01)。ELISA结果发现,与假手术组相比,WT和TLR4-/-模型组TNF-α含量明显升高,TLR4-/-模型组较WT模型组TNF-α含量较低(P<0.05)。Western blot结果显示两模型组细胞间黏附分子ICAM-1表达均增强,WT组ICAM-1的表达高于TLR4-/-组(P<0.05)。结论:心搏骤停/心肺复苏脑损伤过程中TLR4蛋白激活表达可能作为重要炎性受体介导脑缺血炎性损伤。
Objective: To investigate the expression of TLR4 in brain injury after cardiopulmonary resuscitation and its significance. METHODS: Cardiac arrest / cardiopulmonary resuscitation (CA / CPR) was established by intravenous injection of potassium chloride to induce cardiac arrest in wild type (WT: C3H / HeN) and mutant (TLR4 - / -: C3H / Mouse model, divided into sham operation group and model group. Immunohistochemistry was used to observe the distribution of TLR4 and NF-κB in brain tissue and to determine the average optical density of the positive reaction. The contents of TNF-α in brain tissue were detected by enzyme-linked immunosorbent assay (ELISA), and the expression of ICAM-1 was detected by SDS-polyacrylamide gel electrophoresis (SDS-PAGE). Results: The expression of TLR4 protein in brain tissue of WT mice was significantly higher than that in sham-operated group (P <0.01). Compared with sham operation group, NF-κB positive cells in WT and TLR4 - / - model groups were significantly increased, the difference was statistically significant (P <0.01). The results of ELISA showed that TNF-αlevels in WT and TLR4 - / - model groups were significantly increased compared with those in sham-operation group, while TNF-αlevel in TLR4 - / - model group was lower than that in WT group (P <0.05). Western blot results showed that ICAM-1 expression was increased in both models and ICAM-1 in WT group was higher than that in TLR4 - / - group (P <0.05). Conclusion: Activation of TLR4 expression during cardioversion / CPR may play an important inflammatory role in the pathogenesis of cerebral ischemic injury.