NLRP3炎症复合体信号通路分子在幽门螺杆菌感染小鼠模型中表达的研究

发布时间：2018-09-14 21:06

【摘要】：目的：通过构建幽门螺杆菌(Helicobacter pylori, H. pylori)感染C57BL/6小鼠动物模型，检测NLRP3炎症复合体信号通路相关分子在小鼠胃组织的表达情况，综合分析NLRP3炎症复合体信号通路及其分子在H. pylori感染引起的炎症反应中的作用。方法：将6-8周龄C57BL/6小鼠随机分为3组，即感染组、对照组和治疗组。感染组和对照组小鼠分别以H. pylori和PBS灌胃，1次/48h，共5次。4周后，培养法鉴定H. pylori感染是否成功，并设立治疗组，采用三联疗法对小鼠进行治疗，为期1周，治疗后4周处死。对照组和感染组小鼠于感染后不同时间点处死，无菌取胃组织和血液进行进一步实验： (1)HE染色法鉴定感染小鼠胃组织炎症程度的变化； (2)RT-PCR检测小鼠胃组织中NLRP3(NACTH domain-leucine-rich repeat-,andPYD-containing protein, NLRP3)、 ASC(apoptosis-associated speck-like proteincontaining a CARD, ASC)、caspase-1(cysteine aspartic acid specific protease-1,Caspase-1)、IL-1β、IL-18、IL-33mRNA的表达情况； (3)ELISA检测小鼠血清中细胞因子IL-1β、IL-18、IL-33的含量； (4)Western blot检测小鼠胃组织中Caspase-1的表达情况。结果： (1)对照组小鼠胃组织正常，，感染组小鼠胃组织炎症程度随感染时间的延长而不断加重，治疗组小鼠胃组织炎症程度明显减轻； (2)感染组小鼠胃组织中NLRP3炎症复合体信号通路相关蛋白(NLRP3、ASC、caspase-1)和细胞因子(IL-1β、IL-18、IL-33)mRNA表达水平显著升高(P0.05)； (3)H. pylori感染的小鼠血清中IL-1β、IL-18、IL-33的表达均显著增高(P0.05), (4)H. pylori感染的小鼠胃组织中caspase-1的蛋白含量随感染时间延长而增加； (5)治疗组小鼠NLRP3、ASC、caspase-1、IL-1β、IL-18、IL-33的表达量与治疗前相比有所下降。结论： (1) H. pylori感染后小鼠胃组织中NLRP3炎症复合体信号通路相关分子表达升高，表明H. pyori感染可以激活NLRP3炎症复合体信号通路。 (2) H. pylori感染后小鼠的胃炎程度与NLRP3炎症复合体信号通路相关分子的表达呈正相关。
[Abstract]:Objective: to investigate the expression of NLRP3 inflammatory complex signaling pathway related molecules in gastric tissues of C57BL/6 mice infected with Helicobacter pylori (Helicobacter pylori, H. pylori). The role of NLRP3 inflammatory complex signaling pathway and its molecules in the inflammatory response induced by H. pylori infection was comprehensively analyzed. Methods: C57BL/6 mice aged 6 to 8 weeks were randomly divided into three groups: infection group, control group and treatment group. Mice in infected group and control group were perfused with H. pylori and PBS for 1 / 48 h for 5 times. After 4 weeks, culture method was used to determine the success of H. pylori infection, and the treatment group was set up. The mice were treated with triple therapy for 1 week. The mice were killed 4 weeks after treatment. The mice in the control group and the infected group were killed at different time points after infection, and the gastric tissue and blood were collected aseptically for further experiments: (1) HE staining method was used to identify the degree of inflammation in the gastric tissue of the infected mice; (2) RT-PCR was used to detect the expression of NLRP3 (NACTH domain-leucine-rich repeat-,andPYD-containing protein, NLRP3), ASC (apoptosis-associated speck-like proteincontaining a CARD, ASC) caspase-1 (cysteine aspartic acid specific protease-1,Caspase-1) interleukin-18 (IL-33) mRNA in mouse gastric tissue, (3) ELISA was used to detect the content of IL-18 IL-33 in serum of mice. (4) Western blot was used to detect the expression of Caspase-1 in gastric tissues of mice. Results: (1) the gastric tissue in the control group was normal, and the degree of gastric inflammation in the infected group was aggravated with the prolongation of the infection time, and the degree of gastric inflammation in the treatment group was obviously reduced. (2) the expression of NLRP3 signaling pathway related protein (NLRP3,ASC,caspase-1) and cytokine (IL-1 尾 -IL-18 IL-33) in gastric tissues of infected mice were significantly increased (P0.05). (3) the expression of IL-18 IL-33 in serum of mice infected with) H. pylori increased significantly (P0.05) the protein content of caspase-1 in gastric tissues of mice infected with), (4) H. pylori increased with the time of infection. (5) the expression of NLRP3,ASC,caspase-1,IL-1 尾 -IL-18 + IL-33 in treatment group was lower than that before treatment. Conclusion: (1) after) H. pylori infection, the expression of NLRP3 inflammatory complex signal pathway related molecules in the gastric tissues of mice was increased. The results indicated that H. pyori infection could activate the NLRP3 inflammatory complex signaling pathway. (2) the degree of gastritis in mice after) H. pylori infection was positively correlated with the expression of NLRP3 inflammatory complex signal pathway related molecules.
【学位授予单位】：南华大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R-332

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