TOLL样受体7及TOLL样受体9在脂多糖诱导的AR42J细胞炎症效应中的表达及作用

发布时间：2018-05-28 02:28

本文选题：Toll样受体7 + Toll样受体9　；参考：《广西医科大学》2014年硕士论文

【摘要】：目的观察TOLL样受体7(Toll Like Receptor7, TLR7)、TOLL样受体9(Toll Like Receptor9, TLR9)、核因子-κB P65（Nuclear Factor-κB P65, NF-κBP65）、肿瘤坏死因子-α（Tumor Necrosis Factor-α，TNF-α）在脂多糖诱导的AR42J细胞急性胰腺炎模型中的表达变化，探讨TLR7、TLR9在急性胰腺炎发病机制中的作用及可能的作用机制。方法取对数生长期的AR42J细胞用6孔细胞培养板传代培养24小时后，分别用含脂多糖浓度为0mg/L、1mg/L、10mg/L、100mg/L的培养液刺激AR42J细胞，构建体外急性胰腺炎细胞模型,孵育24小时后，收集细胞培养上清液，离心留取上清，用ELISA法检测细胞培养上清液中TNF-α的含量；收集细胞，提取细胞总RNA，用RT-PCR法分别检测细胞中TLR7、TLR9、NF-κB P65mRNA的表达量变化；提取细胞总蛋白，用Western Blot法分别检测细胞中TLR7、TLR9、NF-κB P65蛋白质表达水平变化。结果与脂多糖浓度为0mg/L的空白对照组相比，其余各组TLR7、TLR9、NF-κB P65的mRNA和蛋白表达量都随着脂多糖浓度升高而增加，脂多糖浓度为100mg/L时达到最大值，各组间的差异有统计学意义（均P＜0.05）。TLR7、TLR9与NF-κB P65在mRNA、蛋白的表达水平上皆有正相关性（mRNA：r=0.786，0.762；蛋白：r=0.801，，0.720；均P＜0.05）。细胞培养上清液中TNF-α的含量呈脂多糖浓度依赖性增加，当脂多糖浓度为100mg/L时达到最大值，各组间的差异有统计学意义（P＜0.05）。结论（1）脂多糖诱导的AR42J细胞急性胰腺炎模型中，TLR7及TLR9表达明显上调，提示两者可能在急性胰腺炎疾病发展中发挥重要作用。（2）TLR7、TLR9可能通过作用于NF-κB上调下游炎症因子的产生，而参与急性胰腺炎的炎症反应。
[Abstract]:Objective to observe the expression of TOLL like receptor 7(Toll Like Receptor7, TLR7TLL-like receptor 9(Toll Like Receptor9, nuclear factor- 魏 B P65(Nuclear Factor- 魏 B P65, NF- 魏 BP65, tumor necrosis factor- 伪 tumor Necrosis factor- 伪 in the model of AR42J cell acute pancreatitis induced by lipopolysaccharide (LPS). To investigate the role and possible mechanism of TLR7 and TLR9 in the pathogenesis of acute pancreatitis. Methods AR42J cells in logarithmic growth phase were cultured on a 6-well cell culture plate for 24 hours. The AR42J cells were stimulated with a culture medium containing a concentration of 0 mg / L of lipopolysaccharide at a concentration of 10 mg / L ~ (10) mg 路L ~ (-1). The acute pancreatitis cell model was established in vitro, and incubated for 24 hours. We collected the supernatant of cell culture, centrifuged the supernatant, detected the content of TNF- 伪 in the supernatant of cell culture by ELISA method, collected the cells, extracted the total RNAs of the cells, detected the changes of the expression of NF- 魏 B P65mRNA in the cells by RT-PCR method, and extracted the total cell protein. The expression level of NF- 魏 B p65 in TLR7 and TLR9 was detected by Western Blot assay. Results compared with the control group with lipopolysaccharide concentration of 0mg/L, the mRNA and protein expression of NF- 魏 B p65 in other groups increased with the increase of lipopolysaccharide concentration, and reached the maximum when the concentration of LPS was 100mg/L. There were significant differences among the three groups (P < 0.05). TLR7, TLR9 and NF- 魏 B p65 were all positively correlated with mRNAs in mRNAs, the protein expression levels were all positively correlated with mRNA-0.7860.762protein: rr-0.801and 0.720; all of them were all P < 0.05. The content of TNF- 伪 in the supernatant of cell culture was increased in a dose-dependent manner, and reached the maximum when the concentration of lipopolysaccharide was 100mg/L (P < 0.05). Conclusion the expression of TLR7 and TLR9 in acute pancreatitis model of AR42J cells induced by lipopolysaccharide is obviously up-regulated, which suggests that the expression of TLR7 and TLR9 may play an important role in the development of acute pancreatitis, which may be related to the up-regulation of down-stream inflammatory factor production by NF- 魏 B. And participate in the inflammatory response of acute pancreatitis.
【学位授予单位】：广西医科大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：R57

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