转染窖蛋白1基因对脂多糖诱导的人支气管上皮细胞Toll样受体4表达的影响及大黄素的干预效应
发布时间：2018-09-17 12:22
【摘要】：目的研究转染窖蛋白1（Caveolin-1，CAV-1）基因对脂多糖（Lipopolysaccharide，LPS）诱导的人支气管上皮细胞（Humanbronchialepithelialcells，HBE）Toll样受体4（Toll-likereceptor4，TLR-4）表达的影响，以及大黄素(Emodin)对LPS诱导的HBE细胞CAV-1和TLR-4表达的干预效应。 方法本实验分为两部分：第一部分，对LPS诱导的HBE细胞给予不同浓度的Emodin干预，并设立正常对照组、单纯LPS干预组、阳性对照组（Atorvastatin,阿托伐他汀）和阴性对照组（溶媒DMSO），用免疫印记法（Westernblot）检测并比较各组细胞CAV-1的表达水平；第二部分，构建Cav-EGFP重组质粒，瞬时转染HBE细胞，设立正常对照组和空质粒转染组，Westernblot检测并比较三组细胞CAV-1的表达；于质粒转染HBE细胞48h后给予LPS刺激，Westernblot检测并比较CAV-1过表达组与空质粒转染组TLR-4的表达，并给予Emodin干预转染后LPS诱导的HBE细胞，Westernblot检测比较各组细胞TLR-4的表达。 结果第一部分：与正常对照组相比，单纯LPS干预组HBE细胞经LPS刺激后，CAV-1蛋白的表达量明显上调（P0.05)；中、高剂量Emodin干预组、阳性对照组与单纯LPS干预组相比,CAV-1蛋白的表达量均明显下降（P0.05）；第二部分：CAV-1-EGFP重组质粒转染组与正常对照组、空质粒转染组比较，CAV-1的表达量明显上调（P0.05）,而空质粒转染组与正常对照组相比，，CAV-1的表达量无统计差异（P0.05）；与空质粒转染组相比，CAV-1过表达组能上调LPS诱导的HBE细胞TLR-4的表达,Emodin能抑制该效应(P0.05)。结论Emodin能抑制LPS诱导的HBE细胞CAV-1的表达；转染CAV-1基因能 上调LPS诱导的HBE细胞TLR-4的表达，Emodin能抑制该效应。
[Abstract]:Objective to study the effect of cellar protein 1 (Caveolin-1,CAV-1) gene transfection on the expression of Toll like receptor 4 (Toll-likereceptor4,TLR-4) in human bronchial epithelial cells (Humanbronchialepithelialcells,HBE) induced by lipopolysaccharide (Lipopolysaccharide,LPS) and the effect of emodin (Emodin) on LPS induced CAV-1 and TLR-4 expression in HBE cells. Methods the experiment was divided into two parts: in the first part, the HBE cells induced by LPS were treated with different concentrations of Emodin, and the normal control group was set up, and the control group was treated with LPS alone. The positive control group (Atorvastatin, Atto vastatin) and the negative control group (the solvent DMSO), was used to detect and compare the expression of CAV-1 by (Westernblot). The expression of CAV-1 was detected by Western blot in the normal control group and the empty plasmid transfection group, and the expression of TLR-4 in the CAV-1 overexpression group was detected and compared with that in the empty plasmid transfection group after 48 hours of transfection of the plasmid into HBE cells. The expression of TLR-4 was detected by Western blot in HBE cells induced by LPS after Emodin intervention. Results: the first part: compared with the normal control group, the expression of CAV-1 protein in HBE cells stimulated by LPS was significantly up-regulated in the LPS intervention group (P0.05), while in the middle and high dose Emodin intervention group, the expression of CAV-1 protein was significantly increased in the HBE cells treated with LPS (P0.05). The expression of CAV-1 protein in the positive control group was significantly lower than that in the control group (P0.05). The expression of CAV-1 was significantly up-regulated in the empty plasmid transfection group (P0.05), but there was no statistical difference between the empty plasmid transfection group and the normal control group (P0.05). Compared with the empty plasmid transfection group, the expression of TLR-4 in HBE cells induced by LPS could be upregulated in the overexpression group of CAV-1. Emodin could inhibit this effect (P0.05). Conclusion Emodin can inhibit the expression of CAV-1 in HBE cells induced by LPS, and transfection of CAV-1 gene can up-regulate the expression of TLR-4 in HBE cells induced by LPS.
【学位授予单位】：华中科技大学
【学位级别】：硕士
【学位授予年份】：2013
【分类号】：R563.8
[Abstract]:Objective to study the effect of cellar protein 1 (Caveolin-1,CAV-1) gene transfection on the expression of Toll like receptor 4 (Toll-likereceptor4,TLR-4) in human bronchial epithelial cells (Humanbronchialepithelialcells,HBE) induced by lipopolysaccharide (Lipopolysaccharide,LPS) and the effect of emodin (Emodin) on LPS induced CAV-1 and TLR-4 expression in HBE cells. Methods the experiment was divided into two parts: in the first part, the HBE cells induced by LPS were treated with different concentrations of Emodin, and the normal control group was set up, and the control group was treated with LPS alone. The positive control group (Atorvastatin, Atto vastatin) and the negative control group (the solvent DMSO), was used to detect and compare the expression of CAV-1 by (Westernblot). The expression of CAV-1 was detected by Western blot in the normal control group and the empty plasmid transfection group, and the expression of TLR-4 in the CAV-1 overexpression group was detected and compared with that in the empty plasmid transfection group after 48 hours of transfection of the plasmid into HBE cells. The expression of TLR-4 was detected by Western blot in HBE cells induced by LPS after Emodin intervention. Results: the first part: compared with the normal control group, the expression of CAV-1 protein in HBE cells stimulated by LPS was significantly up-regulated in the LPS intervention group (P0.05), while in the middle and high dose Emodin intervention group, the expression of CAV-1 protein was significantly increased in the HBE cells treated with LPS (P0.05). The expression of CAV-1 protein in the positive control group was significantly lower than that in the control group (P0.05). The expression of CAV-1 was significantly up-regulated in the empty plasmid transfection group (P0.05), but there was no statistical difference between the empty plasmid transfection group and the normal control group (P0.05). Compared with the empty plasmid transfection group, the expression of TLR-4 in HBE cells induced by LPS could be upregulated in the overexpression group of CAV-1. Emodin could inhibit this effect (P0.05). Conclusion Emodin can inhibit the expression of CAV-1 in HBE cells induced by LPS, and transfection of CAV-1 gene can up-regulate the expression of TLR-4 in HBE cells induced by LPS.
【学位授予单位】：华中科技大学
【学位级别】：硕士
【学位授予年份】：2013
【分类号】：R563.8
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