滴滴涕对人大肠癌DLD1细胞上皮间充质转化的影响

发布时间：2018-02-25 03:13

  本文关键词： 滴滴涕 大肠癌 上皮间充质转化 信号转导和转录激活因子　出处：《中国药理学与毒理学杂志》2017年02期 　论文类型：期刊论文

【摘要】：目的探究滴滴涕(DDT)对人结直肠腺癌上皮细胞(DLD1)上皮间充质转化的影响及机制。方法DLD1细胞用DDT 0.01,0.1,1.0,10.0和100.0 nmol·L~(-1)处理48 h后,倒置显微镜下观察细胞形态;实时荧光定量PCR法检测E-钙黏着蛋白、N-钙黏着蛋白、波形蛋白和锌指转录因子Snail1的mRNA表达。Western蛋白质印迹法检测信号转导和转录激活因子3(STAT3)信号通路主要蛋白STAT3和p-STAT3的蛋白水平。用STAT3抑制剂WP1066(5μmol·L~(-1))处理,通过Western印迹法和实时荧光定量PCR法检测其对DDT诱导的STAT3/Snail1信号通路中p-STAT3、STAT3的蛋白水平和上皮间充质转化关键因子E-钙黏着蛋白、N-钙黏着蛋白、波形蛋白和锌指转录因子Snail1的mRNA水平的影响。结果与正常对照组相比,DLD1细胞在DDT处理48 h后,细胞形态由卵圆形逐渐变为长梭形,E-钙黏着蛋白mRNA相对表达显著降低(P0.01),为正常对照组的(42.4±2.8)%。N-钙黏着蛋白和波形蛋白mRNA相对表达显著提高(P0.01),为正常对照组的1.91±0.1倍和(1.5±0.2)倍。STAT3信号通路蛋白STAT3和p-STAT3蛋白表达均升高(P0.01),为正常对照组的2.1和1.8倍。锌指转录因子Snail1的mRNA相对表达显著升高(P0.01),是正常对照组的(1.5±0.1)倍。STAT3抑制剂WP1066 5μmol·L~(-1)处理后,锌指转录因子Snail1 mRNA的表达明显下调(P0.01),为DDT 1.0 nmol·L~(-1)处理组的(56.3±0.9)%,同时抑制DDT诱导的E-钙黏着蛋白mRNA表达升高(P0.01),为DDT 1.0 nmol·L~(-1)处理组的2.5±0.1倍,N-钙黏着蛋白和波形蛋白mRNA表达降低(P0.01),分别为DDT 1.0 nmol·L~(-1)处理组的(50.2±2.9)%和(61.6±6.1)%。结论 DDT可能通过STAT3/Snail1信号通路改变上皮间充质转化子E-钙黏着蛋白、N-钙黏着蛋白和波形蛋白的表达,进而促进大肠癌细胞上皮间充质转化。
[Abstract]:Objective to investigate the effect and mechanism of DDT on the epithelial mesenchymal transformation of human colorectal adenocarcinoma cell line DLD1.Methods DLD1 cells were treated with DDT 0.01C 1.0 and 100.0 nmol 路L ~ (-1) for 48 h, and the morphology of the cells was observed under inverted microscope. Real time fluorescence quantitative PCR assay was used to detect E-cadherin (E-cadherin). MRNA expression of vimentin and zinc finger transcription factor Snail1. Western blot assay was used to detect the protein levels of STAT3 and p-STAT3 in signal transduction and transcription activator 3- STAT3 signaling pathway. STAT3 inhibitor WP1066(5 渭 mol. The protein level of p-STAT3- STAT3 in STAT3/Snail1 signaling pathway induced by DDT and the key factor of epithelial mesenchymal transformation, E-cadherin, were detected by Western blot and real-time fluorescence quantitative PCR. The effects of vimentin and zinc finger transcription factor (Snail1) on mRNA levels were observed. Results compared with the control group, DLD1 cells were treated with DDT for 48 h. The relative expression of E-cadherin mRNA decreased significantly from oval to fusiform, which was significantly increased by 1.91 卤0.1 times and 1.5 卤0.2 times of the normal control group (P = 42.4 卤2.8) and vimentin mRNA (P < 0.01). The expression of STAT3 and p-STAT3 protein were increased by 2.1 and 1.8 times as compared with those of the normal control group. The mRNA expression of zinc finger transcription factor Snail1 was significantly higher than that of the normal control group, and was 1.5 卤0.1 times higher than that of the normal control group. After treatment with WP1066 5 渭 mol 路L ~ (-1), a STAT3 inhibitor, the relative expression of zinc finger transcription factor Snail1 was significantly higher than that of the control group. The expression of zinc finger transcription factor Snail1 mRNA was significantly down-regulated in the DDT 1.0 nmol 路L ~ (-1) group, which was 56.3 卤0.9% of the DDT 1.0 nmol 路L ~ (-1) group. At the same time, it inhibited the increase of DDT induced mRNA expression of E-cadherin, which was 2.5 卤0.1 times lower than that of DDT 1.0 nmol 路L ~ (-1) group. Conclusion DDT may change the expression of E-cadherin and vimentin by STAT3/Snail1 signaling pathway. And then promote the epithelial mesenchymal transformation of colorectal cancer cells.
【作者单位】： 山西大学生物技术研究所化学生物学与分子工程教育部重点实验室;山西医科大学第一医院;
【基金】：国家自然科学基金(21207084);国家自然科学基金(31271516) 山西省自然科学基金(2014011027-5) 高等学校科技创新项目(2016122) 山西省回国留学人员科研资助项目(2016-115)~~
【分类号】：R735.34
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