酪酸梭菌对HT-29细胞水通道蛋白3表达的影响及JNK和p38信号通路对其调控作用

发布时间：2018-02-24 21:18

本文关键词： 酪酸梭菌水通道蛋白 HT-细胞 JNK/p信号通路　出处：《中国药物与临床》2014年12期 　论文类型：期刊论文

【摘要】：目的明确酪酸梭菌对人结肠腺癌细胞HT-29细胞水通道蛋白3(AQP3)m RNA和蛋白表达的影响以及c-Jun氨基末端激酶(JNK)和p38信号转导通路对其表达的调控作用。方法用酪酸梭菌上清液作用于HT-29细胞后,应用实时定量反转录聚合酶链反应(RT-PCR)和蛋白印迹方法检测AQP3 m RNA和蛋白表达水平。AQP3表达水平有差异后测定总JNK、p38以及磷酸化的JNK(P-JNK)和磷酸化的p38(P-p38)蛋白表达水平。后分别加用JNK抑制剂SP600125和p38抑制剂SB203580后测定AQP3蛋白表达量,观察JNK和p38对AQP3表达的调控作用。统计学处理采用单因素方差分析。结果实时定量RT-PCR和蛋白印迹法结果显示酪酸梭菌上清液不同浓度(1∶4和1∶8组)作用HT-29细胞12 h以及同一浓度1∶4作用不同时间(24、12、6 h组)的AQP3 m RNA和蛋白表达水平均较无酪酸梭菌上清液作用组明显上调(P均0.05)。蛋白印迹法结果显示酪酸梭菌上清液组的Pp38/p38和P-JNK/JNK蛋白相对量较空白对照组表达水平明显上调(F=26.065和65.034,P0.05)。蛋白印迹法结果显示加入p38抑制剂SB203580 10μg/ml组和20μg/ml的AQP3蛋白表达相对量为(0.7±0.4)和(0.6±0.6)、加入JNK抑制剂SP600125 10μg/ml组和20μg/ml的AQP3蛋白表达相对量为(0.7±0.4)和(0.5±0.4)均较未加入抑制剂组(1.1±0.7)AQP3蛋白表达水平明显下调(F=36.225,P0.05)。结论酪酸梭菌上清液能够上调HT-29细胞的AQP3表达进而调节肠道的水代谢。JNK和p38信号转导通路可能参与调控AQP3的表达。
[Abstract]:Objective to investigate the effect of Clostridium caseinate on the expression of aquaporin 3AQP3M RNA and protein in human colon adenocarcinoma cell line HT-29 and the regulation of c-Jun amino-terminal kinase (c-Jun) and p38 signal transduction pathway on its expression. After the solution was treated with HT-29 cells, Real time quantitative reverse transcription polymerase chain reaction (RT-PCR) and Western blotting were used to detect the expression level of AQP3 m RNA and protein. AQP3 expression level was different. The expression levels of total JNKP 38, phosphorylated JNKN P P K NKK and phosphorylated p38 P P 38 protein were measured. JNK inhibitor SP600125 and p38 inhibitor SB2035 80 were not used to detect the expression of AQP3 protein. The effect of JNK and p38 on the expression of AQP3 was observed. Univariate ANOVA was used in statistical analysis. Results the results of real-time quantitative RT-PCR and Western blotting showed that the supernatants of Clostridium caseinate at different concentrations (1: 4 and 1: 8) treated HT-29 cells for 12 h. The expression levels of AQP3 m RNA and protein in the same concentration of 1: 4 at different time points were significantly higher than those in the control group without Clostridium butyricum supernatant. The results of Western blot showed that Pp38/p38 and P-JNK-JNK eggs in the supernatant group of Clostridium caseinate were significantly up-regulated than those in the control group without Clostridium caseate supernatant. Compared with the blank control group, the relative dose of white increased the expression level of FN 26.065 and 65.034% P0.050.The results of Western blot showed that the relative levels of AQP3 protein expression in the p38 inhibitor SB203580 10 渭 g / ml group and 20 渭 g / ml group were 0.7 卤0.4) and 0.6 卤0.6 渭 g / ml, respectively. The AQP3 protein expression in the 10 渭 g / ml and 20 渭 g / ml JNK inhibitor SP600125 group was 10 渭 g / ml and 20 渭 g / ml respectively. Compared with the control group, the expression level of AQP3 protein was significantly down-regulated. Conclusion the supernatant of Clostridium caseinate can upregulate the expression of AQP3 in HT-29 cells and regulate the water metabolism. JNK and p38 signal transduction pathway of intestinal tract. Participate in the regulation of AQP3 expression.
【作者单位】：山西大学化学化工学院;山西省人民医院消化科;
【基金】：山西省卫生厅科技攻关项目(201201054)
【分类号】：R329.2

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