【摘要】 目的:探讨脾气虚肺癌小鼠miR-574-5p对Wnt/β-catenin信号转导途径基因表达影响及加味黄芪建中汤干预作用。方法:从50只健康清洁级C57BL/6小鼠中随机选取30只,采用苦寒泻下法+饥饿法+疲劳法建立脾气虚证模型,大黄浓缩液灌胃,同时每日游泳至力竭,并间隔1日节食,连续14d,第15天分别从正常和模型小鼠中随机各取10只小鼠,采用比色法和ELISA法测定血清D-木糖、胃泌素水平,验证造模成功。通过接种传代肿瘤细胞悬液接种Lewis肿瘤组织后,随机将实验小鼠分为单纯肺癌组、脾气虚肺癌组和中药组,中药组给予加味黄芪建中汤灌胃,其他两组给予0.9%氯化钠溶液灌胃,连续15d。脱臼处死小鼠后,完整分离皮下肿瘤称重,完整分离肺脏,计算转移结节数,RT-PCR检测肿瘤组织miR-574-5p、QKI6、β-catenin、p27Kip1 mRNA含量,Western Blot检测肿瘤组织QKI6、β-catenin、p27 Kip1蛋白的表达。结果:脾气虚证模型组小鼠血清D-木糖和胃泌素均较正常组有明显下降(P<0.05);与单纯肺癌组比较,脾气虚肺癌组皮下肿瘤明显增大,肺转移结节数明显增多(P<0.05),肿瘤组织中miR-574-5p、β-catenin mRNA和蛋白的表达均明显升高,而QKI6、p27Kip1 mRNA和蛋白的表达均显著降低(P<0.05);与脾气虚肺癌组比较,中药组肿瘤明显缩小,肺转移结节数明显减少(P<0.05),中药组肿瘤组织中miR-574-5p、β-catenin mRNA和蛋白表达均明显降低,而QKI6、p27Kip1 mRNA和蛋白表达明显升高(P<0.05)。结论:加味黄芪建中汤对脾气虚证肺癌具有较好的治疗效果,其机制可能与降低miR-574-5p的表达进而抑制Wnt/β-catenin信号转导途径的激活有一定的联系。更多还原

【Abstract】 Objective: To investigate the changes of Wnt/β-catenin signaling pathway in lung carcinoma mice with spleen deficiency syndrome induced by miR-574-5 p and the intervention of modified Huangqi Jianzhong Decoction. Methods: Thirty healthy and clean C57 BL/6 mice were randomly selected from 50 mice. Spleen qi deficiency syndrome model was established by bitter cold diarrhea plus starvation and fatigue. Mice were administrated with rhubarb concentrated solution, and swimming to exhaustion daily and dieting one day at intervals for 14 consecutive days. On the 15 th day, 10 mice were randomly selected from normal and model mice group respectively. Colorimetric and ELISA were used to determine the level of D-xylose and gastrin in serum. The mice were randomly divided into lung carcinoma group, lung carcinoma with spleen qi deficiency syndrome group and traditional Chinese medicine group after inoculating Lewis tumor tissue with passaged tumor cell suspension. The Chinese medicine group was given modified Huangqi Jianzhong Decoction for gastric lavage, while the lung carcinoma group and lung carcinoma with spleen qi deficiency syndrome group were given saline for 15 consecutive days. After the mice were killed by dislocation, the subcutaneous tumors were completely separated and weighed, the lungs were completely separated, and the number of metastatic nodules was calculated. The contents of microRNA-574-5 p, QKI6, β-catenin, and p27 Kip1 in tumor tissues were detected by RT-PCR, and the expressions of QKI6, β-catenin and p27 Kip1 protein in tumor tissues were detected by Western Blot. Results: The levels of D-xylose and gastrin in serum of spleen qi deficiency syndrome group were significantly lower than those in normal group(P<0.05); compared with lung carcinoma group, the number of subcutaneous tumors and pulmonary metastatic nodules increased significantly in lung carcinoma group with spleen qi deficiency syndrome(P<0.05); the mRNA and protein expressions of miR-574-5 p and β-catenin increased significantly in lung cancer tissue, while the QKI6 and p27 Kip1 decreased significantly in lung carcinoma group with spleen qi deficiency syndrome(P<0.05); the volume of tumor, the number of pulmonary metastatic nodules and the expression of microRNA-574-5 p and β-catenin were significantly decreased in the Chinese medicine group, while the expression of QKI6 and p27 Kip1 were significantly increased in the Chinese medicine group(P<0.05). Conclusion: Modified Huangqi Jianzhong Decoction has a good therapeutic effect on the lung carcinoma with spleen qi deficiency syndrome. Its mechanism may be related to the reduction of the expression of microRNA-574-5 p and the inhibition of the activation of Wnt/β-catenin signaling pathway.更多还原