健脾化瘀解毒方调控Wnt/β-catenin/GSK3β通路抑制胃癌前病变大鼠Lgr5^+癌干细胞早期转移机制研究

Study on the mechanism of Jianpi Huayu Jiedu Formula on regulating Wnt/β-catenin/GSK3β pathway to inhibit the early metastasis of Lgr5^+ cancer stem cells in rats with GPL

目的:探讨健脾化瘀解毒中药胃炎1号抑制胃癌前病变(GPL)大鼠Lgr5^+癌干细胞早期转移的分子学机制。方法:SD大鼠随机均分为空白组、模型组、维酶素组和胃炎1号组。MNNG水溶液自由饮用+饥饱失常+耗气泻下法复制GPL大鼠模型。连续灌胃给药10周后检测GPL大鼠胃黏膜上皮细胞β-catenin、Lgr5^+、MMP-7、GSK3β的表达。结果:与空白组比较,模型组大鼠胃黏膜上皮的β-catenin与MMP-7蛋白累积面积、光密度、积分光密度显著升高(P<0.05),GSK3β蛋白面积、光密度、灰度及Lgr5^+累积面积、积分光密度显著升高(P<0.05),GPL大鼠Lgr5^+胃癌干细胞由基底侧向胃腔侧侵袭迁移。与模型组比较,胃炎1号GPL大鼠胃黏膜上皮细胞β-catenin蛋白累积面积、光密度、积分光密度,GSK3β蛋白面积、光密度、灰度,Lgr5^+表达累积面积、积分光密度,MMP-7积分光密度显著降低(P<0.05),胃炎1号组Lgr5^+表达面积和强度均减少,散在分布于胃黏膜的固有层和非固有层。结论:胃炎1号可通过下调Wnt信号通路β-catenin、GSK3β、MMP-7的表达,抑制GPL大鼠Lgr5^+胃癌干细胞增殖、侵袭迁移。

Objective: To explore the molecular mechanism of Weiyan Yihao, a Chinese medicine formula with the effect of invigorating spleen and expelling blood stasis for removing toxicity, in inhibiting the early metastasis of Lgr5^+ cancer stem cells in rats with gastric precancerous lesions(GPL). Methods: The SD rats were randomized divided into four groups, including normal group,model group, Vitacoenzyme group and Weiyan Yihao group. The rat model of GPL was replicated by the method of MNNG aqueous solution drinking freely, irregular diet and consuming Qiandpurgation. The expression of β-catenin, Lgr5^+, MMP-7 and GSK3β in gastric epithelial cells of GPL rats were detected after continuous intragastric administration for 10 weeks. Results: Compared with thenormal, in the model mice, the protein accumulation area, the optical density, the integral optical density of β-catenin and MMP-7 were significantly increased(P<0.05). The protein area,the optical density the grayscale of GSK3β significantly increased(P<0.05); the accumulation area, the integral optical density of Lgr5^+ significantly increased(P<0.05), Lgr5^+ gastric cancer stem cells invaded and migrated to the stomach cavity side form stomach recess bottom. Compared with the model group, weiyanyihao could inhibit the expression of β-catenin of the protein accumulation area, the optical density, the integral optical density; Weiyan Yihao inhibites the experssion of GSK3β protein area, light density, gray. The accumulated area and integral optical density of Lgr5^+ expression, MMP-7 integral optical density were significantly reduced(P<0.05). The area and intensity of expression of Lgr5^+ in Weiyan Yihao group were reduced(P<0.05), scattered in the lamina propria and non lamina propria of gastric mucosa. Conclusion: Our findings suggested that the therapeutic mechanisms of Weiyan Yihao in treating GPL might be related with its inhibitory effects on the expressions of β-catenin, GSK3β, MMP-7and the aberrantactivation of Wnt pathway, so that the formular can inhibite the proliferation, invasion and migration of Lgr5^+ gastric cancer stem cells in GPL rats.