内异康复片对EMT大鼠VEGF、MMP-9、HGF及Wnt4、Wnt5α的调控机制研究

Regulatory Mechanism Study of Neiyi Kangfu Tablet on VEGF,MMP-9,HGF,Wnt4 and Wnt5α of Endometriosis Rats

目的:分析内异康复片对子宫内膜异位症(EMT)大鼠异位内膜黏附、浸润及增生的调控作用,并探讨其作用机制。方法:建立子宫内膜异位症大鼠模型,随机分为模型对照组、阳性对照组、中药低、中、高剂量组,每组10只。治疗3周后,测量各组大鼠异位内膜体积;另取模型动物的异位内膜组织,分离异位内膜细胞,体外培养,设空白对照组、阳性对照组、中药低剂量组、中药中剂量组、中药高剂量组,每组4个复孔。采用ELISA法测定血管内皮生长因子(VEGF)、基质金属蛋白酶(MMP)、肝细胞生长因子(HGF)含量;Western Blot检测Wnt4、Wnt5α的表达。结果:动物实验结果:与模型对照组比较,中药各剂量组、阳性对照组的异位内膜体积显著缩小,异位内膜厚度变薄,腺体面积减少,差异均有统计学意义(P<0.01,P<0.05);中药高剂量组、阳性对照组的腺体数目显著少于模型对照组,差异均有统计学意义(P<0.05);中药高剂量组VEGF、MMP-9含量显著低于模型对照组,差异均有统计学意义(P<0.05)。细胞实验结果:中药低、中、高各剂量组培养上清液的VEGF含量显著低于空白对照组,中药高剂量组MMP-9含量显著低于空白对照组,中药高剂量组和阳性对照组Wnt4、Wnt5α表达水平均显著低于空白对照组,差异均有统计学意义(P<0.05,P<0.01)。结论:内异康复片治疗EMT的机理可能与抑制异位内膜细胞分泌VEGF、MMP-9,以及调节Wnt信号转导通路有关。

Objective： To analyze the regulatory function of Neiyi Kangfu tablet on adhesion, infiltration and proliferation of Ectopic endometrial in rats with endometriosis（EMT） and to discuss its mechanism. Methods： Established rat models with endometriosis, and divided them into model control group, positive control group, Chinese medicine （CM） low-dose, mid-dose, high-dose group, 10 cases in each group. After 3 week of treatment, endometriosis volume of rats in every group was measured. In addition, took endometriosis tissue and separated endometriosis cell, then cultured in vitro, divided into blank control group, positive control group, CM low-dose, mid-dose, high-dose group. Detected levels of vascular endothelial growth factor（VEGF）, matrix metalloproteinase （MMP） and hepatocyte growth factor（HGF） by ELISA method, detected expression of Wnt4 and Wnt5α by Western Blot. Results： Comparing with model control group, endometriosis volume of CM every dose group and positive control group was decreased obviously, endometriosis thickness was thinned, gland area was reduced, the differences being significant（P〈 0.01 or P〈 0.05）. Gland numbers in CM high-dose group and positive control group were lower than that in model control group, the difference being significant（P〈 0.05）. Levels of VEGFand MMP-9 in CM high-dose group were lower than those in model-dose group, the differences being significant（P〈 0.05）. The levels of VEGF in culture supernate of CM low-dose, mid-dose, high-dose group were all lower than those in blank control group, the level of MMP-9 in CM high-dose group was lower than that in blank control group, expression levels of Wnt4 and Wnt5α were all lower than those in blank control group, the differences being significant（P 〈0.05 or P 〈0.01）. Conclusion： The mechanism of Neiyi Kangfu tablet for EMT can be related to inhibit endometriosis cell to secrete VEGF and MMP-9 and regulate Wnt signal transduction pathway.