补肾、活血对超促排卵小鼠着床期间子宫内膜MMP-2、MMP-9、TIMP-3表达的影响

Effect of Bu-Shen-An-Tai Recipe and Its Two Components on Expression of MMP-2,MMP-9 and TIMP-3 in Endometrium of Controlled Ovarian Hyperstimulation Mice

目的观察中药补肾安胎方及其拆方对超促排卵模型小鼠子宫内膜基质金属蛋白酶-2、-9(MMP-2,MMP-9),基质金属蛋白酶抑制因子-3(TIMP-3)表达的影响。方法以昆明种小鼠为研究对象,并以孕马血清(PMSG)诱发超促排卵模型,将小鼠随机分为正常组(自然周期)、模型组(PMSG+HCG)、活血组(模型+活血中药)、补肾组(模型+补肾中药)、补肾安胎组(模型+补肾安胎中药),每组24只,自开始腹腔注射PMSG或生理盐水的当天清晨分别予双蒸水或中药灌胃,直至处死小鼠的当天清晨。于妊娠第5、7天取小鼠子宫,免疫组化法检测MMP-2、MMP-9、TIMP-3的表达部位,实时定量PCR检测MMP-2、MMP-9、TIMP-3的mRNA表达。结果免疫组化显示:MMP-2、MMP-9及TIMP-3蛋白在妊娠第5天表达在初级蜕膜带,随着蜕膜化发展,逐渐表达于整个蜕膜化间质细胞中。实时定量PCR显示:妊娠第5天,与正常组相比,模型组MMP-9、TIMP-3mRNA表达均升高,差异有统计学意义(P<0.05)。妊娠第7天,与正常组相比,模型组MMP-2、MMP-9及TIMP-3mRNA表达均明显降低(均P<0.05);各中药组均高于模型组,差异有统计学意义(均P<0.05),其中活血组差异更显著(P<0.01)。结论补肾安胎方及其拆方均能对超促排卵小鼠基质金属蛋白酶及其抑制因子的表达有促进作用,未发现活血药对着床有负面影响。

Objective To examine the effect of Bu-Shen-An-Tai Recipe（BSATR）and its two components（Bushen recipe and Huoxue recipe）on the expression of matrix metalloproteinase-2（MMP-2）,MMP-9 and tissue inhibitor of metalloproteinase-3（TIMP-3）in controlled ovarian hyperstimulation（COH）mice.Methods Pregnant mare serum gonadotropin（PMSG）was used for establishment of COH models in Kunming female mice.The mice were randomly divided into 5 groups,including normal group（N）,COH model group（M）,Bushen group（BS）,Huoxue group（HX）and BSATR group（BH）（n=24each）.Double distilled water or traditional Chinese medicine（TCM）decoction was intragastrically given from the early morning of the day when PMSG or normal saline were intraperitoneally injected till the sacrifice of the mice.Uterus samples were obtained at 5 and 7 days of pregnancy.The expression of MMP-2,MMP-9 and TIMP-3was detected in endometrial tissues by immunohistochemistry and real-time PCR.Results Immunohistochemistry showed that MMP-2,MMP-9and TIMP-3 were expressed in the primary decidual zone at 5 days of pregnancy and gradually in all the decidualized stromal cells with the decidualization.Real-time PCR showed that at 5 days of pregnancy,the expression of MMP-9and TIMP-3 mRNA was significantly increased in the group M as compared with the group N（P0.05）;it was significantly reduced at 7days in the group M（P0.05）.The groups BS, HX and BH had significantly higher expression of MMP-2,MMP-9and TIMP-3in endometrium than the group M at 7days of pregnancy（P0.05）.And the expression in the group HX was strongest（P0.01）.Conclusion BSATR and its two components can improve COH-induced poor endometrial development by increasing the expression of MMP-2,MMP-9 and TIMP-3.Moreover,Huoxue recipe was not found to have adverse effects on implantation.