二硫化碳通过改变卵巢黄体巨噬细胞极化和功能导致胚胎植入障碍

Carbon disulfide induced embryo implantation by changing the polarization and functional of ovarian corpus luteum macrophages

目的探讨二硫化碳(CS2)暴露对孕鼠卵巢黄体巨噬细胞极化和功能的影响,从免疫损伤角度解析巨噬细胞在CS2致胚胎植入障碍中的作用。方法建立围植入期不同时间点暴露于CS2的小鼠模型,3个染毒组分别于受孕第3(GD3)、4(GD4)和5(GD5)天经腹腔染毒CS2(0.4 LD50,631.4 mg/kg)1次,各组设立平行溶剂对照(等体积橄榄油)。各组动物染毒后于不同时间点设立连续观察终点,取卵巢组织进行HE染色、免疫组织化学染色及实时荧光定量PCR实验,检测孕鼠卵巢组织中巨噬细胞极化和功能的改变。结果 (1)卵巢黄体巨噬细胞极化平衡在CS2染毒后表现为M1型巨噬细胞为主导(M1>M2,如GD3染毒组GD4观察终点,t=10.000,P<0.001),后期转变为M2型巨噬细胞为主导(M1<M2,如GD5染毒组GD6观察终点,F=10.235,P=0.003);(2)TNF-α和Vegfa的mRNA表达水平升高(如Vegfa在GD4染毒组GD7观察终点,F=6.801,P=0.007);孕鼠卵巢黄体表现为血管扩张与充血以及结构改变(如GD4染毒组GD5观察终点的结构评价,F=8.638,P=0.019);Cox2和Hif1-α的mRNA表达均呈先升高后下降的趋势(如Cox2在GD3染毒组GD5观察终点,F=6.184,P=0.035)。结论围植入期CS2暴露致胚胎植入障碍的机制可能是卵巢黄体巨噬细胞极化平衡与功能改变,引发并加重卵巢黄体结构和功能受损。

Objective To investigate the polarization and functional changes of ovarian corpus luteum macrophages in pregnant mice exposed to carbon disulfide（ CS2） and to explore the role of macrophages in implantation failure induced by CS2 from the perspective of immune injury. Methods The animal model of exposure to CS2 at different time points was established. Single dose of CS2（ 0.4 LD50,631.4 mg/kg） was injected intraperitoneally at the third（ GD3）,fourth（ GD4）,and fifth（ GD5） day of gestation,and the solvent control group was set up（ equal volume of olive oil）. The continuous observation endpoints were designed at different time points after exposure. Ovarian tissues were collected for HE staining,immunohistochemistry,and real-time quantitative PCR detection,and the polarization and functional changes of macrophages were also determined. Results（1） The polarization balance of ovarian corpus luteum macrophages changed from the M1 type of macrophages（ M1〉M2,such as GD3-treated group at GD4 observation endpoint,t =10.000,P〈0.001） to the M2 type of macrophages（ M1〈M2,such as GD5-treated group at GD6 observation endpoint,F = 10.235,P = 0.003）.（2） The expression of TNF-α and Vegfa mRNA increased significantly（ such as Vegfa in GD4-treated group at GD7 observation endpoint,F = 6.801,P = 0.007）. The ovarian corpus luteum mainly characterized by vasodilatation and congestion,and structural changes（ such as the structure evaluation of GD4-treated group at GD5 observation endpoint,F = 8.638,P = 0.019）. The expression of Cox2 and Hif1-α mRNA showed a tendency of initial increase and then decrease（ such as Cox2 in GD3-treated group at GD5 observation endpoint,F = 6. 184,P = 0. 035）.Conclusion The mechanism of embryo implantation failure induced by CS2 exposure may be due to the polarization and functional changes of ovarian corpus luteum macrophages,which triggers and aggravates the damage in the structure and function of ovarian corpus luteum.