二补助育汤对高龄小鼠子宫内膜容受性及子宫内膜间充质干细胞衰老的影响  

Effects of Erbu Zhuyu Decoction on endometrial receptivity and EMSCs senescence in aged mice

作　　者：郑志博 刘雁峰[2] 陈洁 李影 孔鑫靓 侯婷婷 丁彩飞[1] 江雪娟 ZHENG Zhibo;LIU Yanfeng;CHEN Jie;LI Ying;KONG Xinliang;HOU Tingting;DING Caifei;JIANG Xuejuan(Department of Reproductive,Zhejiang Hospital of Integrated Traditional Chinese and Western Medicine,Hangzhou 310003,China)

机构地区：[1]浙江省中西医结合医院生殖医学科,杭州310003 [2]北京中医药大学东直门医院妇科 [3]浙江省人民医院中西医结合科 [4]青岛大学附属山东省妇幼保健院中医科

出　　处：《环球中医药》2024年第10期1963-1972,共10页Global Traditional Chinese Medicine

基　　金：国家自然科学基金面上项目(82074480、82274567);杭州市红十字会医院博士科研启动基金项目(HHBJ2024001)。

摘　　要：目的 探讨二补助育汤对高龄小鼠子宫内膜容受性及子宫内膜间充质干细胞(endometrial mesenchymal stem cells, EMSCs)衰老的影响。方法 将8~9月龄妊娠小鼠随机分为模型组、阿司匹林组、补佳乐组和二补助育汤低、中、高剂量组,另选择6~8周龄妊娠小鼠为空白组,每组10只。空白组和模型组小鼠每日灌服0.5%羧甲基纤维素钠溶液,其余各组小鼠每日灌服相应药物,连续灌胃5天。于妊娠第5天下午脱颈处死。裸眼观察各组小鼠子宫形态,统计妊娠率及平均胚胎着床位点数。取各组小鼠子宫内膜组织,实时荧光定量PCR、蛋白免疫印迹法检测同源盒基因A10(homeobox gene A10,HOXA10)、白血病抑制因子(leukemia inhibitory factor, LIF)、整合素αvβ3 mRNA及蛋白的表达;分离、提取各组小鼠EMSCs细胞,免疫荧光法进行细胞鉴定,Real-time PCR、蛋白免疫印迹法检测EMSCs衰老指标P53、P21、P16 mRNA及蛋白的表达。结果 与空白组比较,模型组小鼠子宫颜色淡黯,着床部位宫体较细;妊娠率下降,但差异无统计学意义(P>0.05);平均胚胎着床位点数减少(P<0.05);子宫内膜HOXA10、LIF、整合素αvβ3 mRNA及蛋白表达水平显著下降(P<0.05)。与模型组比较,不同剂量的二补助育汤组小鼠子宫色泽红润,着床部位宫体明显膨大,妊娠率升高,但差异无统计学意义(P>0.05);平均胚胎着床位点数增加(P<0.05);子宫内膜HOXA10、LIF、整合素αvβ3 mRNA及蛋白表达均呈上升趋势(P>0.05)。分离、提取各组小鼠EMSCs,免疫荧光显示细胞高表达CD29、CD44,低表达CD34、CD45,提示成功获得EMSCs。与空白组比较,模型组EMSCs衰老指标P53、P21、P16 mRNA及蛋白表达显著升高(P<0.05);与模型组比较,二补助育汤中、高剂量组及各西药组P53、P21、P16 mRNA及蛋白表达水平显著下降(P<0.05)。结论 二补助育汤可上调高龄小鼠子宫内膜HOXA10、LIF、整合素αvβ3的表达,降低EMSCs衰老指标P53、P21�Objective To investigate the effects of Erbu Zhuyu Decoction on endometrial receptivity and EMSCs senescence in aged mice.Methods Pregnant mice aged 8~9 months were randomly divided into 6 groups:the model group,the low-dose EBZY group,the medium-dose EBZY group and the high-dose EBZY group,the aspirin group and the progynova group,with 10 mice in each group.In addition,10 pregnant mice aged 6~8 weeks were selected as the blank group.Mice in the blank group and the model group were perfused with 0.5%Caboxy Methyl Cellulose solution every day,and mice in the other groups were perfused with corresponding interventional medicines every day for 5 days.On the afternoon of the 5th day of pregnancy,uterine morphology of mice in each group were observed,pregnancy rate and average number of embryos were calculated and the mRNA and protein expressions of endometrial HOXA10,LIF and integrinαvβ3 were detected.At the same time,EMSCs cells of mice in each group were separated and extracted and the cells were identified by immunofluorescence.The mRNA and protein expressions of P53,P21 and P16 in EMSCs were detected.Results Compared with the blank group,uterus in the model group were pale,uterine of implantation site were thinner,pregnancy rate was decreased,but the difference was not statistically significant(P>0.05),average number of embryos was decreased(P<0.05)and the mRNA and protein expression of endometrial HOXA10,LIF and integrinαvβ3 were significantly decreased(P<0.05).Compared with the model group,uterus in different doses of Erbu Zhuyu Decoction groups were red,uterine of implantation site was significantly enlarged,pregnancy rate was increased,but the difference was not statistically significant(P>0.05),average number of embryos was increased(P<0.05)and the mRNA and protein expressions of HOXA10,LIF and integrinαvβ3 in endometrium had upward trend(P>0.05).The results of immunofluorescence showed that CD29 and CD44 were highly expressed,while CD34 and CD45 were low,indicating that EMSCs were successfully isolate

关 键 词：二补助育汤 子宫内膜容受性 高龄 子宫内膜间充质干细胞 衰老 

分 类 号：R285.5[医药卫生—中药学]