不同雄性淋巴细胞及其Exosomes过继转输改善妊娠丢失孕鼠胚胎发育的比较分析

摘要

目的：分析雄性T淋巴细胞及其Exosomes（Exo）过继转输对妊娠丢失孕鼠胚胎发育的保护作用。方法：分别将健康男性外周血单个核细胞（ PBMC）、BALB／c及DBA／2雄鼠脾细胞体外诱导增殖，蔗糖密度梯度离心联合超滤制备Exo。CBA／J（♀）×BALB／c（♂）为正常妊娠对照，CBA／J（♀）×DBA／2（♂）为妊娠丢失URSA模型。将URSA孕鼠随机分组，经尾静脉或皮下注射，分别予以父方、非父方、无关雄鼠脾细胞或其Exo，以及男性PBMC来源Exo过继转输。观察胚胎发育，计算胎盘体积、胚胎吸收率及妊娠丢失率。结果：URSA组胎盘体积明显缩小，胚胎吸收率、妊娠丢失率显著升高（均P＜0.0005）；尾静脉或皮下注射不同来源细胞及Exo后，孕鼠胎盘体积显著恢复，胚胎吸收率、妊娠丢失率显著下降（均P＜0.0005）。不同治疗组内两种注射途径产生的疗效无差异（均P＞0.05）；不同细胞来源Exo过继转输后，胚胎吸收率的下降幅度显著强于细胞治疗组（均P＜0.05）；男性Exo过继转输后，妊娠丢失率的下降幅度显著强于细胞治疗组及鼠源Exo治疗组（均P＜0.05）。结论：过继转输不同来源雄性T淋巴细胞及其Exo均可明显改善胚胎发育。 Exo作为非细胞成分生物学制剂，有望取代传统细胞过继免疫治疗。%Objective:To analyze the protective effects of adoptively transferring different patrilineal lymphocytes and their Exosomes ( Exo) on fetation of mice with pregnancy loss comparatively.Methods: The peripheral blood mononuclear cells ( PBMC) from healthy men and the splenocytes from BALB/c and DBA/2 male mice were induced in vitro ,and their Exo were isolated through sucrose gradient ultra-centrifugation combined with ultrafiltration.The mice of CBA/J (♀) mated with BALB/c (♂) were enrolled as control group of normal pregnancy ,and the CBA/J (♀) mated with DBA/2 (♂) as URSA of pregnancy loss experimental animal model.The mice in URSA group were randomly divided into each group with treatment through adoptively transferring , which were injected intravenously or subcutaneously with splenocytes or splenocytes -derived Exo from mated DBA/2,unmated DBA/2 or unrelated BALB/c,also PBMC-derived Exo from men,respectively.And then,the placenta volumes,rates of fetal absorption and pregnancy loss were calculated to observe the fetation of embryos.Results:Compared with the group of normal pregnancy ,the placenta volumes from URSA group decreased greatly ,and rates of fetal absorption and pregnancy loss elevated greatly ( all P<0.000 5 ).After transferring different sources of cells and their Exo through different injection ,the placenta volumes resumed to the level of normal pregnancy ,and the rates of fetal absorption and pregnancy loss decreased significantly ( all P<0.000 5 ).No differences were observed after treatment through injecting intravenously or subcutaneously ( all P>0.05 ).After transferring the Exo derived from either male mice or healthy men,the level of decreased fetal absorption rates were more than that in cellular-therapy groups ( all P<0.05 ).After transferring the Exo derived from men ,the level of decreased pregnancy loss rates were more than that in cellular -therapy groups and mice splenocytes-derived Exo group ( all P<0.05 ).Conclusion:Adoptively transferring patrilineal T lymphocytes and their Exo can greatly improve the fetation.Exo should become a non-cellular bio-remedy,which is expected to replace traditional immunotherapy of adoptively transferring lymphocytes.