Toder V, Savion S, Gorivodsky M, Shepshelovich J, Zaslavsky Z, Fein A, Torchinsky A
Department of Embryology and Teratology, Sackler School of Medicine, Tel-Aviv University, Israel.
J Reprod Immunol. 1996 May;30(2-3):173-85. doi: 10.1016/0165-0378(96)00960-6.
Intra-uterine immunization of mice with paternal allogeneic or xenogeneic (rat) splenocytes was found to increase embryo tolerance to cyclophosphamide (CP)-induced teratogenesis. As the CP-induced teratogenic effect was shown to be associated with apoptosis, the present study was designed to investigate whether the protective effect of immunopotentiation may be realized via an alteration of CP-induced apoptosis. Various doses of CP were injected intraperitoneally into ICR mice on day 12 of pregnancy. Intra-uterine immunization with xenogeneic rat splenocytes was carried out 3 weeks before mating. Implantation sites, resorptions, live and dead fetuses, as well as soft tissue anomalies and external malformations, were recorded to evaluate the CP-induced embryotoxic effect. In parallel, flow cytometric analysis and DNA fragmentation assay were used for evaluation of CP-induced apoptosis in limbs, tail and whole embryos. The treatment of mothers with a high dose of CP induced the death of almost all embryos and striking fetal growth retardation in survivors. This strong embryotoxic effect was accompanied by very prominent DNA degradation in cells collected from whole embryos. Immunostimulation caused a dramatic decrease of embryonal loss (by approximately 50%) and a significant (about 30%) increase in fetal weight. Such an increase in fetal survival and in fetal weight was found to be accompanied by a clear decrease in apoptosis level in embryo cell population as judged by DNA gel electrophoresis with subsequent quantitation of DNA fragmentation in negatives by an image analysis technique. After treatment with a low dose of CP, a decrease in the proportion of fetuses with limb and tail anomalies in immunized females was accompanied by a decrease in the proportion of apoptotic nuclei in cells taken from limbs and tails. The results of this study suggest that the teratogen-induced apoptosis may, at least partly, be dependent on fetomaternal immune interactions.
用父本同种异体或异种（大鼠）脾细胞对小鼠进行宫内免疫，发现可增强胚胎对环磷酰胺（CP）诱导的致畸作用的耐受性。由于已表明CP诱导的致畸作用与细胞凋亡有关，本研究旨在调查免疫增强的保护作用是否可通过改变CP诱导的细胞凋亡来实现。在妊娠第12天，将不同剂量的CP腹腔注射到ICR小鼠体内。在交配前3周用异种大鼠脾细胞进行宫内免疫。记录着床部位、吸收情况、活胎和死胎，以及软组织异常和外部畸形，以评估CP诱导的胚胎毒性作用。同时，采用流式细胞术分析和DNA片段化测定来评估CP在四肢、尾部和整个胚胎中诱导的细胞凋亡。用高剂量CP处理母体导致几乎所有胚胎死亡，存活者出现明显的胎儿生长迟缓。这种强烈的胚胎毒性作用伴随着从整个胚胎收集的细胞中非常显著的DNA降解。免疫刺激导致胚胎损失显著减少（约50%），胎儿体重显著增加（约30%）。通过DNA凝胶电泳及随后用图像分析技术对阴性条带中的DNA片段进行定量分析判断，发现胎儿存活率和体重的这种增加伴随着胚胎细胞群体凋亡水平的明显降低。用低剂量CP处理后，免疫雌性小鼠中四肢和尾部异常胎儿的比例降低，同时从四肢和尾部获取的细胞中凋亡细胞核的比例也降低。本研究结果表明，致畸剂诱导的细胞凋亡可能至少部分取决于母胎免疫相互作用。
