| The purpose of this experiment is to test the effect of CD58 on the secretion of IL-2,IL-4,TGF-β1andTGF-β2 by goat EML.In trial experiment endometrial lymphocyte(EML) from pregnant rabbit were obtained by digestion with trypsin and then by decenfugation. The activation of EML cultured in vitro were tested with MTT assay. In formal experiments CD4-γδT cells were obtained from the EML isolated from pregnant goat. The in vitro activation of goat EML and γδT cells by CD58 (inhibitory rate of total resotte formation 58.89%), progesterone (10ng/mL) and estrone (40pg/mL) was monitored by MTT assay. Additionally, IL-2, IL-4, TGFβ1 and TGFβ2 secretion levels of EML and γδT cells were measured with IL-2 sensitive cell line (CTLL) and ELISA assay kits respectively. The trial experiment firstly proved that normal rabbit EML could be isolated optimally through digestion with 1.25g/L trypsin at 0-4℃ for 7h 30 min to 8h 55min and their activation be tested reliably with MTT assay in vitro . Formal experiment showed that progesterone at physiological concentration activated EML from goat at early gestation (20d-30d) in a dose-dependent manner. But the EML from goat pregnant for 76d were activated more by progesterone at lower concentration than by normal concentration. CD58 at certain range of concentration activated EML of pregnant goat (16d-32d) significantly in dose-dependent manner. Estrone at lower concentration activated EML from goat in early gestation to some extent. Around implantation EML from pregnant goat produced small amount of IL-2, benefiting to maintain of gestation. The IL-4 secreted by EML can be stimulated by CD58 during implantation period, promoting the histological adaptation and immunological tolerance of endometrium.The average proportion of γδT cells in EML from goat (20d-32d after pregnancy) mounted to 54%,γδT cells from different gestation stage could be activated by PHA-P, progesterone and CD58 to different extent. PHA-P uniformly activated endometrial γδT cells from 20d-32d pregnant goat. γδT cells from 20d pregnant goat could be activated significantly by progesterone and insignificantly by CD58. But the γδT cells from 32d pregnant goat could be significantly activated both by progesterone and by CD58 .Goat γδT cells in maternal-fetal interface in early pregnancy could form TGFβs and the secretion be significantly increased by PHA-P. TGFβ1 and TGFβ2 produced at 1.44 ng/mL and 0.77 ng/mL respectively by endometrial γδT cells could be co-stimulated by CD58 and progesterone, showing positive effect on immunological tolerance in goat endometrium. It could be concluded from the results of these experiments and hormone assay that progesterone at the first surge in 10d-32d pregnant goat stimulated EML to secret IL-2 at low concentration and co-stimulated EML to produce IL-4 with CD58 and that progesterone and CD58 could stimulate the secretion of TGFβs either individually or together and the TGFβ1 level is higher than TGFβ2, demonstrating that CD58 and progesterone primarily activated Th2 cells and benefited the endometrial immuno-tolerance.All the finds were of great theoretical significance to immunologically adjusting mechanism in pregnant endometrium and had practical value to success rate of embryo-transplantation and to prevention against fetal loss due to immunological factors. |
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