环孢素A对围着床胚胎与纤连素粘附的调节研究

It is important for embryos to implant in uterus that embryonic integrin αvβ3 adhere to endometrial extracellular matrix (ECM) such as fibronectin (Fn). Researchers found that cyclosporine A (CsA) suppressed immune function, and up-regulated trophoblast proliferation and invasion in early pregnancy at the same time. These implied the potential value of CsA in reproductive medicine. Our former research found that CsA up-regulated integrin αvβ3 expression in peri-implantion mice embryos, enhanced their stretch growing and adhesion on laminin coated dishes. But whether CsA improving embryo adhesion to endometrium is still unknown. In this proposal, mice will be used as model. To research the effect of CsA on adhesive function of embryos to ECM, Fn 120 coated fluorescent green polystyrene microspheres are going to be used as a parameter to test adhesion function of embryos, the cytoplasm of trophoblast cells are going to be photographed with transmission electron microscope to check the ultrastructure changes related to adhesion, the mRNA and protein expression of itgαv and itgβ3 are going to be tested to observe molecular effects. Further more, Ca2+ sensitive fluorescent agent, fluo-3 acetoxymethyl ester, and blockers in cell signal transduction pathway are going to be used to analyse the Ca2+ and the pathway for CsA to regulate embryo's function on adhesion. At last, embryo transfers are going to be introduced with CsA treated embryos to testify its adhesive improving effect in vivo. All the work will apply evidences of adhesion improving effect of CsA on peri-implantation embryos and may shine the light of its use in IVF-ET program on improving embryo implantation.

胚胎整合素αvβ3与子宫内膜细胞外基质纤粘素的粘附在胚胎着床粘着中具有重要的意义。研究发现环孢素A在免疫抑制的同时，可促进早孕期滋养细胞的增殖和侵袭能力，预示着它在生殖医学应用的潜在价值。我们前期研究发现环孢素A上调小鼠围着床胚胎的整合素αvβ3的表达，促进胚胎的伸展生长和与层粘蛋白的粘附，但能否改善胚胎生理粘附的功能尚缺乏证据。本研究以小鼠为模型，拟通过胚胎粘附纤粘素包被的绿荧光聚苯乙烯微球的能力作为功能指标、滋养细胞超微结构为形态学指标、itgαv和itgβ3为分子表达指标，来分析和判断环孢素A对围着床胚胎粘附子宫内膜细胞外基质的功能影响。进一步，通过钙敏荧光试剂分析细胞内游离Ca2+水平、细胞信号传导阻断等手段，分析环孢素A调节围着床胚胎与纤粘素粘附功能的机制。最后通过体外胚胎培养和胚胎移植，验证环孢素A在体内有改善胚胎着床的可能，为临床改善胚胎着床奠定科学依据。