| Objectives:1 To investigate the expression and localization of calcitonin(CT) in human endometrium during menstrual cycle and evaluate its significance in the implantation of the blastocyst.2 To study the expression of CT in human uterine epithelia cells in vitro culture and its differential regulation by estradiol , progesterone and the progesterone receptor antagonist,RU486.3 To study the effects of exposure to CT on different stages of preimplantation embryos and uterine epithelia or stroma cells intracellular Ca 2+ ,and to evaluate the mechanism of CT during the implantation.Materials and Methods:1 Thirty-four samples of endometrium from fertile women were detected.The endometrium was fixed in 10% buffered formalin(PH 7.4) for immunohistochemistry.The expression and location of CT was examined by immunohistochemical method(SABC) in human endometrium during the proliferative and secretory stages.2 Eight samples of endometrium from fertile women in the secretory phase were detected.The samples were put into D^Hank's to experiment for cell culture.Stromal and epithelial cells were separated by enzymatic digestion method.The epithelial cells were cultured in vitro.and passaged to Corning culture dishes to grow for 24 hours. Then E2 and P were added into the cultured system for 24,48hours and in different concentrations of E^llb-estradiol,0,1O'9,10'8,10-7mol/ml),P(progesterone,0,10'10,1O'9,1 Q-'mol/ml) and RU486(10"*mol/L),The variations of the fluorescence on cells were detected with detector of the Flow cytometer .3 LSCM was used to examine the effect of CT on the changes of intracellular Ca2+ in the preimplantation embryos and uterine endometrial cells .Human embryos were thawed and incubated for 1 hour in culture medium containing fluo-3AM.Prior to use,each embryo was drawn through a micropiprtte with a bore measuring two-thirds of the embryo diameter to collapse the zip.The endometrial cells cultured in vitro were passaged into the Petri dish. After scanning 5 min,CT was added into the medium at different concentration(2,5,10nM).The fluorescence was evaluated at 30-second intervals.Result:1 CT was detected in the human endometrium in epithelial cells. CT protein located in the cytoplasm,and the expression of CT in human endometrium had periodical change during the menstrual cycle.The staining of CT protein was mainly shown in secretory phase endometrium with its peak in the implantation window stage.2 P significantly promoted the expression of CT in human endometrium epithelial cells.at the concentrations of 10"6mol/L,the fluorescence intensity increased after P treatment and reached the maxism 48 hours later.E2 holdbacked the expression of CT while it was at high concentration.3 CT increased the concentration of intracellular Ca2v in the stroma cells and preimplantation embryos ,but had no effect on the uterine endometrial epithelial cells.Conclusions:1 CT was synthesized and secreted in human endometrium.CT wouldplay a regulative role in the induction of human endometrial proliferation and differentiation ,and have an important function in the implantation of the human blastocyst.2 The expression of CT in the endometrial epithelial cells would be promoted by P in vitro and inhabited by E2 at high concentration.3 CT would have an important role in human embryos growth, development , implantation and promote endometrium decidual by inducing Ca2+ transient and increasing the concentrations of the intracellular Ca2+.
|
PDF Full Text Request