The Effects Of Low Dose Mifepristone On Population And Subsets Of NK Cells In Human Endometrium During Receptive Phase.

Background:Along with the development of recent life science, new drug contraceptive methods have emerge unceasingly. Mifepristone (RU486) is the first progesterone receptor antagonist for clinical use, and widely used in terminating early pregnancy recently. In 1990' s, both domestic and abroad researchs have reported that mifepristone is widespreadly used as an emergency contraception drug. The dose of mifepristone is gradually reducing. For the past few years, investigations show daily administration of low dose mifepristone can inhibit the development of endometrium to achieve the purpose of contraception,so called "endometrium contraception" . Low dose mifepristone is an effective contraceptional regimens with few side effects.In physiology of gestation, the endometrial receptivity to embryo is one criticality factor for estabilishing pregnancy. The uterus is ready to accept the implanting embryo only during a limited period of time known as the "window of implantation" (days 20-24 of an ideal cycle), outside of which the endometrium may be indifferent or even hostile to the embryo. During this time, the endometrium undergoes a complex temporal sequenceof distinct cells and molecules' changes that render it receptive to the conceptus, under the influence of a series of factors. Uterine NK cells are the most important immune cells in pregnant uterus. They play an important role in embryo implantation and placentation, involving cytokine production,the control of trophoblast invasion,spiral artery remodelling and to keep the immunologic balance between the feto-maternal interface. A lot of investigations showed that the disturbance of uterine NK cells during receptive phase may lead to implantation failure and recurrent miscarriage.Our aim was to determine the effect of low dose mifepristone on the subsets of NK cell' s in human endometrium during receptive phase, in order to investigate the immunologic mechanism of low dose mifepristone as an anti-implantation contraception drug.Methods And Materials:Endometrial biopsies were obtained from fourteen normally cycling IVF-ET patients for tubal resection or man infertility. The criteria of study subjects: the patients were reproductive age (aged 25-35 years) with mormal menstrual cycles (26-31days), and had no hormone or immunosuppressant therapy for the last 3 months. Endometrial biopsies were obtained at the "window of implantalion" (days 20-24 of an ideal cycle). Each endometrial tissue was chopped into 1mm³ pieces and divided into three equal parts, then assigned to three groups (control group,65nmol/L group and 200nmol/L group). The first part of the explant was treated with 2ml DMEM/F-12 containing 10^-9 mol/L estrin + 10^-7 mol/L progesterone. The other two parts of the endometriai explant was incubated with the same medium to which 65nmol/Land 200nmol/L mifepristone was added respectively. Incubations were performed in a humidified atmosphere at 37° C in 5% CO₂. Immunohistochemisty was used to analyze the location and numbers of CD56+ cells in each groups. The percentages of uNK cell subsets(CD3-CD56+CD16-%and CD3-CD56+CD16+ %)within the endometrial lymphocyte suspension were analysed by using flow cytometry.Results:1. HE-stained sections showed no significantly morphologic distinction between pre-cultured and post-cultured endometrial tissue.2. The quantitive variation of CD56+ cells in endometrium:CD56 staining was brown, localized on the cellular membrane. CD56+ cells was found in endometrial stroma of all three groups and in proximity of the gland and the small blood vessels.The mean number of CD56+ cells was 112. 67 + 7. 75 in control group, 145. 50 ±9. 43 in 65nmol/L group, 149. 33 + 4. 47 in 200nmol/L group. The numbers of CD56+ cells in two mifepristone-treated groups were significantly higher when compared to the control group(P<0.05). There was no significantly difference between the two mifepristone-treated groups(P>0. 05).3. The variation of uNK cell subsets in endometrium:The percentage of CD3-CD56+cells was (37. 79 + 6. 32) % in control group, (47. 50 + 7. 71)% in 65nmol/L group and (51. 41 ±7. 61) % in 200nmol/L group. The percentages of CD3-CD56+cells in two mifepristone-treated groups were significantly higher when compared to the control groups(P<0. 05). There was no significantly difference in percentage between the two mifepristone-treated groups(P>0. 05).The percentage of CD3-CD56+CD16-subset was (35. 19 + 6. 23) %in control group, (45.16± 7.33) %in 65nmol/L group and (48. 18±7. 25) in 200nmol/L group. The percentages of CD3-CD56+CD16-subset in two mifepristone—treated groups were significantly higher when compared to the control group (P<0. 05). There was no significantly difference in percentage between the two mifepristone-treated groups (P>0.05).The percentage of CD3-CD56+CD16+ subset was (2. 60±0. 76) % in control group, (2. 34 + 0. 67) % in 65nmol/L group and (3. 24 + 0. 84) % in 200nmol/L group. There was no significantly difference in percentage among the three groups(P=0.39). Conclusion:1. Low dose mifepristone increased the number of CD56+NK cells in endometrium during receptive phase. This suggested that low dose mifepristone promote the excessive proliferation of uNK cells, which might result in the disturbance of human endometrial immuno- microenvironment during receptive phase.2. Low dose mifepristone increased the percentage of CD3-CD56+NK cells and CD3-CD56+CD16-NK subset in endometrium during receptive phase, and had no significant effect on the CD3-CD56+CD16+ subset. This suggested that low dose mifepristone promote the excessive proliferation of CD56+CD16-uNK subset, which might result in the disturbance of human endometrial immunomicroenvironment during receptive phase and lead to implantation failure.