内源性大麻素系列对不明原因性不孕患者种植窗期子宫内膜的影响及其作用机制

发布时间：2018-09-04 07:14

【摘要】：目的:观察不明原因性不孕(unexplained infertility,UI)患者种植窗期子宫内膜中内源性大麻素系统(endocannabinoid system,ECS)的表达情况,并研究内源性大麻素物质花生四烯酸乙醇胺(anandamide,AEA)对子宫内膜中白血病抑制因子(leukemia inhibitory factor,LIF)水平的影响,从而探讨内源性大麻素系统影响不明原因不孕患者种植窗期子宫内膜的作用机制。方法:前瞻性研究2014年3月至2014年12月间就诊于河北医科大学第二医院生殖中心接受IVF助孕符合以下标准的患者资料28例,作为UI组:年龄38岁;月经周期25-35d,可自发排卵;腹腔镜或子宫输卵管造影提示双侧输卵管通畅,无盆腔粘连;行宫腔内人工授精(IUI)≥3周期未孕;排除子宫性、内分泌性、免疫性及男方因素所致的不孕,临床无子宫内膜异位症的症状;近3个月未使用过性激素类药物。选取同期因男方因素或输卵管阻塞(除外输卵管积水)因素预于本院行体外受精-胚胎移植(IVF-ET)/卵胞浆内单精子注射(ICSI)助孕符合下述标准患者资料32例作为对照组:年龄38岁;有妊娠史;月经规律,周期25-35d;有自发排卵;基础内分泌正常;3月内未用激素药物;并排除合并多囊卵巢综合征、子宫内膜异位症、子宫肌瘤、子宫内膜息肉等疾病。两组患者分别于IVF/ICSI助孕前一月经周期行阴道B超监测排卵,确定排卵日。研究两组实验对象排卵后7-9天血清孕酮水平、子宫内膜厚度及子宫内膜组织形态和LIF、内源性大麻素1型受体(cannabinoid type 1,CB1)、脂肪酸酰胺水解酶(fatty acid amide hydrolase,FAAH)表达情况,培养人子宫内膜腺癌细胞RL95-2,分别于培养基中添加AEA、AM251(CB1受体拮抗剂)、AM630(CB2受体拮抗剂)后,研究细胞上清液中LIF水平的变化情况。分析内源性大麻素对种植窗期子宫内膜的影响,推测其作用机制。结果:两组实验对象在体重指数、年龄、不孕年限、基础窦泡数、基础FSH、LH、E2值等基本资料上没有统计学差异。Gn天数、Gn总量、HCG日直径≥16mm卵泡数、获卵数、受精率、卵裂率两组间没有统计学差异。但胚胎种植率(21.74%vs.56.25%)与可移植胚胎率(43.79%vs.67.38%)两组间有统计学差异(P0.05),UI组胚胎种植率及可移植胚胎率低于对照组;UI组新鲜周期临床妊娠率低于对照组(33.33%vs.78.57%),其差异有统计学意义(P0.05)。UI组与对照组黄体中期血清P水平、HCG日血清E2水平、HCG日血清P水平相比较,两组间差异没有统计学意义。两组研究对象间子宫内膜厚度在黄体中期(10.43±1.34mmvs.11.87±1.36mm)及移植日(11.14±1.03mm vs.12.00±1.25mm)的差异有统计学意义(P0.05),UI组在两时期子宫内膜厚度均低于对照组。两组黄体中期子宫内膜中CB1表达(IOD值)相比,UI组略高于对照组(126.86±19.03 vs.111.68±28.21),但两组间差异无统计学意义。两组黄体中期子宫内膜FAAH(101.58±28.78 vs.128.14±24.88)及LIF(120.32±19.61 vs.136.53±18.97)表达比较中,UI组IOD值均低于对照组,两组间有统计学差异(P0.05)。体外培养子宫内膜细胞RL95-2经24h、48h、72h分别于培养基中添加1μmol/L、5μmol/L、10μmol/L、50μmol/LAEA的各组及空白对照组中LIF水平有差异,且差异有统计学意义(F=605.381,P0.05)。其中5μmol/L48h组与10μmol/L48h组相比差异无统计学意义;1μmol/L72h组与10μmol/L72h组相比差异无统计学意义;其余各组间差异均有统计学意义(P0.05)。在24h及48h组中,随着AEA浓度的升高,LIF水平随之升高。在72h组中,5μmol/LAEA组LIF水平高于1μmol/LAEA组(2003.87±37.85 ng/L vs.1948.20±21.81ng/L),而10μmol/LAEA组LIF水平低于5μmol/LAEA组(1916.60±14.05ng/L vs.2003.87±37.85ng/L),而50μmol/LAEA组LIF水平低于上述三种AEA浓度组,且差异均有统计学意义(P0.05)。另于培养基中添加同浓度(5μmol/L)AM251、AM630后各组间LIF水平差异有统计学意义(F=930.121,P0.05)。其中AM251组与AM630组相比无统计学差异;AEA+AM251组与AEA+AM630组相比无统计学差异。同AEA组相比,AEA+AM251组与AEA+AM630组LIF水平均降低,差异有统计学意义(P0.05);AM251组与AEA+AM251组相比(1358.04±18.42ng/L vs.1510.37±16.77ng/L),LIF水平降低,有统计学差异(P0.05);AM630组与AEA+AM630组相比(1353.73±20.64ng/L vs.1508.20±21.50ng/L),LIF水平降低,差异有统计学意义(P0.05)。但各组同空白对照组相比,LIF水平均高于空白对照组,差异有统计学意义(P0.05)。结论:不明原因不孕患者种植窗期子宫内膜中FAAH、LIF表达均低于继发不孕者,而其CB1受体表达有升高趋势,且种植率、新鲜周期妊娠率低于对照组。体外培养子宫内膜细胞RL95-2模拟子宫内膜种植窗期内环境,LIF水平随着AEA浓度或作用时间的递增而升高,但过高浓度的AEA或作用时间过长,LIF水平反而降低,添加CB1及CB2受体拮抗剂后LIF水平均有降低。表明一定浓度的AEA对种植有积极作用,而高剂量的AEA对其产生不利影响,CB1、CB2受体可能参与其调节。内源性大麻素系统的表达异常可能影响不明原因不孕妇女子宫内膜容受性,降低其妊娠率。
[Abstract]:Objective: To investigate the expression of endocannabinoid system (ECS) in endometrium of unexplained infertility (UI) patients during implantation window and the effect of endocannabinoid ethanolamine (AEA) on leukemia inhibitory fac in endometrium. METHODS: A prospective study was conducted to investigate the effects of the endocannabinoid system on the endometrium of unexplained infertility patients during implantation window period. Age 38 years; menstrual cycle 25-35 days, spontaneous ovulation; laparoscopic or hysterosalpingography suggests bilateral tubal patency, no pelvic adhesion; intrauterine insemination (IUI) > 3 cycles of infertility; exclusion of uterine, endocrine, immune and male factors caused by infertility, clinical symptoms of endometriosis; nearly three months not used Thirty-two cases of in vitro fertilization-embryo transfer (IVF-ET) / intracytoplasmic sperm injection (ICSI) pregnant in our hospital due to male factors or oviduct obstruction (except hydrosalpinx) were selected as the control group: age 38 years; pregnancy history; menstrual regularity, 25-35 days; spontaneous ovulation Ovulation was monitored by transvaginal B-ultrasound before IVF/ICSI, and ovulation days were determined. Serum progesterone levels were measured 7-9 days after ovulation in the two groups. Level, endometrial thickness, endometrial tissue morphology and LIF, endogenous cannabinoid type 1 receptor (CB1), fatty acid amide hydrolase (FAAH) expression, cultured human endometrial adenocarcinoma cell line RL95-2, added AEA, AM251 (CB1 receptor antagonist), AM630 (CB2 receptor antagonist) to the medium, respectively. Results: There were no significant differences in body mass index, age, infertility, number of basal sinus vesicles, basal FSH, LH, E2 values between the two groups. There was no significant difference between the two groups in the number of follicles (> 16mm) per day, the number of eggs retrieved, the fertilization rate and the cleavage rate, but there was significant difference between the embryo implantation rate (21.74% vs. 56.25%) and the transplantable embryo rate (43.79% vs. 67.38%) between the two groups (P 0.05). There was no significant difference between the UI group and the control group in the serum P level in the middle luteal phase, E2 level on the day of HCG and P level on the day of HCG. Compared with the control group, the expression of CB1 in the middle luteal endometrium (IOD value) in the UI group was slightly higher than that in the control group (126.86 + 19.03 vs. 111.68 + 28.21), but there was no significant difference between the two groups. Compared with the control group, the IOD value of UI group was lower than that of the control group (P There was no significant difference between 5-micromol/L 48h group and 10-micromol/L 48h group; there was no significant difference between 1-micromol/L 72h group and 10-micromol/L 72h group; there was significant difference between the other groups (P 0.05). In 24-hour and 48-hour groups, the level of LIF increased with the increase of AEA concentration. In 72 h group, the level of LIF in 5-micromol/LAEA group was higher than that in 1-micromol/LAEA group (2003.87+37.85 ng/L vs.1948.20+21.81 ng/L), while that in 10-micromol/LAEA group was lower than that in 5-micromol/LAEA group (1916.60+14.05 ng/L vs.2003.87+37.85 ng/L), and that in 50-micromol/LAEA group was lower than that in the above three AEA groups (P 0.05). There was no significant difference between AM251 group and AM630 group (P 0.05). There was no significant difference between AM251 group and AM630 group. There was no significant difference between AEA + AM251 group and AEA + AM630 group. Compared with AEA+AM 251 group (1358.04+18.42 ng/L vs.1510.37+16.77 ng/L), the level of LIF in AM 251 group was significantly lower than that in AEA+AM 630 group (1353.73+20.64 ng/L vs.1508.20+21.50 ng/L), but the level of LIF in each group was higher than that in blank control group (P 0.05). CONCLUSION: The expression of FAAH and LIF in endometrium of unexplained infertility patients during implantation window is lower than that of secondary infertility patients, and the expression of CB1 receptor is increasing, and the implantation rate and the pregnancy rate in fresh cycle are lower than that of control group. The level of LIF increased with the increase of AEA concentration or acting time, but decreased with the increase of AEA concentration or acting time, and decreased with the addition of CB1 and CB2 receptor antagonist. Regulation. Abnormal expression of endocannabinoid system may affect endometrial receptivity and reduce pregnancy rate in women with unexplained infertility.
【学位授予单位】：河北医科大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R711.6

【相似文献】