摘要
目的观察雌二醇对子宫腺肌病患者子宫内膜-肌层交界区（EMI）平滑肌细胞游离Ca^2＋浓度的影响，并探索其作用模式。方法选择2011年3月至10月在首都医科大学附属北京妇产医院因子宫腺肌病行子宫全切除术的患者16例，其中子宫内膜增殖期9例、分泌期7例。取其EMI平滑肌组织进行原代细胞培养，以Ca^2＋荧光探针负载平滑肌细胞，用激光共聚焦显微镜观察1×10^2、1×10^3、1×10^4、1×10^5pmol／L浓度的雌二醇作用后平滑肌细胞内Ca^2＋浓度（以Ca^2＋荧光强度表示）的变化，选择最佳浓度雌二醇。以最佳浓度雌二醇分别联合17β雌二醇-牛血清白蛋白复合物（17β-E2-BSA）、ER拮抗剂——氟维司群（ICI182780）、蛋白合成抑制剂——放线菌酮（CHX）、G蛋白活化抑制剂——百日咳毒素（PTX）处理者分别为17β-E2-BSA组、ICI182780组、CHX组和PTX组，以仅用最佳浓度雌二醇处理者为上述各组的相应对照组，观察雌二醇作用后增殖期、分泌期EMI平滑肌细胞以及各组EMI平滑肌细胞内Ca^2＋荧光强度的变化。结果（1）EMI平滑肌细胞培养24h后即贴壁生长，状态良好。（2）1×10^2～1×10^5pmoL／L浓度的雌二醇均能在1min内引起细胞内Ca^2＋荧光强度迅速升高，分别为275±16、449±18、615±36、641±47，其中1×10^4pmo]／L与1×10^5pmol／L浓度的雌二醇作用下细胞中Ca^2＋荧光强度增幅最明显，但两者比较，差异无统计学意义（P〉0．05），因此选择1×10^4pmol／L为最佳雌二醇浓度。（3）1×10^4pmol／L浓度雌二醇作用于增殖期与分泌期EMI平滑肌细胞，两者Ca^2＋荧光强度比较，差异无统计学意义（P〉0．05）；17β-E2-BSA组与CHX组细胞内Ca^2＋荧光强度分别为646±32和602±31，与各自相应对照组（分别为513±26和617±35）比较，差异均无统计学意义（P〉0．05）；而PTX组与ICI182780组分别为188±20和302±11，与各自相应对照组（分别为632±33和635±24）比较，差异均有统计学意义（P〈0．01）。结论雌二醇对子宫腺肌病患者EMI平滑肌细胞内游离Ca^2＋浓度的调节符合膜受体介导的非基因转录作用模式。
Objective To investigate the effect and mechanism of estrodial （E2 ） on intracellular free calcium in the endometrial-myometrial interface （EMI） smooth muscle cells from uteri with adenomyosis. Methods From March 2011 to October 2011, 16 uterus specimens were collected from patients with adenomyosis undergoing hysterectomy in Beijing Obstetrics and Gynecology Hospital, which included 9 proliferative endometrium and 7 secretory endometrium. EMI smooth muscle cells from the uterus were cultured and loaded with calcium ion （ Ca^2＋ ） fluorescent probe fluo-4/AM. The labeled cells were stimulated with the various concentration of E2 （ 1 ×10^2, 1×10^3 , 1 ×10^4, 1×10^5 pmol/L, respectively） ,then the changes of intracellular Ca^2＋ fluorescence intensity were measured by laser scanning microscopy. The most suitable concentration of E2 was selected, and the reaction difference between the EMI smooth muscle ceils of two menstrual phases were also investigated ; The changes of intracellular Ca^2 ＋ fluorescence intensity were detected proliferative and secretory smooth muscle cells in E2 conjugated to bovine serum albumin （17β-E2-BSA） group, cycloheximide （CHX） group, fulvestrant （ICI182780） group and pertussis toxin （PTX） group. Results （ 1 ） The cell viability of primary cultured EMI smooth muscle cells was well at 24 hours culture. （2） 1×10^2 - 1 ×10^5 pmol/L E2 can rapidly increase the intracellular Ca^2＋ fluorescence intensity within 1 rain （P 〈0. 01 ） ;The increased amplitudes caused by 1 ×10^4 pmol/L and 1×10^5 pmol/L E2 were the most significant,but there was no significant difference between them （P 〉0. 05）. 1 ×10^4 pmol/L was the most suitable concentration. （3）With the 1 ×10^4 pmol/L E2, the Ca^2＋ fluorescence intensity changes showed no significant difference between the EMI smooth muscle cells from the proliferative phase and secretory phase uterus （P 〉0. 05）. The Ca^2＋ fluorescence intensity changes were 646 ± 32 in 17β-E2-BSA group and 602±31 in CHX group, when compared with 513±26 and 617±35 in respective control group, no significant difference was observed （P 〉 0. 05 ）. The increased amplitude of 188 ± 20 in the PTX group and 302 ± 11 in ICI182780 group exhibited significant difference with 632 ±33 and 635±24 in respective control group （ P 〈 0. 01 ）. Conclusion E2 could increase the intracellular Ca^2 ＋ of EMI through a membrane receptor dependent and nongenomic mechanism of action.
出处
《中华妇产科杂志》
CAS
CSCD
北大核心
2012年第5期351-354,共4页
Chinese Journal of Obstetrics and Gynecology
基金
国家自然科学基金（30872744）
