【摘要】 目的 明确组织选择性雌激素复合物(TSEC)对子宫平滑肌瘤细胞增殖及凋亡能力的影响，为TSEC作为绝经激素治疗(MHT)的新方案及其对围绝经期子宫平滑肌瘤患者治疗的安全性和有效性提供参考。方法 选取2021年6月至8月在西安交通大学第一附属医院因子宫平滑肌瘤行核除术的患者，对手术切除的子宫平滑肌瘤组织通过酶消化法进行原代培养，提取子宫平滑肌瘤细胞，经苏木素-伊红(HE)染色、α-肌动蛋白(α-Actin)染色、雌激素受体(ER)染色进行细胞验证，以不同药物对子宫平滑肌瘤细胞进行刺激干预，设立雌孕激素连续联合组(17-β雌二醇+安宫黄体酮，E+P组)、TSEC组(17-β雌二醇+雷洛昔芬)、单独雌激素组(17-β雌二醇，E组)和对照组(无药物刺激干预)。采用四甲基偶氮唑盐比色法(MTT)检测细胞增殖能力的变化，检测细胞周期及细胞凋亡的变化；通过免疫印迹法检测增殖及凋亡相关蛋白PCNA、Bcl-2的表达，并进行比较分析。结果 MTT实验显示，TSEC组细胞增殖较对照组无差异，E+P组、E组促进了细胞增殖(24h时间点吸光度值比较：TSEC组vs.对照组，t=-1.984,P=0.071;E+P组vs.对照组，t=-30.113,P<0.001;E组vs.对照组，t=-16.148,P<0.001。48h时间点吸光度值比较：TSEC组vs.对照组，t=-1.997,P=0.069;E+P组vs.对照组，t=-23.553,P<0.001;E组vs.对照组，t=-16.432,P<0.001)。免疫印迹法检测显示，TSEC组PCNA蛋白表达水平与对照组无明显差异，E+P组、E组则增加了PCNA蛋白的表达水平(TSEC组vs.对照组，t=-2.449,P=0.627;E+P组vs.对照组，t=-23.267,P<0.001;E组vs.对照组，t=-13.472,P<0.001)。细胞周期检测显示，与对照组相比，E+P组、E组细胞G1/S时相转换减弱，TSEC组对其无明显影响。TSEC组Bcl-2蛋白表达水平与对照组无明显差异，E+P组、E组则增加了Bcl-2蛋白的表达水平(TSEC组vs.对照组，t=1.225,P=0.289;E+P组vs.对照组，t=-66.136,P<0.001;E组vs.对照组，t=-50.215,P<0.001)。细胞凋亡实验显示，与对照组相比，E+P组、E组细胞凋亡减少，而TSEC组对细胞凋亡无明显影响。结论 TSEC对子宫平滑肌瘤细胞的增殖及凋亡无明显影响，与雌孕激素连续联合等MHT方案相比，TSEC在围绝经期子宫平滑肌瘤患者的治疗中可能具有更好的安全性。更多还原

【Abstract】 Objective To clarify the proliferation and apoptosis of uterine leiomyoma cells under the impact of tissue selective estrogen complex(TSEC),and provide references for the new program of TSEC in menopausal hormone therapy(MHT) and its safety and efficacy in the treatment of perimenopause patients with uterine leiomyoma.Methods Patients who conducted nuclear excision for uterine leiomyoma in the First Affiliated Hospital of Xi′an Jiaotong University from June to August 2021 were selected.The surgically excised uterine leiomyoma tissue was primary cultured by enzymatic digestion, and the uterine leiomyoma cells were extracted.Hematoxylin-eosin(HE) staining, α-Actin staining, estrogen receptor(ER) staining were used for cell verification.Different drugs were used to stimulate and interfere with the uterine leiomyoma cells.The continuous combination of estrogen and progesterone group(17-β estradiol+medroxyprogesterone acetate, E+P group),the TSEC group(17-β estradiol+raloxifene),the estradiol alone group(17-β estradiol, E group),and the control group(without the stimulation and intervention of drug) were set.The changes of cell proliferation function, cell cycle and cell apoptosis were observed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide(MTT).Western-blot was done to detect the protein expression of PCNA and Bcl-2 related to proliferation and apoptosis.They were compared and analyzed.Results MTT showed that there was no difference of cell proliferation between the TSEC group and the control group, and the E+P group and the E group promoted cell proliferation(Comparison of absorbance values at 24 h time point: the TSEC group vs. the control group, t=-1.984,P=0.071;the E+P group vs. the control group, t=-30.114,P<0.001;the E group vs. the control group, t=-16.148,P<0.001.Comparison of absorbance values at 48 h time point: the TSEC group vs. the control group, t=-1.997,P=0.069;the E+P group vs. the control group, t=-23.553,P<0.001;the E group vs. the control group, t=-16.432,P<0.001).Western-blot showed there was no significant difference of the expression level of PCNA protein between the TSEC group and the control group, and the E+P group and the E group increased the expression level of PCNA(the TSEC group vs.the control group, t=-2.449,P=0.627; the E+P group vs. the control group, t=-23.267,P<0.001;the E group vs. the control group, t=-13.472,P<0.001).The detection of cell cycle showed that compared with the control group, cellular G1/S phase transition was attenuated in the E+P group and the E group, but had no significant effect in the TSEC group.There was no significant difference of the expression level of Bcl-2 protein between the TSEC group and the control group, and the E+P group and the E group increased the expression level of Bcl-2(the TSEC group vs. the control group, t=1.225,P=0.289;the E+P group vs. the control group, t=-66.136,P<0.001;the E group vs. the control group, t=-50.215,P<0.001).Cell apoptosis experiment showed that compared with the control group, cell apoptosis decreased in the E+P group and the E group, but had no significant effect cell apoptosis on in the TSEC group.Conclusion TSEC shows no effects on proliferation and apoptosis on uterine leiomyoma cells.TSEC may have better safety than the MHT regimens such as continuous combination of estrogen and progesterone in the treatment of perimenopause patients with uterine leiomyoma.更多还原