摘要
目的探讨子宫内膜来源间充质干细胞(endometrium-derived mesenchymal stem cells,EMSCs)对子宫内膜癌HEC-1B细胞上皮-间质转化的影响及对Notch1信号通路的作用。方法子宫肌瘤患者12例和子宫内膜腺癌10例,分别取其正常子宫内膜组织和子宫内膜腺癌组织,分离培养正常EMSCs和子宫内膜腺癌来源MSCs(endometrial adenocarcinoma derived mesenchymal stem cells,ECMSCs),应用流式细胞仪检测不同EMSCs表面抗原表达情况进行表型鉴定,浓缩EMSCs和ECMSCs培养上清。取对数生长期子宫内膜癌HEC-1B细胞,随机分为对照组(采用DMEM培养基培养)、EMSCs组(采用DMEM培养基+EMSCs浓缩上清培养)和ECMSCs组(采用DMEM培养基+ECMSCs浓缩上清培养)。培养48 h后,观察细胞形态和生长情况,采用划痕实验检测细胞迁移率,采用Transwell小室实验检测侵袭细胞数,采用Western blot法检测细胞Notch1、Jagged1、发状分裂相关增强子1(hairy and enhancer of split homolog-1,Hes1)、E-钙黏蛋白(E-cadherin)、波形蛋白(vimentin)相对表达量。结果获得EMSCs和ECMSCs,形状呈长梭形;均表达CD73、CD105、CD106,均不表达或低表达CD34、CD45,符合充质干细胞特征;3组HEC-1B细胞均生长良好,形态未发生明显变化,对照组、EMSCs组、ECMSCs组细胞密度依次增大;ECMSCs组细胞迁移率[(76.84±7.58)%]和侵袭细胞数[(146.70±14.57)个]大于EMSCs组[(52.26±6.35)%、(96.50±12.42)个]和对照组[(21.23±5.51)%、(61.20±9.25)个](P<0.05),EMSCs组大于对照组(P<0.05);ECMSCs组细胞E-cadherin蛋白相对表达量(0.32±0.06)低于对照组(1.12±0.11)和EMSCs组(0.71±0.08)(P<0.05),EMSCs组低于对照组(P<0.05)。ECMSCs组细胞vimentin(1.03±0.10)、Notch1(1.15±0.11)、Jagged1(0.97±0.09)、Hes1(0.83±0.08)蛋白相对表达量高于对照组(0.23±0.06、0.31±0.06、0.27±0.05、0.18±0.04)和EMSCs组(0.65±0.08、0.83±0.09、0.78±0.09、0.57±0.05)(P<0.05),EMSCs组高于对照组(P<0.05)。结论EMSCs和ECMSCs均可诱导子宫内膜癌HEC-1B细胞发生上皮-间质转化,提高细胞生长、迁移和侵袭能力,可能与激活Notch1通路有关。
Objective To investigate the influence of endometrium-derived mesenchymal stem cells(EMSCs)on the epithelial-mesenchymal transition of endometrial carcinoma HEC-1 B cells and the role in Notch1 signaling pathway.Methods The normal endometrial tissue and endometrial adenocarcinoma tissue were taken from 12 cases of uterine fibroids and 10 cases of endometrial adenocarcinoma,respectively.The EMSCs and endometrial adenocarcinoma-derived mesenchymal stem cells(ECMSCs)were isolated and cultured.Flow cytometry was used to detect the surface antigen expression of mesenchymal stem cells from different endometrial sources for phenotypic identification.The EMSCs and ECMSCs were concentrated for culture supernatant.HEC-1 B cells in logarithmic growth phase were randomly divided into control group(cultured with DMEM medium),EMSCs group(cultured with DMEM medium+EMSCs concentrated supernatant)and ECMSCs group(cultured with DMEM medium+ECMSCs concentrated supernatant).After 48 h,the cell morphology and growth were observed.The scratch test was used to detect the cell migration rate,Transwell chamber test was used to detect the number of invaded cells,and Western blot was used to detect the relative expressions of Notch1,Jagged1,Hes1,E-cadherin and vimentin proteins.Results EMSCs and ECMSCs were obtained,in a shape of long spindle.CD73,CD105 and CD106 were expressed,and CD34 and CD45 were not expressed or lowly expressed in EMSCs and ECMSCs,which were in line with the characteristics of mesenchymal stem cells.The HEC-1 B cells in three groups grew well with no significant changes in morphology.The cell density increased gradually in turn in control group,EMSCs group and ECMSCs group.The cell migration rate and the number of invaded cells decreased gradually in turn in ECMSCs group((76.84±7.58)%,146.70±14.57),EMSCs group((52.26±6.35)%,96.50±12.42)and control group((21.23±5.51)%,61.20±9.25)(P<0.05).The relative expression of E-cadherin protein increased gradually in turn in ECMSCs group(0.32±0.06),EMSCs group(0.71±0.08)and control group(1.21±0.11)(P<0.05).The relative expressions of vimentin,Notch1,Jagged1 and Hes1 were higher in ECMSCs group(1.03±0.10,1.15±0.11,0.97±0.09,0.83±0.08)than those in control group(0.23±0.06,0.31±0.06,0.27±0.05,0.18±0.04)and EMSCs group(0.65±0.08,0.83±0.09,0.78±0.09,0.57±0.05)(P<0.05),and higher in EMSCs group than those in control group(P<0.05).Conclusion Both EMSCs and ECMSCs can induce the epithelialmesenchymal transition of endometrial carcinoma HEC-1 Bcells and increase the cell growth,migration and invasion ability,which may be correlated with the activation of the Notch1 pathway.
作者
牛丽佳
夏义欣
宋琪
张晓莉
蔡海瑜
NIU Lijia;XIA Yixin;SONG Qi;ZHANG Xiaoli;CAI Haiyu(Department of Obstetrics and Gynecology,the Third Medical Center of PLA General Hospital,Beijing 100039,China;Department of Gynecology,Zhengzhou Maternal and Child Health Care Hospital,Meternity Hospital Affiliated to Henan University,Zhengzhou 450012,China)
出处
《中华实用诊断与治疗杂志》
2020年第12期1195-1200,共6页
Journal of Chinese Practical Diagnosis and Therapy
基金
河南省医学科技攻关计划项目(201602346)。
关键词
子宫内膜癌
子宫内膜来源间充质干细胞
上皮-间质转化
迁移
侵袭
endometrial carcinoma
endometrium-derived mesenchymal stem cells
epithelial-mesenchymal transition
migration
invasion
