摘要
背景:研究表明,过氧化物酶体增殖物激活受体γ共激活因子1β(peroxisome proliferator-activated receptor gamma coactivator 1beta,PGC-1β)通过PI3K/AKT/mTOR信号通路影响乳腺癌细胞的增殖、迁移及凋亡。而PGC-1β能否通过PI3K/AKT/mTOR信号通路影响乳腺癌干细胞的干性及其影响机制还不清楚。目的:探究PGC-1β对乳腺癌干细胞干性的影响及调控机制。方法:将PGC-1β空载体、过表达载体(Lv-PGC-1β)、干扰载体(Sh-PGC-1β)分别转染乳腺肿瘤MCF-7细胞,荧光显微镜观察转染效果;qRTPCR及Western blot验证慢病毒转染后PGC-1βmRNA和蛋白的相对表达。在干性培养基中分别培养未转染组、PGC-1β空载体组、PGC-1β过表达组及PGC-1β干扰组的MCF-7细胞使其成为乳腺癌干细胞,显微镜下观察并记录干细胞球体的形成过程,Western blot验证干性标志物(ABCG2、ALDH1、OCT4)、上皮-间充质转化标记物(E-Cadherin、N-Cadherin、Vimentin、Slug和ZEB1)及PI3K/AKT/mTOR通路核心蛋白的相对表达。结果与结论:①MCF-7贴壁细胞经干性培养基培养形成了折光性好、中间密度高的致密球干细胞;②乳腺癌干细胞干性蛋白的表达明显高于其贴壁细胞(P<0.01);③与未转染组、空载体组相比,PGC-1β过表达组乳腺癌干细胞的形成时间缩短,细胞球体数目增多、球体直径增大(P<0.01),而PGC-1β干扰组细胞球体数目减少、球体直径减小(P<0.01);④PGC-1β过表达组干性相关蛋白的表达高于未转染组、空载体组(P<0.01),上皮-间充质转化相关蛋白E-Cadherin表达低于未转染组、空载体组(P<0.01),而N-Cadherin、Vimentin、Slug和ZEB1的表达高于未转染组、空载体组(P<0.01);PI3K/AKT/mTOR相关蛋白及其磷酸化相关蛋白表达高于未转染组、空载体组(P<0.01),PGC-1β干扰组结果则与PGC-1β过表达组相反;⑤结果提示,PGC-1β能够通过激活PI3K/AKT/mTOR信号通路影响乳腺癌干细胞上皮-间充质转化,从而促进乳腺癌�
BACKGROUND:Previous studies have shown that peroxisome proliferator-activated receptor gamma coactivator 1 beta(PGC-1β)affected the proliferation,migration and apoptosis of breast cancer cells through PI3K/AKT/mTOR signaling pathway.However,whether PGC-1βcan affect the stemness of breast cancer stem cells through PI3K/AKT/mTOR signaling pathway and its influencing mechanism has not been elucidated.OBJECTIVE:To investigate the effect and regulatory mechanism of PGC-1βon stemness of breast cancer stem cells.METHODS:MCF-7 cells were transfected with PGC-1βempty vector,PGC-1βoverexpression vector(Lv-PGC-1β)and interference vector(Sh-PGC-1β),respectively.The transfection effect was observed under fluorescence microscope.The relative expression of PGC-1βmRNA and protein after lentivirus transfection was verified by qRT-PCR and western blot assay.The cells of non-transfection group,PGC-1βempty vector group,PGC-1βoverexpression vector group and PGC-1βinterference vector group were cultured with stemness medium to foster breast cancer stem cells.The formation process of the spheres was observed and recorded under the microscope.Western blot assay was used to verify the relative expression of stemness proteins(ABCG2,ALDH1 and OCT-4),epithelial-mesenchymal transition proteins(E-cadherin,N-cadherin,Vimentin,Slug and ZEB1)and PI3K/AKT/mTOR pathway related proteins.RESULTS AND CONCLUSION:(1)MCF-7 adherent cells were cultured in stemness medium to form dense sphere stem cells with good refractive index and high intermediate density.(2)The expression of stemness protein in breast cancer stem cells was significantly higher than that in adherent cells(P<0.01).(3)Compared with the non-transfection and PGC-1βempty vector groups,the formation time of breast cancer stem cells was shorter,and the number and diameter of spheres were increased in the PGC-1βoverexpression vector group(P<0.01);however,the number of spheroids and the diameter of spheroids decreased in the PGC-1βinterference vector group(P<0.01).(4)The expressi
作者
赵菊芬
马荣
曹佳
于传扬
陶翔
王嘉
王立斌
Zhao Jufen;Ma Rong;Cao Jia;Yu Chuanyang;Tao Xiang;Wang Jia;Wang Libin(Clinical Medicine College,Ningxia Medical University,Yinchuan 750004,Ningxia Hui Autonomous Region,China;National Biochip Research Center Sub-Center in Ningxia,General Hospital of Ningxia Medical University,Yinchuan 750004,Ningxia Hui Autonomous Region,China)
出处
《中国组织工程研究》
CAS
北大核心
2023年第1期59-65,共7页
Chinese Journal of Tissue Engineering Research
基金
国家自然科学基金(81860470),项目负责人:王立斌
宁夏高层次科技创新领军人才项目(KJT2019003),项目负责人:王立斌
宁夏重点研发计划对外科技合作专项项目(2019BFH02012):项目负责人:王立斌。
