阿霉素诱导MCF10A上皮细胞衰老模型的构建  

An Epithelial Senescence Model Induced by Doxorubicin in MCF 10A Cells

作　　者：王增盛 牛祖彪 张波 郝佳慧 朱一超 杨瑞刚 任禾 刘辰瑜 孙强[2] 任立成 WANG Zeng-Sheng;NIU Zu-Biao;ZHANG Bo;HAO Jia-Hui;ZHU Yi-Chao;YANG Rui-Gang;REN He;LIU Chen-Yu;SUN Qiang;REN Li-Cheng(Key Laboratory of Tropical Translational Medicine of Ministry of Education,Department of Biology,School of Basic Medicine and Life Sciences,Hainan Medical University,Haikou 571199,China;Laboratory of Advanced Biotechnology,Academy of Military Medical Sciences,Beijing 100071,China;Department of Oncology,Beijing Shijitan Hospital of Capital Medical University,Beijing 100038,China)

机构地区：[1]海南医科大学基础医学与生命科学学院热带转化医学教育部重点实验室,生物学教研室,海口571199 [2]军事科学院军事医学研究院前沿生物技术实验室,北京100071 [3]首都医科大学附属北京世纪坛医院肿瘤治疗性疫苗北京市重点实验室,北京100038

出　　处：《中国生物化学与分子生物学报》2025年第1期147-155,共9页Chinese Journal of Biochemistry and Molecular Biology

基　　金：国家重点研发计划(No.2022YFC3600100)资助。

摘　　要：本研究旨在构建稳定的上皮细胞衰老模型,以期用于衰老细胞清除剂(senolytics)药物的筛选和评价研究。探究了阿霉素(doxorubicin)诱导人非转化乳腺上皮细胞系MCF 10A衰老的最佳条件,包括最佳诱导浓度、最佳干预时长和最佳衰老天数,并多维度验证了MCF 10A作为细胞衰老模型的可行性。阿霉素诱导MCF 10A细胞衰老的最佳条件是:用0.6μmol/L的阿霉素处理16 h在第8 d MCF 10A细胞达到最佳衰老状态。在最佳诱导条件下,加药组的衰老相关-β-半乳糖苷酶(senescence-associated β-galactosidase,SA-β-gal)染色阳性率达到97%;同时,检测mRNA、蛋白质和免疫荧光(immunofluorescence,IF)的表达,结果显示:加药组细胞相对于正常细胞的衰老相关分泌表型(senescence-associated aecretory phenotype,SASP)、p16、p21和p53蛋白质的表达量均显著升高,核纤层蛋白B1(lamin B1)显著下降(P<0.001)。加药组细胞与衰老细胞特有特征一致。本研究用阿霉素成功构建了MCF 10A上皮细胞衰老模型,为基于衰老上皮细胞的衰老细胞清除剂的筛选奠定了研究基础。This research aims to construct a stable epithelial cell senescence model for screening and evaluation of senolytics.We explored the optimal conditions for doxorubicin-induced senescence of non-transformed epithelial cells MCF 10A,including the optimal induction concentration,the optimal intervention time,and the optimal senescence duration,and confirmed the feasibility of MCF 10A as an epithelial senescence model by multiple ways.The optimal condition for Doxorubicin-induced senescence of MCF 10A cells was treatment with 0.6μmol/L Doxorubicin for 16 h to achieve the best senescence state on the 8th day.Under the optimal induction conditions,the positive rate of senescence-associated β-galactosidase(SA-β-gal)staining in the treated group reached 97%.At the same time,biochemical results of detecting the expression of mRNA,proteins,and immunofluorescence demonstrated that the expression levels of senescence-associated secretory phenotype(SASP),p16,p21 and p53 in the treated group were significantly higher than those in the control cells,and Lamin B1 was significantly decreased(P<0.001),which were consistent with the specific characteristics of senescence.In summary,an epithelial senescence model was successfully induced in MCF 10A cells by Doxorubicin in this study,which will promote the screening of senolytics for senescent epithelial cells.

关 键 词：上皮细胞衰老模型 阿霉素 人非转化乳腺上皮细胞系MCF 10A 衰老相关-β-半乳糖苷酶 衰老相关分泌表型 

分 类 号：Q71[生物学—分子生物学] Q255