小鼠成纤维细胞衰老模型建立
Establishment of the senescence cell model of mouse fibroblast
马珂 1洪诗瑶 1张燕红 1张聪聪1
作者信息
- 1. 100029 首都医科大学附属北京安贞医院-北京市心肺血管疾病研究所 心血管生物学研究室
- 折叠
摘要
目的:构建小鼠成纤维细胞的衰老模型.方法:利用小鼠3T3成纤维细胞系,给予P53激动剂Nutlin3a(5 µ mol/L)诱导衰老成纤维细胞.使用β-半乳糖苷酶(senescence associated β-galactosidase staining,SA-β-gal)衰老染色评估衰老细胞比例,使用实时定量聚合酶链反应(quantitative realtime polymerase chain reaction,qRT-PCR)的方法检测细胞中衰老标志分子 P16 和 Gls的mRNA表达变化.利用GLS1抑制剂BPTES处理衰老成纤维细胞评估对衰老细胞的清除作用,并评估了不同浓度的BPTES对衰老成纤维细胞的影响.结果:Nutlin3a可诱导3T3成纤维细胞衰老,与对照组细胞相比,成纤维细胞中SA-β-gal阳性的衰老细胞比例增加(11.8％vs.92.8％,P＜0.01).qRT-PCR结果显示诱导衰老的成纤维细胞中衰老标志分子P16和Gls的表达显著上调(P均＜0.01).BPTES处理也可显著减少Nutlin3a诱导的衰老成纤维细胞的数量(P＜0.01),并且BPTES对衰老细胞的杀伤作用呈剂量依赖性(P＜0.01).结论:使用P53激动剂Nutlin3a可成功建立成纤维细胞的细胞衰老模型,BPTES可有效清除衰老成纤维细胞.
Abstract
Objective:To establish a senescence cell model of mouse fibroblast.Methods:Mouse 3T3 fibroblasts were stimulated with 5pmol/L P53 agonist Nutlin3a to induce senescence.The proportion of senescent cells was evaluated by senescence associated β-galactosidase staining(SA-β-gal),and the mRNA expression changes of senescence markers P16 and Gls were detected by real-time quantitative polymerase chain reaction(qRT-PCR).Senescent fibroblasts were treated with BPTES to assess the clearance of senescent cells,and the effects of different concentrations of BPTES on senescent fibroblasts were evaluated.Results:Nutlin3a induced senescence in 3T3 fibroblasts,and the proportion of SA-β-gal positive cells was increased in the senescence fibroblasts compared with the control cells(11.8％vs.92.8％,P＜0.01).Compared with control cells,Nutlin3a treated fibroblasts exhibited increased ratio of SA-β-gal positive cells,accompanied with the up-regulation of senescence related genes P16 and Gls(P＜0.01).In addition,BPTES treatment significantly reduced the number of senescent fibroblasts induced by P53 activator,and the killing effect of BPTES on senescent cells was dose dependent(P＜0.01).Conclusions:Using Nutlin3a can be successfully established senescence model of fibroblasts,and BPTES can effectively eliminate senescent fibroblasts.
关键词
3T3成纤维细胞系/细胞衰老/衰老细胞清除Key words
3T3 fibroblast cell line/Cell senescence/Senescent cell clearance引用本文复制引用
基金项目
北京市自然科学基金资助项目(Z200026)
出版年
2024
NETL
NSTL
万方数据