TNF-α诱导B16细胞自体吞噬的作用研究  

Autophagic cell death in mouse melanoma B16 cells induced by TNF-α

作　　者：鄢文慧[1] 赵立平[1] 乔海晅[1] 

机构地区：[1]天津医科大学生物医学工程学院,300070

出　　处：《国际生物医学工程杂志》2012年第3期151-154,I0003,共5页International Journal of Biomedical Engineering

基　　金：天津市科技计划项目（09JCYBJC13300）

摘　　要：目的 研究肿瘤坏死因子a（TNF-a）诱导小鼠黑色素瘤B16细胞自体吞噬和自噬性死亡的作用.方法体外培养小鼠黑色素瘤B16细胞,TN F-a（20ng/ml）处理后不同时间点（12、24h）分别收取细胞,相差显微镜观察TNF-α处理前后B16细胞的生存状态,透射电子显微镜（TEM）观察TNF-a处理前后B16细胞内双层膜结构自体吞噬泡的情况,反转录聚合酶链式反应（RT-PCR）、蛋白免疫印迹分析自噬标记物微管相关蛋白1轻链3（LC3]和Beclin1以及自噬相关基因（Atg）Atg5、Atg7、Atg12表达的变化,以LC3-Ⅱ/LC3-Ⅰ的比值作为自噬活性的指标,以Beclin1作为自噬性死亡的指标.结果TNF-a处理B16细胞24 h后自体吞噬泡数量增多.与对照组相比,TNF-α持续作用可以诱导自噬标记物LC3-Ⅱ和自噬性死亡标记物Beclin1蛋白表达增高,并且呈时间依赖性；同时自噬相关基因Atg5、Atg7 、Atg12 mRNA的表达在TNF-a处理12、24h后显著增高.结论TNF-α除了可以诱导B16细胞凋亡,还可诱导B16细胞发生自体吞噬并导致自噬性死亡.Objective To analyze the autophagy of mouse melanoma B16 cells induced by TNF-a. Methods Mouse melanoma B 16 cells treated with TNF-a were used in this study. The expression levels of LC3- II and Beelinl were measured by western blot and mRNA levels of autophagy related gene atgS, atg7 and atgl2 were measured by reverse transcription polymerase chain reaction （RT-PCR） after TNF-a treatment. Results The viability of B16 cells were obviously inhibited after incubation with TNF-a. The expression levels of autophagy related protein LC3- II and Beclinl were elevated in TNF-a treated B16 cells compared with in control B16 cells. The mRNA expression levels of autophagy related gene Atg5, atg7 and atgl2 showed consistent up regulation in TNF-a treated B16 cells compared with in control B16 cells. Conclusion TNF-a can induce autophagic cell death in B16 cells.

关 键 词：肿瘤坏死因子-Α B16细胞 自体吞噬 

分 类 号：R730.2[医药卫生—肿瘤]