PK15细胞凋亡过程中角蛋白中间纤维的变化  

CHANGES IN KERATIN DURING APOPTOSIS OF PK15 (PORCINE KIDNEY-15) CELLS

作　　者：陈丹英[1] 高云飞[1] 李丽霞[1] 翟中和[1] 

机构地区：[1]北京大学生命科学学院,北京100871

出　　处：《动物学报》2002年第1期58-63,共6页ACTA ZOOLOGICA SINICA

基　　金：国家重点基础研究发展规划项目 (No G19990 5 39)~~

摘　　要：用放线菌素D诱导上皮型猪肾细胞PK15 (PorcrneKidney 15 )发生凋亡。细胞在凋亡诱导过程中经历了从铺展、皱缩变圆至脱落的形态变化。凝胶电泳证明被诱导细胞的DNA发生降解 ,形成明显的DNAladder。免疫荧光显示凋亡过程中细胞角蛋白网状结构发生改变 ;应用选择性抽提结合整装电镜技术 ,观察到凋亡细胞中仍有中间纤维网络存在 ,这一现象前人未曾报道。免疫印迹反应进一步证明 ,凋亡细胞中部分Ⅱ型角蛋白发生降解 。Intermediate filament (IF) is a major component of cytoskeleton.Recent advances suggest that IFs not only play a cytoskeletal role in cooperation with microfilaments and microtubules, but are also involved in cell differentiation, signal transduction and gene expression.Apoptosis is a physiological form of cell death, it describes the orchestrated collapse of a cell, staging membrane blebbing, cell shrinkage, protein fragmentation, chromatin condensation and DNA degradation.How do IFs change in apoptotic process and what kind of role do they play? The answers to these questions would be of great importance to further study of their functions.Epithelial cell PK15(Porcine Kidney-15) was induced to apoptosis by actinomycin D.The cell shapes changed from spreading to contracting and rounding during the induction process, and most of the cells dropped from the wall after induction for 96 h.DAPI staining showed the chromatin condensation and apoptotic bodies formation characterized by apoptosis in this process.At the same time “DNA ladder” was detected by electrophoresis.Indirect immunofluorescence results showed that the fluorescence of keratin distributed unevenly in the apoptotic cells, and in some cells the keratin fluorescence contracted around the condensed chromatin.Using selective extraction combined with whole mount electron microscopy, we detected the intact keratin filament network in some area of apoptotic cells, and the filaments were obviously shortened.In consistent with the result of immunofluorescence, the keratin filaments around chromatin were condensed in comparison with filaments in other areas.Western blot analysis further showed that part of type Ⅱ keratin protein degraded only partially during apoptosis, whereas type Ⅰ keratin protein molecules were intact.These results suggested that in apoptotic processing the structure changes of keratin network might lead to the changes of cell shape, and might participate the formation of apoptotic bodies.

关 键 词：猪 猪肾细胞 PK15 角蛋白 中间纤维 放线菌素D 细胞凋亡 

分 类 号：Q255[生物学—细胞生物学] Q26