敲低细胞角蛋白18对氧化应激诱导细胞凋亡的影响

发布时间：2018-01-11 16:26

本文关键词：敲低细胞角蛋白18对氧化应激诱导细胞凋亡的影响　出处：《军事医学》2013年07期 　论文类型：期刊论文

【摘要】：目的研究氧化应激条件下中间纤维蛋白细胞角蛋白(cytokeratin)的抗凋亡作用。方法利用RNA干扰(RNAi)技术敲低细胞内细胞角蛋白18(CK18)的表达,免疫印迹法检测细胞角蛋白的表达水平,并通过A549细胞建立细胞角蛋白敲低稳定细胞系。利用免疫荧光染色技术观察细胞角蛋白对细胞骨架的影响。利用流式细胞技术检测细胞角蛋白对细胞内ROS(reactive oxygen species)水平及氧化应激诱导细胞凋亡的影响。结果筛选获得3个干扰效果较好的RNAi序列,成功利用A549细胞建立细胞角蛋白敲低的稳定细胞系(A549-KD)。CK18表达敲低导致细胞内多种细胞角蛋白表达广泛下调,并破坏细胞微丝及微管骨架结构。氧化应激条件下A549-KD细胞抗凋亡活性显著降低。结论细胞角蛋白参与维持细胞骨架结构,对抗ROS诱导的细胞凋亡,提示其在细胞的氧化应激过程中发挥保护作用。
[Abstract]:Objective to study the antiapoptotic effect of cytokeratin in intermediate fibrin cells under oxidative stress. The expression of cytokeratin 18 (CK18) in the cells was reduced. The expression of keratin was detected by Western blot. The cytokeratin knockdown stable cell line was established by A549 cell line. The effect of cytokeratin on cytoskeleton was observed by immunofluorescence staining. Flow cytometry was used to detect the effect of keratin on cell cytoskeleton. OSS (. The level of reactive oxygen speciesand the effect of oxidative stress on apoptosis. Results three RNAi sequences with good interference effect were obtained. The successful use of A549 cell line to establish a stable cell line with low keratin knockdown induced a wide down-regulation of keratin expression in a variety of cells. The anti-apoptotic activity of A549-KD cells was significantly decreased under oxidative stress. Conclusion cytokeratin is involved in the maintenance of cytoskeletal structure. Antagonizing apoptosis induced by ROS may play a protective role in oxidative stress.
【作者单位】：安徽大学生命科学学院;军事医学科学院生物工程研究所;
【分类号】：R363
【正文快照】： 真核生物细胞骨架由微丝(microfilaments)、微管(microtubules)及中间纤维(intermediate fila-ments)组成。这些骨架结构在细胞内形成网络,彼此相互联系而各自又具有独特的结构与功能。细胞角蛋白(cytokeratin)属于中间纤维蛋白的一种,可进一步分为呈酸性的Ⅰ型(K9～K20)及呈

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