Caveolin-1在超微细炭黑颗粒入胞途径中的作用  

The role of caveolin-1 for carbon black nanoparticles uptake in vitro

作　　者：余珉[1] 陈日萍[1] 贾振宇[1] 陈钧强[1] 蒋兆强[1] 冯玲芳[1] 张幸[1] 

机构地区：[1]浙江省医学科学院卫生学研究所,杭州310013

出　　处：《中华劳动卫生职业病杂志》2013年第3期161-165,共5页Chinese Journal of Industrial Hygiene and Occupational Diseases

基　　金：浙江省科技计划项目（2008C13029.2）;浙江省医学支撑学科劳动卫生学（11-ZC02）;浙江省医学科学院青年基金（PT1131E）

摘　　要：目的探讨超微细炭黑颗粒（Carbonblack nanoparticles，CBNPs）对肺泡Ⅱ型上皮细胞（A549）Caveolin-1蛋白的表达的影响及其在人胞过程中的作用。方法A549细胞接受0、25、50、100、200、400μg／mlCBNPs染毒24h，以台盼蓝染色法检测CBNPs对细胞活性的影响；0、25、50、100μg/mlCBNPs染毒A549细胞24h，应用流式细胞术细胞检测侧向角散射参数（sidescatter，SSC）变化，用透射电镜观察细胞形态学变化；采用免疫蛋白印迹法（Westernblot）检测CBNPs对细胞Caveolin-1蛋白表达水平的影响。A549细胞接受1μg／mlFilipin及100Ixg／mlCBNPs共同处理24h，观察细胞SSC的变化。结果与对照组比较，200和400μg／ml剂量组细胞存活率分别下降38．2％和46．6％，差异有统计学意义（P〈0．05），25、50和100μg／ml剂量组细胞活性无明显降低，差异无统计学意义（P〉0．05）。50、100μg／ml剂量组细胞Caveolin-1蛋白表达水平较对照组明显增加，差异有统计学意义（P〈0．05）。透射电镜结果显示，50和100μg／ml处理组细胞胞浆出现质膜囊泡，内含黑色颗粒状物质。流式细胞术检测结果显示，随着cBNPs染毒剂量（0.100μg／ml）的增加，细胞SSC比值由1．007升至1.331；1μg／mlFilipin＋100μg／mlCBNPs剂量组细胞SSC比值由1．331降至1．25。结论A549能胞吞超微细炭黑颗粒，但Caveolin依赖性胞吞机制在介导超微细炭黑颗粒胞吞过程中不起主要作用。Objective To investigate the protein expression of caveolin-1 in type II alveolar epithelial cells （A549） exposed to carbon black nanoparticles （CB NPs） and the role of caveolin in the endocytosis of CB NPs. Methods A549 cells were exposed to 0, 25, 50, 100, 200, and 400 μg/ml CB NPs for 24 h; then, trypan blue assay was applied to determine the cell viability. A549 cells were also exposed to 0, 25, 50, and 100 μg/ml CB NPs for 24 h, then, transmission electron microscopy （TEM） and flow cytometry were applied to observe the morphological change of cells and cellular side scatter （SSC）, and Western blot was used to analyze the effect of CB NPs on the protein expression of caveolin-1. A549 cells were co-exposed to 1 μg/ml filipin and 100 μg/ml CB NPs for 24 h, then, the cellular SSC was observed. Results Compared with controls, the A549 cells exposed to 200 and 400 μg/ml CB NPs had the cell viability decreased by 38.2% and 46.6%, respectively （P〈 0.05）, while those exposed to 25, 50, and 100 μg/ml CB NPs showed no significant decrease in cell vitality （P〉 0.05）. The protein expression of caveolin-1 was significantly higher in the cells exposed to 50 and 100 μg/ml CB NPs than in controls （P〈0.05）. The TEM showed that plasmalemmal vesicles containing black particles were found in the cytoplasm of the cells exposed to 50 and 100 μg/ml CB NPs. The flow cytometry showed that the cellular SSC ratio increased from 1.007 to 1.331 as the dose of CB NPs rose within 0-100 μg/ml and fell to 1.25 after the cells were co-exposed to 1μg/ml filipin and 100 μg/ml CB NPs. Conclusion Carbon black nanoparticles can be transferred into A549 cells by endocytosis, but caveolin-mediated endocytic pathway plays a minor role in this process.

关 键 词：超微细炭黑颗粒 肺泡 上皮细胞 CAVEOLIN-1 蛋白 

分 类 号：R1[医药卫生—公共卫生与预防医学]