整合素beta1亚单位对肝癌细胞与IV型胶原黏附与趋化行为的影响  

Integrin beta1 Mediates Hepatocellular Carcinoma Cells Adhesion & Chemotaxis to Type IV Collagen

作　　者：傅遍红[1] 吴泽志[1] 董澄 秦建[1] 

机构地区：[1]重庆大学生物工程学院生物力学与组织工程教育部重点实验室 [2]美国宾夕法尼亚州州立大学生物工程系

出　　处：《生物医学工程学杂志》2004年第5期741-745,共5页Journal of Biomedical Engineering

基　　金：国家自然科学基金资助项目 ( 3 9970 198);教育部重点实验室访问学者基金资助 (校科字 [2 0 0 2 ] 4号 )

摘　　要：采用微吸管实验技术 ,测定肝细胞癌 (Hepatocellular carcinom a,HCC)细胞与 IV型胶原裱衬表面的黏附力 ;进一步加入针对整合素 beta1亚单位 (CD2 9)的单克隆抗体 (Anti- CD2 9)处理 HCC细胞 ,观察 Anti- CD2 9对细胞与 IV型胶原裱衬表面的黏附力的影响。采用双微吸管实验法进行 HCC细胞趋化实验 ,在两侧微管内加入相同浓度 (6 0 0μg/ ml )的 IV型胶原 ,并引导微管尖端与同一细胞紧密接触 ,动态观察细胞两侧伪足形成过程 ;在一侧微吸管内加入 2 0μg/ ml Anti- CD2 9,考察 beta1整合素亚单位阻断对 HCC细胞伪足形成的影响。利用流式细胞仪对 HCC细胞表面整合素 beta1亚单位的表达进行分析。结果表明 ,HCC细胞与 5μg/ ml IV型胶原裱衬表面之间的黏附力为 932± 134(× 10 - 1 0 N ,n=6 0 ) ,加入 5μg/ ml Anti- CD2 9黏附力减小到 4 4 9± 119(× 10 - 1 0 N,n=6 0 ) ;加入 10 μg/ ml Anti- CD2 9时黏附力减小到 2 2 0± 78(× 10 - 1 0 N,n=5 5 )。双微吸管趋化实验表明 :两侧微吸管加入相同浓度 IV型胶原 ,细胞向两侧微吸管均有伪足形成 ;在此基础上 ,微吸管加入 Anti- CD2 9的一侧 ,HCC细胞伪足生长曲线呈现明显的抑制 ,而未加入 Anti- CD2 9的微吸管一侧与加入的对侧相比 ,该侧细胞的伪足明显增?A micropipette technique was adopted to investigate the effect of blockade of integrin beta1 on adhesion of hepatocellular carcinoma(HCC)cells onto type IV collagen(Col IV) coated surfaces and pseudopod protrusion of HCC cells in response to Col IV stimulation. Adhesion strength was expressed as an adhesion force, which was defined as the product of the cross sectional area and critical negative pressure needed to detach single cell away from the substrate. Chemotactic pseudopod protrusion of an HCC cell was evaluated using a dual-pipette set-up, in which two pipettes filled with Col IV solution were positional in close contact with the same cell and pseudopod protrusion into each pipette was viewed dynamically and recorded with a tape recorder. The lengths of pseudopods were measured and plotted against time to obtain a pseudopod growth curve. The integrin beta1 subunit on the surfaces of HCC cells was analyzed by flow cytometry. The results showed that the adhesion forces for HCC cells adhering on 5 μg/ml Col IV coated surfaces were 932±134(×10 -10N,n=60). Upon treatment of HCC cells with Anti-CD29 in a protein concentration of 5μg/ml and 10μg/ml , the value decreased significantly to 449±119(×10 -10N,n=60) and 220±78(×10 -10N,n=55), respectively. In dual pipette chemotaxis experiment, when the two pipettes were filled with Col IV in an identical concentration of 600μg/ml, pseudopods extended from the HCC cell into each of the pipettes nearly symmetrically, i.e., with nearly identical maximum pseudopod length and similar pseudopod growth curves. Upon addition of Anti-CD29 to one of the pipettes in a protein concentration of 20μg/ml, pseudopod protrusion was blocked nearly completely while protrusion into the opposite pipette became more evidently, with larger maximum length. Expression of integrin beta1 was up to 95.78% to cells chosen in the experiment. These results suggested that integrin beta1 subunit was important constituent receptor subunit for mediating HCC cell adhesion and chemotactic

关 键 词：IV型胶原 HCC CD29 细胞 整合素 亚单位 黏附力 微管 生长曲线 表达率 

分 类 号：R735.7[医药卫生—肿瘤] R687.34[医药卫生—临床医学]