金属蛋白酶抑制剂对外源Rac1表达介导的细胞侵袭胶原屏障的影响

目的 体外研究金属蛋白酶(MMPs)抑制剂对外源Rac1基因表达介导的HT1080纤维肉瘤细胞侵袭胶原屏障的影响.方法 分别转染显性负调控突变体Rac1V12N17(HN)、持续活化型Rac1V12(HV)或空载体(HW)于纤维肉瘤HT1080细胞,G418筛选.蛋白印迹分析检测外源性Rac1基因表达,并在含胶原蛋白凝胶的3D基质中培养,用得克萨斯红结合的鬼笔环肽染色显示细胞肌动蛋白骨架构型.在含胶原蛋白凝胶覆盖滤膜的Transwell小室进行细胞侵袭胶原屏障实验,并观察MMPs抑制剂和抑肽酶对上述细胞侵袭实验的影响.结果 HV细胞伸出明显的突起,HN细胞形态紧凑,HV细胞侵袭胶原屏障的能力大于HW细胞,而后者又强于HN细胞,这种差别在应用广谱MMPs抑制剂后消失,而抑肽酶则无影响.结论 外源活化型Rac1基因在纤维肉瘤HT1080细胞内稳定表达可诱发肌动蛋白纤维聚集,并可增加HT1080纤维肉瘤细胞侵袭胶原屏障的能力,但这种细胞侵袭胶原蛋白能力的增强可被MMPs抑制剂阻断.

Effects of MMPs Inhibitor on HT1080 Fibrosarcoma Cell Invasion Across Collagen Barrier Mediated by Exogenous Rac1 Gene Expression

Objective To investigate the effect of MMPs inhibitor on HT1080 fibrosarcoma cell invasion across collagen barrier mediated by exogenous Rac1 gene expression. Methods HT1080 fibrosarcoma cell line that stably expressed transfected dominant negative (Rac1V12N17-HN), constitutively active Rac1(Rac1V12-HV) and vector(HW) were used. Exogenous Rac1 gene expression was detected by western blot analysis. Structure of actin cytoskeleton was stained with Texas Red-conjugated phalloidin to show the morphological char...