【摘要】 目的研究抑制人肝癌细胞 MHCC97-H 中黏着斑激酶(FAK)表达对细胞黏附、侵袭和骨架重塑的影响。方法利用脂质体转染 FAK siRNA 至 MHCC97-H 细胞内。Western blot 法检测RNA 干扰前后 MHCC97-H 细胞中 FAK 蛋白的表达水平。黏附试验和侵袭试验分别检测 MHCC97-H细胞黏附和侵袭能力在干扰前后的变化。明胶酶谱试验检测 MHCC97-H 细胞分泌 MMPs 的变化。用激光共聚焦显微镜观察免疫荧光标记细胞的骨架重塑的变化。结果特异性 FAK siRNA 明显抑制 MHCC97-H 细胞中 FAK 表达。干扰后 MHCC97-H 细胞与细胞外基质纤维连接蛋白的黏附率为35.8%,低于对照组的57.3%(P<0.05)。干扰后 MHCC97-H 细胞穿透 Tanswell 小室的数目由对照组的(31.3±2.6)个减少至(14.5±3.1)个(P<0.05)。干扰后 MHCC97-H 细胞分泌 MMPs 的量明显减少。FAK 表达抑制影响骨架蛋白 Vinculin 在 PDGF-BB 诱导细胞骨架重塑中分布,阻碍片状伪足形成,延迟黏着斑形成时间。结论 FAK 表达受到抑制后,通过减少 MMPs 分泌和影响细胞骨架重塑可降低 MHCC97-H 细胞黏附、侵袭能力。更多还原

【Abstract】 Objective To study the role of inhibition FAK expression by FAK siRNA in liver cancer cell(MHCC97-H)adhesion,invasion and cytoskeleton rearrangement.Methods FAK siRNA was transfected into MHCC97-H cell by Lipofectamine 2000;then,FAK expression was detected by Western blot analysis.The change of cell adhesive and invasive ability after RNAi was checked by cell adhesive assay and cell invasive assay respectively.Meanwhile,matrix metalloproteinase-2 secretion was checked by gelatin zymography.Cytosekeleton rearrangement labeled by immunofluorescence antibody was examined by confocal laser scanning microscope.Results FAK expression in MHCC97-H cell was obviously inhibited by specific FAK siRNA;However,it was not inhibited by negative siRNA.Adhesion rate between MHCC97-H cell and extraeellular matrix decreased from 57.3% to 35.8% after RNA interference(P<0.05).Compared with untreated group,the number of cell penetrating matrigel also decreased from 31.3±2.6 to 14.5±3.1 after transfection(P<0.05).Besides,matrix metalloproteinase-2 secretion was significantly reduced for FAK expression inhibited by FAK siRNA.FAK inhibition influenced Vinculin rearrangement,blocked the formation of lamellipodium,delayed the time of focal adhesion formation.Conclusion Down-regulation the expression of FAK can reduce adhesive rate and invasive number of MHCC97-H cell by influencing cytoskeleton rearrangement and descreasing matrix metalloproteinase-2 secretion.更多还原