【摘要】 目的应用RNA干扰技术沉默信号转导及转录激活因子3(STAT3)后,观察对喉鳞状细胞癌2(Hep-2)黏附、迁移、侵袭能力的影响及细胞内细胞黏附分子1(ICAM-1)和血管内皮生长因子(VEGF)蛋白表达的改变,探讨STAT3作用于喉癌的可能侵袭机制。方法在前期试验筛选出稳定表达siRNA-STAT3的Hep-2细胞系的基础上,应用MTT比色法、划痕法、Transwell法,分别检测其黏附力、迁移力及侵袭力的变化;Western印迹法检测细胞ICAM-1和VEGF蛋白的表达。以siRNA-shNC组和空白组作为对照。结果①接种20min后,各组细胞的黏附率差异不大(P=0.886);40min后,siRNA-STAT3组的黏附率小于siRNA-shNC组和空白对照组(P=0.02);60min后,黏附抑制效应更加明显(P=0.012)。②接种48h后,空白对照组和siRNA-shNC组的Hep-2细胞向划痕区生长速度明显快于siRNA-STAT3组。③接种24h后,200倍光学显微镜下显示:siRNA-STAT3组穿过Matrigel基质和滤膜的细胞数明显少于空白对照组和siRNA-shNC组。④Western印迹法结果显示:siRNA-STAT3组ICAM-1和VEGF蛋白的表达量低于siRNA-shNC组和空白对照组。结论稳定转染重组STAT3基因能显著抑制喉癌Hep-2细胞的黏附、迁移及侵袭能力。STAT3在喉癌的侵袭中起着重要的作用,可能与其调控ICAM-1和VEGF蛋白的表达有关。更多还原

【Abstract】 Objective To observe the changes of adhesive, migratory and invasive ability of Hep-2 cells after signal transducers and activators of transcription 3(STAT3) expression was blocked by RNA interference technology, and of the expression of intercelluar adhesion the molecule 1(ICAM-1) and vascular endothelial growth factor(VEGF), and to discuss the invasive mechanism of STAT3 on laryngeal cancer. Methods The adhesive ability by MTT assay,migratory ability by cell scratch assay and invasive ability by Transwell chamber was observed on the basis of the previous experiments where the authors selected Hep-2 cell line stably expressing siRNA-STAT3. Results ①Hep-2 cells were inclined to be adhered to matrigel matrix, but the adhesion rate didn’t decline sharply after they were inoculated in 96-well culture plate for 20 minutes(P=0.886). After 40 minutes, the adhesion rate in the siRNA-STAT3 group was much less than that of shNC group and blank control group (P=0.02). The adhesive ability was more inhibited after 60 minutes (P=0.012). ②The movement to scratch zone in the two control groups was faster than that in siRNA-STAT3 group after the cells were inoculated in culture plate for 48 hours. ③After cells in each group were inoculated in transwell chamber for 24 hours, under optical microscope(10×20), the number of cells passing through the matrigel matrix and filter membrane in siRNA-STAT3 group was found less than those in the two control groups. ④ ICAM-1 and VEGF expressions by Western Blot in siRNA-STAT3 group were found less than those in the two control groups. Conclusions The adhesive, migratory and invasive ability of Hep-2 cells was significantly inhibited by stable transfection of recombinant STAT3 gene. Activation of the STAT3 signaling pathway played a vital role in the invasive progression of laryngeal cancer possibly owing to its precise regulation towards ICAM-1 and VEGF.更多还原