中文摘要：

目的 了解外源性NO对HaCaT细胞迁移功能的影响及其机制.方法 以硝普钠作为NO的供体,在HaCaT细胞培养液中加入0.1、1.0、10.0、100.0、1000.0 μmol/L硝普钠,分别于划痕0 h（划痕即时）及划痕后6、12、24、48 h观察并计算细胞迁移率.根据该结果筛选出硝普钠最佳作用浓度及作用时间,以此为实验刺激条件,应用激光共聚焦显微镜观察细胞骨架的改变,采用蛋白质印迹法、PCR等方法检测实验组（加入10.0 μmol/L硝普钠培养24 h）及阴性对照组整合素β_1、RhoA、cdc42、Rac1蛋白及RhoA、cdc42、Rac1 mRNA表达情况.对各实验结果行单因素方差分析和重复测量的方差分析.结果 加入硝普钠处理后各浓度组细胞迁移率随划痕时间的延长而逐渐增加.加入10.0 μmoL/L的硝普钠划痕后6～48 h与划痕0 h比较,差异均有统计学意义（F=31.002,P值均小于0.05）.激光共聚焦显微镜显示,阴性对照组细胞纤毛稀少,胞内应力纤维束纤细,而实验组纤毛明显增多,胞内应力性纤维束增粗.实验组整合素β1蛋白表达水平明显低于阴性对照组（F=8.25,P=0.015）;而RhoA的蛋白表达水平为0.92±0.04,高于阴性对照组的0.64±0.04（F=7.25,P〈0.05）,cdc42、Rac1蛋白表达也高于阴性对照组（F值分别为14.10、6.50,P值均小于0.05）.实验组RhoA、cdc42、Rac1的mRNA水平均高于阴性对照组（F值分别为23.67、10.39、9.52,P值均小于0.05）.结论 适宜浓度的外源性NO可促进HaCaT的增殖及迁移,提示其在皮肤创面修复中起重要作用.其机制可能与整合素β_1表达下降、细胞骨架的改变有关.

英文摘要：

Objective To investigate the effect of exogenous nitric oxide （ NO） on the migration of HaCaT cell and its possible mechanism. Methods Sodium nitroprussiate （SNP） was used as the donor of NO. Different concentrations of SNP （0. 1, 1.0, 10.0, 100.0, 1000.0 μmol/L ） were added into nutrient culture medium of HaCaT cells. Cell migration rate was observed and calculated at post scratching hour （PSH） 0 （immediately after scratching） , 6, 12, 24, 48. The most suitable concentration of SNP and culture duration were selected as stimulation condition. Cytoskeletons of HaCaT cells were observed under con-focal laser scanning microscope. The expressions of integrin beta 1, RhoA, Racl and Cdc42 of cells in experiment group （cultured with 10.0 μmol/L SNP for 24 hours） and negative control group were determined at mRNA and protein levels with RT-PCR and Western blot respectively. Data were processed with one-way analysis of variance （ANOVA） and repeated measure ANOVA. Results Migration rate of HaCaT cells in each group increased gradually as time after scratching went on. There were significant differences between PSH 6-48 and PSH 0 in cells cultured with 10. 0 μmol/L SNP （ F = 31.002, P values all below 0. 05 ）. Pili were rarely observed in negative control group with slender stress fibers in cells. In comparison, the amount of pili amount increased obviously in experiment group with thickened stress fibers. Compared with those of cells in control group （ RhoA protein expression =0.64 ±0.04） , integrin beta 1 expression decreased obviously （ F =8.25, P =0.015）, RhoA （0.92 ±0.04）, Cdc42 and Racl were up-regulated at both protein （ with F value respectively 7. 25 , 14. 10, 6. 50, P values all below 0.05） and mRNA levels （ with F value respectively 23. 67 , 10. 39, 9. 52, P values all below 0. 05 ）. Conclusions Exogenous NO in suitable concentration can promote the proliferation and migration of HaCaT cell, suggesting it exerts significant effect in wound repair. The chang