FK866对非小细胞肺癌细胞迁移的影响

目的: 探讨低浓度烟酰胺磷酸核糖基转移酶（NAMPT）抑制剂FK866对人非小细胞肺癌细胞株（A549细胞）迁移的影响及作用机制。方法: MTT法检测不同浓度FK866对A549细胞增殖的影响；划痕实验检测1.0 nmol/L和10.0 nmol/L FK866对A549细胞迁移的影响；实时定量RT-PCR检测上皮间质转化相关蛋白上皮钙黏素和波形蛋白mRNA的表达量；蛋白质印迹法检测细胞外调节蛋白激酶1/2（ERK1/2）和磷酸化ERK1/2蛋白的表达量。结果: FK866作用时间越长、浓度越高，对A549细胞增殖的抑制作用越明显。FK866作用72 h时的半抑制浓度（IC₅₀）为9.55 nmol/L。以1.0、10.0 nmol/L的FK866预处理细胞48 h，划痕后继续给药48 h，两种浓度的FK866均能抑制A549细胞迁移；如不进行预处理，仅10.0 nmol/L浓度的FK866对划痕愈合有一定的抑制作用；1.0 nmol/L的FK866处理细胞72 h可上调上皮钙黏素和波形蛋白mRNA表达，并激活ERK1/2；以1.0 mmol/L的烟酰胺单核苷酸（NMN）或10.0 μmol/L的ERK1/2抑制剂U0126预处理，能够逆转FK866引起的上皮钙黏素和波形蛋白表达上调。结论: 低浓度FK866能够通过减少细胞内烟酰胺腺嘌呤二核苷酸和激活ERK1/2，增加上皮钙黏素表达，进而抑制细胞迁移，对肿瘤转移可能有一定的抑制作用；但其同时促进了波形蛋白的表达，不利于肿瘤的治疗。

Effects of FK866 on migration of A549 cells and related mechanism

Objective: To investigate the effect of nicotinamide phosphoribosyltransferase (NAMPT) inhibitor FK866 on the migration of human non-small cell cancer A549 cells and related mechanism. Methods: The inhibition effect of FK866 on A549 cells was tested by MTT assay. A549 cells were treated with 1.0 and 10.0 nmol/L FK866, and the cell migration was evaluated by modified wound scratch assay. The mRNA expression of E-cadherin and vimentin was detected by real-time RT-PCR, and the expression of ERK1/2 and pERK1/2 was determined by Western blotting. Results: FK866 inhibited the proliferation of A549 cells in a time- and concentration-dependent manner; after treatment for 72 h, the IC₅₀ of FK866 was 9.55 nmol/L. When 1.0 nmol/L or 10.0 nmol/L FK866 was continuously applied 48 h before and 48 h after a scratch was made in wound scratch assay, the migration of A549 cells was significantly inhibited. However, when the FK866 was applied only 48 h after the scratch, the migration of A549 cells was inhibited by 10.0 nmol/L but not by 1.0 nmol/L FK866. The mRNA expression of E-cadherin and vimentin, and the activated ERK1/2 were significantly increased after 1.0 nmol/L FK866 treatment for 72 h. The pretreatment with nicotinamide adenine dinucleotide (NAD) precursor nicotinamide mononucleotide(1.0 mmol/L) or ERK1/2 inhibitor U0126 (10.0 μmol/L) reversed the up-regulation of E-cadherin and vimentin expression induced by FK866. Conclusions: Low concentration of FK866 decreases the migration of A549 cells through the inhibition of NAD level, activation of ERK1/2 and up-regulation of E-cadherin expression. However, it also up-regulates the expression of vimentin, indicating that it may have dual effects on the migration of tumor cells.