肿瘤坏死因子α诱导蛋白2对非小细胞肺癌细胞株H1299的增殖、凋亡、迁移和侵袭能力的影响

Effect of TNF-α induced protein-2 on the proliferation, apoptosis, migration,and invasion of non-small cell lung cancer H1299 cells

摘要:

目的：观察肿瘤坏死因子α诱导蛋白2（TNF-α-induced protein 2，TNFAIP2）在非小细胞肺癌（non-small cell lung can－cer，NSCLC）中的表达以及对细胞增殖、凋亡、迁移和侵袭能力的影响。方法：RT-PCR检测NSCLC中TNFAIP2 mRNA表达情况；免疫组化检测NSCLC中TNFAIP2蛋白表达情况。构建TNFAIP2基因短发夹RNA（short hairpin RNA，shRNA）干扰慢病毒载体（shRNA-TNFAIP2）和阴性对照慢病毒载体（shRNA-NC）分别感染H1299细胞后采用RT-PCR和Western blot检测感染效率。用Transwell实验检测细胞的迁移、侵袭能力；用CCK-8检测细胞的增殖能力；用流式细胞术检测细胞的凋亡能力；用Western blot检测细胞蛋白E-cadherin、N-cadherin、Vimentin表达的变化。结果：TNFAIP2 mRNA和蛋白质在NSCLC癌组织中的表达明显高于癌旁组织中的表达。慢病毒下调TNFAIP2表达后，shRNA-TNFAIP2组细胞迁移数（170±11）较shRNA-NC组（329±31）明显减少（P=0.001）；侵袭细胞数（75±10）较shRNA-NC组（135±8）明显减少（P=0.001）。CCK-8结果提示，shRNA-TNFAIP2组的吸光度值明显低于shRNA-NC组（P=0.000）；流式细胞术结果显示，shRNA-TNFAIP2组的凋亡率（18.730±0.490）%明显高于shRNA-NC组（6.570±0.870）%（P=0.000）；Western blot结果显示，shRNA-TNFAIP2组较shRNA-NC组细胞蛋白N-cadherin、Vimentin表达降低，E-cadherin表达增加。结论：TNFAIP2在NSCLC中存在高表达，下调能够抑制细胞的增殖、迁移和侵袭能力，促进细胞凋亡，逆转上皮间质转化。

Abstract:

Objective：To investigate the expression of TNF-α-induced protein 2（TNFAIP2） in non-small cell lung cancer （NSCLC） and its effect on cell proliferation，apoptosis，migration，and invasion. Methods：RT-PCR was used to measure the mRNA expression of TNFAIP2 and immunohistochemistry was used to measure the protein expression of TNFAIP2 in NSCLC. The lentivirus-mediated short hairpin RNA of TNFAIP2（shRNA-TNFAIP2） and negative control lentivirus（shRNA-NC） were constructed and were used to infect H1299 cells，and then RT-PCR and Western blot were used to measure the efficiency of infection. Transwell assay was used to measure cell migration and invasion abilities，CCK-8 assay was used to measure proliferative capacity，flow cytometry was used to measure cell apoptosis，and Western blot was used to measure the changes in the protein expression of E-cadherin，N-cadherin，and vimentin. Results：The mRNA and protein expression of TNFAIP2 in NSCLC tissue was significantly higher that that in adjacent tissue. After TNFAIP2 was downregulated by lentivirus，compared with the shRNA-NC group，the shRNA-TNFAIP2 group had significant reductions in the number of migrated cells（170±11 vs. 329±31，P=0.001） and the number of invasive cells（75±10 vs. 135±8，P=0.001）. CCK-8 assay showed that the shRNA-TNFAIP2 group had a significantly lower absorbance value than the shRNA-NC group（P=0.000）；flow cytometry showed that the shRNA-TNFAIP2 group had a significantly higher apoptosis rate than the shRNA-NC group[（18.730±0.490）% vs. （6.570±0.870）%，P=0.000]；Western blot showed that compared with the shRNA-NC group，the shRNA-TNFAIP2 group had reductions in the protein expres－sion of N-cadherin and vimentin and an increase in the protein expression of E-cadherin. Conclusion：TNFAIP2 is highly ex－pressed in NSCLC. Downregulation of TNFAIP2 can inhibit cell proliferation，migration，and invasion，promote cell apoptosis，and re－verse epithelial-mesenchymal transition.