IL-1β促进肺癌细胞增殖的作用机制研究

Study on mechanisms of IL-1β promoted lung cancer cells proliferation

目的:研究IL-1β在巨噬细胞促进肺癌细胞增殖的作用机制。方法:采用Transwell插入式细胞培养皿共培养人肺癌细胞株A549、NCI-H520细胞和巨噬细胞。Brd U-ELISA法检测巨噬细胞对肺癌细胞的增殖能力。采用Real-time PCR法检测巨噬细胞和A549、NCI-H520细胞中IL-1βmRNA的表达。应用IL-1β的中和抗体抑制IL-1β的活性,观察IL-1β在巨噬细胞促进肺癌细胞增殖中的作用。利用Western blot法检测肺癌细胞中自噬标志物Beclin 1的表达。结果:Brd UELISA检测结果:A549细胞与巨噬细胞以1∶0.5的比例共培养,A549细胞的OD值从(0.41±0.06)增加到(1.13±0.10),差异有统计学意义(P<0.05),且A549细胞的OD值随共培养巨噬细胞比例的增加而升高(P<0.05)。NCI-H520细胞与不同浓度的巨噬细胞共培养后,NCI-H520细胞的OD值变化趋势与A549细胞相似。Real-time PCR法检测结果:在共培养后,巨噬细胞中IL-1βmRNA的表达显著上调,且有时间依赖性,并显著高于肺癌细胞中的表达。加入IL-1β中和抗体后,Brd U ELISA法检测的细胞增殖,结果显示IL-1β中和抗体可明显抑制巨噬细胞对肺癌细胞A549和NCI-H520的增殖促进作用。Western blot法检测结果显示IL-1β可显著抑制肿瘤细胞Beclin 1的表达。结论:巨噬细胞和肺癌细胞通过增强IL-1β的表达促进肿瘤细胞的增殖。抑制肿瘤细胞的自噬可能是IL-1β促进肺癌发展的作用机制。

Objective： To investigate the mechanisms of IL-1β promoted lung cancer cells proliferation. Methods： The＂Transwell Inserts＂system was used to coculture lung cancer cells A549,NCI-H520 with macrophages. Brd U ELISA used to measure the effect of macrophages promoted lung cancer cells proliferation. Expression of mRNA of IL-1β in A549 and NCI-H520 cells were analysed by Real-time PCR analysis. IL-1β was responsible for macrophage-promoted lung cancer cells growth,IL-1β neutralizing antibody was added. The autophagy marker Beclin1 protein was detected by Western blot. Results： The Brd U ELISA assay showed that after coincubation with macrophages in the proportion of 1∶ 0. 5,the OD value of A549 increased from（ 0. 41 ± 0. 06） to（ 1. 13 ± 0. 10）. There was statistical significance（ P〈0. 05）. It also showed that the growth of the A549 cell was dependent on the macrophage number（ P〈0. 05）. The OD value variability of NCI-H520 cells was as same as A549 cell upon cocultured with macrophages. Real-time PCR results showed that the expression of IL-1β mRNA in macrophages was remarkably enhanced in a time dependent manner upon coincubated with lung cancer cell,and the expression level was higher than lung cancer cells. Addition of IL-1β neutralizing antibody markedly inhibited macrophage-promoted lung cancer cells proliferation. The OD value of these two cells were decreased from（ 3. 63 ± 0. 33） to（ 1. 46 ± 0. 18）,from（ 2. 94 ± 0. 38） to（ 1. 53 ± 0. 20）,respectively（ P〈0. 05）. After treatment with IL-1β,the expression of Beclin1 was significantly inhibited in tumor cells. Conclusion： Over-expression of IL-1β from macrophages and lung cancer cells is responsible for proliferation of tumor cells in coculture condition. Inhibition of autophagy in tumor cells may be the important mechanisms of IL-1βpromotes lung cancer cells proliferation.