染色质重塑因子1在肺癌中过表达并促进肺癌增殖能力

Remodeling and Spacing Factor 1 is Overexpressed and Promotes the Tumor Cell Proliferation in Lung Cancer

目的检测染色质重塑因子1(Rsf-1)对肺癌增殖能力的影响,初步探讨Rsf-1促进肺癌增殖的分子机制。方法应用Western blot方法筛选Rsf-1高表达人肺癌H1299、H460细胞系,通过小干扰RNA(si RNA)方法干扰内源性Rsf-1表达,并检测干扰效率。应用集落形成、流式细胞术检测Rsf-1干扰前后肺癌细胞增殖能力的变化。应用Western blot检测干扰内源性Rsf-1表达前后肺癌细胞中增殖相关因子的变化。结果 Western blot结果显示:肺癌细胞中Rsf-1蛋白表达水平明显高于正常支气管上皮HBE细胞系,其中H1299、H460细胞中存在非常明显的Rsf-1高表达。转染Rsf-1特异的si RNA于高表达Rsf-1的H1299和H460细胞后,其克隆形成明显少于对照组(H1299细胞中对照组和干扰组分别为123±15和83±9;H460细胞中对照组和干扰组分别为218±18和112±17)(P<0.05)。流式细胞术结果显示:干扰内源性Rsf-1表达后,H1299和H460细胞中G1期细胞比例增多(H1299细胞中对照组和干扰组分别为56%±5%和71%±7%;H460细胞中对照组和干扰组分别为53%±4%和70%±6%),S期细胞比例减少(H1299细胞中对照组和干扰组分别为17%±2%和11%±5%;H460细胞中对照组和干扰组分别为19%±2%和10%±5%)。细胞周期相关蛋白检测结果:干扰内源性Rsf-1表达后,H1299和H460细胞中cyclin D1和磷酸化细胞外调节蛋白激酶(p ERK)水平明显下调。干扰Rsf-1在下调H1299和H460细胞p ERK蛋白表达的作用结果与应用细胞外调节蛋白激酶(ERK)抑制剂U0126相似。结论 Rsf-1可通过cyclin D1/ERK相关通路促进肺癌细胞的增殖。

Objective To study the relationship between remodeling and spacing factor 1 （Rsf-1 ） expression and tumor cell proliferation in lung cancers. Methods Small interfering RNA （siRNA） was used to knockdown the high endogenous Rsf-1 expression in H1299 and H460 cell lines. Colony formation assays and flow cytometry were employed to detect the change of cell proliferation ability of H1299 and H460 cells. The expression of proliferation related factor changes after interference was evaluate by Western blot. Results Knockdown of Rsf- 1 expression in H1299 and H460 cells resulted in a decrease of clone formation ability （H 1299 control vs siRNA ： 123±15 vs 83±9 ; H460 control vs siRNA ： 218±18 vs 112±17, P 〈 0.05 ）. Rsf- 1 knockdown induced ceil cycle arrest at the G1/S boundary to inhibit cell proliferation, G, phase cells （H1299 control vs siRNA ： 56%± 5% vs 71%±7% ; H460 control vs siRNA ： 53%±4% vs 70%±6% ）, S phase cells （H1299 control vs siRNA ： 17%±2% vs 11%±5% ; H460 control vs siRNA ： 19%±2% vs 10%±5% ） （P 〈 0.05）. Western blot results revealed that knockdown of Rsf-1 decreased cyclinD1 and phosphorylation of extraceilular-regulated protein kinase （pERK） protein level in both cell lines. At the same time, Rsf-1 knockdown showed the same effect with U0126 in down-regulating the levels of pERK in both cell lines. Conclusion Rsf-1 contributes to malignant cell growth by modulating cyclinD1 ex- pression and ERK activity, which makes Rsf- 1 a potential therapeutic target in lung cancer.