LPS-induced The Proliferation Of Lung Cancer A549 Cells:Role Of Hippo Signaling Pathway

Objective:Cancer-associated inflammation was considered the "seventh hallmark of cancer";Many chronic inflammatory lung diseases could increase the risk of lung cancer,and to a certain extent,promote the progress of lung cancer.Lipopolysaccharide(LPS)was one of an improtant pro-inflammatory cytokines released by gram-negative bacillus and played an important role both in inflammatory immune response and endotoxic shock.Some Previous study showed that Hippo signaling pathway was involved the the proliferation of lung cancer cells.Our aim was to study the function and mechanism of Hippo signaling pathway which was involved in LPS-induced the proliferation of lung adenocarcinoma A549 cells.Methods:1.A549 were cultured and stimulated with LPS,and the cell proliferation of A549 was detected by CCK-8.2.Hippo signaling pathway associated protein in A549((Mstl、SAV1、LATS1、Mob1、P-mob1、YAP and P-YAP(serl27))were measured by western blot analysis after stimulated with LPS in different concentrations and different time points.3.A549 were transfected with siRNA marked by cy3 and the efficiency of transfection was observed by immunofluorescence microscopy;Inhibition rate of target protein was measured by WB and RT-PCR.4.A549 were transfected with Mstl siRNA,YAP siRNA,for inhibiting the expression of Mstl,YAP,and the cell proliferation was detected by CCK-8 experiment after A549 were stimulated with LPS;5.A549 were transfected with Mstl siRNA,YAP siRNA,for inhibiting the expression of Mstl,YAP,and the expression of Mstl,LATS1,YAP and P-YAP(serl27)were measured by WB.Then the expressed location of YAP inA549 were measured by immunofluorescence.Results:1.0.1 μg/ml of LPS markedly promoted the proliferation of A549;2.LPS decreased the expression of Mstl,LATS1 and p-mobl total protein in A549,enhanced the expression of SAV1,Mob1,YAP and P-YAP total protein and promoted the nuclear translocation of YAP protein;The localization of YAP protein had no obvious effect;3.Mstl siRNA and YAP siRNA transfected into A549 successfully,and they strongly inhibited the expression of Mstl and YAP in A549;4.After transfection with Mstl siRNA,LPS had no effects on the proliferation of A549 cells;But after transfection with YAP siRNA,the effect of LPS on promoting the proliferation of A549 was markly weaken;5.After transfection with Mstl siRNA,LPS could reduce the expression of Mstl in A549 further,and strongly increase the expression of YAP;After transfection with YAP siRNA,LPS had no obvious effect on the expression of Mstl inA549,but the promotion of YAP expression was significantly weaken.6.After transfection with Mstl siRNA and YAP siRNA,the localization of YAP protein had no obvious effect in A549.Conclusion:1.Low concentration of LPS promoted the proliferation of A549 cells by enhancing the expression of YAP;2.YAP was involved in the regulation of A549 cell proliferation induced by LPS not entirely through the Hippo-dependent way.