S100P对肺癌细胞增殖和凋亡的影响及其机制

Effect of S100P on proliferation and apoptosis of lung cancer cells and its mechanism

目的探究S100P对人肺癌细胞增殖和凋亡的影响及作用机制。方法qRT-PCR检测人正常肺上皮细胞和4种人肺癌细胞A549、95D、H1975和LTEP-a-2中S100P的表达。将处于对数生长期的A549细胞分为两组:对照组(感染含有无意义链Plvx-shRNA2-shNC的慢病毒载体)和敲低组(感染含有Plvx-shRNA2-S100P的慢病毒载体),然后利用qRT-PCR检测A549细胞中S100P基因的水平。MTT及克隆形成实验检测敲低S100P后A549细胞的增殖能力,流式细胞术检测敲低S100P后A549细胞的凋亡水平,Western blot检测增殖相关蛋白PCNA及凋亡相关蛋白cleaved Caspase-8、Caspase-8、cleaved PARP、PARP及过氧化物酶体增殖物激活受体γ(PPAR-γ)蛋白的表达情况。结果人肺癌细胞系中S100P基因的相对表达量高于人正常肺上皮细胞(P<0.01)。成功感染慢病毒后,敲低组A549细胞S100P基因的相对表达量较对照组明显降低(P<0.01)。与对照组相比,敲低组A549细胞增殖能力显著下降(P<0.01),凋亡率显著上升(P<0.01)。与对照组相比,敲低组A549细胞中PCNA的蛋白含量显著降低(P<0.01),Caspase-8和PARP的蛋白含量无明显变化,cleaved Caspase-8、cleaved PARP及PPAR-γ的蛋白含量显著增高(P<0.01)。结论S100P在肺癌细胞中表达上调,沉默S100P后可通过抑制PPAR-γ信号通路,从而抑制肺癌细胞的增殖并促进肺癌细胞的凋亡。

Objective To investigate the effect and mechanism of S100P on proliferation and apoptosis of human lung cancer cells.Methods The expression of S100P was detected in human normal lung epithelial cells and four human lung cancer cells A549,95D,H1975 and LTEP-a-2 by qRT-PCR.A549 cells in the logarithmic growth phase were divided into two groups:control group and knockdown group,and the cells were infected with lentiviral vectors containing nonsense strands Plvx-shRNA2-shNC and lentiviral vectors containing Plvx-shRNA2-S100P,respectively.Then qRT-PCR was used to verify the level of S100P gene in A549 cells.MTT and clonogenesis experiments were used to detect the proliferation ability of A549 cells after knocking down S100P.Flow cytometry was used to detect the apoptosis of A549 cells after knocking down S100P.Western blot was used to detect the expression of proliferation-related protein PCNA,apoptosis-associated proteins cleaved Caspase-8,Caspase-8,cleaved PARP and PARP proteins,and peroxisome proliferator-activated receptorγ(PPAR-γ)protein.Results The relative expression of S100P in human lung cancer cell lines was higher than that of human normal lung epithelial cells(P<0.01).After successful lentivirus infection,the relative expression of S100P gene in A549 cells in knockdown group was significantly lower than that in control group(P<0.01).Compared with control group,the proliferation capacity of A549 cells was significantly decreased and the apoptosis rate was significantly increased in knockdown group(P<0.01).Compared with control group,the protein level of PCNA in A549 cells in knockdown group was significantly reduced(P<0.01),the protein levels of Caspase-8 and PARP showed no significant change,and the protein levels of cleaved Caspase-8,cleaved PARP and PPAR-γwere significantly increased(P<0.01).Conclusion The expression of S100P is upregulated in lung cancer cells,and S100P silencing can inhibit the proliferation of lung cancer cells and promote the apoptosis by inhibiting the PPAR-γsignaling pathway.