SOX9基因下调JAK2/STAT3信号诱导肺癌细胞凋亡及降低免疫逃逸相关因子表达的研究
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摘要
目的:探讨SOX9基因对肺癌细胞凋亡、免疫逃逸相关因子HIF-1α和VEGF表达及JAK2/STAT3信号通路的影响。方法:以脂质体Lip-2000为载体,将针对SOX9特异性靶点的siRNA转染肺腺癌A549细胞,Western blot检测SOX9的蛋白表达; CCK8法检测细胞活力;流式细胞术检测细胞凋亡率; DCFH-DA检测ROS水平; Western blot检测免疫逃逸相关因子HIF-1α、VEGF及JAK2/STAT3信号通路p-JAK2、p-STAT3和下游相关基因survivin的蛋白表达。结果:转染SOX9的siRNA的肺癌细胞SOX9的表达明显降低;与阴性对照组比较,si-SOX9组细胞活力显著降低,细胞凋亡率显著升高,ROS水平显著升高,HIF-1α、VEGF、p-JAK2、p-STAT3和survivin的蛋白表达均显著降低。结论:siRNA抑制SOX9基因可降低肺癌细胞活力,诱导细胞凋亡,降低免疫逃逸相关因子HIF-1α和VEGF表达,机制可能与提高细胞ROS水平和下调JAK2/STAT3信号通路有关。
Objective: To investigate the effect of SOX9 gene on the apoptosis of lung cancer cells,the expression of immune escape related factors HIF-1α and VEGF and the JAK2/STAT3 signaling pathway. Methods: Lip-2000 liposomes as the carrier,the SOX9 specific target siRNA was transfected into lung adenocarcinoma A549 cells,the expression of SOX9 protein was detected by Western blot; cell viability was measured by CCK8; cell apoptosis was detected by flow cytometry; the ROS level was detected through DCFHDA; Western blot was used to detect the protein expression of immune escape related factors HIF-1α,VEGF,JAK2/STAT3 signaling pathways p-JAK2,p-STAT3 and downstream related genes survivin. Results: The expression of SOX9 in lung cancer cells transfected with SOX9 siRNA was significantly reduced; compared with the negative control group,cell viability decreased significantly in si-SOX9 group,the apoptosis rate was significantly increased,ROS level was significantly increased,expression of HIF-1α,VEGF,p-JAK2,pSTAT3 and survivin protein were significantly decreased. Conclusion: Inhibition of SOX9 gene expression by siRNA can reduce lung cancer cell viability,induce cell apoptosis,and reduce the expression of HIF-1α and VEGF of immune escape related factors,which may be related to the improvement of ROS level and down regulation of JAK2/STAT3 signaling pathway.
Objective: To investigate the effect of SOX9 gene on the apoptosis of lung cancer cells,the expression of immune escape related factors HIF-1α and VEGF and the JAK2/STAT3 signaling pathway. Methods: Lip-2000 liposomes as the carrier,the SOX9 specific target siRNA was transfected into lung adenocarcinoma A549 cells,the expression of SOX9 protein was detected by Western blot; cell viability was measured by CCK8; cell apoptosis was detected by flow cytometry; the ROS level was detected through DCFHDA; Western blot was used to detect the protein expression of immune escape related factors HIF-1α,VEGF,JAK2/STAT3 signaling pathways p-JAK2,p-STAT3 and downstream related genes survivin. Results: The expression of SOX9 in lung cancer cells transfected with SOX9 siRNA was significantly reduced; compared with the negative control group,cell viability decreased significantly in si-SOX9 group,the apoptosis rate was significantly increased,ROS level was significantly increased,expression of HIF-1α,VEGF,p-JAK2,pSTAT3 and survivin protein were significantly decreased. Conclusion: Inhibition of SOX9 gene expression by siRNA can reduce lung cancer cell viability,induce cell apoptosis,and reduce the expression of HIF-1α and VEGF of immune escape related factors,which may be related to the improvement of ROS level and down regulation of JAK2/STAT3 signaling pathway.
引文
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[16]于秀艳,李铤,吴雪峰,等.异土木香内酯通过介导ROS产生及线粒体损伤诱导人宫颈癌Hela细胞凋亡[J].中国免疫学杂志,2016,32(10):1467-1471.Yu XY,Li T,Wu XF,et al.Isoalantolactone induces apoptosis in human cervical cancer Hela cells through ROS generation and Mitochondrial dysfunction[J].Chin J Immunol,2016,32(10):1467-1471.
[17]Seino M,Okada M,Shibuya K,et al.Differential contribution of ROS to resveratrol-induced cell death and loss of self-renewal capacity of ovarian cancer stem cells[J].Anticancer Res,2015,35(1):85-96.
[18]Wu L,Guo L,Liang Y,et al.Curcumin suppresses stem-like traits of lung cancer cells via inhibiting the JAK2/STAT3 signaling pathway[J].Oncol Reports,2015,34(6):3311-3317.
[19]Wan J,Wu W,Che Y,et al.Low dose photodynamic-therapy induce immune escape of tumor cells in a HIF-1αdependent manner through PI3K/Akt pathway[J].Int Immunopharmacol,2015,28(1):44-51.
[20]Sahin E,Baycu C,Koparal A T,et al.Resveratrol reduces IL-6and VEGF secretion from co-cultured A549 lung cancer cells and adipose-derived mesenchymal stem cells[J].Tumor Biol,2016,37(6):7573-7582.
[2]Luanpitpong S,Li J,Manke A,et al.SLUG is required for SOX9stabilization and functions to promote cancer stem cells and metastasis in human lung carcinoma[J].Oncogene,2016,35(22):2824-2833.
[3]Ma F,Ye H,He HH,et al.SOX9 drives WNT pathway activation in prostate cancer[J].J Clin Investigation,2016,126(5):1745-1758.
[4]李天客,陈中,包阳,等.RNA干扰SBF2对人口腔癌细胞株SCC15增殖,凋亡及侵袭的影响[J].中国老年学杂志,2016,36(21):5260-5262.Li TK,Chen Z,Bao Y,et al.Effects of RNA interference SBF2 on proliferation,apoptosis and invasion of human oral cancer cell line SCC15[J].Chin J Gerontol,2016,36(21):5260-5262.
[5]Mansoori B,Shotorbani S S,Baradaran B.RNA interference and its role in cancer therapy[J].Advanced Pharmaceutical Bulletin,2014,4(4):313-321.
[6]楚广民,张建波,孙淼淼.SOX9基因沉默对膀胱癌EJ细胞株生物学特性的影响及其机制研究[J].癌症进展,2018,16(3):286-289.Chu GM,Zhang JB,Sun MM.Effects of SOX9 silencing on biological characteristics of bladder cancer EJ cells and the mechanism[J].Oncol Prog,2018,16(3):286-289.
[7]Li Z,Li B,Niu L,et al.miR-592 functions as a tumor suppressor in human non-small cell lung cancer by targeting SOX9[J].Oncol Reports,2017,37(1):297-304.
[8]Zhang YJ,Xu F,Zhang YJ,et al.miR-206 inhibits non small cell lung cancer cell proliferation and invasion by targeting SOX9[J].Intern J Clin Exp Med,2015,8(6):9107-9113.
[9]Cui J,Xi H,Cai A,et al.Decreased expression of Sox7 correlates with the upregulation of the Wnt/β-catenin signaling pathway and the poor survival of gastric cancer patients[J].Int J Mol Med,2014,34(1):197-204.
[10]Wang J,Zhang S,Wu J,et al.Clinical significance and prognostic value of SOX7 expression in liver and pancreatic carcinoma[J].Mol Med Reports,2017,16(1):499-506.
[11]Shen F,Cai W S,Feng Z,et al.MiR-492 contributes to cell proliferation and cell cycle of human breast cancer cells by suppressing SOX7 expression[J].Tumor Biol,2015,36(3):1913-1921.
[12]Burdelski C,Bujupi E,Tsourlakis MC,et al.Loss of SOX9 expression is associated with PSA recurrence in ERG-positive and PTEN deleted prostate cancers[J].PLo S one,2015,10(6):e0128525.
[13]Wang H,Lian P,Zheng PS.SOX9,a potential tumor suppressor in cervical cancer,transactivates p21WAF1/CIP1 and suppresses cervical tumor growth[J].Oncotarget,2015,6(24):20711-20722.
[14]Wang X,Liu Y,Liu X,et al.miR-124 inhibits cell proliferation,migration and invasion by directly targeting SOX9 in lung adenocarcinoma[J].Oncol Reports,2016,35(5):3115-3121.
[15]Lei Z,Shi H,Li W,et al.miR 185 inhibits non small cell lung cancer cell proliferation and invasion through targeting of SOX9and regulation of Wnt signaling[J].Mol Med Reports,2018,17(1):1742-1752.
[16]于秀艳,李铤,吴雪峰,等.异土木香内酯通过介导ROS产生及线粒体损伤诱导人宫颈癌Hela细胞凋亡[J].中国免疫学杂志,2016,32(10):1467-1471.Yu XY,Li T,Wu XF,et al.Isoalantolactone induces apoptosis in human cervical cancer Hela cells through ROS generation and Mitochondrial dysfunction[J].Chin J Immunol,2016,32(10):1467-1471.
[17]Seino M,Okada M,Shibuya K,et al.Differential contribution of ROS to resveratrol-induced cell death and loss of self-renewal capacity of ovarian cancer stem cells[J].Anticancer Res,2015,35(1):85-96.
[18]Wu L,Guo L,Liang Y,et al.Curcumin suppresses stem-like traits of lung cancer cells via inhibiting the JAK2/STAT3 signaling pathway[J].Oncol Reports,2015,34(6):3311-3317.
[19]Wan J,Wu W,Che Y,et al.Low dose photodynamic-therapy induce immune escape of tumor cells in a HIF-1αdependent manner through PI3K/Akt pathway[J].Int Immunopharmacol,2015,28(1):44-51.
[20]Sahin E,Baycu C,Koparal A T,et al.Resveratrol reduces IL-6and VEGF secretion from co-cultured A549 lung cancer cells and adipose-derived mesenchymal stem cells[J].Tumor Biol,2016,37(6):7573-7582.