幽门螺杆菌感染激活NF-κB信号通路促进胃癌SGC-7901细胞炎症因子释放
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摘要
为了探讨幽门螺杆菌对胃癌SGC-7901细胞炎症因子释放的影响,本研究将幽门螺杆菌感染SGC-7901细胞后,采用细胞计数盒(CCK-8)检测SGC-7901细胞活力,酶联免疫吸附实验(ELISA)检测炎症因子TNF-α、IL-1β以及IL-8的水平,Real-time PCR检测细胞TNF-α、IL-1β以及IL-8 m RNA的表达,蛋白免疫印迹法(Western blotting)检测NF-κB信号通路相关蛋白NF-κB p65蛋白表达以及IκBα磷酸化水平。研究结果表明,幽门螺杆菌感染后,SGC-7901细胞活力显著增加;幽门螺杆菌感染明显上调SGC-7901细胞TNF-α、IL-1β以及IL-8 mRNA的表达;本研究还进一步发现幽门螺杆菌感染显著增加SGC-7901细胞TNF-α、IL-1β以及IL-8的水平;此外,幽门螺杆菌处理的SGC-7901细胞,其NF-κB p65的蛋白表达以及IκBα磷酸化水平均显著上调。本研究的结论初步表明,幽门螺杆菌感染促进胃癌SGC-7901细胞炎症因子的释放,其机制可能涉及激活NF-κB信号通路。
In order to investigate the effect of Helicobacter pylori on the release of inflammatory factors in gastric cancer SGC-7901 cells, we infected SGC-7901 cells with Helicobacter pylori. Then the SGC-7901 cell viability were detected by Cell Counting Kits(CCK-8), and the levels of inflammatory factors TNF-α, IL-1β and IL-8 were detected by enzyme-linked immunosorbent assay(ELISA). The expressions of TNF-α, IL-1β, IL-8 m RNA were detected by real-time PCR, and NF-κB pathway related protein, including NF-κB p65 protein expression and IκBα phosphorylation, were detected by western blotting. The results showed that the cell viability in SGC-7901 cells was significantly increased after H. pylori infection. H. pylori infection obviously increased the expressions of TNF-α、IL-1β and IL-8 mRNA in SGC-7901 cells. Furthermore, H. pylori infection markedly elevated the levels of TNF-α、IL-1β and IL-8 in SGC-7901 cells as well. In addition, the protein expressions of NF-κB p65 and the phosphorylation of IκBα in SGC-7901 cells were obviously upregulated after H. pylori infection. The conclusions of this study preliminary showed that H. pylori infection might promote the release of inflammatorycytokine in gastric cancer SGC-7901 cells and the potential mechanism might be involved in activation of NF-κB signaling pathway.
In order to investigate the effect of Helicobacter pylori on the release of inflammatory factors in gastric cancer SGC-7901 cells, we infected SGC-7901 cells with Helicobacter pylori. Then the SGC-7901 cell viability were detected by Cell Counting Kits(CCK-8), and the levels of inflammatory factors TNF-α, IL-1β and IL-8 were detected by enzyme-linked immunosorbent assay(ELISA). The expressions of TNF-α, IL-1β, IL-8 m RNA were detected by real-time PCR, and NF-κB pathway related protein, including NF-κB p65 protein expression and IκBα phosphorylation, were detected by western blotting. The results showed that the cell viability in SGC-7901 cells was significantly increased after H. pylori infection. H. pylori infection obviously increased the expressions of TNF-α、IL-1β and IL-8 mRNA in SGC-7901 cells. Furthermore, H. pylori infection markedly elevated the levels of TNF-α、IL-1β and IL-8 in SGC-7901 cells as well. In addition, the protein expressions of NF-κB p65 and the phosphorylation of IκBα in SGC-7901 cells were obviously upregulated after H. pylori infection. The conclusions of this study preliminary showed that H. pylori infection might promote the release of inflammatorycytokine in gastric cancer SGC-7901 cells and the potential mechanism might be involved in activation of NF-κB signaling pathway.
引文
Ang T.L.,and Fock K.M.,2014,Clinical epidemiology of gastric cancer,Singapore Med.J.,55(12):621-628
C ao Z.,Zhu J.,Lu P.,and Shen J.,2015,The investigation of the relationship between HSP70 and TLR-NF-κBduring the process of acute lung injury,Linchuang Yu Bingli Zazhi(International Journal of Pathology and Clinical Medicine),35(3):406-411(曹征,朱健,陆平,盛净,2015,HSP70与TLR-NF-κB信号途径在急性肺损伤发病机制中作用的实验研究,临床与病理杂志,35(3):406-411)
Ding J.N.,Xiang L.J.,Yu L.R.,and Chen F.Z.,2015,Analysis of Helicobacter pylori infection in patients with upper gastrointestinal disease,Zh onghua Yiyuan Ganranxue Zazhi(Chinese Journal of Nosocomiology),24(19):4356-4364(丁金囡,项利娟,余利荣,陈方志,2015,上消化道疾病患者幽门螺杆菌感染调查分析,中华医院感染学杂志,24(19):4356-4364)
De Simone V.,Franze E.,and Ronchetti G.,2015,Th17-type cytokines,IL-6 and TNF-alpha synergistically activate STAT3and NF-κB to promote colorectal cancer cell growth,Oncogene,34(27):3493-3503
Fernandes J.V.,Cobucci R.N.,Jatoba C.A.,Fernandes T.A.,de Azevedo J.W.,and de Araújo J.M.,2015,The role of the mediators of inflammation in cancer development,Pathol.Oncol.Res.,21(3):527-534
Guggenheim D.E.,and Shah M.A.,2013,Gastric cancer epidemiology and risk factors,J.Surg.Oncol.,107(3):230-236
Guo Y.X.,Wang Y.J.,and Zhu Y.,2014,The correlational study of gastric carcinoma and Helicobacter pylori infection,Zhonghua Yixue Ganranxue Zazhi(Chinese Journal of Nosocomiology),24(7):1700-1704(郭云霞,王郁杰,朱莹,2014,幽门螺杆菌感染与胃癌的相关性探讨,中华医院感染学杂志,24(7):1700-1704)
Hatakeyama M.,2014,%Helicobacter pylori CagA and gastric cancer:a paradigm for hit-and-run carcinogenesis,Cell Host Microbe,5(3):306-316
Jin R.,Xia Y.,and Chen Q.,2016,Da0324,an inhibitor of nuclear factor-kappaB activation,demonstrates selective antitumor activity on human gastric cancer cells,Drug Des.Devel.Ther.,(10):979-995
Karimi P.,Islami F.,Anandasabapathy S.,Freedman N.D.,and Kamangar F.,2014,Gastric cancer:descriptive epidemiology,risk factors,screening,and prevention,Cancer Epidemiol Biomarkers Prev.,23(5):700-713
Merchant J.L.,2017,NF-kappaB mediated transcription of DARPP-32 prevents Helicobacter pylori-induced cell death,Gut,66(5):761-762
Nie Y.,Wu K.,Yu J.,Cai X.,Shang Y.,Zhou J.,Pan K.,Sun L.,Fang J.,Yuan Y.,You W.,and Fan D.,2017,A global burden of gastric cancer:the major impact of China,Expert.Rev.Gastroenterol Hepatol.,11(7):651-661
Sanchez-Zauco N.,Torres J.,Gomez A.,Camorlinga-Ponce M.,Mu觡oz-Pérez L.,Herrera-Goepfert R.,Medrano-Guzmán R.,Giono-Cerezo S.,and Maldonado-Bernal C.,2017,Circulating blood levels of IL-6,IFN-gamma,and IL-10 as potential diagnostic biomarkers in gastric cancer:a controlled study,BMC Cancer,17(1):384
Wang Y.,and Xu H.F.,2015,Serum inflammatory cytokine levels in patients with gastric cnacer,Xiandai Zhongliu Yixue(Journal of Modern Oncology)23(21):3136-3139(万瑶,徐海峰,2015,胃癌患者血清炎性因子水平的检验分析,现代肿瘤医学,23(21):3136-3139)
Wu X.W.,Ji H.Z.,Yang M.F.,Wu L.,and Wang F.Y.,2015,Helicobacter pylori infection and inflammatory bowel disease in Asians:a meta-analysis,World J.Gastroenterol.,21(15):4750
Xia Z.B.,Meng F.R.,Fang Y.X.,Wu X.,Zhang C.W.,Liu Y.,Liu D.,Li G.Q.,Feng F.B.,and Qiu H.Y.,2018,Inhibition of NF-KappaB signaling pathway induces apoptosis and suppresses proliferation and angiogenesis of human fibroblast-like synovial cells in rheumatoid arthritis,Medicine(Baltimore),97(23):e10920
Zuo T.T.,Zhen R.S.,Zeng H.M.,Zhang S.W.,and Chen W.Q.,Epidemiology of stomach cancer in China,Zhongguo Zhongliu Linchuang(Chinese Journal of Clinical Oncology),44(1):52-58(左婷婷,郑荣寿,曾红梅,张思维,陈万青,2017,中国胃癌流行病学现状,中国肿瘤临床,44(1):52-58)
Zou Z.,Zhao L.,and Su S.,2018,The plasma levels of 12 cytokines and growth factors in patients with gastric cancer,Medicine(Baltimore),97(19):e0413
C ao Z.,Zhu J.,Lu P.,and Shen J.,2015,The investigation of the relationship between HSP70 and TLR-NF-κBduring the process of acute lung injury,Linchuang Yu Bingli Zazhi(International Journal of Pathology and Clinical Medicine),35(3):406-411(曹征,朱健,陆平,盛净,2015,HSP70与TLR-NF-κB信号途径在急性肺损伤发病机制中作用的实验研究,临床与病理杂志,35(3):406-411)
Ding J.N.,Xiang L.J.,Yu L.R.,and Chen F.Z.,2015,Analysis of Helicobacter pylori infection in patients with upper gastrointestinal disease,Zh onghua Yiyuan Ganranxue Zazhi(Chinese Journal of Nosocomiology),24(19):4356-4364(丁金囡,项利娟,余利荣,陈方志,2015,上消化道疾病患者幽门螺杆菌感染调查分析,中华医院感染学杂志,24(19):4356-4364)
De Simone V.,Franze E.,and Ronchetti G.,2015,Th17-type cytokines,IL-6 and TNF-alpha synergistically activate STAT3and NF-κB to promote colorectal cancer cell growth,Oncogene,34(27):3493-3503
Fernandes J.V.,Cobucci R.N.,Jatoba C.A.,Fernandes T.A.,de Azevedo J.W.,and de Araújo J.M.,2015,The role of the mediators of inflammation in cancer development,Pathol.Oncol.Res.,21(3):527-534
Guggenheim D.E.,and Shah M.A.,2013,Gastric cancer epidemiology and risk factors,J.Surg.Oncol.,107(3):230-236
Guo Y.X.,Wang Y.J.,and Zhu Y.,2014,The correlational study of gastric carcinoma and Helicobacter pylori infection,Zhonghua Yixue Ganranxue Zazhi(Chinese Journal of Nosocomiology),24(7):1700-1704(郭云霞,王郁杰,朱莹,2014,幽门螺杆菌感染与胃癌的相关性探讨,中华医院感染学杂志,24(7):1700-1704)
Hatakeyama M.,2014,%Helicobacter pylori CagA and gastric cancer:a paradigm for hit-and-run carcinogenesis,Cell Host Microbe,5(3):306-316
Jin R.,Xia Y.,and Chen Q.,2016,Da0324,an inhibitor of nuclear factor-kappaB activation,demonstrates selective antitumor activity on human gastric cancer cells,Drug Des.Devel.Ther.,(10):979-995
Karimi P.,Islami F.,Anandasabapathy S.,Freedman N.D.,and Kamangar F.,2014,Gastric cancer:descriptive epidemiology,risk factors,screening,and prevention,Cancer Epidemiol Biomarkers Prev.,23(5):700-713
Merchant J.L.,2017,NF-kappaB mediated transcription of DARPP-32 prevents Helicobacter pylori-induced cell death,Gut,66(5):761-762
Nie Y.,Wu K.,Yu J.,Cai X.,Shang Y.,Zhou J.,Pan K.,Sun L.,Fang J.,Yuan Y.,You W.,and Fan D.,2017,A global burden of gastric cancer:the major impact of China,Expert.Rev.Gastroenterol Hepatol.,11(7):651-661
Sanchez-Zauco N.,Torres J.,Gomez A.,Camorlinga-Ponce M.,Mu觡oz-Pérez L.,Herrera-Goepfert R.,Medrano-Guzmán R.,Giono-Cerezo S.,and Maldonado-Bernal C.,2017,Circulating blood levels of IL-6,IFN-gamma,and IL-10 as potential diagnostic biomarkers in gastric cancer:a controlled study,BMC Cancer,17(1):384
Wang Y.,and Xu H.F.,2015,Serum inflammatory cytokine levels in patients with gastric cnacer,Xiandai Zhongliu Yixue(Journal of Modern Oncology)23(21):3136-3139(万瑶,徐海峰,2015,胃癌患者血清炎性因子水平的检验分析,现代肿瘤医学,23(21):3136-3139)
Wu X.W.,Ji H.Z.,Yang M.F.,Wu L.,and Wang F.Y.,2015,Helicobacter pylori infection and inflammatory bowel disease in Asians:a meta-analysis,World J.Gastroenterol.,21(15):4750
Xia Z.B.,Meng F.R.,Fang Y.X.,Wu X.,Zhang C.W.,Liu Y.,Liu D.,Li G.Q.,Feng F.B.,and Qiu H.Y.,2018,Inhibition of NF-KappaB signaling pathway induces apoptosis and suppresses proliferation and angiogenesis of human fibroblast-like synovial cells in rheumatoid arthritis,Medicine(Baltimore),97(23):e10920
Zuo T.T.,Zhen R.S.,Zeng H.M.,Zhang S.W.,and Chen W.Q.,Epidemiology of stomach cancer in China,Zhongguo Zhongliu Linchuang(Chinese Journal of Clinical Oncology),44(1):52-58(左婷婷,郑荣寿,曾红梅,张思维,陈万青,2017,中国胃癌流行病学现状,中国肿瘤临床,44(1):52-58)
Zou Z.,Zhao L.,and Su S.,2018,The plasma levels of 12 cytokines and growth factors in patients with gastric cancer,Medicine(Baltimore),97(19):e0413