摘要
该文探讨了幽门螺杆菌(Helicobacter pylori,HP)炎症微环境对结肠癌SW620细胞发生上皮-间质转化(epithelial mesenchymal transformation,EMT)的影响。使用ELISA方法检测HP干预U937细胞后上清中炎症因子MIF、IL-1β等的变化;采用RT-PCR方法检测MIF、IL-1β、NF-κB基因水平的变化;细胞划痕实验检测炎症上清处理SW620细胞后SW620细胞的侵袭迁移能力;CCK-8方法检测HP处理U937细胞后炎症上清对SW620细胞增殖活性的影响;免疫荧光实验检测炎症上清处理SW620细胞后SW620细胞EMT相关蛋白Vimentin的荧光变化;Western blot实验检测脂多糖(LPS)上清干预SW620细胞后EMT相关蛋白Vimentin、E-cadherin和N-cadherin的变化;Western blot实验检测HP干预U937细胞后NF-κB蛋白,炎症上清处理SW620细胞后EMT相关蛋白Vimentin、E-cadherin和N-cadherin的变化。结果显示,HP干预U937细胞后引起巨噬细胞移动抑制因子(MIF)、白细胞介素-1β(IL-1β)和NF-κB等相关炎症因子及基因表达升高;CCK-8结果表明HP干预U937细胞后,U937细胞毒性增强;划痕实验结果表明在12 h、24 h、36 h时,随着时间点的推移,SW620细胞的侵袭迁移能力有所增强;免疫荧光实验结果显示SW620细胞Vimentin绿色点状聚集显著增加;Western blot结果显示在16 h、24 h时间点U937细胞NF-κB蛋白表达明显升高;LPS上清干预SW620细胞后Vimentin蛋白表达无明显变化,N-cadherin蛋白24 h组表达减少,E-cadherin蛋白增多,HP炎症微环境上清干预SW620细胞后12 h、24 h、36 h时,随着时间点推移SW620细胞Vimentin蛋白表达显著增加,E-cadherin和N-cadherin蛋白表达显著减少。该实验研究结果表明,幽门螺杆菌致炎症微环境可以促进SW620细胞发生EMT。
This article explores the effects of HP inflammation microenvironment on EMT of colon cancer SW620 cells.The ELISA method was used to detect the changes of inflammatory factors MIF and IL-1βin the supernatant after HP intervention in U937 cells.RT-PCR was used to detect the changes of MIF,IL-1β,NF-κB gene levels.Scratch assay was used to detect the invasion and migration ability of SW620 cells after treatment with inflammatory supernatant.Effect of inflammatory supernatant on proliferation of SW620 cells after HP-treated U937 cells was observed by CCK-8 method.Changes of EMT-related proteins Vimentin,E-cadherin and N-cadherin in SW620 cells treated with LPS supernatant were detected by Western blot assay.Immunofluorescence assay was used to detect the fluorescence changes of EMT-related protein Vimentin in SW620 cells treated with inflammatory supernatant.Western blot was used to detect the changes of NF-κB protein in U937 cells treated with HP,and EMTrelated protein Vimentin,E-cadherin and N-cadherin in SW620 cells treated with inflammatory supernatant.The results showed that HP could increase the expression of MIF,IL-1βand NF-κB in U937 cells.Scratch test showed that the invasion and migration ability of SW620 cells increased with the passage of time points at 12 h,24 h and 36 h.Immunofluorescence test showed that Vimentin green dot aggregation of SW620 cells increased significantly.Western blot analysis showed that the expression of NF-κB protein in U937 cells was significantly increased at 16 h and 24 h.There was no significant change in the expression of Vimentin protein after intervention of SWPS cells in LPS supernatant,the expression of N-cadherin protein in 24 h group was decreased and E-cadherin protein was increased.The expression of Vimentin protein in SW620 cells increased significantly and the expression of E-cadherin and N-cadherin protein decreased significantly at 12 h,24 h and 36 h after intervention of HP inflammatory microenvironment supernatant.The results of this experimental study indicated that H.pylori-induced microenvironment can promote the occurrence of EMT in SW620 cells.
作者
宋光永
张晓荣
李梦俊
高艳萍
SONG Guangyong;ZHANG Xiaorong;LI Mengjun;GAO Yanping(Shanxi Medical University,Taiyuan 030001,China;Fenyang College of Shanxi Medical University,Fenyang 032200,China)
出处
《中国细胞生物学学报》
CAS
CSCD
2019年第11期2152-2159,共8页
Chinese Journal of Cell Biology
基金
山西省自然科学基金(批准号:201601D102077)
山西省青年科技研究基金(批准号:201601D021108)资助的课题.
关键词
幽门螺杆菌
炎症微环境
上皮–间质转化
SW620细胞
Helicobacter pylori
inflammatory microenvironment
epithelial-mesenchymal transformation
SW620 cells
