机构地区:[1]苏州大学附属第三医院肿瘤生物诊疗中心,江苏省肿瘤免疫治疗工程技术研究中心,苏州大学细胞治疗研究院,常州213003 [2]苏州大学附属第三医院临床营养科,常州213003
出 处:《中华实验外科杂志》2024年第2期209-213,共5页Chinese Journal of Experimental Surgery
基 金:国家自然科学基金(32270955、81972869);江苏省重点研发计划专项资金项目临床前沿技术(BE2022719);江苏省医学重点学科(YXZDXK202236);江苏省老年医学临床技术应用研究培养对象(LR2021042);常州市卫生健康委员会科技重大专项(ZD202102);常州市卫生健康委员会青年人才发展计划(2020-233)(CZQM2020044);江苏省研究生科研与实践创新计划项目(KYXC23_3265)。
摘 要:目的探讨人白细胞介素-1受体Ⅱ(IL-1R2)对胃癌细胞生物学行为的影响及其对血管的影响。方法用反转录-聚合酶链反应(RT-PCR)、蛋白免疫印迹法检测胃癌细胞株(AGS、MGC-803、BGC-823、SGC-7901、HGC-27、HSC-39)及人胃黏膜细胞(GES-1)中IL-1R2的mRNA、蛋白水平, 选择MGC-803、BGC-823构建下调及过表达IL-1R2细胞模型。细胞计数试剂盒(CCK-8)细胞增殖、划痕实验、Transwell侵袭实验明确IL-1R2对胃癌细胞增殖、迁移、侵袭能力的影响。细胞周期及凋亡实验探究IL-1R2对细胞周期及凋亡的作用。RT-PCR检测下调及过表达IL-1R2的MGC-803、BGC-823细胞中血管内皮生长因子(vascular endothelial growth factor, VEGF) mRNA水平。下调、过表达IL-1R2细胞组及其对照组与血管内皮细胞(HMEC-1)共培养, 明确IL-1R2对HMEC-1增殖、迁移及VEGF、IL-6 mRNA水平的影响。两组间差异表达采用t检验方法比较;多组间采用方差分析(ANOVA)和Tukey trend检验。结果 RT-PCR、蛋白免疫印迹实验证明胃癌细胞株(AGS、MGC-803、BGC-823、SGC-7901、HGC-27)中IL-1R2的mRNA及蛋白水平较GES-1显著升高, 差异有统计学意义(mRNA:1.944±0.310、4.777±0.320、2.214±0.100、3.235±0.121、4.809±0.190比1.000±0.118, t=2.971、11.54、10.01、16.06、18.81, P<0.05, P<0.01;蛋白:2.010±0.151、4.445±0.362、1.988±0.113、2.821±0.083、4.777±0.280比1.000±0.087, t=5.973、11.54、10.01、16.06、18.81, P<0.01, P<0.01)。细胞增殖、划痕实验、Transwell侵袭实验提示IL-1R2可促进胃癌细胞的增殖(下调组48、72、96 h:0.298±0.028比0.438±0.030、0.418±0.030比0.592±0.026、0.599±0.025比0.779±0.014, t=3.415、4.396、6.193, P<0.05, P<0.01;过表达组72、96 h:0.745±0.051比0.562±0.009、0.886±0.025比0.790±0.015, t=3.577、3.306, P<0.05, P<0.01)、迁移(下调组:0.620±0.070比1.000±0.080, t=3.587, P<0.05;过表达组:1.643±0.116比1.000±0.067, t=4.810, P<0.01)和侵袭(下调组:76.670±4.631比144.300±4.978, t=9Objective To investigate the effect of human interleukin-1 receptorⅡ(IL-1R2)on biological behavior of gastric cancer cells and its effect on blood vessels.Methods mRNA and protein levels of IL-1R2 in various gastric cancer cell lines(AGS,MGC-803,BGC-823,SGC-7901,HGC-27,HSC-39)and human gastric mucosa cells(GES-1)were detected by reverse transcriptase-polymerase chain reaction(RT-PCR)and Western blotting.MGC-803 and BGC-823 were selected to construct down-regulated and overexpressed IL-1R2 cell models.The effects of IL-1R2 on the proliferation,migration and invasion of gastric cancer cells were determined by cell counting kit-8(CCK-8)assay,scratch test and Transwell invasion test.The effects of IL-1R2 on cell cycle and apoptosis were investigated by cell cycle and apoptosis experiment.Vascular endothelial growth factor(VEGF)mRNA levels in MGC-803 and BGC-823 cells with down-regulated or overexpressed IL-1R2 were detected by RT-PCR.The cells in the down-regulated and overexpressed IL-1R2 groups and the control group were co-cultured with vascular endothelial cells(HMEC-1)to determine the effects of IL-1R2 on the proliferation and migration of HMEC-1 and the mRNA levels of VEGF and IL-6.The difference expression between the two groups was compared by t test.Analysis of variance(ANOVA)and Tukey trend test were used among the groups.Results RT-PCR and Western blotting showed that the mRNA and protein levels of IL-1R2 in gastric cancer cell lines(AGS,MGC-803,BGC-823,SGC-7901,HGC-27)were significantly higher than GES-1(mRNA:1.944±0.310,4.777±0.320,2.214±0.100,3.235±0.121,4.809±0.190 vs.1.000±0.118,t=2.971,11.54,10.01,16.06,18.81,P<0.05,P<0.01;Protein:2.010±0.151,4.445±0.362,1.988±0.113,2.821±0.083,4.777±0.280 vs.1.000±0.087,t=5.973,11.54,10.01,16.06,18.81,P<0.01,P<0.01).CCK-8 assay,scratch test and Transwell invasion test suggested that IL-1R2 could promote the proliferation of gastric cancer cells(48,72,96 h in down-regulated group:0.298±0.028 vs.0.438±0.030,0.418±0.030 vs.0.592±0.026,0.599±0.025 vs.
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