MMPs抑制剂对结肠癌细胞凋亡、免疫功能及炎性因子的影响

Effects of MMPs Inhibitors on Apoptosis,Immune Function and Inflammatory Factors of Colon Cancer Cells

目的:探究基质金属蛋白酶(Matrix metalloproteinases,MMPs)抑制剂对结肠癌细胞凋亡、免疫功能及炎性因子的影响。方法:取SW480结肠癌细胞,随机分为低表达组(将MMPs抑制剂慢病毒质粒与SW480细胞混合培养),空白对照组(SW480细胞与慢病毒包装质粒混合培养)。采用流式细胞法、免疫细胞化学法、酶联免疫吸附(Enzyme-linked immunosorbent assay,ELISA)法、实时荧光定量PCR(real-time fluorescence quantitative PCR,qRT-PCR)法、Hoechst 33258荧光染色法检测SW480细胞凋亡,TNF-α、IL-6蛋白阳性表达、免疫球蛋白表达水平、MMP-9 mRNA表达量。结果:空白对照组与低表达组相比较,低表达组SW480细胞内MMP-9 mRNA表达量显著下降(P<0.05),说明转染成功。与空白对照组相比较,低表达组SW480细胞凋亡数量显著上升(P<0.05)。低表达组细胞的细胞核出现核固缩的概率显著高于空白对照组(P<0.05)。与空白对照组相比较,低表达组SW480细胞IL-2表达水平显著升高(P<0.05),IL-4、TGF-β1、TIM-1表达水平显著降低(P<0.05)。SW480细胞内的MMP-9 mRNA与IL-2呈现负相关性(r=-0.723,P=0.007),MMP-9 mRNA与IL-4、TGF-β1及TIM-1均呈现负相关性(均P<0.05)。空白对照组SW480细胞中TNF-α、IL-6蛋白阳性数最高,低表达组SW480细胞中TNF-α、IL-6蛋白阳性数最低,与空白对照组相比较,低表达组SW480细胞中TNF-α、IL-6阳性数显著下降(均P<0.05)。结论:MMPs抑制剂可促进SW480细胞凋亡,改善免疫功能,降低炎性介质的表达水平。

Objective:To explore the effects of Matrix metalloproteinases(MMPs)inhibitors on apoptosis,immune function and inflammatory factors of colon cancer cells.Methods:SW480 colon cancer cells were randomly divided into low expression group(mixed culture of MMPs inhibitor lentivirus plasmid and SW480 cells)and blank control group(mixed culture of SW480 cells and lentivirus packaging plasmid).Flow cytometry,immunocytochemistry,Enzyme-Linked immunosorbent assay were used.SW480 cell apoptosis was detected by ELISA,real-time fluorescence quantitative PCR(QRT-PCR)and Hoechst 33258 fluorescence staining.TNF-α,IL-6 protein expression,immunoglobulin expression level,MMP-9 mRNA expression level.Results:Compared with the blank control group and the low expression group,the expression level of MMP-9 mRNA in SW480 cells in the low expression group was significantly decreased(P<0.05),indicating that the transfection was successful.Compared with blank control group,the number of apoptosis of SW480 cells in low expression group was significantly increased(P<0.05).The probability of nuclear pyknosis in low expression group was significantly higher than that in blank control group(P<0.05).Compared with blank control group,the expression level of IL-2 in SW480 cells in low expression group was significantly increased(P<0.05),and the expression levels of IL-4,TGF-β1 and TIM-1 in SW480 cells were significantly decreased(P<0.05).There was a negative correlation between MMP-9 mRNA and IL-2 in SW480 cells(r=-0.723,P=0.007),MMP-9 mRNA and IL-4,TGF-β1 and TIM-1 showed negative correlation(all P<0.05).The positive number of TNF-αand IL-6 protein in SW480 cells in the blank control group was the highest,and the positive number of TNF-αand IL-6 protein in SW480 cells in the low expression group was the lowest.Compared with the blank control group,the positive number of TNF-αand IL-6 protein in SW480 cells in the low expression group was significantly decreased(all P<0.05).Conclusion:MMPs inhibitors can promote apoptosis of SW480 cells,improve immune function,and reduce the expression level of inflammatory mediators.