SDF-1／CXCR7对胃癌SGC-7901细胞合成及分泌炎症因子的影响

Effect of SDF-1/CXCR7 on inflammatory cytokine synthesis and secretion in gastric cancer SGC-7901 cells

目的探讨SDF-1/CXCR7对胃癌SGC-7901细胞合成及分泌炎症因子的影响及其机制。方法采用CXCR7的shRNA慢病毒表达载体感染人胃癌SGC-7901细胞,Real-time PCR及Western blot检测CXCR7mRNA和蛋白表达;应用SDF-1干预,分为4组:阴性对照组(LV-shRNA-NC),LV-shRNA-NC+SDF-1组,CXCR7沉默组(LV-shRNACXCR7),LV-shRNA-CXCR7+SDF-1组。采用Real-time PCR检测炎症因子TNF-α、IL-1β、IL-6、IL-8的mRNA表达;采用ELISA检测细胞上清液TNF-α、IL-1β、IL-6、IL-8的蛋白含量;采用Western blot检测NF-κB通路的变化。结果 1Real-time PCR及Western blot结果提示,CXCR7沉默组SGC-7901细胞CXCR7mRNA及蛋白的表达水平较阴性对照组显著降低(P均<0.01),阴性对照组与空白对照组间无明显差异。2与LV-shRNA-NC组相比,LV-shRNANC+SDF-1组IL-6及IL-8的mRNA表达及分泌明显增加(P<0.01),LV-shRNA-CXCR7组IL-6及IL-8mRNA表达及分泌减少(P<0.05);与LV-shRNA-NC+SDF-1组相比,LV-shRNA-CXCR7+SDF-1组IL-6及IL-8mRNA表达及分泌明显减少(P<0.01)。而TNF-α及IL-1β的mRNA表达及分泌在4组间无明显差异。3NF-κB p65核内表达、t-IκBα及p-IκBα表达在4组间均无明显差异。结论 SDF-1可能通过CXCR7促进SGC-7901细胞合成及分泌炎症因子IL-6及IL-8,但不依赖于NF-κB通路。

Objective To investigate the effect of SDF-1/CXCR7 on inflammatory cytokine synthesis and secretion in gastric cancer SGC-7901 cells. Methods CXCR7 gene in SGC-7901 cells was silenced by shRNA lentiviral vector and the expression of CXCR7 was detected using Western blot and Real-time PCR. There were four groups as follows.. LV-shRNA-NC, LV-shRNA-NC＋ SDF-1, LV-shRNA-CXCRT, and LV-shRNA-CXCR7-F SDF-1 groups. Real-time PCR was used to detect the mRNA expressions of TNF-α, IL-1β, IL-6 and IL-8. ELISA was used to detect the protein levels of TNF-α, IL-1β, IL-6 and IL-8 in the culture supernatant. Western blot was used to detect the protein expressions of NF-kB pathway. Results （1） Transfection of SGC-7901 cells with CXCR7-shRNA lentiviral vector resulted in a significantly decreased expression of CXCR7 at both mRNA and protein levels （all P〈 0.01）. （3） Compared with those in LV-shRNA-NC group, IL-6 and IL-8 mRNA expressions and protein levels in the culture supernatant were increased in LV-shRNA-NC＋ SDF-1 group （P〈0.01） and decreased in LV-shRNA- CXCR7 group （P〈0.05）. Compared with those in LV-shRNA-NC＋SDF-1 group, the expressions of IL-6 and IL-8 at mRNA and protein levels in the culture supernatant were significantly cut down in LV-shRNA-CXCR7＋SDF-1 group （P〈0.01）. However, the expressions of TNF-α and IL-1β at mRNA and protein levels in the culture supernatant were not significantly changed by SDF-1 and CXCR7 shRNA. （3） The protein expressions of nuclear NF-kB p65, t-IkBα and p-IkBα exhibited no significant differences among the four groups. Conclusion SDF-1/CXCR7 can promote the synthesis and secretion of inflammatory cytokines IL-6 and IL-8 in gastric cancer SGC-7901 ceils through an NF-kB-independent pathway.