IL-6诱导小鼠骨髓中性粒细胞中MMP-9的生成与释放的实验

目的 探讨IL-6诱导小鼠骨髓中性粒细胞（PMN）生成和释放基质金属蛋白酶-9（MMP-9）的作用及激活途径。方法 分离小鼠骨髓PMN，将其分为空白对照组、IL-6刺激组、IL-6+DMSO组、IL-6+STAT3抑制剂Ⅷ组。实时荧光定量聚合酶链式反应检测细胞内MMP-9 mRNA表达，明胶酶谱检测PMN释放的MMP-9活性，Western blot检测PMN中STAT3蛋白磷酸化改变。结果 IL-6刺激后，PMN中MMP-9 mRNA水平明显升高（P=0.0050），明胶酶谱检测的释放到胞外MMP-9活性明显高于对照组（P=0.0087），IL-6刺激后PMN中STAT3磷酸化水平明显升高（P=0.0089）。在IL-6作用时加入特异性STAT3抑制剂Ⅷ后，MMP-9 mRNA水平、培养上清液中MMP-9活性以及PMN中磷酸化STAT3蛋白水平与IL-6刺激组相比均明显下降（P=0.0067, 0.048和0.0098）。结论 IL-6刺激骨髓PMN中MMP-9的生成和释放，该作用可能是通过诱导STAT3信号通路磷酸化实现的。

IL-6 Induces Production and Releasing of MMP-9 in Murine Bone Marrow Neutrophils

Objective To investigate the effect of IL-6 inducing murine bone marrow neutrophils MMP-9 production and releasing and its mechanism of activation. Methods Murine bone marrow neutrophils were separated and cultured in vitro and randomly divided into four groups: control group, IL-6 50ng/ml stimulation group, IL-6 50ng/ml+DMSO group and IL-6 50ng/ml+STAT3 inhibitor Ⅷ 50μmol/L group. RT-PCR method was used to evaluate the mRNA level of intracellular MMP-9 of neutrophils. Gelatin zymography was performed to detect the activity of MMP-9 released from neutrophils. Western blot was applied to test the phosphorylation levels of STAT3 pathway in neutrophils. Results Compared with control group, the administration of IL-6 enhanced the mRNA expression of MMP-9(P=0.0050); the activity of MMP-9 released from neutrophils was elevated(P=0.0087) and the p-STAT3 levels in neutrophils was also increased (P=0.089). After the STAT3 inhibitor Ⅷ was added, the mRNA expression of MMP-9, the activities of MMP-9 in the supernatant and the p-STAT3 levels in neutrophils were decreased significantly than those in the IL-6 group (P=0.0067, 0.048 and 0.098, respectively). Conclusion IL-6 induces the production and releasing of MMP-9 in neutrophils and the effect may be induced by the activation of STAT3 pathway.