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本研究探讨Elmo1对PMA刺激的中性粒细胞产生ROS的调控作用。分离WT(Elmo1+/+)和Elmo1-/-小鼠骨髓来源的中性粒细胞后,FACS法检测中性粒细胞的纯度。用PMA刺激中性粒细胞,通过分子探针标记检测ROS的产量,利用亲和沉淀GTP-Rac的方法研究Rac的激活情况,进而探索Elmo1对ROS和Rac活性的影响以及其中的作用机制。结果显示:小鼠中性粒细胞表面标志物Gr-1和CD11b的双阳性率高达90%以上。PMA刺激后,Elmo1-/-小鼠中性粒细胞ROS的产量明显低于WT(P<0.05),且Rac1和Rac2的激活程度也显著弱于WT(P<0.05)。使用Rac抑制剂后,Elmo1+/+和Elmo1-/-的中性粒细胞ROS产量之间的差异消失了。以上结果揭示了Elmo1通过激活Rac促进中性粒细胞产生ROS。
This study was to investigate the regulatory effect of Elmo1 on ROS production by PMA-stimulated neutrophils. After separation of bone marrow-derived neutrophils from WT (Elmo1 + / +) and Elmo1 - / - mice, the purity of neutrophils was measured by FACS. Neutrophils were stimulated with PMA, the production of ROS was detected by molecular probe labeling, and the activation of Rac was studied by affinity precipitation of GTP-Rac. Then the effect of Elmo1 on ROS and Rac activity and the mechanism of action were explored. The results showed that the double positive rate of mouse neutrophil surface markers Gr-1 and CD11b was over 90%. After PMA stimulation, the production of ROS in Elmo1 - / - mice was significantly lower than WT (P <0.05), and the activation of Rac1 and Rac2 was also significantly weaker than WT (P <0.05). The difference between the neutrophil ROS production of Elmo1 + / + and Elmo1 - / - disappeared with Rac inhibitors. The above results reveal that Elmo1 promotes neutrophil production of ROS by activating Rac.