Wang Zhong-Ying, Xie Ru-Feng, Yang Jie, Ren Ya-Na, Yang Yi-Ming, Fan Hua-Hua
Department of Biology and Medicine, East China Normal University, Shanghai, China.
Zhongguo Shi Yan Xue Ye Xue Za Zhi. 2012 Aug;20(4):989-94.
The aim of this study was to examine the priming effect of sphingosine 1-phosphate (S1P) on fMLP-activated neutrophils, mainly to detect the neutrophil respiratory burst products, and to investigate the signaling pathway involved in S1P activity. Flow cytometry was used to evaluate the new isolated neutrophil; the superoxide anion output was detected indirectly by cytochrome C reduction in respiratory burst; the dihydro-rhodamine 123 was used to detect the intensity of respiratory burst; the signal transduction pathways of neutrophil respiratory burst were explored by Western blot. The results showed that after pretreated with S1P, the level of superoxide anion released by fMLP-activated neutrophils significantly increased; the Rhodamine 123 mean fluorescence intensity in S1P primed fMLP-activated neutrophils group was significantly higher than that in fMLP treatment group; PI3K and Akt proteins involved in the signal pathway of neutrophil respiratory burst. It is concluded that S1P is a new priming reagent, which primes respiratory burst of fMLP-activated neutrophils; this signal pathway may be that S1P interacts with its receptor, activates PI3K, then activates Akt-transmitting signals through NADPH oxidase, finally results in the respiratory burst.
本研究的目的是检测1-磷酸鞘氨醇（S1P）对fMLP激活的中性粒细胞的启动作用，主要检测中性粒细胞呼吸爆发产物，并研究S1P活性涉及的信号通路。采用流式细胞术评估新分离的中性粒细胞；通过呼吸爆发中细胞色素C还原间接检测超氧阴离子输出；用二氢罗丹明123检测呼吸爆发强度；通过蛋白质免疫印迹法探索中性粒细胞呼吸爆发的信号转导途径。结果显示，用S1P预处理后，fMLP激活的中性粒细胞释放的超氧阴离子水平显著升高；S1P启动的fMLP激活的中性粒细胞组中罗丹明123平均荧光强度显著高于fMLP处理组；PI3K和Akt蛋白参与中性粒细胞呼吸爆发信号通路。结论是，S1P是一种新的启动试剂，可启动fMLP激活的中性粒细胞的呼吸爆发；该信号通路可能是S1P与其受体相互作用，激活PI3K，然后通过NADPH氧化酶激活Akt传递信号，最终导致呼吸爆发。
