1-磷酸鞘氨醇对粒细胞功能的影响及其信号通路的初步研究

发布时间：2018-01-07 08:23

本文关键词：1-磷酸鞘氨醇对粒细胞功能的影响及其信号通路的初步研究　出处：《华东师范大学》2012年硕士论文　论文类型：学位论文

【摘要】：本文主要研究1-磷酸鞘氨醇(Sphingosine-1-phosphate, S1P)对人外周血中性粒细胞(Neutrophils)活化的影响,首次将S1P作为中性粒细胞活化的引发剂,研究其对中性粒细胞功能的影响及其引发中性粒细胞呼吸爆发的信号通路。中性粒细胞,又称多形核嗜中性粒细胞(polymorphonuclear neutrophil, PMN),是机体防御细菌和真菌第一道防线中的关键成分。它们通过快速地移位到炎症部位并活化激发灭菌机制,构成了早期的固有免疫反应,其中一种方式就是快速地发生呼吸爆发,产生大量的活性氧分子(ROS)。ROS具有很强的抗菌能力,但也可能会引起周围的组织损伤或诱导其他免疫细胞的凋亡。中性粒细胞的呼吸爆发是清除进入体内的微生物所必须的防御过程。呼吸爆发活性的降低与杀灭细菌能力的降低以及感染性疾病的易感性增加有关。本实验用流式细胞术检测新鲜分离PMN的状态、活化前后CDllb表达的变化；利用高铁细胞色素C还原法间接计算PMN呼吸爆发超氧阴离子(O2-)释放量；二氢若丹明123(Dihydrorhodamine123, DHR)流式检测粒细胞的呼吸爆发程度；并利用RT-PCR方法检测S1PR的表达。结果显示,新鲜分离的PMN状态良好,S1P对粒细胞CDllb的表达影响不显著；S1P预处理后,甲酰甲硫氨酰亮氨酰苯丙氨酸(fMLP)活化的PMN所释放的O2量明显提高,且若丹明123的平均荧光强度明显高于fMLP单独处理组。本研究通过RT-PCR检测方法证明PMN普遍表达S1PR1、4、5,而受体2和3仅在少数人PMN上表达。S1P与PMN表面S1PR结合后可能激发了一条或几条以P13K为连接点的信号通路,并进一步改变PMN的功能。因此我们进一步用Western Blot方法证明了S1P会通过P13K传递信号,进而导致细胞内信号蛋白蛋白激酶B(Akt)的磷酸化,并进一步影响PMN呼吸爆发。本研究针对S1P对PMN功能影响的结果可为今后PMN炎症引起的组织损伤、输血相关急性肺损伤发生机制和S1P类似物免疫抑制剂化学治疗的研究提供实验依据和新的治疗方向。
[Abstract]:The effect of 1 - phosphate - 1 - phosphate ( S1P ) on the activation of neutrophils ( Neutrophils ) in human peripheral blood was studied in this paper . The effect of S1P on the activation of neutrophils was first studied . Neutrophils , also known as multinucleated neutrophils ( PMN ) , are the key components of the body ' s defense against bacteria and fungus ' s first line of defense . They form an early innate immune response by rapidly shifting to the site of inflammation and activating the challenge sterilization mechanism , one of which is the rapid onset of respiratory burst , producing a large number of reactive oxygen molecules ( ROS ) . ROS possess strong antibacterial ability , but may also cause surrounding tissue damage or induce apoptosis of other immune cells . The respiratory burst of neutrophils is a necessary defense process to remove microorganisms entering the body . The decrease of respiratory burst activity is related to the decrease of the ability of killing bacteria and the increase of susceptibility to infectious diseases . The expression of superoxide anion ( O2 - ) is detected indirectly by flow cytometry . In this study , the expression of S1P on PMN was demonstrated by RT - PCR . The expression of S1P and the surface S1PR of PMN could stimulate the phosphorylation of signal protein kinase B and further change the function of PMN . The results of this study suggested that the effect of S1P on PMN ' s function could provide experimental basis and new treatment direction for the study of the effect of S1P on PMN function .

【学位授予单位】：华东师范大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R392.1

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