紫杉醇对小鼠T细胞CD69、CD25表达及增殖作用的流式细胞术分析

Flow cytometric analysis of effects of paclitaxel on the expression of CD69,CD25 and proliferation on murine T cells

目的研究紫杉醇对多克隆刺激剂作用下小鼠T细胞CD69、CD25表达及增殖的影响,并探讨其机制。方法利用荧光标记的单克隆抗体染色结合流式细胞术,检测多克隆刺激剂佛波醇酯(PDB)或刀豆蛋白(ConA)刺激下小鼠T细胞表达活化抗原CD69、CD25的百分率;以羧基荧光素乙酰乙酸琥珀酰亚胺酯(CFDASE)标记技术结合流式细胞术分析PDB+Ion(Ionomycin)或ConA刺激下T细胞增殖指数。结果PTX无论对ConA或PDB刺激组T细胞CD69表达均无抑制作用(P>0.05),而对T细胞CD25表达有明显抑制作用(P<0.05),且呈剂量依赖关系;PTX无论对ConA或PDB+Ion刺激组T细胞增殖均有抑制作用(P<0.05),且呈剂量依赖关系。但在ConA及PDB+Ion刺激初立即或是24h后加入不同浓度的PTX,PTX抑制T细胞增殖作用无差异(P>0.05)。结论PTX可明显抑制多克隆刺激剂诱导的T细胞中后期活化及增殖,其作用机制可能与早期活化相关蛋白PTK及PKCθ无关,而与PKCθ下游活化信号相关。

Aim To investigate the effects of paclitaxel(PTX) on the expression of CD69, CD25 and proliferation of T cells by polyclonal stimulas in vitro, and explore the molecular mechanism of paclitaxel. Methods Fluorescence conjugated monoclonal antibodies and flow cytometry were used to detect the express of CD69 and CD25 by activated T cells in vitro in response to Concanavalin(Con A) and Phorbol 12,13-dibutyrate(PDB) or T cell proliferation index stained by CFDA-SE in response to PDB+Ion or Con A. Results Paclitaxel had no effect on the expression of CD69, but inhibited the expression of CD25 in activated T cells in response to Con A or PDB in a concentration-dependent manner. Paclitaxel caused a dose-dependent suppression of T cell proliferation to Con A as well as to PDB+Ion. Whether added at the beginning or after 24 h of stimulation by Con A or PDB+Ion, paclitaxel had identical effects. Conclusion The mid and later activation and proliferation of murine T cells stimulated by Con A or PDB+Ion were significantly inhibited by paclitaxel, suggesting that paclitaxel acts on the downstream signaling pathways of PKCθ,and not act on the intitial activated associated proteins such as PTK and PKCθ.