CD69的表达在小鼠嗜酸细胞的活化与凋亡中的作用

Role of CD69 expression in activation and apoptosis of mouse eosinophils

目的:观察在体内、体外不同条件下小鼠嗜酸细胞(EOS)表面CD69的表达与细胞存活率,探讨CD69的表达在小鼠EOS的活化、凋亡中的作用。方法:提纯IL-5高分泌转基因小鼠外周血中的EOS,测定其表面CD69的表达,体外以PMA+MA刺激EOS,在1 h、12 h、18 h、24 h测定细胞表面CD69的表达与细胞存活率;分别以1μg/L的IL-4、IL-5、IL-12、IL-13、IFN-γ、GM-CSF培养EOS18 h后测定细胞的存活率与表面CD69的表达。制备小鼠哮喘模型,观察CD69在小鼠BALF、外周血EOS中的表达。结果:新鲜提纯的小鼠外周血EOS不表达CD69,PMA+MA刺激的EOS在1 h后即有CD69的表达,12 h表达至高峰,至少持续24 h以上;但细胞的存活率快速下降。不同细胞因子对EOS的培养均可诱导CD69的表达,其中IL-13、IFN-γ、GM-CSF对此影响明显,且GM-CSF可显著抑制EOS的凋亡;哮喘小鼠外周血EOS无CD69的表达,而BALF中EOS可有CD69的表达。结论:静止小鼠EOS表面不表达CD69,但在体内、体外不同条件下激活的EOS表面均有CD69的表达;同时,体外实验显示CD69的表达与细胞凋亡密切相关。结果提示CD69既可作为EOS激活的表面标记物,同时又可诱导EOS的凋亡,这为哮喘治疗提供新的思路。

AIM： To investigate the expression of CD69 on mouse eosinophils （EOS） and cell viability under different status in vitro or in vivo, and discuss the role of CD69 expression in mouse EOS activation and apoptosis. METHODS： Peripheral blood EOS from IL -5 transgenic mice were purified, which constitutively expressed this cytokine exclusively from mature T cells. CD69 expression on the cells was assayed by FACs. Purified EOS were stimulated with 10^-7 mol/L phorbol 12 - myristate 13 - acetate and ionomycin （ PMA ＋ MA）. The expressions of CD69 on cell surface and cell viability at 1 h, 12 h, 18 h, 24 h were assayed. EOS were cultured with IL-4, IL-5, IL-12, IL-13, IFN-γ, GM -CSF for 18 h at concentration of 1 μg/L and CD69 expression and cell viability were also determined. Meanwhile, the expressions of CD69 on EOS from blood and BALF of asthmatic mice were detected. RESULTS： Fresh purified EOS didn＇t express CD69. At 1 h, the expression of CD69 with PMA ＋ MA stimulation increased a little, at 12 h the expression of CD69 approached peak and lasted at least 24 h. The cell viability decreased rapidly after cultured with PMA ＋ MA. Interestingly, EOS cultured in media also expressed CD69 and died spontaneously though slower than the cells stimulated with PMA ＋ MA. The cytokines tested induced the expression of CD69 on EOS, though it was more obvious in cells cultured with IFN - 21, IL - 13, GM - CSF, which also inhibited the apoptosis of EOS induced by activation at the same time. EOS from blood didn＇t express CD69, while EOS from BALF of asthmatic mice expressed CD69. CONCLUSION： Fresh EOS from peripheral blood of mice do not express CD69. Activated EOS cultured with PMA ＋ MA, different cytokines or from BALF of asthmatic mice express CD69. CD69 is an activation marker of mouse EOS, it also induces the apoptosis of EOS after activation.