CD25含量与T辅助细胞对IL-2的促增殖反应性

CD25 content variation with the responses of T helper cell to IL-2 induced proliferation

目的研究不同激活状态下T辅助细胞（Th）中IL－2R各亚基的表达变化及对IL-2反应的相关性。方法以包被的抗CD3抗体激活Thl细胞，同时设立未激活对照。’H掺入法测定其对IL-2的促增殖反应；real－timePCR检测IL-2R各亚基编码基因的表达；流式细胞术检测CD25及CDl22的含量；以I125标记的IL-2检测不同状态的Thl细胞对IL-2的亲和力。脂质体法转染IL-2RotsiRNA至激活的Thl细胞，比较不同CD25含量时Thl细胞对IL-2的反应性。自小鼠体内分离CD4细胞，以抗CD3抗体激活后在不同时间点收集细胞，比较它们的CD25含量及对IL-2的反应性。结果较未激活对照相比，激活的Thl细胞中IL-2Rα亚基，即CD25的表达显著增加，而其他两个亚基则无变化；同时伴有对IL－2亲和力的升高，但对IL-2的促增殖反应性却显著降低。以siRNA适当下调IL-2Rα基因的表达则可显著提高激活的Thl细胞对IL-2的反应性。虽然激活后的naiveCIM细胞对IL-2的反应性明显增加，但却不与CD25的含量及激活度正相关。最高的IL-2反应性出现在低度激活的CD4细胞中。结论CD25在激活后的Th细胞中显著增多，并可通过自身数量的变化来调节所在细胞对IL-2的促增殖反应性。适度高表达的CD25可使靶细胞具有最高的IL-2反应性，过度表达的CD25则可导致对IL－2反应性的降低。

Objective To elucidate the expression diversity of IL-2R subunits in T helper（Th） cells under different activation condition and clarify its relationship with the response to IL-2 induced proliferation. Methods In vitro cultured Thl cells were activated by plate bound anti-mouse CD3 McAb. In both activated and inactivated Thl cells the expression of different IL-2R subunit was evaluated by real-time PCR and fluorescence staining, 3H incorporation assay was adopt to measure the proliferation of the cells in response to IL-2 treatment, IL-2 affinity was determined via 1125 labeled IL-2. IL-2Rα siRNA transfection was applied to knock- down CD25 expression in activated Thl cells and its effect was confirmed by Western blot, IL-2 induced proliferation in IL-2Rα siRNA transfected Thl cells were subsequently detected. CD4 ceils were isolated from naive BALB/c mice and were also stimulated by plate bound anti-CD3 McAb. Cells were harvested at different days posterior to the stimulation, CD25 content and IL-2 induced proliferation were determined by flow cytometry and 3 H incorporation assay respectively. Results In comparison with inactivated cells, only the expression of CD25 but not other IL-2R subunits was significantly increased in activated Thl cells. In accordance with elevated CD25, IL-2 affinity was also increased in activated Thl cells, however, which resulted in decreased cell proliferation in response to IL-2 treatment. In activated Thl cells, when CD25 expression was differently knockdown by different-dosed IL-2Rct siRNA transfection, the highest IL-2 response was observed in the cells with partially CD25 depression. For in vitro activated naive CD4 cells the elevated CD25 expression gradually decreased and the highest proliferation response was detected at day 8 post stimulation upon IL-2 treatment. Conclusion Although CD25 is necessary for IL-2 induced proliferation, however, the excessively expressed CD25 lead to lower proliferation response. Not fully activated Thl or CD4 ceils, but partially activated ceils with lightly increased CD25 content possessed highest proliferation rate in response to IL-2 treatment.