蛋白转导结构域介导BCR/ABL对慢性粒细胞白血病患者T细胞的活化作用

目的　研究蛋白转导结构域 (PTD)介导的BCR ABL融合蛋白对慢性粒细胞白血病(CML)患者T细胞的特异性活化作用。方法　利用基因工程技术将PTD基因与CMLb3a2bcr abl基因融合并原核表达 ,纯化的PTD BCR ABL融合蛋白与CML患者外周血单个核细胞 (PBMNC)体外共孵育 ,用流式细胞仪分别检测CD4+ 、CD8+ T细胞的早期活化抗原CD6 9的表达。结果　PTD BCR ABL抗原体外激活CD8+ T细胞的最佳浓度为 10 0 μg ml,时间为 3d。在此刺激条件下 ,15例CML患者中 ,10例表现CD8+ T细胞早期活化 ,CD6 9表达率为 (15 .0 1± 3.75 ) % ;4例表现CD4+ T细胞早期活化 ,CD6 9表达率为 (10 .32± 3.0 8) % ;其中有 3例CD8+ 和CD4+ T细胞同时活化 ;而对照的BCR ABL抗原刺激组无一例表现CD8+ 或CD4 + T细胞活化 ,其CD6 9表达率分别为 (1.36± 0 .31) %和 (1.4 1± 0 .4 3) % ,与PBS空白对照组相比差异无显著性 (P >0 .0 5 )。结论　PTD能将外源性BCR ABL抗原转导入抗原呈递细胞内 ,加工呈递后激活抗原特异性CD8+ 及CD4+ T细胞。

Activating effects of protein transduction domain mediated BCR/ABL protein on CML T cells

OBJECTIVE: To study the activating effect of protein transduction domain (PTD) mediated BCR/ABL protein on T cells from CML patients. METHODS: The plasmid containing PTD and b3a2 bcr/abl of CML was constructed by genetic engineering and expressed in E. coli. The peripheral blood mononuclear cells from CML patients were stimulated in vitro with purified PTD-BCR/ABL protein and the expression of the early activation antigen CD(69) on CD(8)(+) and CD(4)(+) T cells was detected by flow cytometry (FCM). RESULTS: The optimal concentration of PTD-BCR/ABL protein for activating CD(8)(+) T cells in vitro was 100 micro g/ml, CD(69) expression peaked in three days stimulation. CD(8)(+) T cells were activated in 10 of 15 CML patients, the expression rate of CD(69) was (15.01 +/- 3.75)%. CD(4)(+) T cells were activated in 4 of 15 patients, the expression rate of CD(69) was (10.32 +/- 3.08)%. Both CD(8)(+) and CD(4)(+) T cells were activated simultaneously in 3 of them. However, neither CD(4)(+) nor CD(8)(+) T cells was activated by stimulation with BCR/ABL protein in all 15 specimens, the expression rate of CD(69) on CD(8)(+) and CD(4)(+) T cells was (1.36 +/- 0.31)% and (1.41 +/- 0.43)%, respectively. There was no difference compared with that of PBS control group (P > 0.05). CONCLUSION: By using a PTD-mediated antigen delivering system, exogenous BCR/ABL protein can be delivered into APC, processed and presented onto surface of APC to activate Ag-specific CD(8)(+) and CD(4)(+) T cells in vitro.