多肽类结核杆菌抗原激活的γδ T细胞表达抗原提呈细胞表型和功能

Phenotype expression and function of antigen presenting cells in huaman γδ T cells activated by peptide antigen from Mycobacterium tuberculosis

目的:观察用多肽类结核杆菌耐热抗原(Mtb-HAg)激活的人外周血γδT细胞是否具有抗原提呈细胞的表型和抗原提呈作用。方法:外周血单个核细胞(PBMC)经贴壁法获取单核细胞,加GM-CSF,IL-4和LPS培养诱导为成熟DC。PBMC用Mtb-HAg优势扩增γδT细胞后用流式分选纯化γδT(Vδ2+)细胞。PBMC经尼龙毛柱法获得纯化T细胞,用CFSE标记后单独加Mtb分泌性抗原(Mtb-SAg),或加单核细胞和Mtb-SAg共同培养,或与经Mtb-SAg预处理的成熟DC或活化γδT细胞共同培养。第11天时用流式细胞术检测CD4+T细胞的增殖。Mtb-HAg活化的γδT细胞与FITC标记的Mtb-SAg孵育不同时间后用流式细胞术和激光共聚焦显微镜检测摄入情况。结果:Mtb-HAg活化的γδT细胞CD80,CD86和MHC-Ⅱ类分子的表达显著增加。Mtb-SAg预处理活化的γδT细胞诱导初始T细胞增殖数和CD4+T细胞增殖指数的活性与成熟DC相似。Mtb-HAg活化的γδT细胞能摄入FITC标记的Mtb-SAg。结论:用多肽类Mtb-HAg激活的γδT细胞具有抗原提呈细胞的表型,并具有摄入和提呈可溶性抗原给初始T淋巴细胞的能力。

AIM： To explore whether the human peripheral γδT cells stimulated by polypeptide heat resistant antigen from Mycobacterium tuberculosis （Mtb-HAg） express phenotype and have the fuction of antigen presentation. METHODS： The monocytes isolated by adhesion method from peripheral blood mononuclear cells （PBMCs） were cultured with GM-CSF, IL-4 and induced into the mature dendritic cells by adding LPS. PBMCs were stimulated with Mtb-HAg and IL-2 to expand predominantally γδT cells that were further purified by flow sorting. The pure T cells were isolated from PBMCs using adhesion revomal and nylon column method, labeled with CFSE, and cultured in alone with Mtb secretory antigen （Mtb-SAg）, in monocytes with Mtb-SAg, or with Mtb-SAg pre-pulsed dendritic cells or Mtb-HAg activated γδT cells for 11 days. The proliferation of the CD4^＋ T cells were measured by flowcytometry. The Mtb-HAg activated γδT cells were incubated with FITC labelled Mtb-SAg for different time, and the ingestion of Mtb-SAg by γδT cells was measured by flowcytometry and observed with laser confocal microscopye. RESULTS： The expressions of MHC-Ⅱ and co-stimulatory molecules CD80 and CD86 on γδT cells that activated by Mtb-HAg marketly incrased in comparison with that on the resting γδT cells. The expanded cell counts and proliferation index of the pure naive T cells that induced by Mtb-SAg pretreated activated γδT cells were similar to that induced by mature DC. By using flowcytometer and laser confocal microscope, FITC labellaed Mtb-SAg was ingested by the activated γδT cells. CONCLUSION：γδT cells stimulated by polypeptide Mtb-HAg express the phenotype of professional antigen-presen- ting cells and they are able to present Mtb-SAg to naive T cells to induce the proliferation of CD4^＋ T cells.