目的:探讨肿瘤引流淋巴结(TDLNs)内调节性T细胞(Tregs)对局部免疫效应细胞的调节作用。方法:建立小鼠肝癌TDLNs模型,通过免疫组织化学染色和流式细胞仪检测TDLNs内Foxp3+Tregs和CD4+及CD8+T细胞的数量。实时定量PCR测定Foxp3mRNA表达水平。应用酶联免疫斑点法(ELISPOT)检测TDLNs内CD8+T细胞分泌IFN-γ的功能。结果:TDLNs内Tregs和效应性T细胞均明显扩增,Tregs弥散分布于CD8+T细胞聚居区。TDLNs内Foxp3mRNA表达水平显著高于同一接种肿瘤小鼠腹股沟淋巴结(P〈0.01)和脾脏(P〈0.01)。Tregs趋向于在TDLNs内聚集,而非其它外周淋巴结位点。荷瘤小鼠的脾脏Foxp3mRNA表达明显高于注射LPS小鼠脾脏。Tregs抑制TDLNs内已初始化的CD8+T细胞分泌IFN-γ的功能,经anti-CD3刺激激活后,CD8+T细胞分泌IFN-γ的功能可恢复。结论:TDLNs内Tregs通过调控CD8+T细胞功能而发挥重要作用,清除Tregs是发挥特异性肿瘤免疫治疗的关键。
AIM:To investigate the roles of regulatory T cells (Tregs) on the function of effector T lymphocytes in tumor-draining lymph nodes (TDLNs). METHODS:The number expansion of Foxp3 + Tregs and CD4 + or CD8 + T cells in the TDLNs from mouse hepatocellular carcinoma model was detected by immunohistochemical staining and flow cytometry. Foxp3 mRNA expression was determined by real -time quantitative PCR. The ability of IFN-γ secretion in CD8 + T cells in the TDLNs was measured by enzyme-linked immunosorbent spot technique(ELISPOT). RESULTS:The expansions of Tregs and effector T cells were significantly increased in the TDLNs during tumor development. Tregs diffusely distributed in the CD8 + T cells occupancy area. The level of Foxp3 mRNA expression was significantly higher in the TDLNs than that in the inguinal lymph node (P 0. 01) and spleen (P 0. 01) from the same mouse inoculated Hepa1-6 cells. Tregs trended to accumulate within the TDLNs exclusively,but not in other peripheral lymph nodes(LNs) of the same host. Foxp3 mRNA expression was significantly higher in the spleen from the tumor mice than that from mice injected with LPS (P 0. 01). Tregs suppressed the CD8 + T cells primed in the TDLNs that retained the ability to secrete IFN-γ via anti -CD3 stimulation. CONCLUSION:Tregs play an important role in regulating the function of CD8 + T cells. Deletion of Tregs could be crucial for establishment of tumor-specific immunotherapy.