目的 研究头颈肿瘤中浸润的Foxp3+γδT细胞的数量与头颈肿瘤临床分期的关系.方法 用组织研磨法分离患者肿瘤浸润淋巴细胞(TILs),用密度梯度离心法分离患者外周血单个核细胞(PBMCs);用固相化anti-panTCRγδ单克隆抗体对TIL和PBMC进行2～4周的体外扩增,免疫荧光染色和流式细胞仪测定扩增前后的细胞;对头颈肿瘤组织芯片进行免疫组织化学染色.结果 15例头颈肿瘤组织标本TIL中γδT占CD3+T细胞的0.37%-12.35%,其中0.13%～34.38%为Foxp3+γδT.扩增后TIL中γδT占CD3+T细胞的20.38%～82.87%,且以Vδ1亚型为主.肿瘤组织芯片按照肿瘤TNM分期中的T分期分为4组,Foxp3+γδT细胞的比例随肿瘤分期的提高而增加.结论 现有证据证明了头颈肿瘤组织中存在Foxp3+γδT细胞的浸润,并提示肿瘤组织中调节表型T细胞的数量可能与头颈肿瘤的预后相关.%Objective To explore the relationship between infiltrated Foxp3 + γδT cells and the clinical stage of head and neck tumor. Methods Head and neck tumor biopsies were mechanically minced to get tumor infiltrating lymphocytes (TILs). Peripheral blood mononuclear cells (PBMCs) from the patients were isolated by density gradient centrifugation. TILs and PBMCs were stimulated by immobilized anti-pan-TCR｜δ in vitro for two to four weeks. Freshly isolated and expanded cells underwent immunofluorescence staining and flowcytometry analysis.Commercial head and neck tumor tissue chips were used for immunohistochemical staining. Results Immunofluorescence staining of TILs freshly isolated from 15 head and neck tumor samples showed that the γδT/CD3 + T ratio was O. 37％ ～ 12. 35％, Foxp3 + γδT/γδT ratio was 0. 13％ ～34. 38％. Followingin vitro expansion, γδT/CD3 +T cellratio increased to 20. 38％ ～ 82. 87％. Expanded cells primarily belonged to the Vδ1 subset. Tumor tissue chips were divided into four groups according to the T stage of the clinical TNM staging system. Subsequent immunohistochemical results indicated a correlation between the quantity of Foxp3 + ｜δT cells and tumor TNM stage.Conclusion Our data demonstrate the existence of Foxp3 + γδT cells in the head and neck tumor tissue, and suggest that the quantity of tumor infiltrating ｜δT regulatory cells might be associated with the prognosis of head and neck tumor.
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