输注体外扩增的同源CD4^+CD25^+调节性T细胞增进小鼠肿瘤易感性  

Infusion of ex vivo expanded homologous CD4^+CD25^+ regulatory T cells promotes the susceptibility to tumor in mice

作　　者：郭涛[1] 罗诗樵[1] 许继凡[1] 戴珏[1] 刘钊[1] 高祥[1] 

机构地区：[1]重庆医科大学附属第一医院肝胆外科,重庆400016

出　　处：《细胞与分子免疫学杂志》2014年第5期466-470,共5页Chinese Journal of Cellular and Molecular Immunology

基　　金：国家自然科学基金(H1006/30972789)

摘　　要：目的研究过继输注体外扩增同源调节性T细胞(Treg)对小鼠抗肿瘤免疫的影响,探索过继输注Treg治疗方法可能存在的风险。方法免疫磁珠分离法分离小鼠脾脏内CD4+CD25+Treg,流式细胞术测定其纯度后予以CD3/CD28单克隆抗体磁珠和大剂量IL-2(1 000 U/mL)刺激,进行2周2轮次扩增后收集Treg,混合淋巴细胞培养测定其体外抑制功能;后将1×107Treg静脉注射BALB/c小鼠,24 h后再注射B16F10肿瘤细胞,同时设置单独接种肿瘤细胞组,14 d后计数肺部移植瘤数目,测定外周血Treg比例。结果新鲜CD4+CD25+Treg纯度大于95%,平均96.3%±2.88%,体外扩增后纯度大于85%,平均87.73%±2.35%;与新鲜Treg相比,扩增后抑制功能未受损(P>0.05);给BALB/c小鼠注射1×106B16F10细胞后14 d肺部肿瘤结节数为(14±5)个,先注射1×107体外扩增Treg后再注射1×106B16F10细胞,肿瘤结节数增多为(73±9)个(P=0.007),与单独注射2×106B16F10细胞相当(86±8)个(P=0.230);给C57BL/6小鼠输注5×105B16F10细胞后结节数为(70±15)个,预先输入8×106体外扩增Treg后再注射5×105B16F10细胞,肺部肿瘤结节数明显增多,大于300个,与前者相比,差异有统计学意义(P<0.01),同时荷瘤小鼠外周血Foxp3+Treg比例上升更为显著(P<0.05)。结论过继输注体外扩增Treg诱导移植物免疫耐受的同时,可能抑制机体的抗肿瘤免疫,因此存在一定风险。Objective To study the effects of adoptive transfer of ex vivo expanded homologous CD4＋CD25 ＋ regulatory T cells （Tregs） on antitumor immunity in mice and explore the potential risk of this therapeutic method. Methods Marine CD4＋ CD25 ＋ Tregs were separated from spleens by magnetic activated cell sorting （MACS） and expanded by anti-CD3/ CD28-coated microbeads and 1 000 U/mL interleukin 2 （IL-2） in vitro. The purity of fresh and expanded Tregs was determined by FACS. The cells were collected after two rounds of expansion （7 days each）, and suppressive activity of expanded Tregs was tested by the mixed lymphocyte reaction （MLR） in vitro. Thereafter, the BALB/c mice were intravenously injected with 1 × 10^7expanded Tregs and 24 hours later, they were inoculated intravenously with 1 × 10^6 B16 Fl0 tumor cells. The mice injected with tumor cells only were set as the control group. Fourteen days later, the percentage of CD4 ＋ CD25 ＋ Foxp3 ＋ Tregs in peripheral blood was analyzed by flow cytometry and the numbers of pulmonary metastases were counted. Results The purity of expanded Tregs decreased from （96.3 ± 2.88 ）% to （87.73 ± 2.35）% compared with fresh Tregs, but there was no significant difference in the suppressive activity between fresh Tregs and expanded Tregs （ P 〉 0.0.5）. The number of tumor nodules in lung of BALB/c mice was significantly elevated from 14 ± 5 to 73 ± 9 after injected ] x ]07 expanded Tregs compared with the control group （ P = 0.007）, and the same with the mica that were inoculated with 2 × 10^6 B16 F10 cells alone （P = 0.230）. What＇s more, the number of pulmonary metastases was significantly raised from 70 ± 15 to over 300 （ P 〈 0. Ol ） in 5 × 10^5 B16 F10 inoculated C57BL/6 mica if they were injected previously with 8 × 106 expanded Tregs. The percentage of Foxp3＋ Tregs in peripheral blood significantly increased in the experimental group as compared with normal mica and control group （P〈0.05）. Condusien Adopti

关 键 词：调节性T细胞 肿瘤免疫 FOXP3 移植免疫 

分 类 号：R392.9[医药卫生—免疫学] R730.5[医药卫生—基础医学]