结直肠癌组织中LAP^+CD4^+T细胞免疫调节功能的研究  

A study on immune regulation mechanism of LAP^+CD4^+T cells in colorectal cancer

作　　者：曹云飞[1] 江志远[1] 钟武[1] 唐卫中[1] 张森[1] 高枫[1] 郭云[1] 刘小健[1] 

机构地区：[1]广西医科大学第一附属医院结直肠肛门外科,南宁市530021

出　　处：《实用医学杂志》2016年第21期3498-3502,共5页The Journal of Practical Medicine

基　　金：国家自然科学基金资助项目(编号:81250316)

摘　　要：目的:探讨结直肠癌组织中LAP^+CD4^+T细胞对效应T细胞的免疫抑制途径及作用机制。方法:利用免疫磁珠法分选35例结直肠肿瘤组织中LAP^+CD4^+T细胞和LAP-CD4^+T细胞,流式细胞仪检测其纯度;CCK-8法检测其分选细胞的增殖能力,Transwell小室检测LAP^+CD4^+T细胞对LAP-CD4^+T细胞抑制途径;ELISA法检测细胞上清液中IL-10及TGF-β含量。结果:免疫磁珠法分选获得的LAP^+CD4^+T细胞纯度为(95.10±2.87)%;CCK-8法结果显示LAP^+CD4^+T细胞分选前后细胞均保持较高存活率(P=0.547);两种细胞单独体外培养72 h后,与LAP-CD4^+T细胞组相比,LAP^+CD4^+T细胞组OD值差异有统计学意义(P=0.000);两种细胞混合培养组OD值为0.51±0.35,LAP-CD4^+T细胞单独培养组OD值为0.74±0.18,两组间比较差异有统计学意义(P=0.001);Transwell小室结果显示LAP^+CD4^+T细胞可通过非接触抑制作用于LAP-CD4^+T细胞。ELISA检测结果显示LAP^+CD4^+T细胞较LAP-CD4^+T细胞分泌更高浓度IL-10、TGF-β(P=0.000),两组相比差异有统计学意义。结论 :肠癌组织中的LAP^+CD4^+T细胞具有免疫无能性及免疫抑制性,可能通过接触抑制和分泌IL-10及TGF-β两种作用机制参与抑制效应T细胞的反应。Objective To investigate the immune regulation mechanism of LAP＋CD4＋T cells and explore its immunosuppressive pathways and mechanism on effector T cells. Methods LAP＋CD4＋T cells and LAP-CD4＋T cells were isolated from colorectal tumor tissues in 35 patients with different stages of CRC by MACS （magnetic cell sorting）. The purity of LAP＋CD4＋T cells was analyzed by flow cytometry （FCM）. CCK-8 was used to detect the proliferative capacity of these cells. Transwell chamber was used to detect the immunosuppressive pathways of LAP＋CD4＋T cells on LAP-CD4＋T cells. The concentration of IL-10 and TGF-β was measured by ELISA. Results The purity of isolated LAP＋CD4＋T cells was （95.10 ± 2.87）%. LAP＋CD4＋T cells maintained a higher survival rate before and after sorting （P = 0.547）. 72 hours after culture in vitro, the OD value of LAP＋CD4＋T cells was significantly higher than that of LAP-CD4＋Tcells （P = 0.000）. The OD value of single culture of LAP＋CD4＋T cells was significantly higher than that of co-culture group （P = 0.001）. Transwell chamber showed that LAP＋CD4＋T cells inhibited the proliferation of LAP-CD4＋T cells in a non-contact way. ELISA showed that LAP＋CD4＋T cells secreted more IL-10 and TGF-β than LAP-CD4＋T cells （P = 0.000）. Conclusions LAP＋CD4＋T cells have the characteristics of immune anergy and immune inhibition. The suppressive function of CD4 ＋LAP＋T cells was dependent on both cell contact and secretion of IL-10 and TGF-β.

关 键 词：结直肠癌 LAP+CD4+T细胞 IL-10 TGF—β 免疫抑制 

分 类 号：R735.34[医药卫生—肿瘤]