Objective:To observe the influence of cyclical low-dose cyclophosphamide (CTX) administration on regulatory T cell of melanoma-bearing mice, and the anti-tumor effects of combination with CTLs adoptive transfusion, explore its possible mechanism. Methods: Models of tumor-bearing mice were established by subcutaneous inoculation with melanoma cells. The changes of the CD4 + CD25 + Foxp3 + Treg in spleens were detected by flow cytometry. The dendritic cells derived from the bone marrow of mice were cultured, and the tumor antigen-specific CTLs were prepared by cultivation in vitro. Tumor-bearing mice were injected intraperitoneally with CTX every 7 days, then received CTLs by veinous transfusion 4 days later, which were repeated 3 cycles. The growth curves of tumor were drawn. Results: With time prolonged after tumor inoculation, the proportion of CD4 + CD25 + Foxp3 + /CD4 + was increased gradually in mice. Single administration of CTX only inhibited Tregs in short time, while cyclical administration of CTX could prolong the period in which Tregs was kept at low level. Cyclical administration of CTX could not delay the growth of tumor. But the tumor growth was delayed more significantly in cyclical CTX + CTLs group (P < 0. 05) , autoimmune vitiligo and other adverse side effect were not observed in mice treated with CTX. Conclusion: Cyclical administration of low-dose CTX could enhence the anti-tumor effect of CTLs adoptive transfusion by regulating Tregs.%目的:观察循环应用低剂量环磷酰胺(CTX)对荷瘤鼠调节性T细胞(Tregs)的影响,及联合细胞毒性T细胞(CTLs)过继免疫治疗的实际抗瘤效果并探讨其中可能的机制.方法:通过皮下接种瘤细胞制备黑色素瘤荷瘤鼠模型;腹腔注射CTX,每隔7天给药一次,共3次;应用流式细胞术检测小鼠脾脏中CD4+CD25+Foxp3+调节性T细胞(Tregs)的变化.体外培养小鼠骨髓来源的树突状细胞(DCs),制备肿瘤抗原特异性的CTLs.在腹腔注射CTX后4天,鼠尾静脉注射CTLs,每隔7天治疗一次,共3次.绘制各组肿瘤生长曲线.结果:随着荷瘤时间的延长,荷瘤鼠CD4+CD25+Foxp3+/CD4+比例逐渐升高.循环应用CTX 能够延长对Tregs的抑制,维持其在相对较低水平.循环CTX+CTLs疫苗治疗组的抑瘤作用最为明显(P＜0.05);治疗组中未见免疫性白斑等自身免疫病及明显化疗毒副反应.结论:循环应用低剂量CTX能够规律有效地调控Tregs,显著提高CTLs过继免疫治疗的疗效.
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