论文部分内容阅读
目的观察树突状细胞(DC)分泌的外泌体(exosomes)与环磷酰胺(CTX)、PolyⅠ∶C联合应用的抗肿瘤效果。方法重组鼠粒细胞-单核细胞集落刺激因子诱导骨髓来源的造血干细胞分化为髓系树突状细胞,肿瘤细胞L1210的冻融抗原负载树突状细胞,以超速离心结合膜过滤的方法提取外泌体;用得到的外泌体在DC存在的情况下刺激T细胞,应用cck-8试剂盒测量T细胞的增殖;制备抗原特异性的细胞毒性T淋巴细胞(CTL),通过DIOC-18联PI的方法测定CTL对肿瘤的杀伤作用;L1210细胞接种小鼠,制备荷瘤小鼠模型,分为实验组(exosomes联合CTX和PolyⅠ∶C)和对照组(1.PBS组,2.CTX组,3.CTX+PolyⅠ∶C组4.exosomes组)作治疗,观察肿瘤的生长速度,小鼠的生存期。结果T细胞的增殖实验中,平均吸光度:实验组为0.90±0.22,对照组为0.53±0.13,差异具有统计学意义(P<0.05)。负载肿瘤抗原的exosomes刺激的CTL在体外能够特异性杀伤肿瘤细胞L1210,在效靶比分别为5∶1和10∶1时实验组的杀伤率分别为(21.19±3.99)%、(30.56±3.85)%,对照组的杀伤率分别为(17.54±4.48)%、(19.38±4.68)%,10∶1时实验组与对照组相比杀伤率差异具有统计学意义(P<0.05);与对照组相比,实验组荷瘤小鼠的肿瘤生长速度明显降低,小鼠的生存期延长。结论经过肿瘤抗原负载的exosomes刺激的T细胞体外能够杀伤肿瘤细胞;exosomes联合CTX和PolyⅠ∶C能显著抑制荷瘤小鼠体内肿瘤的生长,延长荷瘤小鼠的生存期。
Objective To observe the antitumor effects of exosomes secreted by dendritic cells (DCs) in combination with cyclophosphamide (CTX) and PolyⅠ: C. Methods Recombinant murine granulocyte-monocyte colony-stimulating factor induced bone marrow-derived hematopoietic stem cells to differentiate into myeloid dendritic cells. The frozen-thawed antigens of tumor cells L1210 were loaded with dendritic cells and were isolated by ultracentrifugation and membrane filtration The exosomes were stimulated with exosomes in the presence of DC to stimulate T cells. T cell proliferation was measured using the cck-8 kit. Antigen-specific cytotoxic T lymphocytes (CTLs) were prepared by DIOC-18 PI to determine the killing effect of CTL on tumor. L1210 cells were inoculated into mice to prepare tumor-bearing mice model and divided into experimental group (exosomes combined with CTX and PolyⅠ: C) and control group (1. PBS group, 2.CTX group , 3.CTX + PolyⅠ: C group 4.exosomes group) for treatment, observe the growth rate of tumor, mouse survival. Results The average absorbance in T cell proliferation experiment was 0.90 ± 0.22 in the experimental group and 0.53 ± 0.13 in the control group, with statistical significance (P <0.05). Exosomes stimulated with tumor antigen loaded CTL can kill tumor cell L1210 specifically in vitro, and the killing rates of the experimental group were (21.19 ± 3.99)% and (30.56 ± 3.85) at the target ratios of 5:1 and 10:1, respectively ), And the killing rates of the control group were (17.54 ± 4.48)% and (19.38 ± 4.68)%, respectively. The kill rate of the experimental group was significantly different from that of the control group (P <0.05) Compared with the control group, the tumor growth rate of the tumor-bearing mice in the experimental group was significantly decreased, and the survival of the mice was prolonged. Conclusion Exosomes stimulated by tumor antigen can kill tumor cells in vitro. Exosomes combined with CTX and PolyⅠ: C can significantly inhibit the growth of tumor in tumor-bearing mice and prolong the survival of tumor-bearing mice.