不同大鼠胶质瘤抗原致敏的DC体外诱导CTL活性的研究

Induction of tumor-specific cytotoxic T lymphocytes in vitro by dendritic cells pulsed with different glioma antigens

目的 :比较大鼠骨髓来源的树突状细胞 (dendriticcells,DC)体外经由大鼠C6胶质瘤细胞由不同方式制备的不同抗原致敏后 ,对特异性细胞毒性T淋巴细胞 (cytotoxicTlymphocyte ,CTL)的诱导作用。方法 :自大鼠骨髓分离DC前体细胞 ,经重组大鼠粒细胞巨噬细胞集落刺激因子 (rrGM -CSF) +白细胞介素 4 (rrIL - 4 )诱导培养、扩增 ;由C6胶质瘤细胞经由反复冻融、煮沸灭活及超声破碎细胞抽提其总蛋白的方法制备各种不同抗原致敏DC ,致敏的DC与T淋巴细胞进行共培养诱导CTL ;以ELISA法检测CTL诱导过程中淋巴细胞趋化因子 (lymphocytechemoattractantfactor)及细胞因子IFN -γ分泌水平 :以 [3 H]-TdR掺入法检测DC诱导T细胞增殖及其特异性CTL杀伤活性。结果 :体外应用煮沸灭活瘤细胞制备的肿瘤抗原致敏DC ,能诱导更强的刺激T细胞增殖的能力、并且可以诱导杀伤活性更强的CTL。结论 :应用煮沸灭活的瘤细胞制备瘤抗原负载DC获得瘤苗可获得更强的抗肿瘤保护作用。

AIM: The goal of this study was to compare different methods for tumor antigen preparation, to observe the induction of tumor-specific cytotoxic T lymphocytes in rats by dendritic cells (DCs) pulsed with different tumor antigens. METHODS: The precursors of dendritic cells were isolated from bone marrow of rats, stimulated in vitro with recombinent rat granulocyte-macrophage colony-stimulating factor (rrGM-CSF) and interleukin-4 (rrIL-4). Then rat DCs were pulsed with C6 tumor cell antigens prepared with different methods: freeze-thaw, boiling or total protein extracted from ultrasonic crushed tumor cell. Subsequently primed DCs were cocultured with T lymphocytes isolated from spleen to induce CTL. Lymphocyte chemoattractant factor from DCs and cytokine IFN-γ release were determined by ELISA, the cytotoxicity of CTL was assayed by JAM test. RESULTS: DCs pulsed with boiled tumor cell in vitro induced an enhanced ability of T-cell proliferation and cytotoxic T lymphocyte activity.CONCLUSION: Our results demonstrated that DCs primed with boiled tumor cell may represent a method for inducing immune responses against the entire repertoire of tumor antigens of malignancies.