论文部分内容阅读
目的:探讨细胞因子信号转导抑制因子-1(suppressors of cytokine signaling1,SOCS1)拮抗物pJAK2多肽(氨基酸序列号为1001-1013)参与树突状细胞(dendritic cells,DCs)的体外诱导培养后对DCs抗肿瘤作用的影响。方法:采集健康人外周血,离心获得单个核细胞,用重组人粒细胞巨噬细胞集落刺激因子(recombinant human granulocyte-macrophage colony stimulating factor,rhGM-CSF)及重组人白细胞介素-4(recombinant human interleukin-4,rhIL-4)诱导DCs,第5天分为4组:单纯DCs培养(对照)组、抗原负载(Lysate-DCs)组、多肽修饰(pJAK2-DCs)组和抗原与多肽共培养(Lysate+pJAK2-DCs)组,第6天各组加入肿瘤坏死因子-α(tumor necrosis factor-alpha,TNF-α)促成熟。倒置显微镜下观察DCs形态;FCM法检测DCs表型;乳酸脱氢酶(lactate dehydrogenase,LDH)细胞毒实验检测各组细胞毒性T淋巴细胞(cytotoxic T lymphocyte,CTL)对胃癌细胞BGC-823的靶向杀伤作用;ELISA法检测白细胞介素-12(interleukin-12,IL-12)和γ干扰素(interferon-γ,IFN-γ)的水平。结果:与未加入诱导剂组相比,各组均成功诱导出成熟DCs,均高表达CD80、CD83、CD86和人类白细胞DR抗原(human leukocyte antigen DR,HLA-DR),但以Lysate+pJAK2-DCs组的表达水平最高。在10:1～30:1的效靶比范围内,CTL杀伤作用与效靶比呈正相关。当效靶比为30:1时,对照组的CTL杀伤率达(19.77±2.34)%,低于其他3组(P<0.01),而Lysate+pJAK2-DCs组较Lysate-DCs组及pJAK2-DCs组都高(P<0.05)。Lysate+pJAK2-DCs组培养上清液中IL-12及IFN-γ的分泌水平明显高于对照组(P<0.01)。结论:SOCS1拮抗物pJAK2多肽(1001-1013)可增强DCs对胃癌细胞的抗原递呈及特异性抗肿瘤作用。
Objective: To investigate the effect of pJAK2, a suppressor of cytokine signaling1 (SOCS1) antagonist, on the induction of dendritic cells (DCs) DCs antitumor effect. Methods: Peripheral blood was collected from healthy volunteers and centrifuged to obtain mononuclear cells. Recombinant human granulocyte-macrophage colony stimulating factor (rhGM-CSF) and recombinant human interleukin-4 DCs were induced on the 5th day after DCs induction. The DCs were cultured in the DCs (control group), Lysate-DCs group, pJAK2-DCs group and co-cultured with antigen (Lysate + pJAK2-DCs). On the 6th day, tumor necrosis factor-alpha (TNF-α) was added into each group to induce maturation. The morphological changes of DCs were observed under an inverted microscope. The phenotype of DCs was detected by FCM. The cytotoxic T lymphocytes (CTLs) were detected by cytotoxic T lymphocyte (LDH) cytotoxicity assay in gastric cancer cells BGC-823 The killing effect was detected by ELISA. The levels of interleukin-12 (IL-12) and interferon-γ (IFN-γ) were detected by ELISA. RESULTS: Mature DCs were successfully induced in all the groups compared with those without inducing agent, all of which expressed CD80, CD83, CD86 and HLA-DR highly, but Lysate + pJAK2- DCs group the highest level of expression. In the range of 10: 1 ~ 30: 1, the killing effect of CTL was positively correlated with the target ratio. When the target ratio was 30: 1, the killing rate of CTL in the control group was (19.77 ± 2.34)%, which was lower than that in the other three groups (P <0.01), while Lysate + pJAK2- DCs group were high (P <0.05). The secretion of IL-12 and IFN-γ in Lysate + pJAK2-DCs group was significantly higher than that in control group (P <0.01). CONCLUSIONS: SOCS1 antagonist pJAK2 polypeptide (1001-1013) can enhance antigen presentation and specific anti-tumor effect of DCs on gastric cancer cells.