He Song-bing, Wang Liang, Zhang Yan-yun
Department of General Surgery, the First Affiliated Hospital of Soochow University, Suzhou 215006, China.
Zhonghua Zhong Liu Za Zhi. 2009 May;31(5):330-4.
To investigate the anti-gastric carcinoma immunological efficacy of dendritic cells (DC) precursors, that were mobilized into the peripheral blood by injection of macrophage inflammation protein-1 alpha (MIP-1 alpha), and induced by DC vaccine expressing melanoma antigen gene-3 (MAGE-3) ex vivo and in vivo.
615 mice were injected with MIP-1 alpha via the tail vein. Freshly isolated B220(-) CD11c+ cells were cultured with cytokines and assayed by phenotype analysis and mixed lymphocyte reaction (MLR). For adenoviral (Ad)-mediated gene transduction, cultured B220(-) CD11c+ cells were incubated with Ad-melanoma antigen gene-3. MIP-1 alpha-mobilized B220(-) CD11c+ cells pulsed MFC cells tumor lysate were used as positive control. The stimulated DC vaccination-induced T lymphocytes, and the killing effect of the T cells on gastric carcinoma cells were assayed by MTT. INF-gamma production was determined with the INF-gamma ELISA kit. To establish the solid tumor model, groups of 615 mice were injected with MFC cells subcutaneously into the abdominal wall. MIP-1 alpha-mobilized DC vaccines expressing MAGE-3 gene were used to immunize the mice after the challenge of MFC cells, then the tumor size and the survival of mice were examined to detect the therapeutic effect of DC vaccines.
B220(-) CD11c+ cells increased obviously after MIP-1 alpha injection, and freshly isolated B220(-) CD11c+ cells cultured with mGM-CSF, IL-4, and mTNF-alpha were phenotypically identical to typical DC, gained the capacity to stimulate allogeneic T cells. These MIP-1 alpha-mobilized DCs were transduced with Ad-MAGE-3, which were prepared for DC vaccines expressing tumor antigen. T lymphocytes stimulated with DC-transduced with Ad-MAGE-3 showed specific killing effect on gastric carcinoma cells and produced high levels of INF-gamma [(1460.00 +/- 16.82) pg/ml]. Five days after the MFC cells challenge, the mice were subsequently injected with DC vaccines. The tumor size of the experimental group was significantly smaller than that in the positive control group and the negative control groups (P<0.01). Kaplan-Meier survival curves showed the survival of the experimental group mice was significantly longer than that of the control groups (P<0.01).
B220(-) CD11c+ DC precursors are rapidly accumulated in the peripheral blood after injection of MIP-1 alpha into mice, which can further differentiate into mature DCs. These MIP-1 alpha-mobilized DCs, when transduced with MAGE-3 gene, can induce specific CTL to gastric carcinoma cells ex vivo, and can generate anti-tumor therapeutic effects on MFC cells loading mice in vivo. The efficiency of anti-tumor therapeutic immunity induced by MIP-1 alpha-mobilized DCs expressing tumor antigen are much more potent than MIP-1 alpha mobilized DCs pulsed MFC cells tumor lysate.
研究通过注射巨噬细胞炎症蛋白-1α（MIP-1α）动员至外周血中的树突状细胞（DC）前体，经体外和体内表达黑色素瘤抗原基因-3（MAGE-3）的DC疫苗诱导后对胃癌的免疫疗效。
615只小鼠经尾静脉注射MIP-1α。将新鲜分离的B220(-)CD11c+细胞用细胞因子培养，并通过表型分析和混合淋巴细胞反应（MLR）进行检测。对于腺病毒（Ad）介导的基因转导，将培养的B220(-)CD11c+细胞与Ad-黑色素瘤抗原基因-3孵育。用MIP-1α动员的B220(-)CD11c+细胞脉冲MFC细胞肿瘤裂解物作为阳性对照。用MTT法检测经DC疫苗刺激诱导的T淋巴细胞以及T细胞对胃癌细胞的杀伤作用。用INF-γ ELISA试剂盒测定INF-γ的产生。为建立实体瘤模型，将615只小鼠分组，经腹壁皮下注射MFC细胞。在MFC细胞攻击后，用表达MAGE-3基因的MIP-1α动员的DC疫苗免疫小鼠，然后检查肿瘤大小和小鼠存活情况以检测DC疫苗的治疗效果。
注射MIP-1α后，B220(-)CD11c+细胞明显增加，用mGM-CSF、IL-4和mTNF-α培养的新鲜分离的B220(-)CD11c+细胞在表型上与典型DC相同，获得了刺激同种异体T细胞的能力。这些MIP-1α动员的DC用Ad-MAGE-3转导，制备用于表达肿瘤抗原的DC疫苗。用Ad-MAGE-3转导的DC刺激的T淋巴细胞对胃癌细胞显示出特异性杀伤作用，并产生高水平的INF-γ[（1460.00±16.82）pg/ml]。在MFC细胞攻击后5天，随后给小鼠注射DC疫苗。实验组的肿瘤大小明显小于阳性对照组和阴性对照组（P<0.01）。Kaplan-Meier生存曲线显示实验组小鼠的生存期明显长于对照组（P<0.01）。
向小鼠注射MIP-1α后，B220(-)CD11c+ DC前体在外周血中迅速积累，可进一步分化为成熟DC。这些MIP-1α动员的DC用MAGE-3基因转导后，可在体外诱导对胃癌细胞的特异性CTL，并可在体内对荷MFC细胞小鼠产生抗肿瘤治疗效果。表达肿瘤抗原的MIP-1α动员的DC诱导的抗肿瘤治疗性免疫效率比MIP-1α动员的DC脉冲MFC细胞肿瘤裂解物更强。
