目的： 以肾癌细胞和G250单克隆抗体（G250 monoclonal antibody, G250 mAb）复合物致敏树突状细胞（dendritic cells，DC）制备肾癌DC疫苗，并检测其活化水平，为临床应用DC肿瘤疫苗提供依据。 方法： 制备凋亡的肾癌细胞与G250 mAb形成的复合物（G250 mAb IgG-complexed apoptotic tumor cell，IC-ATC）。取健康人新鲜外周血分离单个核细胞（peripheral blood mononuclear cell，PBMC），以GM-CSF和IL-4诱导PBMC成未成熟的树突状细胞（immature dendritic cells，iDC），将iDC分别负载凋亡肿瘤细胞(apoptotic tumor cell，ATC)、IC-ATC、G250 mAb，并均以TNF-α诱导成熟树突状细胞（mature dendritic cells，mDC），将未负载的DC作为对照组。流式细胞术检测各组mDC的免疫表型、 ELISA试剂盒检测IL-12分泌水平，CCK-8法检测mDC刺激淋巴细胞增殖的能力。 结果： 成功制备IC-ATC致敏的DC疫苗。相对于负载ATC、G250mAb和未负载的DC，负载IC-ATC的DC疫苗显著上调CD86、CD80、CD83、HLA-DR的表达\[(4204±3.42）% vs (28.34±1.16)%、(33.77±1.61)%、(26.52±2.14)%, P<0.05；(38.17±2.55)% vs (23.79±2.41)%、(31.94±3.29)%、(24.32±3.23)%, P<005；(79.39±1.44)% vs (69.06±2.01)%、(7449±1.35)%、(66.71±3.83)%， P<0.05；(35.52±2.72)% vs (26.90±2.82)%、(29.45±158)%、(27.42±2.11)%, P<0.05\]和IL-12分泌水平（25.04 vs 5.27、13.32、7.53， P<0.05），且能更有效地刺激淋巴细胞增殖(4.02 vs 1.73、1.22、1.41， P<0.01）。 结论: IC-ATC可有效促进DC成熟和活化，IC-ATC致敏的DC疫苗诱导淋巴细胞增殖能力显著增强。

Objective : To use G250 monoclonal antibody (G250 mAb) IgG-complexed renal carcinoma cells to acti-vate dendritic cells (DCs), develop the DC vaccine of renal carcinoma and determine its activation for clinical tumor biological therapy. Methods: Prepare apoptotic renal carcinoma cells and induce the G250 mAb-complexed apoptotic renal carcinoma cells (IC-ATC). Immature dendritic cells (iDCs) induced from peripheral blood mononuclear cells of healthy volunteers were cultured and propagated in vitro using rhGM-CSF and rhIL-4. iDCs were loaded with apoptotic renal carcinoma cells (ATC), IC-ATC and G250 mAb. Then mature dendritic cells (mDCs) were induced by TNF-α, while the DCs pulsed with no antigen served as a control group. The immune phenotype of mDCs in different groups was detected by flow cytometry, secretion of IL-12 by DCs was measured by ELISA and the ability of DCs to stimulate lymphocyte proliferation was examined by CCK-8 assay. Results: It was found that when compared with ATC, G250 mAb and the control group, the mDCs pulsed with IC-ATC obriously up-regulated the expressions of CD83, CD80,CD86, and HLA-DR (\[4204±342\]% vs \[28.34±1.16\]%, \[33.77±1.61\]%, \[2652±2.14\]%, P<0.05；\[38.17±2.55\]% vs \[23.79±241\]%, \[31.94±3.29\]%, \[24.32±3.23\]%, P<0.05；\[79.39±1.44\]% vs \[69.06±2.01\]%, \[7449±135\]%, \[66.71±3.83\]%， P<0.05；\[35.52±2.72\]% vs \[26.90±2.82\]%, \[29.45±1.58\]%, \[27.42±211\]%, P<0.05), and secreted higher quantity of IL-12(25.04 vs 5.27, 13.32, 7.53, P＜0.05). Moreover, mDCs loaded by IC-ATC induced multiplication of lymphocytes was more effective (4.02 vs 1.73, 1.22 vs 1.41, P<0.05）. Conclusion: IC-ATC can promote DCs mature effectively, and DCs pulsed with IC-ATC can induce the proliferation and activation significantly.

辽宁省医学高峰建设工程专项基金(No. 2010017)