论文部分内容阅读
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(dendriticcells,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G250mAb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(peripheralblood 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 dendriticcells,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 dendritic cells (DCs) sensitized by a combination of G250 monoclonal antibody (G250 mAb) and human renal cell carcinoma cells (DCs) to prepare a DC vaccine of renal cell carcinoma and to detect its activation level for the clinical application of DC tumor vaccine Provide evidence. Methods: A G250 mAb IgG-complexed apoptotic tumor cell (IC-ATC) was prepared from apoptotic kidney cancer cells. Peripheral blood mononuclear cells (PBMCs) were isolated from fresh peripheral blood of healthy volunteers. PBMCs were induced to immature dendritic cells (iDC) with GM-CSF and IL-4, respectively. The apoptotic tumor cells (ATC), IC-ATC and G250 mAbs were all induced by TNF-α. Mature dendritic cells (mDC) were induced by TNF-α, and DCs without load were used as control group. Flow cytometry was used to detect the immunophenotype of mDC in each group. ELISA kit was used to detect the secretion of IL-12. The CCK-8 assay was used to detect the ability of mDC to stimulate lymphocyte proliferation. Results: IC-ATC primed DC vaccine was successfully prepared. DC vaccine loaded with IC-ATC significantly upregulated the expression of CD86, CD80, CD83 and HLA-DR [(42.04 ± 3.42)% vs (28.34 ± 1.16)% vs (33.77 ± (P <0.05); (31.44 ± 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% respectively, P <0.05; (35.52 ± 2.72)% vs (26.90 ± 2.82)%, (29.45 ± 1.58)%, (27.42 ± 2.11)%, P <0.05] and IL-12 secretion levels (25.04 vs 5.27, 13.32 and 7.53, P <0.05) and more effectively stimulated lymphocyte proliferation (4.02 vs 1.73, 1.22 and 1.41, P <0.01). CONCLUSION: IC-ATC can effectively promote the maturation and activation of DC. IC-ATC-primed DC vaccine can significantly enhance the proliferation of lymphocytes.