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目的:研究人粘蛋白核心肽-连续重复序列(MUC1-VNTR)基因转染的脐血DC疫苗抗乳腺肿瘤的免疫效应。方法:提取人脐带血淋巴细胞,培养脐血树突状细胞(DCs),采用Lipofectamine2000阳离子脂质体法将MUC1-VNTR转染入脐血DC,用Western印迹法检测MUC1-VNTR基因的表达;与自体T细胞共培养,用MTT法检测转染MUC1基因的脐血DC所致敏的细胞毒性淋巴细胞对乳腺癌MCF-7的杀伤活性;ELISA法检测其刺激T细胞分泌IFN-γ的能力。结果:Western印迹法检测到1条MUC1-VNTR基因表达的特殊条带,转染MUC1-VNTR基因的脐血DC对MUC1表达阳性的乳腺癌细胞系MCF-7的杀伤活性显著高于单纯脐血DC组(P<0.05),而且其刺激同种T细胞分泌高水平的IFN-γ,显著高于未转染的脐血DC(P<0.05)。结论:MUC1-VNTR基因转染的人脐血树突状细胞可诱导特异性CTL,产生有效的针对MUC1阳性乳腺癌细胞MCF-7的杀伤效应,并诱导特异性抗乳腺癌免疫应答。
Objective: To study the immune effect of cord blood DC vaccine transfected with human mucin core peptide-MUC1-VNTR gene on breast cancer. Methods: Human umbilical cord blood lymphocytes were isolated and cultured. Umbilical cord blood dendritic cells (DCs) were cultured. MUC1-VNTR was transfected into DC by Lipofectamine 2000 cationic liposome method. The expression of MUC1-VNTR gene was detected by Western blotting. T cells were cultured with autologous T cells. MTT assay was used to detect the killing activity of cytotoxic lymphocytes induced by cord blood DCs transfected with MUC1 gene on breast cancer cells MCF-7. The ability of T cells to stimulate the secretion of IFN-γ was detected by ELISA . Results: A special band of MUC1-VNTR gene was detected by Western blotting. The cytotoxicity of MUC1-VNTR transfected cord blood DC to MUC1-positive breast cancer cell line MCF-7 was significantly higher than that of cord blood alone DC group (P <0.05). Moreover, it stimulated allogeneic T cells to secrete high level of IFN-γ, which was significantly higher than that of untransfected umbilical cord blood (P <0.05). CONCLUSION: Human umbilical cord blood dendritic cells transfected with MUC1-VNTR gene can induce specific CTLs to produce effective killing effect on MUC1-positive breast cancer cells MCF-7 and induce specific anti-breast cancer immune response.