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目的:构建含OVA-Fc融合基因并对树突状细胞具有靶向性的DNA疫苗,评价其在肿瘤治疗中的作用。方法:构建真核表达载体OVA-Fc-pcDNA3.1,以脂质体转染法将其导入CHO细胞,用流式细胞术和ELISA法检测融合蛋白OVA-Fc的表达。建立E.G7-OVA荷瘤小鼠模型,用51Cr释放实验测定免疫小鼠脾脏细胞毒性T淋巴细胞(CTL)的抗肿瘤活性。通过观察荷瘤小鼠肿瘤的体积和生存期评价该肿瘤疫苗的疗效。结果:酶切鉴定和序列测定证明,真核表达载体OVA-Fc-pcDNA3.1构建正确。用流式细胞术和ELISA法均表明,转染的CHO细胞能表达OVA-Fc融合蛋白。OVA-Fc能激发CTL的杀伤活性,发挥抗肿瘤作用,从而减缓肿瘤的生长,延长荷瘤小鼠的生存期。结论:含OVA-Fc-pcDNA3.1的树突状细胞靶向性DNA疫苗能在体内有效地激发抗瘤免疫应答,为进一步开展临床实验奠定了基础。
OBJECTIVE: To construct a DNA vaccine containing OVA-Fc fusion gene and targeting to dendritic cells and to evaluate its role in tumor therapy. Methods: The eukaryotic expression vector OVA-Fc-pcDNA3.1 was constructed and transfected into CHO cells by lipofection method. The expression of OVA-Fc fusion protein was detected by flow cytometry and ELISA. The E.G7-OVA tumor-bearing mouse model was established and the antitumor activity of splenic cytotoxic T lymphocytes (CTL) of immunized mice was determined by 51Cr release assay. The efficacy of the tumor vaccine was evaluated by observing the tumor size and survival of tumor-bearing mice. Results: Enzyme digestion and sequencing proved that the eukaryotic expression vector OVA-Fc-pcDNA3.1 was constructed correctly. Flow cytometry and ELISA showed that transfected CHO cells could express OVA-Fc fusion protein. OVA-Fc can stimulate cytotoxic activity of CTL, play an anti-tumor effect, thereby slowing the growth of tumors and prolonging the survival of tumor-bearing mice. Conclusion: The dendritic cell-targeted DNA vaccine containing OVA-Fc-pcDNA3.1 can effectively stimulate the anti-tumor immune response in vivo, which lays the foundation for further clinical trials.