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目的制备一种能高效激活免疫细胞的肿瘤疫苗,提高免疫细胞杀伤肿瘤细胞的功能。方法同源重组表达含人免疫球蛋白(Ig G)Fc片段的MUC1蛋白作为抗原,并利用DOTAP阳离子脂质体包被抗原呈递给体外诱导培养的树突状细胞(DC),然后用DC细胞激活产生特异性免疫应答的T细胞,检测T细胞对肿瘤细胞的杀伤能力。结果成功构建含人免疫球蛋白(Ig G)Fc片段的MUC1蛋白表达载体,经大肠杆菌成功表达目的蛋白并纯化,经聚丙烯酰胺凝胶电泳验证得到目的蛋白条带。抗原蛋白经DOTAP阳离子脂质体包被,包封率达到97%。包被后的抗原刺激体外培养的免疫细胞,流式细胞术检测发现1~40μg/ml抗原制剂均能有效激活产生免疫应答,并显著增加功能性肿瘤杀伤细胞亚群比例,联合对肿瘤细胞(MCF-7)杀伤功能分析实验,10μg/ml疫苗制剂效果最佳,能提高免疫细胞对肿瘤细胞杀伤率45%以上。结论制备的疫苗制剂能有效被DC细胞捕获,并激活产生特异性应答的功能性肿瘤杀伤细胞,显著提高了免疫细胞对肿瘤细胞的杀伤功能。
Objective To prepare a tumor vaccine that can efficiently activate immune cells and improve the function of immune cells to kill tumor cells. Methods MUC1 protein containing Fc fragment of human immunoglobulin (Ig G) was expressed by homologous recombination as antigen, and then cultured in vitro with DOTAP cationic liposome coated antigen. Dendritic cells (DCs) Activate T cells that produce a specific immune response and test the ability of T cells to kill tumor cells. Results The MUC1 protein expression vector containing Fc fragment of human immunoglobulin (Ig G) was successfully constructed. The target protein was successfully expressed in Escherichia coli and purified. The target protein band was confirmed by polyacrylamide gel electrophoresis. The antigen protein was coated with DOTAP cationic liposomes and the encapsulation efficiency reached 97%. After the coated antigen stimulated immune cells cultured in vitro, the results of flow cytometry showed that 1 ~ 40μg / ml antigenic preparation can effectively activate immune response and significantly increase the proportion of functional tumor killer cell subsets, combined with the tumor cells ( MCF-7) killing function analysis experiments, 10μg / ml vaccine formulations the best, can improve immune cells to tumor cell killing rate of 45%. Conclusion The vaccine preparation can be effectively captured by DCs and activate functional tumor killer cells that produce specific response, which can significantly improve the killing effect of immune cells on tumor cells.