摘要
目的观察真核质粒p MAGEA3-IRES-SEA免疫小鼠后,鼠脾淋巴细胞对喉癌细胞模型B16/MAGEA3的杀伤作用。方法将葡萄球菌肠毒素A(staphylococcal endotoxin A,SEA)和喉癌来源的黑色素瘤抗原A3(melanoma-associated antigen A3,MAGE-A3),构建成基因共表达的真核质粒p MAGEA3-IRES-SEA,用电转染的方法免疫BALB/C小鼠单侧股四头肌。末次免疫2周后,取脾分离淋巴细胞,与B16/MAGE-A3细胞共同培养,MTT法观察活化的淋巴细胞对B16/MAGE-A3的杀伤作用。结果 ptk-IRES-SEA、p IRES2-EGFP/MAGE-A3和p MAGEA3-IRES-SEA质粒组对B16/MAGE-A3细胞特异性杀伤率均高于空白质粒及生理盐水组(P<0.05);p MAGEA3-IRESSEA质粒组杀伤作用大于ptk-IRES-SEA、p IRES2-EGFP/MAGE-A3质粒组(P<0.05)。结论构建的p MAGEA3-IRES-SEA真核质粒可通过电转染的方式输入到小鼠体内,诱导特异性细胞免疫,可提高杀伤B16/MAGE-A3细胞的作用。为DNA疫苗接种途径、方法及抗喉癌疫苗的研究提供了实验依据。
Objective To observe the killing effect of lymphocyte from spleen of mice immunized by plasmid expressing MAGE-A3 antigen and staphylococcal endotoxin A （pMAGEA3-IRES-SEA） on laryngeal carcinoma cell model of B16/MAGE-A3. Methods The eukaryotic expression vector was constructed to express the Staphylococcal endotoxin A （SEA） and human melanoma-associated antigen gene A3 （MAGE-A3） from laryngocarcinoma. The plasmid was then electrotransfected into unilateral hindlimb skeletal muscles of BALB/C mice. Two weeks after the final immunization, the spleen lymphocytes were separated and cultured together with B16/MAGE-A3 cells for 12 hours. The killing effect of the lymphocytes on laryngeal carcinoma cell model of B16/MAGE-A3 was detected with MTY assay. Results The killing rate of the activated lymphocytes （ 57. 72% ） was significantly higher than that of the control group. Conclusion The constructed pMAGEA3-IRES-SEA eukaryotic expressing plasmid can be effectively transmitted into mice and improve killing ability of spleen lymphocytes to B16/MAGE-A3 cells in vitro, which provides a theoretical basis for approach and method of DNA vaccine inoculation, and for the study of anti-laryngocarcinoma vaccine.
出处
《中国耳鼻咽喉颅底外科杂志》
CAS
2015年第2期119-123,共5页
Chinese Journal of Otorhinolaryngology-skull Base Surgery
基金
广州市科技和信息化局社会发展应用基础研究专项(2013J4100024)
广东省科技厅产业技术研究与开发资金计划项目(2012B031800340)
关键词
喉癌
葡萄球菌肠毒素A
黑色素瘤抗原
体内免疫
体外实验
肿瘤细胞模型
Laryngeal neoplasma
Staphylococcal endotoxin A
Human melanoma-associated antigen
In vivo im-mune
In vitro test
Tumor cell model
