中试生产的结核DNA疫苗免疫效能研究

中国防痨杂志 ›› 2012, Vol. 34 ›› Issue (3): 168-171.

中试生产的结核DNA疫苗免疫效能研究

吴小丽闭兰赵亚杰罗丹齐建新王炯李忠明

430060武汉生物制品研究所免疫学研究室（吴小丽、闭兰、赵亚杰、罗丹、齐建新、王炯）；武汉海规生物科技有限公司（李忠明）

收稿日期:2012-01-29 出版日期:2012-03-10 发布日期:2012-05-03
通信作者: 闭兰 E-mail:bilan630421@163.com
基金资助:
“十一五”国家科技重大专项（2008ZX-10003-013 (2)）

Immune efficacy of tuberculosis DNA vaccine produced by magnifying technology

WU Xiao-li，BI Lan，ZHAO Ya-jie，LUO Dan，QI Jian-xin，WANG Jiong，LI Zhong-ming

Department of Immunology，Wuhan Institute of Biological Products，Wuhan 430060，China

Received:2012-01-29 Online:2012-03-10 Published:2012-05-03
Contact: BI Lan E-mail:bilan630421@163.com

摘要/Abstract

摘要： 目的以300 L发酵罐中试规模生产的冻干Mtb Ag85A质粒DNA疫苗为基础免疫小鼠，对该疫苗的免疫效能进行研究。方法将4~6周龄的BALB/c小鼠20只通过数字表法随机分为2组，一组免疫Mtb Ag85A质粒DNA疫苗（结核DNA疫苗免疫组），另一组免疫PBS作为阴性对照（PBS对照组）；于免疫前采血，肌内注射免疫3次。在第3次免疫后3周摘眼球处死小鼠，收集血清，进行血清特异性抗体检测和细胞因子(IFN-γ、IL-4)检测；同时对每只小鼠取脾细胞进行特异性γ干扰素 (IFN-γ) 分泌检测和脾细胞CD4⁺、CD8⁺百分率检测。结果（1）免疫前后免疫组与对照组抗体水平吸光度A值均小于0.2。（2）结核DNA疫苗免疫组血清细胞因子IL-4水平［（146.2±34.3）pg/ml］低于PBS对照组［（177.7±28.1）pg/ml］，差异有统计学意义（t=2.244，P=0.038），而IFN-γ水平［（129.6±159.0）pg/ml］虽然高于PBS对照组［（76.5±21.5）pg/ml］，但差异无统计学意义（t=－1.047，P=0.309）。（3）酶联免疫斑点检测结果表明结核DNA疫苗免疫组脾细胞特异性IFN-γ分泌水平（分泌IFN-γ细胞个数）（103.60±112.14）高于PBS对照组（5.78±5.83），差异有统计学意义（t=36.538，P=0.018）。（4）结核DNA疫苗免疫组CD4⁺细胞百分率均值（21.57%）高于PBS对照组（12.17%），差异有统计学意义（t=3.043，P=0.038）；CD8⁺细胞百分率均值（13.70%）与PBS对照组（10.57%）相比，差异无统计学意义（t=0.847，P=0.445）。结论 Mtb Ag85A质粒DNA疫苗主要刺激机体Th1型细胞免疫，而对诱导机体产生Th2型细胞免疫应答没有显著的促进作用。

关键词: 疫苗, DNA, 酰基转移酶类, 抗原, 细菌, 免疫活性

Abstract: Objective To study the immunogenicity of Mycobacterial tuberculosis Ag85A DNA vaccine with pilot scale production in mice. Methods Twenty BALB/c mice were randomly divided into 2 groups, and then immunized intramuscularly for 3 times with Mtb Ag85A DNA or PBS respectively. The mice were killed and the whole blood samples were collected at 3 weeks after the last immunization. The specific antibody and cytokines (IL-4) in sera were detected. The Mtb Ag85A specific interferon-γ (IFN-γ) level and the percentages of CD4⁺ or CD8⁺ T cell in mouse spleen cells were also detected. Results The data obtained by statistical analyses demonstrated that: (1) The absorbance values of Ag85A-specific antibody in the sera of each group mice before and after immunization were all less than 0.2. (2) The IL-4 level in sera from Ag85A DNA group［(146.2±34.3) pg/ml］was significantly lower than that from PBS group ［(177.7±28.1) pg/ml］(t=2.244，P=0.038). The IFN-γ level in sera from Ag85A DNA group ［(129.6±159.0) pg/ml］was higher than that from PBS group ［(76.5±21.5) pg/ml］, but there was no significant difference (t=－1.047，P=0.309). (3) The data from the ELISPOT assay for IFN-γ showed that the number of spot-forming spleen cells in Ag85A DNA group(103.60±112.14) was significantly higher than that in PBS group(5.78±5.83) (t=36.538，P=0.018). (4) The percentage (21.57%) of CD4⁺ T cells in Ag85A DNA group was higher than that (12.17%) in PBS group (t=3.043，P=0.038); however, there was no significant difference on the percentage of CD8⁺ T cells between the two groups (13.70% vs. 10.57%, t=0.847，P=0.445). Conclusion The Th1 cellular immunity was mainly stimulated in the mice immunized by Mtb Ag85A DNA vaccine. But Th2 cellular immune response was not significantly induced in these mice.

Key words: Vaccines, , DNA, Acyltransferases, Antigens, bacterial, Immunocompetence

吴小丽闭兰赵亚杰罗丹齐建新王炯李忠明. 中试生产的结核DNA疫苗免疫效能研究[J]. 中国防痨杂志, 2012, 34(3): 168-171.

WU Xiao-li，BI Lan，ZHAO Ya-jie，LUO Dan，QI Jian-xin，WANG Jiong，LI Zhong-ming. Immune efficacy of tuberculosis DNA vaccine produced by magnifying technology[J]. Chinese Journal of Antituberculosis, 2012, 34(3): 168-171.

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