Induction of a Th1-type of immune response but not protective immunity by intramuscular DNA immunisation with Brucella abortus GroEL heat-shock gene
Induction of a Th1-type of immune response but not protective immunity by intramuscular DNA immunisation with Brucella abortus GroEL heat-shock gene
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DOI：
10.1099/0022-1317-51-1-20
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发表时间：
2002-01-01
影响因子：
3
通讯作者：
Oliveira, SC
中科院分区：
文献类型：
--
作者：
Leclerq, S;Harms, JS;Oliveira, SC
用BALB/c小鼠肌肉注射免疫，检测了编码流产布鲁氏菌GroEL热休克基因的DNA疫苗的免疫原性和保护效果。针头注射或皮下注射基因枪。通过聚合酶链式反应扩增了布鲁氏菌GroEL基因，并将其克隆到两种不同的哺乳动物表达载体pCMV-link和pCMV-tPA中。将两种载体分别导入D17细胞，Northern印迹法检测GroEL转录本。为了确定合成的蛋白质水平，将转基因细胞裂解产物提交给Western印迹。由pCMV-LINK构建产生的重组GroEL的非分泌型比由pCMV-tPA构建产生的分泌型蛋白的量要大得多。免疫后，免疫小鼠可检测到较强的抗GroEL抗体。只有当使用pCMV-link/GroEL质粒时，才能注射或基因枪。关于诱导免疫反应的模式，I.M.针刺注射主要引起Th1反应，主要是IgG2a特异性的抗GroEL和脾T细胞产生的高水平的干扰素-γ。GONE枪支免疫在小鼠中诱导了Th0型免疫反应，其特征是高lgG1/IgG2a比率以及IL-4和干扰素(干扰素)-γ的产生。尽管根据所使用的免疫途径产生了不同的免疫反应模式，但这两种方法都没有产生GroEL DNA疫苗显著的保护水平。
The immunogenicity and protective efficacy of a DNA vaccine encoding the GroEL heat-shock gene from Brucella abortus was tested in BALB/c mice immunised by intramuscular (i.m.) needle injection or epidermally by gene gun. The Brucella GroEL gene was amplified by PCR and cloned into two different mammalian expression vectors pCMV-link and pCMV-tPA. The D17 cell line was transfected with both constructs and GroEL transcripts were detected by Northern blot. To determine the level of protein synthesised, transfected cell lysates were then submitted to Western blot. The non-secreted form of the recombinant GroEL produced by the pCMV-link construct was detected in much greater amount than the secreted form of the protein produced by the pCMV-tPA construct. After immunisation, a strong anti-GroEL IgG response was detected in mice vaccinated by i.m. injection or gene gun only when the pCMV-link/ GroEL plasmid was used. Regarding the pattern of immune response induced, i.m. needle injection raised a predominantly Th1 response with mostly IgG2a-specific anti-GroEL and high levels of IFN-gamma produced by splenic T cells. Gone gun immunisation induced a Th0 type of immune response in mice characterised by a high lgG1/IgG2a ratio, and IL-4 and interferon (IFN)-gamma production. Even though a distinct pattern of immune response was generated depending upon the immunisation route used, neither method engendered a significant level of protection with the GroEL DNA vaccine.