Wang X X, Chen X, Li Y Q, Xiao T Y, Jiang Y, Li M C, Liu H C, Wan K L
State Key Laboratory for Communicable Disease Prevention and Control, Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, National Institute for Communicable Disease Control and Prevention, Chinese Center for Disease Control and Prevention, Beijing 102206, China.
School of Public Health, University of South China, Hengyang 421001, China.
Zhonghua Liu Xing Bing Xue Za Zhi. 2017 May 10;38(5):665-669. doi: 10.3760/cma.j.issn.0254-6450.2017.05.022.
To investigate the human T cell epitopes of () Rv0585c protein antigen and their immunogenicity and provide evidence for the development of specific tuberculosis immune diagnostic techniques and tuberculosis vaccine. We synthesized peptides from Rv0585c protein antigen predicted by TE-predict and IEDB human T cell epitope prediction tool. The cellular immunoreactivity of the predicted peptides was evaluated through ELISpot assay with the peripheral blood monouclear cells (PBMC) of clinical tuberculosis patients. In animal experiments, BALB/c mice were respectively immunized with high dose (100 μg/mice) and low dose (50 μg/mice) of the peptides of Rv0585c, at the same time, high dose (50 μg/mice) and low dose (20 μg/mice) of Ag85B protein were used in positive control group. The levels of IFN-γ, IL-2, IL-4 and IL-10 were tested with ELISA kit respectively. By means of bioinformatics technique, 66 human T cell epitopes of Rv0585c were predicted, from which9 peptides concentrated epitopes were synthesized for the animal immune experiments. Peptides P10110, P10112 and P10117 were confirmed to be antigenic. The sensitivity and specificity of P10110, P10112 and P10117 were 14.00, 12.00, 6.00 and 100.00, 100.00, 97.96 respectively when they were used as diagnostic reagents of tuberculosis. The sensitivity and specificity were 22.00 and 97.96 when the epitopes were combined together. The results of animal immunity test showed that high levels of cytokines IFN-γ, IL-2, IL-4 and IL-10 were induced by high and low dose of P10110, and high levels of IFN-γ、IL-2 and IL-10 were induced by high and low dose of P10112, which were much higher than that in negative controls, respectively (<0.001). Rv0585c, including its human T cell epitopes, has good immunogenicity and immunoreactivity, stimulating the body to produce a stronger cellular immune response and has better potential application value in cellular diagnosis of tuberculosis and the development of new type of tuberculosis vaccine.
为研究（）Rv0585c蛋白抗原的人T细胞表位及其免疫原性，为特异性结核病免疫诊断技术的开发和结核病疫苗提供依据。我们合成了由TE-predict和IEDB人T细胞表位预测工具预测的Rv0585c蛋白抗原的肽段。通过用临床结核病患者的外周血单个核细胞（PBMC）进行ELISpot测定来评估预测肽段的细胞免疫反应性。在动物实验中，分别用高剂量（100μg/只小鼠）和低剂量（50μg/只小鼠）的Rv0585c肽段免疫BALB/c小鼠，同时，阳性对照组使用高剂量（50μg/只小鼠）和低剂量（20μg/只小鼠）的Ag85B蛋白。分别用ELISA试剂盒检测IFN-γ、IL-2、IL-4和IL-10的水平。通过生物信息学技术，预测了Rv0585c的66个人T细胞表位，从中合成了9个肽段集中表位用于动物免疫实验。证实肽段P10110、P10112和P10117具有抗原性。当P10110、P10112和P10117用作结核病诊断试剂时，其敏感性和特异性分别为14.00、12.00、6.00和100.00、100.00、97.96。当这些表位组合在一起时，敏感性和特异性为22.00和97.96。动物免疫试验结果表明，高剂量和低剂量的P10110诱导了高水平的细胞因子IFN-γ、IL-2、IL-4和IL-10，高剂量和低剂量的P10112诱导了高水平的IFN-γ、IL-2和IL-10，分别显著高于阴性对照组（<0.001）。Rv0585c，包括其人T细胞表位，具有良好的免疫原性和免疫反应性，刺激机体产生较强的细胞免疫反应，在结核病的细胞诊断和新型结核病疫苗的开发中具有较好的潜在应用价值。
