Diagnostic value of Mycobacterium tuberculosis antigen 85B and early secreted antigenic target 6-kDa antigen fusion protein for tuberculous pleurisy

doi:10.3969/j.issn.1000-6621.2018.11.004

Abstract

Abstract:

Objective To explore the value of Mycobacterium tuberculosis antigen 85B (Ag85B) and early secreted antigenic target 6-kDa antigen (ESAT6) fusion protein in the diagnosis of tuberculous pleurisy.Methods Inpatients and healthy controls were enrolled in Hebei Chest Hospital from January 2014 to January 2015, including 45 patients with tuberculous pleurisy, 26 patients with non-tuberculous pleurisy, and 25 cases of healthy controls. Flow cytometry was used to detect the IFN-γ concentrations in the serum, and in the pleural effusion with and without the Ag85B-ESAT6 fusion protein stimulation. The number of CD4 ⁺ and CD8 ⁺cells in tuberculous pleurisy group and non-tuberculous pleurisy group patients was detected by immunohistochemical staining. The statistic analysis was performed by SPSS 13.0 software by t test, P<0.05 was considered as statistically significant.Results With and without stimulation of Mycobacterium tuberculosis Ag85B-ESAT6 fusion protein, the concentrations of IFN-γ in the blood of tuberculous pleurisy group were (42.63±10.51)pg/ml and (401.90±72.54)pg/ml, respectively, and non-tuberculous pleurisy group were (38.97±7.08)pg/ml and (40.04±6.80)pg/ml, respectively, and healthy control-group were (39.61±7.28)pg/ml and (39.86±6.97)pg/ml, respectively. Under the same condition, the concentrations of IFN-γ in tuberculous pleurisy group pleural effusion was (411.91±41.56)pg/ml and (1342.67±167.96)pg/ml, respectively, and non-tuberculous pleurisy group were (47.99±11.49)pg/ml and (48.76±11.25)pg/ml, respectively. The concentration of pleural effusion IFN-γ without stimulation was significantly higher in tuberculous pleurisy group than that in non-tuberculous pleurisy group(t=55.194, P=0.000). The stimulated concentration of blood IFN-γ in tuberculous pleurisy group was significantly higher than those in the other groups whether stimulated or not(t=32.879, 33.211, 33.204, P=0.000, 0.000, 0.000). The stimulated concentration of pleural effusion IFN-γ was significantly higher than those in the other groups whether stimulated or not(t=36.085, 51.478, 51.499, P=0.000, 0.000, 0.000). Immunohistochemical staining showed that the numbers of CD4 ⁺ and CD8 ⁺ cells in tissues between tuberculous pleurisy group (16349.91±2376.36, 10525.77±1164.86) and non-tuberculous pleurisy group (1853.64±670.40, 1327.15±175.55) was significantly different (t=14.381, 19.127, all P=0.000). Conclusion Mycobacterium tuberculosis Ag85B-ESAT6 fusion protein significantly increased the concentration of IFN-γ after stimulation of peripheral blood and pleural effusion, and showed potential clinical value for the diagnosis of tuberculous pleurisy.

Key words: Tuberculosis, pleural, Interferon-gamma, Laboratory techniques and procedures, Evaluation studies, Ag85B-ESAT6 fusion protein

AN Xiao-ying,YANG Yong-hui,ZHU Gui-yun,FANG Peng,BAI Hong-zhong,ZHANG Zhi-hua. Diagnostic value of Mycobacterium tuberculosis antigen 85B and early secreted antigenic target 6-kDa antigen fusion protein for tuberculous pleurisy[J]. Chinese Journal of Antituberculosis, 2018, 40(11): 1164-1169. doi: 10.3969/j.issn.1000-6621.2018.11.004

Figures/Tables 6

组别	性别(例或名)		χ²值	P值	年龄 (岁, $x ˉ$ ±s)	t值	P值
组别	男	女	χ²值	P值	年龄 (岁, $x ˉ$ ±s)	t值	P值
结核组	31	14	1.607^a	0.205	35.13±6.77	0.960^a	0.340
非结核组	14	12	1.163^b	0.281	33.46±7.56	-0.232^b	0.818
健康组	14	11	0.024^c	0.877	33.96±7.81	0.658^c	0.513

组别	例数(例或名)	IFN-γ(pg/ml, $x ˉ$ ±s)	t值	P值
结核组	45	42.63±10.51	1.747^a	0.085
非结核组	26	38.97±7.08	-0.317^b	0.752
健康组	25	39.61±7.28	1.412^c	0.163

组别	总例数	刺激后IFN-γ(pg/ml, $x ˉ$ ±s)	刺激前IFN-γ(pg/ml, $x ˉ$ ±s)	t值	P值
结核组	45	1342.67±167.96	411.91±41.56	36.085	0.000
非结核组	26	48.76±11.25	47.99±11.49	0.244	0.808

组别	例数(例或名)	IFN-γ含量(pg/ml, $x ˉ$ ±s)	t值	P值
结核组	45	401.90±72.54	33.211^a	0.000
非结核组	26	40.04±6.80	0.094^b	0.925
健康组	25	39.86±6.97	33.204^c	0.000

组别	总数(例或名)	刺激后IFN-γ(pg/ml, $x ˉ$ ±s)	刺激前IFN-γ(pg/ml, $x ˉ$ ±s)	t值	P值
结核组	45	401.90±72.54	42.63±10.51	32.879	0.000
非结核组	26	40.04±6.80	38.97±7.08	0.554	0.582
健康组	25	39.86±6.97	39.61±7.28	0.122	0.903

References 21

[1]	Porcel JM . Tuberculous pleural effusion. Lung, 2009,187(5):263-270. doi: 10.1007/s00408-009-9165-3 URL
[2]	孟苏凯, 吴振萍, 钟达 . 全血γ-干扰素释放试验在诊断结核分枝杆菌感染中的应用价值. 中国防痨杂志, 2015,37(12):1217-1220. doi: 10.3969/j.issn.1000-6621.2015.12.009 URL
[3]	Levine H, Szanto PB, Cugell DW . Tuberculous pleurisy. An acute illness. Arch Intern Med, 2016,122(4):329-332. doi: 10.1001/archinte.1968.00300090039009 URL pmid: 5676418
[4]	Nishimura T, Hasegawa N, Mori M , et al. Accuracy of an interferon-gamma release assay to detect active pulmonary and extra-pulmonary tuberculosis. Int J Tuberc Lung Dis, 2008,12(3):269-274. doi: 10.1258/ijsa.2007.007310 URL pmid: 18284831
[5]	宋雪娜, 杨华, 刘君 . Th1/Th2免疫应答平衡在结核性和恶性肿瘤性胸腔积液中的意义. 中国肿瘤临床与康复, 2014,21(9):1052-1054.
[6]	王洁玲, 唐余燕, 张毅 , 等. 结核分枝杆菌抗原Ag85B优势诱导活动性结核患者Th1型细胞免疫反应研究. 医学研究杂志, 2016,45(3):77-82. doi: 10.11969/j.issn.1673-548X.2016.03.021 URL
[7]	Kuo CJ, Bell H, Hsieh CL , et al. Novel mycobacteria antigen 85 complex binding motif on fibronectin. J Biol Chem, 2012,287(3):1892-1902. doi: 10.1074/jbc.M111.298687 URL
[8]	Qiao D, Yang BY, Li L , et al. ESAT-6 and CFP-10-specific Th1, Th22 and Th17 cells in tuberculous pleurisy may contribute to the local immune response against Mycobacterium tuberculosis infection. Scand J Immunol, 2011,73(4):330-337. doi: 10.1111/j.1365-3083.2011.02512.x URL pmid: 21223348
[9]	中华医学会. 临床诊疗指南·结核病分册. 北京: 人民卫生出版社, 2005.
[10]	刘建利, 师长宏, 靳亚平 , 等. 结核分枝杆菌Ag85B与ESAT6融合蛋白的表达、纯化及抗原活性的初步研究. 中国人兽共患病学报, 2009,25(5):449-452. doi: 10.3969/j.issn.1002-2694.2009.05.013 URL
[11]	孔忠顺, 刘京铭, 高孟秋 . 结核性胸膜炎诊断的研究进展. 中国防痨杂志, 2016,38(4):327-330. doi: 10.3969/j.issn.1000-6621.2016.04.020 URL
[12]	Ates G, Yildiz T, Ortakoylu MG , et al. Adapted T cell interferon-gamma release assay for the diagnosis of pleural tuberculosis. Respiration, 2011,82(4):351-357. doi: 10.1159/000323184 URL pmid: 21293103
[13]	Liu F, Gao M, Zhang X , et al. Interferon-gamma release assay performance of pleural fluid and peripheral blood in pleural tuberculosis. PLoS One, 2013,8(12):e83857. doi: 10.1371/journal.pone.0083857 URL pmid: 3873962
[14]	Yamashita Y, Hoshino Y, Oka M , et al. Multicolor flow cytometric analyses of CD4⁺ T cell responses to Mycobacterium tuberculosis-related latent antigens . JPN J Infect Dis, 2013,66(66):207-215. doi: 10.7883/yoken.66.207 URL pmid: 23698481
[15]	Green AM, Difazio R, Flynn JL . IFN-γ from CD4 T cells is essential for host survival and enhances CD8 T cell function during Mycobacterium tuberculosis infection. J Immunol, 2012,190(1):270-277.
[16]	柴雪敏, 黎友伦 . 结核分枝杆菌Ag85复合物相关疫苗研究进展. 中国防痨杂志, 2013,35(4):282-285.
[17]	Ping W, Limei W, Wei Z , et al. Immunotherapeutic efficacy of recombinant Mycobacterium smegmatis expressing Ag85B-ESAT6 fusion protein against persistent tuberculosis infection in mice. Human Vaccines, 2013,10(1):150-158. doi: 10.4161/hv.26171 URL pmid: 23982126
[18]	何宗林, 杜先智 . ESAT6在结核病中的最新研究进展. 国际检验医学杂志, 2011,32(6):663-665.
[19]	van Dissel JT, Soonawala D, Joosten SA , et al. Ag85B-ESAT-6 adjuvanted with IC31 ^® promotes strong and long-lived Mycobacterium tuberculosis specific T cell responses in volunteers with previous BCG vaccination or tuberculosis infection . Vaccine, 2011,29(11):2100-2109. doi: 10.1016/j.vaccine.2010.02.094 URL pmid: 20226890
[20]	姜德华 . 结核分枝杆菌融合蛋白疫苗(Ag85B-ESAT6)的构建及免疫学特性研究. 长春:吉林大学, 2010.
[21]	You Q, Wu Y, Jiang D , et al. Immune responses induced by heterologous boosting of recombinant bacillus Calmette-Guerin with Ag85B-ESAT6 fusion protein in levamisole-based adjuvant. Immunol Invest, 2012,41(4):412-428. doi: 10.3109/08820139.2012.658940 URL pmid: 22360290