肺结核患者全血γ干扰素释放试验影响因素的探讨

Study on the Influencing Factors of Whole Blood Gamma Release Assay in the Patients with Pulmonary Tuberculosis

目的探讨菌量负荷、疾病活动性、影像学累及范围、外周血淋巴细胞计数等因素对全血γ干扰素释放试验结果的影响。方法选取肺结核患者106例（包括44例菌阳肺结核患者、47例菌阴肺结核患者和15例陈旧性肺结核患者）和健康对照组35名，所有患者（健康对照者）抽取外周血并以肝素抗凝分别与结核分枝杆菌特异抗原分泌抗原靶-6（ESAT-6）、ESAT-6／培养滤过蛋白-10（CFP-10）融合抗原、PPD共同孵育，并用ELISA的方法检测血浆中IFN-γ的含量，各组间差异的比较应用Mann-WhitneyU检验，P〈0．05为差异有统计学意义。结果抗原刺激后分泌IFN—γ量的中位数在菌阳组和菌阴组之间差异无统计学意义[ESAT-6（114．7Pg／ml vs 82Pg／ml，Z=-0．500，P〉0．05），ESAT-6／CFP-10（3488Pg／mlvs 2350Pg／ml，Z=-0．949，P〉0．05），PPD（4514Pg／mlvs4326Pg／ml，Z=-0．822，P〉0．05）]；ESAT-6和ESAT-6／CFP-10抗原刺激后分泌IFN-γ量的中位数在菌阴组和陈旧性肺结核组之间差异无统计学意义[ESAT-6（82Pg／mlvs137Pg／ml，Z=-0．781，P〉0．05），ESAT-6／CFP-10（2350Pg／mlvs1784Pg／ml，Z=-1．685，P〉0．05）]，ESAT-6抗原刺激后分泌IFN—γ量的中位数在菌阳组和陈旧性肺结核组之间差异无统计学意义（114．7Pg／mlvs137Pg／ml，Z=-0．757，P〉0．05），但ESAT-6／CFP-10融合抗原刺激后分泌IFN-γ量的中位数在菌阳组和陈旧性肺结核组之间差异有统计学意义（3488Pg／mlvs1784Pg／ml，z=-0．242，P〈0．05）；ESAT-6、EsAT-6／CFP-10融合抗原刺激后分泌IFN—γ量的中位数在肺部病变累及少和肺部病变累及多之间差异有统计学意义[ESAT-6（117Pg／mlvs42Pg／ml，z=-2．341，P〈0．05），ESAT-6／CFP-10（3055Pg／mlvs1562．5Pg／ml，Z=-2．850，P〈0．05）]；ESAT-6、ESAT-6／CFP-10融合抗原、PPD抗原刺激后分泌IFN—γ量的中位数在外周血淋巴细胞≥1．0×10^9／L和〈1．0×10^9／L之间差异有统计学意义[ESAT-6（97．5Pg／mlvs48Pg／ml，Z=-2．745，P〈0．05），ESAT-6／CFP-10（3082Pg／mlvs1190Pg／ml，z=-2．911，P〈0．05），PPD（4322Pg／mlvs3200Pg／ml，z=-2．216，P〈0．05）]。结论基于RDl区的抗原刺激后外周血分泌IFN-γ的量不受菌量负荷的影响，但可能受到外周血淋巴细胞绝对值和病变累及范围的影响。

Objective To evaluate the clinical influencing factors on the whole blood gamma release assay in the patients with pulmonary tuberculosis. Methods One hundred and six clinical diagnosed tuberculosis patients who included 44 smear-or culture-positive cases, 47 smear-or culture-negative cases and 15 cases with non-active pulmonary tuberculosis and 35 healthy controls were enrolled in this study. The heparinized bloods from all participants were incubated with purified protein derivative （PPD） and M. tuberculosis-specific antigens early secretory antigen target 6 （ ESAT- 6） and ESAT-6/CFP-10 fusion protein encoded in the mycobacterial genomic region of difference （RD1） . The production of interferon γ （ IFN- γ ） was detected with ELISA. Results There were no significant difference between smear-positive group and smear-negative group in the IFN- γ median stimulated by ESAT-6 （ 114. 7 pg/ml vs 82 pg/ml, Z = - 0. 500, P 〉 0. 05） , ESAT-6/CFP-10 （ 3488 pg/ml vs 2350 pg/ml, Z = - 0. 949,P 〉 0. 05） and PPD （ 4514 pg/ml vs 4326 pg/ml, Z= - 0. 822, P 〉 0. 05） . The IFN- γ median stimulated by ESAT-6 （ 82 pg/ml vs 137 pg/ml, Z = - 0. 781, P 〉0. 05） and ESAT-6/CFP-10 （ 2350 pg/ml vs 1784 pg/ml, Z = - 1. 685, P 〉 0.05） had not significant difference between the smear negtive pulmonary tuberculosis group and non-active pulmonary tuberculosis group. However, there were statistic difference between smear-positive pulmonary tuberculosis group and non-active pulmonary tuberculosis group in the IFN- γ median stimulated by ESAT-6/CFP-10（ 3488 pg/ml vs 1784 pg /ml, Z = - 0. 242, P 〈 0. 05） . The patients with less than 4 segments of lesions on the X-ray had higher IFN- γ, median than those with more than 4segmentsoflesions [ESAT-6（117pg/mlvs42pg/ml, Z= - 2. 341, P 〈 0.05） , ESAT-6/CFP-10（3055pg/mlvs 1562.5pg/ml, Z = - 2. 850, P 〈 0. 05） ]. There were also significant difference between the patients with higher peripheral blood lymphocyte count （ ≥ 1.0 × 10^9/L） and those with lower peripheral blood lymphocyte count （ 〈 1.0 × 10^9/L） in the IFN-γ median [ESAT-6 （ 97.5 pg/ml vs 48 pg/ml, Z= - 2. 745, P 〈 0. 05）, ESAT-6/CFP-10 （ 3082 pg/ml vs 1190 pg/rnl, Z = - 2. 911, P 〈 0. 05）, PPD（ 4322 pg/ml vs 3200 pg/ml, Z = - 2. 216, P 〈 0.05） ]. Conclusion The production oflFN- γ, stimulated by M. tuberculosis specific antigens was not influenced by the load of bacilli, and probably influenced by accumulated lesion in X-ray and peripheral blood lymphocyte count.