一种新的检测IGRA开发测试的T细胞反应对HBV抗原的慢性乙型肝

StephenW

Research
Development of a novel IGRA assay to test T cell responsiveness to HBV antigens in whole blood of chronic Hepatitis B patients

Werner Dammermann1*, Frank Bentzien2, Eva-Maria Stiel1, Claudia Kühne1, Sebastian Ullrich3, Julian Schulze zur Wiesch145 and Stefan Lüth14   

    * Corresponding author: Werner Dammermann [email protected]

Author Affiliations

1Department of Medicine, University Medical Center Hamburg-Eppendorf, Martinistrasse 52, Hamburg 20246, Germany

2Department of Transfusion Medicine, University Medical Center Hamburg-Eppendorf, Martinistrasse 52, Hamburg 20246, Germany

3Department of Anatomy and Experimental Morphology, University Medical Center Hamburg-Eppendorf, Hamburg, Germany

4German Center for Infection Research (DZIF), partner site Hamburg, Hamburg, Germany

5Heinrich Pette Institute – Leibniz Institute for Experimental Virology, Hamburg, Germany

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Journal of Translational Medicine 2015, 13:157  doi:10.1186/s12967-015-0513-1
Published: 13 May 2015
Abstract (provisional)

Background Interferon gamma release assays (IGRA) have been developed to support easy and fast diagnosis of diseases like tuberculosis, and CMV in transplant patients. IGRAs focus on cellular immunity especially memory T cells and thus also allow rapid screening prior to complex flow cytometric testing. Here, we describe a novel, sensitive whole blood based cytokine release assay capable of assessing T cell responsiveness to HBV antigens in Hepatitis B patients and assessing hepatitis B vaccination status in healthy individuals. Methods Seventy two chronic Hepatitis B patients (CHB), 8 acute hepatitis B patients (AHB) and 80 healthy controls (HC) were tested by ELISA for IFNγ- and IL2-secretion in whole blood after challenge with synthetic peptide libraries of hepatitis B core antigen (HBcAg) or hepatitis B surface antigen (HBsAg). Results The developed IGRA test reliably differentiated between Hepatitis B patients, vaccinees and unvaccinated healthy controls. Treatment naïve and treated CHB patients showed a weaker IFNγ response to HBcAg (16 ± 5 and 35 ± 28 pg/ml, respectively) compared to the AHB group (82 ± 39 pg/ml), whereas HC remained unresponsive (6 ± 1 pg/ml). IL2 levels after HBcAg challenge were also higher in the AHB group compared to naive and treated CHB as well as HC (47 ± 21 vs. 12 ± 3, 15 ± 10 and 12 ± 9 pg/ml, respectively). HBsAg stimulation led to increased IFNγ and IL2 levels in the AHB group (33 ± 12 and 22 ± 12 pg/ml) and even higher levels in HC due to a high hepatitis B vaccination rate (41 ± 10 and 167 ± 58 pg/ml). Naive and treated CHB patients developed no or only weaker IFNγ or IL2 responses to HBsAg (5 ± 2 and 12 ± 7 pg/ml, for naive CHB, 12 ± 10 and 18 ± 15 pg/ml, for treated CHB). For HC, IL2 release after HBsAg stimulation depicted hepatitis B vaccination status with a diagnostic sensitivity and specificity of 85 % and 90 %.
Conclusion Our novel whole blood based cytokine release assay constitutes an easy and robust tool for screening HBV specific cellular immunity as alternative to flow cytometry or ELISPOT assays.
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StephenW

研究
一种新的检测IGRA开发测试的T细胞反应对HBV抗原的慢性乙型肝炎患者的全血

沃纳Dammermann1 *，弗兰克Bentzien2，伊娃玛丽亚Stiel1，克劳迪娅Kühne1，塞巴斯蒂安Ullrich3，朱利安舒尔茨楚Wiesch145和StefanLüth14

    *通讯作者：沃纳Dammermann [email protected]

作者机构

医学大学医学中心汉堡Eppendorf公司，Martinistrasse 52，汉堡20246，德国教研室

输血医学大学医学中心汉堡Eppendorf公司，Martinistrasse 52，汉堡20246，德国教研室

解剖与实验形态学，大学医​​学中心汉堡Eppendorf公司，德国汉堡3Department

4German中心感染研究所（DZIF），合作伙伴网站，汉堡，德国

5Heinrich佩特研究所 - 莱布尼茨研究所实验病毒学，德国汉堡

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杂志 - 转化医学2015年，13：157 DOI：10.1186 / s12967-015-0513-1
发布时间：2015年5月13日
摘要（临时）

背景干扰素γ释放测定（IGRA）已经被开发以支持容易和快速诊断的像肺结核病和CMV移植患者。 IGRAs专注于细胞免疫尤其是记忆性T细胞，从而也让快速筛选之前复杂的流式细胞仪检测。在这里，我们描述了一种新的，敏感的全血基于细胞因子的释放能够评估T细胞的反应，以HBV抗原在乙肝患者和评估健康人接种乙肝疫苗状况分析。方法72只慢性乙肝患者（CHB），8急性乙型肝炎患者（AHB）和80名健康对照（HC）用ELISAIFNγ-和IL2分泌的全血乙肝核心的合成肽库攻击后进行了测试抗原（HBcAg）或乙型肝炎表面抗原（HBsAg）。结果开发IGRA试验乙肝患者，接种和未接种健康对照组之间的差异可靠。初次接受治疗和治疗慢性乙型肝炎患者表现出更弱的IFNγ响应的HBcAg（16±5和35±28微克/毫升，分别）相比，在AHB组（82±39微克/毫升），而HC仍然没有响应（6±1皮克/ ml）中。核心抗原挑战后的IL2水平也越高AHB组相比幼稚和处理CHB以及HC（47±21对12±3 15±10和12±9微克/毫升，分别）。 HBsAg的刺激导致增加IFNγ和IL2水平在AHB组（33±12和22±12微克/毫升），并且由于高的乙肝疫苗接种率（甚至更高的水平以HC 41±10和167±58微克/毫升）。天真和治疗的慢性乙肝患者发展为没有或只有弱IFNγ或IL2反应的HBsAg（5±2和12±7皮克/毫升，对于天真CHB，12±10和18±15微克/毫升，为治疗慢性乙肝）。对HC，乙肝表面抗原刺激后的IL2释放描绘乙型肝炎疫苗接种状态与诊断的灵敏度为85％和90％和特异性。
结论我们新颖的全血细胞因子的基础释放测定构成一个简单而强大的工具筛选HBV特异性细胞免疫为替代流式细胞仪和酶联免疫斑点试验。
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