乙型肝炎病毒亚病毒颗粒与树突状细胞相互作用的特性研究

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Characterization Of The Interaction Between Subviral Particles Of Hepatitis B Virus And Dendritic Cells – In Vitro Study
         

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Authors Farag MMS, Peschel G, Müller M, Weigand K

Received 29 June 2019

Accepted for publication 28 August 2019

Published 7 October 2019 Volume 2019:12 Pages 3125—3135

DOI https://doi.org/10.2147/IDR.S221294

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Peer reviewers approved by Dr Colin Mak

Peer reviewer comments 2

Editor who approved publication: Dr Joachim Wink

Mohamed MS Farag,1 Georg Peschel,2 Martina Müller,2 Kilian Weigand2

1Botany and Microbiology Department, Faculty of Science, Al-Azhar University, Nasr City, Cairo, Egypt; 2Department of Gastroenterology, Endocrinology, Rheumatology and Infectious Diseases, University Hospital Regensburg, Regensburg 93053, Germany

Correspondence: Mohamed MS Farag
Botany and Microbiology Department, Faculty of Science, Al-Azhar University, Nasr City, Cairo 11884, Egypt
Tel +20 2 100 670 4553
Fax +202 22611418
Email [email protected]

Background: During an infection with hepatitis B virus (HBV), infectious particles (Dane particles) can be detected in addition to aggregates of the subviral particles (SVP) which is considered an immune escaping mechanism for the virus. Dendritic cells (DCs) are a specialized type of antigen-presenting cell (APC) that can activate native T-cells to prime an immune response controlling HBV infection. The aim of this study was to characterize the interaction between HBVsvp and DCs in vitro.
Methods: HBVsvp that comprises surface and core proteins were produced in vitro by HepG2.2.15 as a culturing system; DCs derived from the bone marrow of mice were pulsed by HBVsvp. A different pattern of cytokines secreted by bone-marrow-derived dendritic cells from C56BL/6 mice pulsed with HBVsvp were analyzed. The interactions between HBVsvp and DCs were characterized using FACS analysis, protein assay, Western blot, and immunofluorescence staining.
Results: Pulsation of DCs with HBVsvp resulted in strong activation and higher secretion of DC cytokines including INF-α, INF-γ, TNF-α, IL-1α, IL-10, and IL-12; but not for IL-1β, IL-2, IL-6, and IL-15. The production of CXCL-10/IP-10 was increased during the observation period and reached the maximal secretion after 24 hrs (p < 0.001). In total protein assay, we found significantly higher protein concentration in HBVsvp stimulated DC groups compared to not activated DCs (p < 0.001). Both 24 kDa small surface antigen (HBVs) and the 21 kDa core protein (HBVc) were detected in activated DCs. For DCs immunofluorescence staining, our data showed clear differences in the morphology of DCs between negative control and those pulsed with HBVsvp.
Conclusion: Result demonstrates a significant complex interaction between HBVsvp and DCs, in vitro.

Keywords: dendritic cell-based therapy, cytokines, hepatitis B virus subviral particles

StephenW

乙型肝炎病毒亚病毒颗粒与树突状细胞相互作用的特性研究–体外研究


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Farag MMS，Peschel G，MüllerM，Weigand K的作者

2019年6月29日收到

接受发布2019年8月28日

2019年10月7日发布2019年卷：12页3125—3135

DOI https://doi.org/10.2147/IDR.S221294

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许可出版的编辑：Joachim Wink博士

Mohamed MS Farag，1 Georg Peschel，2 MartinaMüller，2 Kilian Weigand2

1Al-Azhar大学理学院植物学与微生物学系，埃及开罗纳斯尔市； 2雷根斯堡大学医院消化内科，风湿病学和传染病科，德国雷根斯堡93053

通讯：Mohamed MS Farag
Al-Azhar大学理学院植物学与微生物学系，埃及纳斯尔市，开罗11884
电话+20 2 100 670 4553
传真+202 22611418
电子邮件[email protected]

背景：在感染乙型肝炎病毒（HBV）期间，除了亚病毒颗粒（SVP）的聚集体之外，还可以检测到传染性颗粒（Dane颗粒），这被认为是该病毒的一种免疫逃逸机制。树突状细胞（DC）是一种特殊类型的抗原呈递细胞（APC），可以激活天然T细胞来引发控制HBV感染的免疫应答。这项研究的目的是表征HBVsvp与DC之间的相互作用。
方法：以HepG2.2.15为培养体系，体外制备包含表面蛋白和核心蛋白的HBVsvp。 HBVsvp对源自小鼠骨髓的DC进行脉冲控制。分析了用HBVsvp脉冲刺激的C56BL / 6小鼠的骨髓来源树突状细胞分泌的不同细胞因子模式。 HBVsvp和DC之间的相互作用通过FACS分析，蛋白质测定，蛋白质印迹和免疫荧光染色进行了表征。
结果：用HBVsvp冲动DC会导致DC细胞因子（包括INF-α，INF-γ，TNF-α，IL-1α，IL-10和IL-12）的强烈激活和更高的分泌。但不适用于IL-1β，IL-2，IL-6和IL-15。在观察期间，CXCL-10 / IP-10的产量增加，并在24小时后达到最大分泌量（p <0.001）。在总蛋白分析中，我们发现与未激活的DC相比，HBVsvp刺激的DC组的浓度更高（p <0.001）。在激活的DC中均检测到24 kDa的小表面抗原（HBV）和21 kDa的核心蛋白（HBVc）。对于DC的免疫荧光染色，我们的数据显示阴性对照和HBVsvp脉冲的DC之间在DC形态上存在明显差异。
结论：结果表明，HBVsvp与DC之间存在明显的复杂相互作用。

关键词：树突状细胞治疗；细胞因子；乙型肝炎病毒亚病毒颗粒