HBsAg诱导TNF-α和IL-10抑制浆样树突状细胞产生α干扰素

肝胆速递

http://www.plosone.org/article/info%3Adoi%2F10.1371%2Fjournal.pone.0044900
HBsAg Inhibits IFN-α Production in Plasmacytoid Dendritic Cells through TNF-α and IL-10 Induction in MonocytesHBsAg诱导单核细胞产生TNF-α和IL-10抑制浆样树突状细胞产生α干扰素


Bisheng Shi 1, Guangxu Ren 1,2, Yunwen Hu 1, Sen Wang 1, Zhanqing Zhang 1, Zhenghong Yuan 1,2*

1 Shanghai Public Health Clinical Center, Fudan University, Shanghai, People’s Republic of China,
2 Laboratory of Molecular Virology, Shanghai Medical College, Fudan University, Shanghai, People’s Republic of China

Abstract

Type I Interferon (IFN) is one of the first lines of defense against viral infection. Plasmacytoid dendritic cells (pDCs) are professional IFN-α-producing cells that play an important role in the antiviral immune response. Previous studies have reported that IFN-α production is impaired in chronic hepatitis B (CHB) patients. However, the mechanisms underlying the impairment in IFN-α production are not fully understood. Here, we report that plasma-derived hepatitis B surface antigen (HBsAg) and HBsAg expressed in CHO cells can significantly inhibit toll like receptor (TLR) 9-mediated Interferon-α (IFN-α) production in peripheral blood mononuclear cells (PBMCs) from healthy donors. Further analysis indicated that monocytes participate in the inhibitory effect of HBsAg on pDCs through the secretion of TNF-α and IL-10. Furthermore, TLR9 expression on pDCs was down-regulated by TNF-α, IL-10 and HBsAg treatment. This down-regulation may partially explain the inhibition of IFN-α production in pDCs. In conclusion, we determined that HBsAg inhibited the production of IFN-α by pDCs through the induction of monocytes that secreted TNF-α and IL-10 and through the down-regulation of TLR9 expression on pDCs. These data may aid in the development of effective antiviral treatments and lead to the immune control of the viral infections.

Introduction

More than 350 million people worldwide are chronically infected with hepatitis B virus (HBV), and chronic infection with HBV causes significant morbidity and mortality [1],[2]. It is estimated that most neonates and approximately 5% of adults progress to chronic HBV infection following acute infection. Antiviral therapy can suppress HBV, but it cannot eliminate HBV infection entirely. Additionally, interferon-α (IFN-α), which is one of the most commonly used treatment for HBV infection, is only effective in half of all chronic HBV-infected patients who received the IFN-α treatment. Accumulating evidence indicates that an inadequate immune response to HBV is responsible for viral persistence [3], [4].

Plasmacytoid dendritic cells (pDCs) contribute to at least 95% of all of the IFN-α production among peripheral blood mononuclear cells (PBMCs). pDCs constitutively express the pattern recognition receptors toll like receptor (TLR) 7 and TLR9 [5], [6], [7]. TLR ligand stimulation or the detection of pathogen associated molecular patterns (PAMPs) can activate the myeloid differentiation factor 88 (MyD88)-interferon regulatory factor 7 (IRF-7) signaling pathway and result in high amounts of IFN-α secretion by pDCs [8]. Previous reports have indicated that both a reduced frequency and an impaired function of pDCs are observed in CHB patients. Following lamivudine treatment, a reduced HBV DNA level was observed to accompany an increased pDCs amounts. This suggested that HBV is important in the observed deficiency in the function of pDCs [9], [10]. However, the mechanism underlying the impairment of pDC function has not been fully elucidated.

It has been reported that HBV and the HBV surface antigen can enter dendritic cells, but the cells do not support viral replication [11]. This suggests a possible role of the HBV surface antigen in the impairment of pDC function. HBV surface antigen (HBsAg) is the subviral form of HBV envelope protein, and it contains multiple lipid modifications from the host hepatocyte and o-glycosylation [12], [13]. HBsAg consists of a small (S), a middle (M) and a large (L) protein that are all encoded by the same open reading frame (ORF), but translation is initiated at three distinct start codons. It has been reported that HBsAg is found in excess concentrations relative to the concentration of the HBV virion in CHB patients [14]. In some CHB patients, HBsAg concentrations have been reported to be as high as 100 µg/ml. However, there are still conflicting reports regarding the role of HBV and HBsAg in the inhibition of pDC function. Our previous study demonstrated that HBsAg could inhibit the IFN-α production of pDCs through the binding of HBsAg to the inhibitory receptor BDCA-2, thus leading to the down-regulation of IRF-7 nuclear translocation [15]. Woltman and colleagues have reported that HBV and HBsAg could abrogate the CpG-A/TLR9-induced mammalian target of rapamycin (mTOR)-mediated S6 phosphorylation, leading to IRF7 phosphorylation and IFN-α gene transcription. Additionally, HBV/HBsAg was also shown to inhibit the upregulation of co-stimulatory molecules, the production of TNF-α, IP-10 and IL-6 and pDC-induced NK cell function [16]. However, Vincent, I.E et al. have reported that HBV, but not HBsAg, can inhibit IFN-α production and TLR9 expression in pDCs [17]. HBsAg has also been reported to inhibit the TLR4-mediated cytokine production and CD80 upregulation in mDCs [18].

Herein, we further demonstrate that HBsAg can inhibit the production of IFN-α by pDCs in an indirect manner. HBsAg induced the secretion of TNF-α and IL-10 in monocytes, and these cytokines down-regulated the expression of TLR9 in pDCs to subsequently inhibit IFN-α production by pDCs.

肝胆速递

乙型肝炎表面抗原（HBsAg）通过TNF-α和IL-10诱导单核细胞，浆细胞树突状细胞抑制IFN-α的生产

Shi 1，光绪REN 1，毕升，森王，马蕴雯HU 1，Zhanqing Zhang 1，正虹媛 1，2 *

1上海公共卫生临床中心，复旦大学，上海，中华人民共和国，
2 中华人民共和国，上海，复旦大学上海医学院分子病毒学实验室

摘要
I型干扰素（IFN）是一种病毒感染的第一道防线。浆树突细胞（PDC）的是专业的IFN-α产生细胞，抗病毒免疫反应中发挥重要的作用。以前的研究报告，IFN-α生产受损，慢性乙型肝炎（CHB）患者。然而，在IFN-α的生产减值的机制并不完全理解。这里，我们报告血浆衍生的B型肝炎表面抗原（HBsAg）和HBsAg在CHO细胞中表达，可以显着抑制Toll样受体（TLR）的9-介导的α-干扰素（IFN-α）的生产在外周血单核细胞（PBMC中）从健康的捐赠者。进一步的分析表明，单核细胞参与的HBsAg上的pDCs通过分泌的TNF-α和IL-10的抑制效果。此外，TLR9表达的pDCs下调TNF-α，IL-10和HBsAg治疗。这种下调可能部分解释了抑制IFN-α的生产pDCs中。总之，我们确定乙肝表面抗原的PDC上抑制了生产的IFN-α通过诱导单核细胞分泌TNF-α和IL-10和TLR9表达的pDCs通过下调。这些数据可有助于有效的抗病毒治疗的发展导致的病毒感染的免疫控制。

说明

全球超过350万人患有慢性B型肝炎病毒（HBV）感染，慢性乙肝病毒感染会导致重大的发病率和死亡率[1]，[2]。据估计，大多数新生儿约5％的成年人进展到慢性HBV感染的急性感染。抗病毒治疗可以抑制乙肝病毒，但它不能完全消除HBV感染。此外，干扰素-α（IFN-α），这是一个最常用的治疗HBV感染，是唯一有效的谁收到的IFN-α处理的所有慢性HBV感染的病人的一半。越来越多的证据表明不足对HBV的免疫应答是负责病毒持续感染[3]，[4]。

浆树突细胞（PDC）的至少95％的所有的外周血单个核细胞（PBMC）中的IFN-α的生产作出贡献。的pDCs组成性表达的模式识别受体Toll样受体（TLR）的7和TLR9的[5]，[6]，[7]。 TLR配体刺激或病原体相关的分子模式（PAMPS）的检测可以激活髓样分化因子88（MyD88的）-干扰素调节因子7（IRF-7），在高量的IFN-α的分泌信号通路和结果的pDCs [8 。以前的报告表明，降频功能受损的pDCs在慢性乙型肝炎患者中观察到。拉米夫定治疗，降低HBV DNA水平，观察到伴随增加PDCS金额的。这表明，在所观察到的缺陷中的功能的pDCs HBV是重要的[9]，[10]。然而，PDC功能的减值机制尚未完全阐明。

已经报道，HBV和HBV表面抗原的树突状细胞可以输入，但这些细胞不支持病毒复制的[11]。这表明乙肝病毒表面抗原的PDC功能的减值可能发挥的作用。乙型肝炎病毒表面抗原（HBsAg）是亚病毒HBV包膜蛋白的形式，并且它包含从宿主肝细胞的多个脂质修改和o-糖基化[12]，[13]。 HBsAg的由一个小的（S），中（M）和一个大蛋白（L）都是由相同的开放阅读框（ORF）编码，但在三个不同的起始密码子的翻译起始。据报道，HBsAg被发现CHB患者HBV病毒粒子的浓度相对于过量的浓度[14]。在一些慢性乙型肝炎患者中，HBsAg浓度已高达100微克/毫升。然而，仍然有矛盾的报告，关于在pDC的功能抑制HBV和HBsAg的作用。我们以前的研究表明，HBsAg的能抑制IFN-α产生的pDCs通过结合的HBsAg的抑制性受体BDCA-2，从而导致IRF-7核易位[15]的向下调节。沃特曼和他的同事报告说，乙肝病毒和乙型肝炎表面抗原（HBsAg）废除CpG-A/TLR9-induced哺乳动物雷帕霉素靶蛋白（mTOR）的介导的S6磷酸化，导致IRF7的磷酸化和IFN-α基因的转录。此外，HBV / HBsAg的也显示出抑制的共刺激分子的上调，肿瘤坏死因子-α，IP-10和IL-6和pDC的诱导NK细胞的功能[16]的生产。然而，文森特，即等。有报道，乙肝病毒，但不HBsAg的，可以抑制pDCs中IFN-α的产生和TLR9的表达[17]。 HBsAg的也有报道[18]中的mDC抑制TLR4介导的细胞因子的生产和CD80的上调。

这里，我们进一步表明的HBsAg可以抑制生产的IFN-α的pDCs以间接的方式。 HBsAg的诱导分泌TNF-α和IL-10在单核细胞中，这些细胞因子下调pDCs中的表达的TLR9随后抑制IFN-α产生的pDCs。