GRP78稳定内质网跨膜信号抗乙肝表面抗原诱导的肝细胞损伤研究及其应用

Hepatitis B surface antigen (HBsAg) is not only the serum marker of chronic hepatitis B (CHB), but also an important causative factor of liver diseases. However，the clearance of HBsAg is one of the most difficult problems in HBV field. The aim of this study is to demonstrate the molecular mechanism of liver injury induced by HBsAg to find out an effective treatment of CHB in early stage. In our previous study, it has been proved that the amount of GRP78 in endoplasmic reticulum (ER) was decreased due to the HBsAg-induced protein secretion, which plays a role in liver injury. It was also reported that GRP78 could bind to ER transmembrane signaling factors and adjust their activities to resist the liver injury. Therefore, we propose that the binding of GRP78 to transmembrane factors could be inhibited by competitive bind of HBsAg. In this study, we will employ the animal model and cell model to investigate the influence of HBsAg to the distribution and binding of GRP78 and ATF-6 or XBP1. Furthermore, we will investigate how ATF-6 and XBP1 induce the entrance of NFκB to nucleus followed by activating the downstream genes. Then, it will be determined whether restoration of GRP78 can block the changes of ER transmembrane signaling. Finally, it will be tested if GRP78 can resist HBsAg induced liver injury via stabilizing endoplasmic reticulum transmembrane signaling factors.

乙肝表面抗原(HBsAg)是慢性乙肝的标志，也是重要的致病因子，对人群危害严重。但HBsAg难以清除是本领域的难点问题。我们的前期工作证实HBsAg能结合并外迁GRP78蛋白，降低内质网内GRP78蛋白，此过程与肝细胞持续损伤密切相关。GRP78能结合内质网跨膜信号分子调控其活化，发挥抗细胞损伤作用。因此我们假设在HBsAg诱导的细胞损伤中，HBsAg竞争性破坏GRP78与跨膜分子的结合，而活化下游分子。本课题将利用细胞模型和小鼠模型研究HBsAg对GRP78和ATF-6，XBP1分布和结合的影响，以及后二者调控NFκB入核及下游基因活化变化；并通过回补GRP78，干扰HBsAg诱发的内质网信号变化，验证GRP78通过稳定内质网跨膜信号发挥抗HBsAg诱导的肝细胞损伤作用的假设，阐明HBsAg导致肝损伤的分子机制，据此寻求对慢性乙肝的早期防治，避免严重肝病的发生。

乙肝病毒表面抗原（HBsAg）研究一直是乙肝病毒研究的重点和难点问题之一。HBsAg致病机制缺乏明确的证据，HBsAg的清除手段更显匮乏，抗乙肝方法多围绕抑制病毒复制，而慢性乙肝的肝损伤发生多围绕HBx、复制的话题展开，并不能完全解释诸多临床现象。通过本项目的支持，对HBsAg可能的致病机制进行了多侧面的研究，并尝试了基于此理论的抗HBsAg的手段，也进一步印证了我们提出的HBsAg致病机制的可靠性。.根据研究计划,着力解决了乙肝表面抗原（HBsAg）通过影响胞内内质网（ER）GRP78水平，进而影响内质网相关的信号通路，参与HBsAg相关致病机制。并在此基础上，为特异性干预乙肝相关疾病提供理论依据。HBsAg通过GRP78-ATF6、XBP1-NF kappa B通路的分子机制，及HBsAg钝化外界刺激对NF kappa B的激活作用。初步提示了HBsAg一种可能的致病机制，即，通过与GRP78结合，促使其大量外迁，这种结合GRP78的作用，竞争了原本与内质网内GRP78结合的ER跨膜分子，导致后者游离入核，并进一步激活其下游效应。而ATF6、XBP1-NF kappa B通路就是其中一条通路。这条通路可激活UPR反应，引起内质网内UPR反应的伴侣分子等上调表达。初步提出HBsAg-GRP78共分泌，UPR反应再激活，GRP78基因表达水平上升的反馈机制。这为HBsAg可以持续大量表达的机制提供了理论支撑。并据此理论，利用环孢霉素A和木犀草素-葡萄糖苷，通过降低GRP78，达到抑制HBsAg的目的，这一发现既验证了GRP78参与HBsAg分泌机制，又为降低或清除HBsAg提供了候选途径。.