Damage-associated Alarmin Key to Immune Responses in Viral Hepatitis

描述（由申请人提供）：在病毒性肝炎中调节免疫控制与疾病发病机制的关键分子事件仍然难以捉摸。然而，最近的报道表明，慢性肝炎患者血清中Alarmin（IL-33）及其诱饵受体sST 2浓度升高。它们的水平与抗病毒治疗引起的肝损害、病毒和临床结果相关。我们假设IL-33可以直接参与肝脏中的先天性和适应性免疫系统，并且还诱导新的第2组先天性淋巴样细胞（ILC 2）的分化。通过这些作用，IL-33可以调节免疫反应并在炎症过程中保护肝脏。提出了两种互补的方法来测试这些假设。1)检测IL-33是否在急性肝炎和持续性病毒感染中保护肝脏。在这个目的中，我们将检测IL-33及其受体ST 2在肝脏和血清中的内源性水平，然后静脉内递送重组腺病毒（rAd）。为了研究IL-33/ST 2通路在持续性病毒感染中的作用，我们还将研究淋巴细胞性脉络丛脑膜炎病毒克隆13（LCVM CL 13）诱导的肝炎和这些动物的抗病毒免疫应答。将在IL-33-/-小鼠中以及在野生型（wt）动物中通过抗ST 2 mAb评估IL-33的功能。相反，我们将用rIL-33治疗感染的动物以评估其保护功能。2)分析IL-33诱导的增强免疫应答和改善肝损伤的免疫机制。我们的工作假设是IL-33可以直接作用于T效应细胞或通过诱导肝脏中的ILC 2。我们将测试IL-33不仅增强T细胞活力， 使用重建的LCMV TCR转基因小鼠（P14）模型扩展其抗原宽度。在初步研究中，我们发现IL-33可以增强IFN-？但抑制TNF-？生产我们推测，前IL-33，一个未加工的核蛋白，可以作为一个转录因子，调节效应细胞因子的T细胞上的表达。为了研究IL-33和DNA的相互作用，我们将使用两步ChIP测定来确定IL-33是否结合和隔离NF-？TNF-α的B信号？表情我们预计，缺乏IL-33基因敲除动物将导致NF-？B信号和TNF-？失调，导致肝损伤加重。这项研究的新证据可以解释临床观察结果，即损伤相关分子如IL-33及其受体与慢性肝炎和药物诱导的肝毒性患者的T细胞反应和临床预后相关。这些发现可能会改变范式。虽然这项研究本质上是探索性的，但它有很大的潜力发展到更详细的研究，涉及未来的治疗候选人和感染肝炎病毒的人类受试者。

DESCRIPTION (provided by applicant): The critical molecular events that regulate immune control vs. disease pathogenesis in viral hepatitis remain elusive. However, recent reports suggested that patients with chronic hepatitis displayed elevated concentrations of alarmin (IL-33) and its decoy receptor sST2 in the serum. Their levels were associated with hepatic damage, viral and clinical outcomes in response to antiviral therapy. We hypothesize that IL-33 can directly engage innate and adaptive immune systems in the liver and also induce differentiation of novel group 2 innate lymphoid cells (ILC2). Through these actions, IL-33 can modulate immune responses and protect the liver during the inflammatory process. Two complementary approaches are proposed to test these hypotheses. 1) To examine if IL-33 protects the liver in acute hepatitis and persistent viral infection. In this Aim, we will examine he endogenous levels of IL-33 and its receptor ST2 in the liver and serum following i.v. delivery of recombinant adenovirus (rAd). To investigate the IL-33/ST2 pathway in persistent viral infection, we will also examine hepatitis induced by lymphocytic choriomeningitis virus clone 13 (LCVM CL13) and antiviral immune responses in these animals. The function of IL-33 will be assessed in IL-33-/- mice as well as by anti-ST2 mAb in wild-type (wt) animals. Conversely, we will treat infected animals with rIL-33 to evaluate its protective functions. 2) To analyze IL-33-induced immune mechanisms responsible for enhanced immune responses and ameliorated liver injury. Our working hypothesis is that IL-33 can directly act on T effector cells or through inducing ILC2s in the liver. We will test the possibility that IL-33 not only enhances T cell vigor but also expands its antigenic breadth using a reconstituted LCMV TCR transgenic mouse (P14) model. In preliminary studies, we found that IL-33 can enhance IFN-? but inhibit TNF-? production. We hypothesize that pro-IL-33, an unprocessed nucleoprotein, can act as a transcriptional factor to modulate the effector cytokines expression on T cells. To examine the IL-33 and DNA interaction, we will use a two-step ChIP assay to determine whether IL-33 binds and sequesters NF-?B signal for TNF-? expression. We anticipate that lack of IL-33 in the knockout animals will lead NF-?B signaling and TNF-? dysregulation, leading to exacerbated liver injury. The new evidence from this study may explain clinical observations that damage-associated molecules like IL-33 and its receptor correlates to T responses and clinical prognosis in patients with chronic hepatitis and drug-induced hepatotoxicity. Such findings would be potentially paradigm-shifting. Although this research is exploratory in nature, it has strong potential to be developed to more detailed studies involving future therapeutic candidates and human subjects infected with hepatitis virus.