A20 Gene Polymorphisms in LDLT

描述(由申请人提供)：我们的长期工作表明，A20(TNFAIP3)可增强肝脏对损伤的抗炎、抗凋亡和抗氧化反应，并促进肝细胞增殖和肝再生。在实验动物模型中，我们已经证明A20在肝脏中的过度表达对根治性肝切除和严重的肝脏缺血再灌注损伤具有保护作用，并允许成功植入边缘肝移植。功能丧失实验进一步证实了A20在支持肝脏再生方面的基本生理作用。事实上，我们发现，在杂合子小鼠(A20)中发现的部分A20基因敲除显著损害了肝再生，并增加了2/3部分肝切除后的致死率。最近，在人类全基因组关联研究中，TNFAIP3基因多态性已被确定为几种炎症性和自身免疫性疾病的易感基因。在这项建议中，我们希望确定A20基因的多态性及其在移植物中的表达水平是否可以作为预测活体肝移植(LDLT)中肝再生和功能的可靠预后指标。LDLT已成为缓解原位肝移植(OLT)可用器官短缺的解决方案。然而，它在成人中的广泛使用受到可安全采集的移植物大小的限制。在提高移植成功率的同时限制供者风险的生物标记物是允许安全扩大移植领域的迫切需要，这将有助于缓解可供OLT使用的器官的严重短缺。我们计划：1.检测所有活体肝供者中TNFAIP3基因的多态性。2.评估A20及其靶基因在肝移植前、预期队列、再灌流后(受体)或手术结束前(供者)肝移植活检组织中的表达水平。作为这一目标的一部分，我们还将使用捐赠者的外周血单核细胞来测定A20mRNA和蛋白水平。3.检测移植肝再灌注(受体)或肝切除(供者)前后受体和供体血清中启动细胞因子(TNF和IL-6)的循环水平。4.通过动态增强多探测器CT(DCE-MDCT)结合随后的定性和定量3D分析和MEVIS重建，将TNFAIP3基因多态性、A20表达水平及其靶基因表达水平与循环启动细胞因子水平和再生状态、肝功能(肝酶和凝血试验)以及早期临床结果(出院时间、原发功能障碍、是否需要再次移植)相关联。这项工作收集的数据将促进使用A20基因多态和表达水平作为LDLT中配对供者和受者的可靠选择标记，以降低供者发病率和改善受者结局。 公共卫生相关性：原位肝移植(OLT)是急性或慢性肝功能衰竭患者的救命措施。然而，需求大大超过了器官的可获得性，导致许多患者在等待名单上死亡。活体供肝移植(LDLT)已成为缓解供肝移植器官短缺的一种解决方案。然而，在成人中，LDLT受到可安全获取的移植物大小的限制。由TNFAIP3/A20基因多态性决定的肝脏保护性蛋白A20的表达水平和功能是一种很有前途的工具，可以作为供受者配对的选择生物标志物，从而降低供者的发病率，改善受者的预后。验证A20作为LDLT中可靠的预后生物标志物可以作为供体充分性的客观衡量标准，并有助于扩大LDLT、分裂肝和OLT的一般领域。

DESCRIPTION (provided by applicant): Our longstanding work demonstrates that A20 (tnfaip3) enhances the liver's combined anti-inflammatory, anti- apoptotic, and anti-oxidant response to injury, and promotes hepatocyte proliferation and liver regeneration. In experimental animal models, we have shown that overexpression of A20 in the liver is protective following radical lethal hepatectomy and severe liver ischemia reperfusion injury, and allows for successful engraftment of marginal liver transplants. Loss-of-function experiments further confirmed the essential physiologic role of A20 in supporting liver regeneration. Indeed, we showed that partial A20 knockdown, as seen in heterozygote mice (A20 ) significantly impairs liver regeneration and increases lethality following 2/3 partial hepatectomy. Recently, in human genome-wide association studies, tnfaip3 gene polymorphisms have been identified as susceptibility loci for several inflammatory and autoimmune diseases. In this proposal, we wish to determine whether A20 gene polymorphisms and the expression level of A20 and of its target genes in the graft could serve as reliable prognostic markers for liver regeneration and function in living donor liver transplantation (LDLT). LDLT has emerged as a solution to ease the shortage of organs available for orthotopic liver transplantation (OLT). However its widespread use in adults is limited by the size of the graft that can be safely harvested. Biomarkers that would limit the risk to the donor while improving graft success are direly needed to allow for safe expansion of the field, which would help in easing the severe shortage of organs that are available for OLT. We plan to: 1. Probe for tnfaip3 gene polymorphisms in all living liver donors. 2. Evaluate the expression levels of A20 and of its target genes in liver transplant biopsies before liver transplantation, and in the prospective cohort of patients, after reperfusion (recipient), or before the end of surgery (donor). As part of this aim, we will also determine A20 mRNA and protein levels using peripheral blood mononuclear cells from the donor. 3. Measure circulating levels of priming cytokines (TNF and IL-6) in recipient and donor sera before and after graft reperfusion (recipient) or liver resection (donor). 4. Correlate tnfaip3 gene polymorphisms, A20 expression levels, and expression levels of its target genes with circulating levels of priming cytokines, and regeneration status, as determined by Dynamic Contrast Enhanced Multi-Detector CT (DCE-MDCT) with subsequent qualitative and quantitative 3D analysis and MeVis reconstruction, liver function (liver enzymes, and coagulation tests), and early clinical outcomes (time to discharge, primary dysfunction, need for re-transplantation). Data gathered in this work will promote the use of A20 gene polymorphisms and expression levels as reliable selection markers for pairing donors and recipients in LDLT to reduce donor morbidity and improve recipient outcomes. PUBLIC HEALTH RELEVANCE: Orthotopic liver transplantation (OLT) is a life-saving measure for patients suffering from acute or chronic liver failure. However, demand greatly exceeds organ availability, causing a number of patients to die while on the waiting list. Living donor liver transplantation (LDLT) has emerged as a solution to ease the shortage in organs that are available for OLT. However, in adults, LDLT is limited by the size of the graft that can be safely harvested. Expression levels and function of the hepatoprotective protein A20, as determined by tnfaip3/A20 gene polymorphisms, are promising tools that can be used as selection biomarkers for pairing donors and recipients in LDLT, which would reduce donor morbidity and improve recipient outcomes. Validation of A20 as a reliable prognostic biomarker in LDLT could serve as an objective measure of donor adequacy and could help to expand the field of LDLT, split livers and OLT in general.