Improved liver function and regeneration with A20

说明(申请人提供)：肝脏具有显著的再生能力，可在损伤或切除后恢复。然而，足够的再生取决于保持健康的残留肝脏肿块，否则就会接踵而至的暴发性肝衰竭。这一问题在肝移植中尤其重要，因为同种异体肝移植面临严重的缺血、炎症和免疫损伤，而在使用“小体积”肝移植的成人活体供肝移植中，这些问题进一步加剧。了解保护肝细胞和促进其增殖的生理保护机制，对于制定旨在改善肝移植预后的治疗策略至关重要。我们已经确定7-锌指蛋白A20是肝细胞保护和再生反应的关键成分，因为它通过改变启动子Caspase 8的表达来保护肝细胞免受凋亡；通过限制氧化损伤来保护肝细胞免于缺血性坏死；通过抑制NFkB的激活来包含肝细胞炎症反应；通过减少细胞周期蛋白依赖的激酶抑制物p21waf1的表达来促进肝细胞的增殖。A20的这些“保肝”功能转化为戏剧性的再生优势，大大提高了小鼠在根治性肝切除或长期肝脏缺血后的存活率。有趣的是，在“小体积”的肝移植中，A20的转录减少，可能是导致损伤增加和再生不足的原因之一。我们最近揭开了A20在肝细胞中的新靶点，这些靶点可能是A20的有益作用：1)增加PPAR1的表达，2)减少p21waf1的表达，3)通过减少细胞因子信号转导抑制物(SOCS)-3的表达来增强信号转导和转录激活因子3(STAT-3)的磷酸化，尽管IL-6水平较低，但我们希望：特定的目的1：阐明A20依赖保护肝细胞免受氧化损伤后坏死细胞死亡的分子基础，并探讨PPAR1参与介导这一效应。具体目的2：通过研究p21waf1和IL-6/STAT-3/SOCS3在支持A20促增殖作用中的作用，探讨A20在肝细胞中促增殖作用的分子基础。具体目标3：评估A20在肝脏中的表达对非优化肝移植存活和功能的影响，包括小体积肝移植和缺血时间延长的移植肝。在我们的实验模型中展示A20‘S的有益作用，应该为将基于A20的疗法转化为临床实践奠定基础。公共卫生相关性：原位肝移植是急性或慢性肝功能衰竭患者的救命措施。然而，需求超过器官供应几个数量级，导致许多患者在等待名单上死亡。我们已经确定A20蛋白是一种理想的“保肝”候选蛋白。A20结合了肝细胞的抗凋亡、抗坏死、抗炎、抗氧化和促增殖功能，在小鼠的根治性致命肝切除和严重的缺血再灌注损伤后，这将转化为显著的存活和再生优势。我们希望测试基于A20的疗法在改善通常不适合移植的次优肝移植物的存活率和功能方面的有效性。A20在这些模型中的任何有益作用都将扩大肝脏捐赠者的池，并有助于缓解肝移植中严重的器官短缺。

DESCRIPTION (provided by applicant): The liver has remarkable regenerative capacity, allowing recovery following injury or resection. Adequate regeneration is, however, contingent upon maintenance of a healthy residual liver mass, otherwise fulminant hepatic failure ensues. This issue is of particular importance in liver transplantation where allografts face substantial ischemic, inflammatory, and immune insults that are further exacerbated in adult living donor liver transplants using "small-for-size" liver grafts. Understanding the physiologic protective mechanisms safeguarding hepatocytes and promoting their proliferation is critical for devising therapeutic strategies aimed at improving outcome of liver transplantation. We have identified the 7-Zinc finger protein A20 as a critical component of the protective and regenerative responses of hepatocytes in that it protects hepatocytes from apoptosis by altering the expression of the initiator Caspase 8; safeguards hepatocytes from ischemic necrosis by limiting oxidative damage; contains hepatocyte inflammatory responses by inhibiting NFkB activation; and promotes hepatocyte proliferation by decreasing the expression of the Cyclin Dependent Kinase Inhibitor p21waf1. These "hepatoprotective" functions of A20 translate into a dramatic regenerative advantage with greatly improved survival following radical lethal hepatectomy or prolonged liver ischemia in mice. Interestingly, transcription of A20 is decreased in "small-for-size" liver grafts, likely contributing to increased damage and inadequate regeneration. We have recently unraveled novel targets for A20 in hepatocytes that may account for its beneficial effects: 1) Increases the expression of Peroxisome Proliferator-Activated Receptor alpha (PPAR1), 2) Decreases the expression of p21waf1 and 3) Enhances Signal Transducer and Activator of Transcription 3 (STAT-3) phosphorylation despite lower IL-6 levels through decreasing Suppressor of Cytokine Signaling (SOCS)-3 expression We wish to: Specific Aim 1: Decipher the molecular basis for the A20 dependent protection of hepatocytes from necrotic cell death following oxidative damage and probe the involvement of PPAR1 in mediating this effect. Specific Aim 2: Explore the molecular basis for the pro-proliferative function of A20 in hepatocytes by investigating the role of p21waf1 and IL-6/STAT-3/SOCS3 in supporting this effect. Specific Aim 3: Evaluate the impact of A20 expression in the liver upon survival and function of non- optimal liver grafts, including small-for-size liver grafts and grafts with prolonged ischemia time. Demonstrating A20's beneficial effects in our experimental models should set the basis for translating A20- based therapies into clinical practice. PUBLIC HEALTH RELEVANCE: Orthotopic liver transplantation is a life-saving measure for patients suffering from acute or chronic liver failure. However, demand exceeds organ availability by several orders of magnitude, causing a number of patients to die while on the waiting list. We have identified the A20 protein as an ideal "hepatoprotective" candidate. A20 combines anti-apoptotic, anti-necrotic, anti-inflammatory, anti-oxidative and pro-proliferative functions in hepatocytes, which translates into significant survival and regenerative advantages following radical lethal hepatectomy and severe ischemia reperfusion injury in mice. We wish to test the effectiveness of A20-based therapies in improving survival and function of sub-optimal liver grafts that are usually unsuitable for transplantation. Any beneficial effects of A20 in these models would expand the pool of liver donors and help ease the severe organ shortage in liver transplantation.