Improved liver function and regeneration with A20

A20 改善肝功能和再生

• 批准号: 6693848
• 负责人: CHRISTIANE FERRAN
• 金额: $ 35.96万
• 依托单位: BETH ISRAEL DEACONESS MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-01-01 至 2007-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6693848
• 关键词: Improved; liver; function; regeneration; A20

DESCRIPTION (provided by applicant): Necrosis and apoptosis of hepatocytes are critical pathologic features associated with liver injury. Hepatocyte apoptosis is a feature of viral hepatitis, ischemic liver injury, sepsis, cholestasis, and a result of exposure to hepatotoxic substances such as ethanol, acetaminophen and cytostatic drugs. Massive hepatocyte apoptosis and necrosis result in fulminant hepatic failure (FHF). Only 14% of patients diagnosed with FHF recover with medical therapy. Orthotopic liver transplantation (OLT) has dramatically improved the fate of these patients (49% undergo OLT), yet 37% die while awaiting OLT. This gloomy picture is balanced by the unique capacity of the liver to regenerate. Hepatocyte replication leads to a full recovery of liver function and mass 1-2 weeks following surgical, viral or chemical hepatic loss. We propose that protecting hepatocytes from apoptosis and promoting their proliferation are two strategies that could beneficially impact FHF. Our preliminary data demonstrate that A20 promotes hepatocyte proliferation and is anti-apoptotic. A20 is part of the physiologic response of hepatocytes to injury. A20 is upregulated in hepatocytes by pro-inflammatory stimuli including TNF and LPS and functions to protect from TNF mediated apoptosis. Gene transfer of A20 to mice livers protects from lethality in the galactosamine and LPS (D-gal/LPS) model of toxic FHF. Adenovirus mediated expression of A20 in livers of BALB/c mice yields an 89% survival rate following administration of D-gal/LPS as compared to 15-20% in control mice. Mice expressing A20 maintain normal liver function as assessed by prothrombin time while controls suffer from a severe bleeding diathesis. Expression of A20 in the liver protects from lethality associated with a subtotal (87%) liver resection (LR). In this model, resection of 87% of the liver mass results in 100% lethality. In contrast, >60% of mice expressing A20 survive the 87% LR and demonstrate increased regenerative capacity as assessed by the number of PCNA (proliferating cell nuclear antigen) positive nuclei in the liver. These results qualify A20 as a critical gene involved in accelerating liver regeneration and promoting hepatocyte survival and function, even when facing extreme metabolic demands. These encouraging results prompted the submission of this proposal. Our specific aims are i) to dissect, in vitro, the molecular basis of the (1) anti-apoptotic and (2) pro-proliferative function of A20 in hepatocytes and ii) to confirm that liver directed gene therapy using A20 will beneficially impact upon toxic, FAS-mediated and surgical experimental models of FHF. From a basic science standpoint, the in vitro work proposed will address the effect of A20 upon transcription factors and expression of genes involved in apoptosis, activation and proliferation of hepatocytes. This should unveil many unknowns in our understanding of hepatocyte biology and could lead to the discovery of novel therapeutic targets. From a therapeutic standpoint, validation of the beneficial effect of A20 in the murine in vivo models of FHF should set the basis for extending this approach to models of FHF in non human primates and potentially to clinical applications. The generation of novel safer and tissue specific viral vectors for gene transfer and the development of non-viral means of protein delivery to cells will facilitate clinical translation of A20 based therapies.

描述（由申请人提供）：肝细胞坏死和凋亡是与肝损伤相关的关键病理特征。肝细胞凋亡是病毒性肝炎、缺血性肝损伤、败血症、胆汁淤积的一个特征，是接触乙醇、对乙酰氨基酚和细胞生长抑制药物等肝毒性物质的结果。大量肝细胞凋亡和坏死导致暴发性肝衰竭（FHF）。诊断为 FHF 的患者中只有 14% 通过药物治疗康复。原位肝移植 (OLT) 显着改善了这些患者的命运（49% 接受了 OLT），但 37% 的患者在等待 OLT 期间死亡。这种悲观的景象被肝脏独特的再生能力所平衡。手术、病毒或化学性肝损失后 1-2 周，肝细胞复制可导致肝功能和质量完全恢复。我们认为，保护肝细胞免于凋亡和促进其增殖是可能有益于 FHF 的两种策略。我们的初步数据表明 A20 促进肝细胞增殖并具有抗凋亡作用。 A20 是肝细胞对损伤的生理反应的一部分。 A20 在肝细胞中通过促炎刺激（包括 TNF 和 LPS）上调，并起到保护免受 TNF 介导的细胞凋亡的作用。将 A20 基因转移至小鼠肝脏可防止半乳糖胺和 LPS (D-gal/LPS) 毒性 FHF 模型死亡。给予 D-gal/LPS 后，BALB/c 小鼠肝脏中腺病毒介导的 A20 表达产生 89% 的存活率，而对照小鼠的存活率为 15-20%。通过凝血酶原时间评估，表达 A20 的小鼠维持正常的肝功能，而对照组则患有严重的出血素质。 A20 在肝脏中的表达可防止与次全 (87%) 肝切除术 (LR) 相关的死亡。在此模型中，切除 87% 的肝脏质量导致 100% 致死率。相比之下，表达 A20 的小鼠超过 60% 在 87% LR 后存活，并通过肝脏中 PCNA（增殖细胞核抗原）阳性细胞核的数量评估，表现出再生能力增强。这些结果使 A20 成为参与加速肝脏再生和促进肝细胞存活和功能的关键基因，即使在面临极端代谢需求时也是如此。这些令人鼓舞的结果促使该提案的提交。我们的具体目标是 i) 在体外剖析 A20 在肝细胞中 (1) 抗凋亡和 (2) 促增殖功能的分子基础，以及 ii) 确认使用 A20 进行肝脏定向基因治疗将产生有益影响FHF 的毒性、FAS 介导和手术实验模型。从基础科学的角度来看，拟议的体外工作将解决 A20 对转录因子和参与肝细胞凋亡、活化和增殖的基因表达的影响。这应该揭示我们对肝细胞生物学理解中的许多未知因素，并可能导致新治疗靶点的发现。从治疗的角度来看，A20 在 FHF 小鼠体内模型中的有益作用的验证应该为将该方法扩展到非人类灵长类动物 FHF 模型以及潜在的临床应用奠定基础。用于基因转移的新型更安全和组织特异性病毒载体的产生以及将蛋白质递送至细胞的非病毒方法的开发将促进基于 A20 的疗法的临床转化。