Pathogenesis and Treatment of Liver Disease in Transaldolase Deficiency

转醛醇酶缺乏性肝病的发病机制和治疗

• 批准号: 8501433
• 负责人: Andras Perl
• 金额: $ 31.62万
• 依托单位: UPSTATE MEDICAL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-07-01 至 2015-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8501433
• 关键词: 4 hydroxynonenal; Acetaminophen; Acetylcysteine; Acute; Acute Liver Failure; Age; Aldehyde Reductase; Aminophenols; Animal Model; Antioxidants; Apoptosis; Blood donor; Cancer Etiology; Case Study; Caucasians; Caucasoid Race; Cause of Death; Cell Death; Cells; Cessation of life; Child; Cirrhosis; Code; Defect; Development; Disease; Dysplasia; Enzymes; Exhibits; Failure; Fatty Liver; Fibroblasts; Gene Mutation; Genes; Genetic; Genomics; Glucose; Glucose-6-Phosphate; Glutathione; Hepatocarcinogenesis; Hepatocyte; Human; Human Cloning; In Vitro; Inflammatory; JUN gene; Link; Lipid Peroxides; Lipids; Liver; Liver Cirrhosis; Liver Failure; Liver diseases; Malignant Neoplasms; Malondialdehyde; Mediating; Membrane Potentials; Metabolic; Metabolic Pathway; Mitochondria; Molecular; Mus; Mutation; NADP; Overdose; Oxidation-Reduction; Oxidative Stress; Oxidoreductase; Pathogenesis; Pathway interactions; Patients; Pentosephosphate Pathway; Phase; Phosphogluconate Dehydrogenase; Phosphorylation; Play; Predisposition; Prevalence; Primary carcinoma of the liver cells; Production; Protein Dephosphorylation; Proteins; Recycling; Relative (related person); Resistance; Role; Secondary to; Siblings; Supplementation; TNFRSF6 gene; Testing; Tissues; Toxic effect; Transaldolase; Tylenol; alpha-Fetoproteins; base; cell type; chronic liver disease; computerized data processing; hepatoma cell; in vivo; inhibitor/antagonist; liver cell proliferation; liver inflammation; lymphoblast; mitochondrial dysfunction; mortality; multicatalytic endopeptidase complex; neoplastic cell; neuronal cell body; non-alcoholic fatty liver; nonalcoholic steatohepatitis; novel; polyol; prevent; public health relevance; receptor; research study; ribose-5-phosphate; sedoheptulose 7-phosphate; sugar; transcription factor; tumor; young adult

DESCRIPTION (provided by applicant): Although oxidative stress is implicated in both acute acetaminophen-induced liver failure and chronic liver diseases, cirrhosis and hepatocellular carcinoma (HCC), no common underlying metabolic pathways have been identified. The preliminary results indicate that deficiency of transaldolase (TAL, encoded by the TALDO1 genomic locus), an enzyme of the pentose phosphate pathway (PPP), is an important cause of oxidative stress in the liver that predisposes to acetaminophen-induced liver failure and hepatocarcinogenesis. Recent case reports suggested a link of homozygous mutations in the TAL gene with liver failure and cirrhosis in children. In fibroblasts and lymphoblasts of the first TAL-deficient patient, we found that deletion of S171 caused mis- folding, proteasome-mediated degradation and complete deficiency of enzymatic activity. TAL deficiency resulted in accumulation of sedoheptulose 7-phoshate (S7P) and depletion of glucose 6-phoshate (G6P), indicating a failure to recycle ribose 5-phosphate (R5P) into G6P through the non-oxidative branch of the PPP, thus reducing NADPH production by the oxidative branch. This patient's cells exhibited mitochondrial dysfunction, increased susceptibility to H2O 2 and resistance to Fas apoptosis. To create an animal model of TAL deficiency, we inactivated the TALDO1 genomic locus in the mouse. Cirrhosis and nodular dysplasia were highly prevalent in mice with partial deficiency (TAL+/-; 62/229, 27.0%) or complete deficiency of TAL (TAL-/-; 77/97, 79.4%), relative to wild-type littermates (TAL+/+; 1/102, 1.0%). Cirrhosis invariably followed the formation of microvesicular and macrovesicular lipid droplets indicating nonalcoholic fatty liver disease (NAFLD) and inflammatory changes resembling non-alcoholic steatohepatitis (NASH). HCC was the leading cause of death in both TAL+/- (39/229; 17%) and TAL-/- mice (45/97; 46%) as compared to TAL+/+ littermates (1/102; 1%). Oxidative stress in TAL-deficient livers was characterized by the accumulation of S7P, failure to recycle R5P for the oxidative PPP, depleted NADPH and glutathione levels, and increased production of lipid hydroperoxides (LPO). Reduced 2-catenin phosphorylation and enhanced c-jun expression in TAL-/- livers reflected adaptation to oxidative stress. TAL-/- hepatocytes were resistant to CD95/Fas-mediated apoptosis in vitro and in vivo. Remarkably, lifelong administration of N-acetylcysteine (NAC) blocked acetaminophen susceptibility, restored Fas-dependent apoptosis, phosphorylation of 2-catenin, activation of c-jun and prevented the development of NASH, cirrhosis, and hepatocarcinogenesis in TAL-deficient mice. We also found an increased prevalence of TAL haploinsufficiency in human livers with cirrhosis and HCC. Thus, the preliminary studies identify TAL deficiency as a novel cause of liver cirrhosis and HCC which are preventable by lifelong supplementation of the potent antioxidant NAC. The proposed studies will test the hypothesis that TAL deficiency leads to oxidative stress, through diminished production of NADPH leading to secondary depletion of GSH that underlies the susceptibility of acetaminophen-induced liver failure and chronic liver disease, progressing from NAFLD to NASH, cirrhosis, and HCC. Under Specific Aim 1, we will determine the molecular mechanisms that cause NADPH depletion in TAL-deficient hepatocytes. Under Specific Aim 2, we will test the hypothesis that hepatocarcinogenesis of TAL-deficient mice is triggered by NADPH depletion- induced oxidative stress, mediated by the over-expression of aldose reductase (AR) and the activation of 2- catenin and c-jun, and prevented by post-natal treatment with N-acetylcysteine or AR blockade. Under Specific Aim 3, we will determine the prevalence and molecular basis of TAL deficiency in patients with acetaminophen-induced liver failure, NAFLD, NASH, cirrhosis and HCC. These studies will generate fundamental new information on the pathogenesis and treatment of liver disease. PUBLIC HEALTH RELEVANCE: Hepatocellular carcinoma (HCC) is the fifth most common cancer and the third leading cause of cancer- related death worldwide. Acetaminophen (also known as Tylenol) overdose is the leading cause of acute liver failure in the US. Although oxidative stress has been implicated in HCC and acetaminophen toxicity, the genetic causes that predispose patients to these diseases are unknown. Recently, deficiency of transaldolase (TAL) was discovered in children with liver failure and cirrhosis. TAL is an enzyme that helps the utilization of the sugar glucose in certain cells of the body, such as liver cells, to produce and sustain normal levels of the compounds NADPH and glutathione which protect the integrity of the cells against oxidative stress. We created an animal model of TAL deficiency by a targeted genetic mutation in the mouse. Although mice partially (TAL+/-) or completely deficient of TAL (TAL-/-) develop normally, they are 27-fold and 79-fold more likely to develop cirrhosis and HCC than their siblings having normal TAL gene and activity. TAL-deficient (TAL-/-) mice are also more susceptible to acetaminophen-induced liver failure and death than their siblings carrying the normal TAL gene. Thus, the preliminary studies identify TAL deficiency as a novel cause of liver cirrhosis and HCC as well as acetaminophen-induced liver failure. We hypothesize that NADPH plays critical roles in the survival and multiplication of tumor cells in the liver. The proposed experiments will determine the mechanism of NADPH depletion, whether a correctable defect is responsible for the tumor-causing proliferation of liver cells, and examine the involvement of TAL deficiency in patients with acetaminophen- induced liver failure, fatty liver inflammation, cirrhosis and HCC. These studies should generate fundamental new information on the cause and treatment of potentially fatal diseases of the liver.

描述（由申请人提供）：尽管氧化应激与急性对乙酰氨基酚诱导的肝衰竭和慢性肝病、肝硬化和肝细胞癌（HCC）有关，但尚未确定共同的潜在代谢途径。初步结果表明，缺乏转醛醇酶（TAL，由TALDO 1基因组位点编码），戊糖磷酸途径（PPP）的酶，是肝脏氧化应激的重要原因，易患对乙酰氨基酚诱导的肝功能衰竭和肝癌。最近的病例报告表明TAL基因的纯合突变与儿童肝衰竭和肝硬化有关。在第一个TAL缺陷患者的成纤维细胞和淋巴母细胞中，我们发现S171的缺失导致错误折叠、蛋白酶体介导的降解和酶活性的完全缺乏。TAL缺乏导致7-磷酸景天庚酮糖（S7 P）积累和6-磷酸葡萄糖（G6 P）消耗，表明5-磷酸核糖（R5 P）未能通过PPP的非氧化分支再循环到G6 P，从而减少氧化分支的NADPH产生。该患者的细胞表现出线粒体功能障碍，对H2 O2的敏感性增加和对Fas凋亡的抵抗。为了建立TAL缺乏症的动物模型，我们灭活了小鼠中的TALDO 1基因组位点。肝硬化和结节性发育不良在TAL部分缺乏（TAL+/-; 62/229，27.0%）或TAL完全缺乏（TAL-/-; 77/97，79.4%）的小鼠中相对于野生型同窝小鼠（TAL+/+; 1/102，1.0%）高度流行。肝硬化总是伴随着微泡和大泡脂滴的形成，表明非酒精性脂肪性肝病（NAFLD）和类似非酒精性脂肪性肝炎（NASH）的炎症变化。与TAL+/+同窝小鼠（1/102; 1%）相比，HCC是TAL+/-（39/229; 17%）和TAL-/-小鼠（45/97; 46%）的主要死亡原因。TAL缺乏的肝脏中的氧化应激的特征在于S7 P的积累，未能回收R5 P用于氧化PPP，耗尽NADPH和谷胱甘肽水平，以及脂质过氧化氢（LPO）的产生增加。TAL-/-肝脏中2-catenin磷酸化减少和c-jun表达增强反映了对氧化应激的适应。TAL-/-肝细胞在体外和体内对CD 95/Fas介导的凋亡具有抵抗性。值得注意的是，终身给予N-乙酰半胱氨酸（NAC）阻断了对乙酰氨基酚的敏感性，恢复了Fas依赖性细胞凋亡，2-连环蛋白的磷酸化，c-jun的激活，并防止了TAL缺陷小鼠中NASH，肝硬化和肝癌发生的发展。我们还发现肝硬化和HCC患者中TAL单倍不足的患病率增加。因此，初步研究确定TAL缺乏是肝硬化和HCC的新原因，可以通过终身补充有效的抗氧化剂NAC来预防。拟议的研究将测试TAL缺乏导致氧化应激的假设，通过减少NADPH的产生导致GSH的二次消耗，这是对乙酰氨基酚诱导的肝衰竭和慢性肝病的易感性的基础，从NAFLD进展到NASH，肝硬化和HCC。在特定目标1下，我们将确定导致TAL缺陷肝细胞中NADPH耗竭的分子机制。在特定目标2下，我们将检验以下假设：TAL缺陷型小鼠的肝癌发生是由NADPH耗竭诱导的氧化应激触发的，由醛糖还原酶（AR）的过表达以及2-连环蛋白和c-jun的活化介导，并通过N-乙酰半胱氨酸或AR阻断的产后治疗来预防。在具体目标3下，我们将确定对乙酰氨基酚诱导的肝衰竭、NAFLD、NASH、肝硬化和肝癌患者中TAL缺乏症的患病率和分子基础。这些研究将为肝病的发病机制和治疗提供基本的新信息。 公共卫生相关性：肝细胞癌（HCC）是世界范围内第五大常见癌症和第三大癌症相关死亡原因。对乙酰氨基酚（也称为泰诺）过量是美国急性肝衰竭的主要原因。虽然氧化应激与肝细胞癌和对乙酰氨基酚毒性有关，但使患者易患这些疾病的遗传原因尚不清楚。近年来，转醛醇酶（TAL）的缺陷被发现在儿童肝功能衰竭和肝硬化。TAL是一种酶，有助于利用身体某些细胞（如肝细胞）中的糖葡萄糖，以产生和维持正常水平的化合物NADPH和谷胱甘肽，保护细胞免受氧化应激的完整性。我们通过在小鼠中进行靶向基因突变来创建TAL缺乏症的动物模型。虽然小鼠部分（TAL+/-）或完全缺乏TAL（TAL-/-）正常发展，他们是27倍和79倍更有可能发展肝硬化和肝癌比他们的同胞具有正常TAL基因和活性。TAL缺陷（TAL-/-）小鼠也比携带正常TAL基因的同胞更容易受到对乙酰氨基酚诱导的肝衰竭和死亡。因此，初步研究确定TAL缺乏是肝硬化和HCC以及对乙酰氨基酚诱导的肝衰竭的新原因。我们假设NADPH在肝脏肿瘤细胞的存活和增殖中起关键作用。所提出的实验将确定NADPH耗竭的机制，可纠正的缺陷是否是导致肝细胞肿瘤增殖的原因，并检查TAL缺乏症在对乙酰氨基酚诱导的肝衰竭、脂肪肝炎症、肝硬化和HCC患者中的参与。这些研究应该产生关于潜在致命肝脏疾病的病因和治疗的基本新信息。