Liver-Enriched Transcription Factors

肝细胞核因子4α (hnf4α)在肝脂质稳态中的作用：肝脏对营养信号的适应是由细胞内代谢物调节的几种转录因子调节的。一个由HNF4alpha控制的转录因子网络，在禁食和摄食条件下协调脂肪酸转运和代谢酶的相互表达。Hes6被确定为一种新的HNF4alpha靶点，在正常喂养的动物中，Hes6与HNF4alpha一起维持低水平的PPARgamma表达，并抑制几种pparalpha调节基因。在禁食期间，Hes6表达减少，过氧化物酶体增殖物激活受体α (ppar α)取代靶基因调控区域的hnf4 α /Hes6复合物来激活转录。基因表达和启动子占用分析证实HNF4alpha在体内是Pparalpha基因的直接激活因子，其表达受Pparalpha和Hes6蛋白的反馈调控。这些结果确立了HNF4alpha、Hes6、PPARalpha和PPARgamma之间的动态调控相互作用在脂肪酸转运和代谢相关基因的协调表达中的基本作用。HNF4alpha在肠道中的作用：HNF4alpha在从十二指肠到结肠和从隐窝到绒毛的肠上皮中高度表达。这种不断更新的上皮细胞的稳态依赖于增殖、分化和凋亡的综合控制，以及上皮细胞的功能结构。为了确定HNF4alpha在成人肠上皮中丢失的后果，我们使用了他莫昔芬诱导的ccr - loxp系统来灭活HNF4alpha基因。hnf4 α缺失导致隐窝增殖增加，Wnt/ β -连环蛋白系统控制的几个基因表达增加。体外实验证实了HNF4alpha对Wnt/ β -catenin信号通路的控制。细胞谱系受到影响，如杯状细胞数量增加和肠细胞和肠内分泌细胞成熟受损所示。在缺乏HNF4alpha的情况下，细胞-细胞连接不稳定，细胞旁肠通透性增加。这些结果表明，hnf4α调节Wnt/ β -连环蛋白信号并控制肠上皮稳态、细胞功能和细胞结构。HNF4alpha调节肠道增殖和分化之间的平衡，并可能在肠道中发挥肿瘤抑制作用。C/EBPalpha在肺发育中的作用：早产儿呼吸系统并发症与慢性阻塞性肺疾病（COPD）易感性之间的联系正受到越来越多的关注。我们之前发现，与健康吸烟者相比，COPD患者气道上皮细胞中的CCAAT/增强子结合蛋白（C/EBP）活性降低，提示C/EBP在COPD发病机制中的作用此前未知。为了研究转录因子C/EBPalpha在肺发育中的作用及其在COPD中的潜在作用，我们制造了肺上皮特异性破坏C/EBPalpha基因的小鼠。Cebpa（dLE）小鼠表现出肺发育和上皮分化受损，以及血管功能受到影响。存活至成年的Cebpa（dLE）小鼠出现不规则肺气肿的严重病理表现；细支气管炎，包括杯状细胞增生、细支气管化生、纤维化和粘液堵塞；炎症细胞和基因表达谱与COPD相似。Cebpa（dLE）小鼠在发育过程中表现出肺不成熟，成年Cebpa（DeltaLE）小鼠表现出COPD的大部分组织病理学和炎症特征。因此，Cebpa（dLE）小鼠可以为理解肺不成熟的长期后果以及与COPD易感性的联系提供新的有价值的见解。芳烃受体（AhR）在肝脂肪变性中的作用：为了确定AhR在肝脂肪变性中的内源性作用，我们使用野生型、组成型激活的AhR转基因、AhR缺失和CD36/脂肪酸转位酶缺失的小鼠来研究AhR是否影响脂肪变性，并确定CD36参与AhR的脂肪变性作用。激活AhR诱导以甘油三酯积累为特征的自发性肝脂肪变性。AhR的脂肪变性作用可能是由于CD36和脂肪酸转运蛋白的上调、脂肪酸氧化的抑制、甘油三酯肝脏输出的抑制、外周脂肪动员的增加以及肝脏氧化应激的增加。启动子分析证实CD36是AhR的一个新的转录靶点。肝细胞中AhR的激活诱导CD36基因表达并增强脂肪酸摄取。AhR激动剂的脂肪变性作用在CD36-/-小鼠中被抑制。结论：我们的研究揭示了ahr诱导的脂肪变性与CD36表达之间的新联系。工业或军事接触二恶英及相关化合物与人类脂肪肝患病率增加有关。本研究结果可能有助于建立AhR及其靶点CD36作为脂肪肝疾病新的治疗和预防靶点。版权所有(c) 2010 AGA研究所。Elsevier Inc.出版。版权所有。AhR在免疫系统中的作用：AhR KO小鼠对脂多糖（LPS）诱导的脓毒性休克过敏，主要是由于其巨噬细胞功能障碍。在LPS的作用下，AhR KO小鼠的骨髓源性巨噬细胞（BMDM）分泌大量的白细胞介素-1 β (il -1 β)。由于通过表达Pai-2的腺病毒转导补充纤溶酶原激活物抑制剂-2 （Pai-2）表达可抑制il -1 β分泌的增强，因此Pai-2表达的降低可能是AhR KO小鼠BMDM分泌il -1 β增加的原因之一。基因表达分析显示，AhR通过NF-kappaB而非Arnt参与的机制，以lps依赖的方式直接调控Pai-2的表达。再加上AhR配体3-甲基胆蒽部分保护野生型AhR小鼠免于内毒素诱导的死亡，这些结果提出了一种合适的AhR配体可能对治疗炎症性疾病患者有用的可能性。AhR在皮肤癌变中的作用：苯并[a]芘（B[a]P）是AhR的配体。为了研究AhR/Arnt在皮肤中的作用，我们培育了在成年皮肤表皮中特异性破坏Arnt基因的小鼠。皮肤中缺乏AhR活性没有明显的病理影响；表皮发育正常。为了研究Arnt在皮肤癌中的作用，将突变体烟酰胺腺嘌呤二核苷酸（磷酸）：醌氧化还原酶（NAPQ1）等位基因引入Arnt皮肤缺失小鼠品系，使其更容易被B[a]P。在肿瘤启动-促进方案中，该菌株表皮缺乏AhR/Arnt活性完全阻止了皮肤肿瘤的诱导，在该方案中，单次局部应用B[a]P作为肿瘤启动事件，而重复局部应用12- o -十四烷醇-13-醋酸酯（TPA）提供肿瘤促进作用。相反，在同样使用TPA作为启动子但使用n -甲基-n '-硝基-n -亚硝基胍作为引发剂的方案中，破坏Arnt并没有阻止皮肤肿瘤的诱导，其活性不太可能受到AhR、Arnt或其靶基因活性的影响。这些观察结果表明，Arnt是皮肤中B[a]P引发肿瘤所必需的。

Role of hepatocyte nuclear factor 4alpha (HNF4alpha) in hepatic lipid homeostasis: Adaptation of liver to nutritional signals is regulated by several transcription factors that are modulated by intracellular metabolites. A transcription factor network under the control of HNF4alpha that coordinates the reciprocal expression of fatty acid transport and metabolizing enzymes during fasting and feeding conditions. Hes6 is identified as a novel HNF4alpha target, which in normally fed animals, together with HNF4alpha, maintains PPARgamma expression at low levels and represses several PPARalpha-regulated genes. During fasting, Hes6 expression is diminished, and peroxisome proliferator-activated receptor alpha (PPARalpha) replaces the HNF4alpha/Hes6 complex on regulatory regions of target genes to activate transcription. Gene expression and promoter occupancy analyses confirmed that HNF4alpha is a direct activator of the Pparalpha gene in vivo and that its expression is subject to feedback regulation by PPARalpha and Hes6 proteins. These results establish the fundamental role of dynamic regulatory interactions between HNF4alpha, Hes6, PPARalpha, and PPARgamma in the coordinated expression of genes involved in fatty acid transport and metabolism. Role of HNF4alpha in the intestine: HNF4alpha is highly expressed in the intestinal epithelium from duodenum to colon and from crypt to villus. The homeostasis of this constantly renewing epithelium relies on an integrated control of proliferation, differentiation, and apoptosis, as well as on the functional architecture of the epithelial cells. In order to determine the consequences of HNF4alpha loss in the adult intestinal epithelium, we used a tamoxifen-inducible Cre-loxP system to inactivate the Hnf4alpha gene. Loss of HNF4alpha led to increased proliferation in crypts and increased expression of several genes controlled by the Wnt/beta-catenin system. This control of the Wnt/beta-catenin signaling pathway by HNF4alpha was confirmed in vitro. Cell lineage was affected, as indicated by an increased number of goblet cells and an impairment of enterocyte and enteroendocrine cell maturation. In the absence of HNF4alpha, cell-cell junctions were destabilized and paracellular intestinal permeability increased. These results showed that HNF4alpha modulates Wnt/beta-catenin signaling and controls intestinal epithelium homeostasis, cell function, and cell architecture. HNF4alpha regulates the intestinal balance between proliferation and differentiation, and that it might act as a tumor suppressor in the gut. Role of C/EBPalpha in lung development: The link between respiratory complications in prematurely born infants and susceptibility for developing chronic obstructive pulmonary disease (COPD) is receiving increasing attention. We have previously found that CCAAT/enhancer binding protein (C/EBP) activity in airway epithelial cells of COPD patients is decreased compared to healthy smokers, suggesting a previously unknown role for C/EBPs in COPD pathogenesis. To investigate the role of the transcription factor C/EBPalpha in lung development and its potential role in COPD, mice with a lung epithelial-specific disruption of the C/EBPalpha gene were generated. Cebpa(dLE) mice exhibit impaired lung development and epithelial differentiation, as well as affected vascularity. Cebpa(dLE) mice that survive until adulthood develop a severe pathological picture with irregular emphysema; bronchiolitis, including goblet cell hyperplasia, bronchiolar metaplasia, fibrosis and mucus plugging; and an inflammatory cell and gene expression profile similar to COPD. Cebpa(dLE) mice display lung immaturity during development, and adult Cebpa(DeltaLE) mice develop a majority of the histopathological and inflammatory characteristics of COPD. Cebpa(dLE) mice could thus provide new valuable insights into understanding the long-term consequences of lung immaturity and the link to susceptibility of developing COPD. Role of the aryl hydrocarbon receptor (AhR) in hepatic steatosis: To determine the endobiotic role of AhR in hepatic steatosis, wild-type, constitutively activated AhR transgenic, AhR null and CD36/fatty acid translocase null mice were used to investigate whether AhR influences steatosis and to determine the involvement of CD36 in the steatotic effect of AhR. Activation of AhR induced spontaneous hepatic steatosis characterized by the accumulation of triglycerides. The steatotic effect of AhR likely is owing to the combined up-regulation of CD36 and fatty acid transport proteins, suppression of fatty acid oxidation, inhibition of hepatic export of triglycerides, increase in peripheral fat mobilization, and increased hepatic oxidative stress. Promoter analysis established CD36 as a novel transcriptional target of AhR. Activation of AhR in liver cells induced CD36 gene expression and enhanced fatty acid uptake. The steatotic effect of an AhR agonist was inhibited in CD36-/- mice. CONCLUSIONS: Our study reveals a novel link between AhR-induced steatosis and the expression of CD36. Industrial or military exposures to dioxin and related compounds have been linked to increased prevalence of fatty liver in human beings. Results from this study may help to establish AhR and its target CD36 as novel therapeutic and preventive targets for fatty liver disease. Copyright (c) 2010 AGA Institute. Published by Elsevier Inc. All rights reserved. Role of AhR in the immune system: AhR KO mice are hypersensitive to lipopolysaccharide (LPS)-induced septic shock, mainly due to the dysfunction of their macrophages. In response to LPS, bone marrow-derived macrophages (BMDM) of AhR KO mice secreted an enhanced amount of interleukin-1beta (IL-1beta). Since the enhanced IL-1beta secretion was suppressed by supplementing plasminogen activator inhibitor-2 (Pai-2) expression through transduction with Pai-2-expressing adenoviruses, reduced Pai-2 expression could be a cause of the increased IL-1beta secretion by AhR KO mouse BMDM. Analysis of gene expression revealed that AhR directly regulates the expression of Pai-2 through a mechanism involving NF-kappaB but not Arnt, in an LPS-dependent manner. Together with the result that administration of the AhR ligand 3-methylcholanthrene partially protected mice with wild-type AhR from endotoxin-induced death, these results raise the possibility that an appropriate AhR ligand may be useful for treating patients with inflammatory disorders. Role of AhR is skin carcinogenesis: Benzo[a]pyrene (B[a]P) is a ligand for the AhR. To study the role of AhR/Arnt is skin, mice were developed in which the Arnt gene was disrupted specifically in adult skin epidermis. There was no overt pathological effect of lack of AhR activity in the skin; the epidermis developed normally. TO study the role of Arnt in skin cancer, the mutant nicotinamide adenine dinucleotide (phosphate): quinone oxidoreductase (NAPQ1) allele was introduced into the Arnt skin-null mouse strain to render it more susceptible to tumor initiation by B[a]P. Lack of AhR/Arnt activity in the epidermis of this strain completely prevented the induction of skin tumors in a tumor initiation-promotion protocol in which a single topical application of B[a]P acted as the tumor-initiating event, and tumor promotion was provided by repeated topical applications of 12-O-tetradecanoyl phorbol-13-acetate (TPA). In contrast, disruption of Arnt did not prevent the induction of skin tumors in a protocol also using TPA as the promoter but using as the initiator N-methyl-N'-nitro-N-nitrosoguanidine, whose activity is unlikely to be affected by the activity of AhR, Arnt or their target genes. These observations demonstrate that Arnt is required for tumor initiation by B[a]P in the skin.