Heme Oxygenase-1 Gene Expression by TGF-Beta 1

TGF-Beta 1 的血红素加氧酶 1 基因表达

• 批准号: 7245147
• 负责人: NATHALIE HILL-KAPTURCZAK
• 金额: $ 21.19万
• 依托单位: UNIVERSITY OF ALABAMA AT BIRMINGHAM
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-06-15 至 2009-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7245147
• 关键词: Age; Animal Model; Apoptosis; Attention; Attenuated; Bilirubin; Binding; Biological; Cell Proliferation; Cells; Complement Factor B; Complex; DNA Sequence; Data; Deposition; Development; Disease; Dominant-Negative Mutation; Electrophoretic Mobility Shift Assay; Enzymes; Epithelial; Epithelial Cells; Equilibrium; Exposure to; Extracellular Matrix; Fibronectins; Fibrosis; Gene Expression; Genes; Homeostasis; Human; In Vitro; Inflammation; Injury; Kidney; Kidney Diseases; Ligation; Mediating; Modeling; Molecular; Mus; Pathogenesis; Polymerase Chain Reaction; Production; Protein Overexpression; Proteins; Reaction; Regulation; Regulatory Element; Reporting; Research Personnel; Role; Signal Transduction; Site; Site-Directed Mutagenesis; Small Interfering RNA; Testing; Tissues; Trans-Activators; Transfection; Transforming Growth Factors; Tubular formation; Ureteral obstruction; Week; cell injury; cytokine; genetic regulatory protein; heme oxygenase-1; human TGFB1 protein; in vivo; in vivo Model; kidney cell; programs; promoter; protective effect; protein expression; response; transcription factor; urinary tract obstruction

DESCRIPTION (provided by applicant): Transforming growth factor-B (TGF-B) is a regulatory cytokine that is implicated in a variety of kidney diseases where it promotes extracellular matrix deposition and fibrosis. Paradoxically, TGF-B has a critical role in normal homeostasis and stabilizes and attenuates tissue injury through mechanisms that are not clearly understood. We previously reported that, in renal tubular epithelial cells, TGF-B1 is a potent inducer of a cytoprotective enzyme, heme oxygenase-1 (HO-1). The overall hypothesis of this proposal is that HO-1 induction is a cytoprotective response to TGF-B1mediated cellular injury in the kidney. The mechanisms underlying the induction of HO-1 by TGF-B as well as the effects of HO-1 expression and its products on the renal epithelial cellular responses to TGF-B1 are not known. In preliminary studies, we have observed that kidneys from HO-1-/- mice (age 24 weeks) show significantly higher fibronectin expression than age matched HO-1 () mice. In contrast, HO-1 induction is associated with decreased levels of fibronectin. Furthermore, bilirubin, 1of the products of the HO-1 catalyzed reaction, significantly lowers TGF-B1-mediated increases in fibronectin levels in renal tubular epithelial cells. Our studies also demonstrate that the inhibitory Smad, Smad7, blocks HO-1 induction by TGF-B1 in renal epithelial cells. The in vivo relevance of the inverse relationship between Smad7 and HO-1 was also observed in kidneys from TGF-p overexpressing mice. TGF-B1-mediated HO-1 induction is transcriptionally regulated and requires a cis-acting region between-9.1 and-9.4kb of the human HO-1 promoter and may involve Sp1-like sequences. This proposal will evaluate the biological role of HO-1 induction in response to TGF-B1 in vitro, using HO-1 deficient cells (primary cells derived from HO-1 -/-, +/+ mice and HO-1 knockdown using siRNA in HK-2 cells) as well as HO-1 overexpressing HK-2 cells (Aim I). HO-1 -/- mice and their littermates will be used to study the role of HO-1 and HO reaction products in the pathogenesis of fibrosis in a model for obstructive uropathy and renal fibrosis, the unilateral ureteral obstruction (DUO) model (Aim II). Finally, the molecular regulation of TGF-B1 -mediated HO-1 gene expression will be characterized (Aim III) and integrated with Aim 1 by testing the effects of overexpression or dominant negative expression of the protein/transcription factor identified on the cellular responses to TGF-B1. Studies to understand the molecular mechanisms involved in the induction of HO-1 by TGF-B1 are timely and relevant for efforts to fine tune endogenous HO-1 activity in renal injury.

描述（由申请人提供）：转化生长因子-B（TGF-B）是一种调节性细胞因子，与多种肾脏疾病有关，它促进了细胞外基质沉积和纤维化。矛盾的是，TGF-B在正常的稳态中具有关键作用，并通过未清楚理解的机制稳定并减轻组织损伤。我们先前报道说，在肾小管上皮细胞中，TGF-B1是细胞保护酶的有效诱导剂，血红素加氧酶-1（HO-1）。该提议的总体假设是HO-1诱导是对肾脏中TGF-B1介导的细胞损伤的细胞保护反应。 TGF-B诱导HO-1的基础机制以及HO-1表达及其产物对TGF-B1肾上皮细胞反应的影响尚不清楚。在初步研究中，我们观察到来自HO-1 - / - 小鼠（24周）的肾脏表达明显高于年龄匹配的HO-1（）小鼠。相反，HO-1诱导与纤连蛋白水平降低有关。此外，HO-1催化反应的产物1的胆红素显着降低了肾小管上皮细胞中纤连蛋白水平的TGF-B1介导的增加。我们的研究还表明，抑制性SMAD SMAD7在肾上皮细胞中阻止了TGF-B1诱导HO-1。在TGF-P过表达小鼠的肾脏中，还观察到Smad7和HO-1之间的反比关系的体内相关性。 TGF-B1介导的HO-1诱导受转录调节，需要人类HO-1启动子的-9.1和-9.4KB之间的顺式作用区域，并且可能涉及SP1样序列。该提案将使用HO-1缺陷细胞（源自HO-1 - / - ， +/ +小鼠衍生出的原代细胞，使用siRNA衍生出HO-1细胞，使用HO-1缺乏细胞评估HO-1诱导对TGF-B1的生物学作用在HK-2细胞中）以及HO-1过表达HK-2细胞（AIM I）。 HO-1 - / - 小鼠及其同窝材料将用于研究HO-1和HO反应产物在纤维化发病机理中的作用，在阻塞性尿道病和肾纤维化模型中，单侧输尿管阻塞（DUO）模型（AIM） ii）。最后，将表征TGF-B1介导的HO-1基因表达的分子调节（AIM III），并通过测试鉴定出对细胞对细胞反应的蛋白质/转录因子的过表达或显性负表达的影响，与AIM 1进行了整合。 TGF-B1。了解TGF-B1诱导HO-1涉及的分子机制的研究及时且与对肾脏损伤中内源性HO-1活性进行微调的努力相关。