Hemochromatosis - Epidemiology and Molecular Mechanisms

描述（由申请人提供）：本项目旨在更好地了解铁体内平衡的调节。铁体内平衡失调导致铁超载或缺铁。铁稳态的主要调节因子是由25个氨基酸组成的肽hepcidin。由HFE、TfR2和HJV基因突变引起的血色素沉着病主要是由于hepcidin转录失调。我们最近发现一种由Tmprss6基因编码的血清蛋白酶是hepcidin的有效下调因子。对铁的强烈转录反应仅在完整的动物中观察到。为了了解铁对hepcidin的正常转录调控，也为了深入了解已知参与其调控的基因产物的作用，我们将研究hepcidin在肝脏中的转录，肝脏是野生型小鼠和缺乏这些基因的突变小鼠的主要合成位点。在双管齐下的方法中，我们将：1)研究从铁刺激的小鼠制备的核提取物；2)研究hepcidin启动子报告结构在我们设计的体内系统中的反应。经典的离体技术包括EMSA， ChIP测定，DNA亲和纯化和质量分光光度鉴定将被用于鉴定核提取物中的转录因子。这些研究将揭示哪些转录因子参与了对铁的反应，以及这种反应如何在突变动物中被修改。在纯合C282Y HFE突变的患者中，血色素沉着症表型的表达有显著的可变性。迄今为止的研究未能确定遗传或环境因素，这些因素只占已知变异性的很小一部分。我们将根据前几代的铁吸收经验，在小鼠中研究是否存在铁吸收的跨代表观遗传修饰。喂养缺铁或含铁过量的小鼠，它们的后代在断奶时吃同样的食物或吃正常的食物。那些断奶后缺乏铁或铁过量的人将会再繁殖几代。如果发生跨代表观遗传修饰，那么铁缺乏小鼠转移到正常饮食的铁负荷将大于来自铁重祖的小鼠。公共卫生相关性：通过研究各种铁超载和铁缺乏的小鼠模型，我们将了解身体如何调节铁。这将为治疗铁超载、慢性炎症贫血以及血色素沉着症本身打开大门。如果我们能够证明铁吸收的调节是表观遗传的，这一发现不仅可以解释血色素沉着症的可变外显率，而且可以极大地改变我们对自然选择如何在几代人的时间内起作用的看法。

DESCRIPTION (provided by applicant): The purpose of this project is to better understand the regulation of iron homeostasis. Dysregulation of iron homeostasis results in iron overload or iron deficiency. The major regulator of iron homeostasis is the 25 amino acid peptide hepcidin. Hemochromatosis caused by genetic mutations in HFE, TfR2 and HJV is due primarily to dysregulation of hepcidin transcription. We have recently discovered that a serum protease encoded by the Tmprss6 gene is a potent downregulator of hepcidin. A robust transcriptional response to iron is only observed in intact animals. To understand normal transcriptional regulation of hepcidin by iron and also to gain insight into the roles of the products of the genes known to be involved in its regulation, we will investigate hepcidin transcription in liver, the primary site of synthesis in wildtype mice and mutant mice lacking these genes. In a two-pronged approach we will: 1) study nuclear extracts prepared from mice stimulated with iron; and 2) investigate the response of hepcidin promoter reporter constructs in an in vivo system that we have devised. Classical ex vivo techniques including EMSA, ChIP assays, DNA affinity purification and mass spectrophotometric identification will be employed to identify transcription factors in nuclear extracts. These studies will reveal which transcription factors are involved in the response to iron and how this response is modified in mutant animals. There is marked variability in the expression of the hemochromatosis phenotype in patients homozygous for the C282Y HFE mutation. Studies to date have failed to identify either genetic or environmental factors that account for any but a very small portion of the known variability. We will investigate, in mice, whether there is transgenerational epigenetic modification of iron absorption, based on the experience of prior generations with iron. Mice receiving iron-deficient diets or diets containing iron excess will be bred and their offspring will either be weaned to the same diet or to a normal diet. Those weaned to a deficient or iron excess diet will be bred for additional generations. If transgenerational epigenetic modification occurs, then the iron burden of iron deficient mice moved to normal diet will be greater than that of mice derived from iron-heavy progenitors. PUBLIC HEALTH RELEVANCE: By studying various mouse models of iron overload and iron deficiency, we will gain an understanding how the body regulates iron. This will open the door to treatments of iron overload, the anemia of chronic inflammation, as well as of hemochromatosis itself. If we are able to demonstrate that the regulation of iron absorption is inherited epigenetically, this finding would not only explain the variable penetrance of hemochromatosis, but would greatly alter our perception of how natural selection functions over the span of only a few generations.