THYROID PHYSIOLOGY STUDIES OF INHERITED DISORDERS

本研究计划的广泛目标是通过研究关键调控过程中的遗传缺陷来促进对甲状腺生理学的理解。除了发现新的综合征和基因缺陷外，研究重点还包括通过研究细胞膜运输、代谢和作用缺陷导致甲状腺激素敏感性降低的综合征，从而介导甲状腺激素（TH）的作用。这些分别是由MCT8（单羧酸转运蛋白8）、SBP2（硒代半胱氨酸插入序列结合蛋白2）和TRIl（核TH受体IJ）基因以及其他尚未确定的基因突变引起的。每一种都通过三种方法进行研究，临床（体内）、组织和基因表达（体外）和动物模型（基因改变的小鼠再现了人类的缺陷）。每种方法都是相互补充的，并弥补了它们固有的局限性。1. 临床研究将有助于更好地表征TH的表型，以确定组织和器官特异性缺陷的转运和代谢。研究将包括产前基因分型和尝试治疗。新的变异和不寻常的表型将被寻找。2. 利用患者皮肤成纤维细胞的原代培养和缺陷基因的异源表达进行体外研究，将有助于确定分离的缺陷分子的特性。在SBP2缺陷的情况下，体外研究将有助于确定各种突变对基因转录和翻译的影响。通过免疫组织化学检查患者组织，将有助于在细胞水平上确定缺陷的后果，并验证动物实验结果的相关性。3. 缺乏被研究物质的小鼠和那些在人类中观察到的突变的小鼠，将允许更好地描述所研究的缺陷，并测试各种可能的治疗方式。Mct8基因缺失的小鼠已被证明对人类甲状腺异常的理解是无价的，现在将用于表征胚胎和产后早期生活中Mct8缺失的后果以及产前治疗干预的潜力。诱导Sbp2K0小鼠的创建将有助于确定硒蛋白的表达时间，硒蛋白对生存和发育至关重要，以及它们缺乏的后果。大规模平行测序将用于鉴定与TRIi基因无关的TH抗性基因。因此，这些新基因的功能将被表征。

The broad objective of this research proposal is to advance understanding of thyroid physiology through study of genetic defects at key regulatory processes. In addition to identification of new syndromes and gene defects, research centers on the mediation of thyroid hormone (TH) effects by studying syndromes of reduced sensitivity to TH due to defects in cell membrane transport, metabolism and action. These are caused, respectively, by mutations in the MCT8 (monocarboxylate transporter 8), SBP2 (selenocysteine insertion sequence-binding protein 2) and TRIl (nuclear TH receptor IJ) genes and others not yet identified. Each is studied by a triple approach, clinical (in vivo), tissue and gene expression (in-vitro) and animal models (gene altered mice that recapitulate the defect in humans). Each approach complements the other and compensates for their inherent limitations. 1. Clinical studies will serve to better characterize the phenotype, to identify tissue and organ specific defects in TH transport and metabolism. Studies will include prenatal genotyping and attempted treatment. New variants and unusual phenotypes will be sought. 2. In vitro studies using primary culture of the patients' skin fibroblasts and heterologous expression of the defective genes will serve to determine the properties of the defective molecules in isolation. In the case of SBP2 defects, in vitro studies will serve to determine the effects of various mutations on gene transcription and translation. Examination of patients' tissues by immunohistochemistry, will serve to identify the consequences of the defects at the cell level and to verify the pertinence of results generated from animal experiment. 3. Mice deficient in the substances being studied and those harboring the mutations observed in humans, will allow for better characterization of the defects under investigation and for the testing of various modalities of potential treatment. Mice deficient in the Mct8 gene have proven invaluable in the understanding of the thyroid abnormalities in humans and will now serve to characterize the consequences of Mct8 deficiency in embryonic and early postpartum life and the potentials of prenatal therapeutic intervention. The creation of inducible Sbp2K0 mice will facilitate determination of the timing of expression of selenoproteins that are crucial to survival and development, as well as the consequences of their deficiency. Massively-parallel sequencing will be applied for the identification of the gene(s) involved in resistance to TH not linked to the TRIi gene. Thus, function of such new genes will be characterized.