Endoplasmic Reticulum stress and thyroid cell death

内质网应激和甲状腺细胞死亡

• 批准号: 10414536
• 负责人: PETER ARVAN
• 金额: $ 39万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-04-01 至 2027-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10414536
• 关键词: Alleles; Animal Model; Animals; Attention; Cell Death; Cell membrane; Cells; Cessation of life; Clinical; Cretinism; Curiosities; Development; Diabetes Mellitus; Disease; Dwarfism; Endoplasmic Reticulum; Engineering; Environment; Epithelial Cells; Evolution; Exhibits; Failure; Frequencies; Gene Mutation; Generations; Genetic; Glycoproteins; Goiter; Grant; Growth; Growth Disorders; Heterozygote; Homozygote; Human; Human Inbreeding; Hyperplasia; Hypothyroidism; Inbreeding; Individual; Infiltration; Inflammatory; Ingestion; Inherited; Intake; Iodides; Iodination reaction; Knock-in; Knock-in Mouse; Knockout Mice; Life; Link; Livestock; Malignant Neoplasms; Malignant neoplasm of thyroid; Mediating; Mind; Modeling; Mus; Mutation; Nerve Degeneration; Pathogenicity; Patients; Persons; Physiology; Population; Proteins; Rattus; Reporting; Role; Serum; Severities; Sheep; System; TG gene; Testing; Thyroglobulin; Thyroid Adenoma; Thyroid Diseases; Thyroid Gland; Thyroid Hormones; Thyroiditis; Thyroxine; Tissues; Travel; Triiodothyronine; apical membrane; autosomal recessive trait; blocking factor; cell growth; cytotoxicity; endoplasmic reticulum stress; human disease; in vivo; misfolded protein; mouse model; mutant; oxidation; protein degradation; proteotoxicity; trafficking

ABSTRACT Efficient generation of the thyroid hormone thyroxine (T4) requires the iodination of thyroglobulin (Tg), which is synthesized in the endoplasmic reticulum (ER) and undergoes folding and trafficking to the thyroid follicle lumen wherein iodination takes place. Endogenous T4 is 100% produced by thyrocytes, and the formation of T4 in Tg has been conserved in evolution for ≥ 500 million years. To date, hundreds of different pathogenic mutations of the TG gene have been found to cause congenital hypothyroidism in humans. Genetic hypothyroidism from homozygous (or compound heterozygous) TG mutation is rare, but the frequency of a single pathogenic TG allele in the human population is very common (≥ 1:200 individuals). All Tg mutants studied to date are misfolded proteins trapped in the ER, causing ER stress. We recently reported that in untreated TG homozygotes, endogenous thyroid hormone synthesis still occurs despite failure of Tg export from the ER. The mechanism involves thyroid epithelial cell death, with extrusion of dead thyrocytes into the lumen of thyroid follicles, leading to the disintegration and iodination of those cells in the follicle lumen. Massive expression of Tg protein in thyrocytes, and the extremely high frequency of heterozygous TG mutations in the population, now leads us to ask whether thyroid epithelial cell death may also be a (heretofore unsuspected) widespread feature across individuals who are simple heterozygotes for mutant TG. This can be easily tested in animal models. Additionally, in homozygous rdw/rdw (Tg-G2298R) rats, thyroid cell death has long been recognized as a factor blocking goiter growth. Remarkably, we now find that in the congenital goiter mouse (cog/cog, Tg-L2263P), there is also widespread thyroid cell death (that is nevertheless outpaced by thyroid growth). In the current proposal, 1) we have engineered a rdw/rdw knockin mouse and will directly compare cell growth and cell death to that seen in cog/cog mice. 2) We will examine thyroid cell death in simple heterozygotes of these two models, which have perfectly normal serum T4 levels but exhibit thyrocyte ER stress. 3) As heterozygous mutant Tg is misfolded in the ER, it may be degraded at least in part by ER-associated protein degradation (ERAD). We find that defective ERAD of Tg leads to a thyroiditis with infiltration of inflammatory cells and follicle involuion. 4) Extrusion of thyrocytes into the follicle lumen may expose non-apical portions of the cell to iodination by surrounding follicular cells as a contributing mechanism of cytotoxicity. To test this, we will examine a TG-KO mouse model in which TSH-stimulated overgrowth (and not ER stress) drives thyrocytes into the follicle lumen, exposing the sensitive basolateral plasma membrane to the oxidative / iodination environment of the lumen, where it may trigger thyroid cell death. In summary, all of the Aims of this proposal will highlight questions of ER stress- dependent and independent mechanisms of thyrocyte death.

摘要 有效地产生甲状腺激素甲状腺激素(T4)需要甲状腺球蛋白(TG)的碘化，这是 在内质网(ER)中合成，并经历折叠和运输到甲状腺滤泡腔 在那里进行碘化作用。内源性T4 100%由甲状腺细胞产生，T4在TG中的形成 已经在进化中保守了≥5亿年。到目前为止，数百种不同的致病突变 已发现TG基因会导致人类先天性甲状腺功能减退症。遗传性甲状腺功能减退症 纯合子(或复合杂合子)TG突变很少见，但单一致病TG的频率 等位基因在人类群体中非常常见(≥1：200个体)。到目前为止，所有研究的TG突变体都被错误折叠 蛋白质被困在内质网，造成内质网应激。我们最近报道，在未经治疗的TG纯合子中， 内源性甲状腺激素合成仍然发生，尽管内质网输出甘油三酯失败。这一机制 涉及甲状腺上皮细胞死亡，死亡的甲状腺细胞被挤出到甲状腺滤泡的管腔内，导致 导致滤泡腔内细胞的解体和碘化。Tg蛋白在大肠杆菌中的大量表达 甲状腺细胞，以及人群中极高频率的杂合子TG突变，现在引导我们 问一问甲状腺上皮细胞死亡是否也是一种(迄今未被怀疑的)普遍特征 在突变TG的单纯杂合子个体中。这可以很容易地在动物模型中进行测试。 此外，在纯合子RDW/RDW(TG-G2298R)大鼠中，甲状腺细胞死亡长期以来一直被认为是一个因素 阻止甲状腺肿的生长。值得注意的是，我们现在发现在先天性甲状腺肿小鼠(cog/cog，TG-L2263P)中，有 还有广泛的甲状腺细胞死亡(然而，甲状腺细胞的生长速度超过了这一速度)。在目前的提案中， 1)我们已经设计了一只RDW/RDW敲击小鼠，并将直接将细胞生长和死亡与之进行比较 可见于啮齿/啮齿小鼠。2)我们将检测这两个模型的单纯杂合子中的甲状腺细胞死亡。 血清T4水平完全正常，但表现出甲状腺细胞内质网应激。3)杂合子突变体Tg折叠错误 在内质网中，它可能至少部分地被内质网相关蛋白降解(ERAD)所降解。我们发现 甘油三酯ERAD缺陷可导致甲状腺炎症，并伴有炎症细胞和卵泡退变。4)拉伸 甲状腺细胞进入滤泡腔可能会使细胞的非根尖部分暴露在碘的作用下 滤泡细胞作为细胞毒性的一个贡献机制。为了测试这一点，我们将检查一个TG-KO小鼠模型 在这种情况下，TSH刺激的过度生长(而不是ER应激)将甲状腺细胞驱动到滤泡腔，暴露了 对管腔的氧化/碘化环境敏感的基侧质膜，在那里它可能 引发甲状腺细胞死亡。总而言之，该提案的所有目标都将突出ER压力的问题- 甲状腺细胞死亡的依赖和独立机制。