Dissecting the Cellular and Molecular Mechanisms Contributing to Craniofacial Fibrous Dysplasia

剖析导致颅面纤维发育不良的细胞和分子机制

• 批准号: 10678639
• 负责人: Kelly Lee Wentworth
• 金额: $ 17.21万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-01 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10678639
• 关键词: Address; Adrenal Gland Neoplasms; Adult; Affect; Area; Automobile Driving; Blindness; CREB1 gene; Cells; Characteristics; Childhood; Clustered Regularly Interspaced Short Palindromic Repeats; Cyclic AMP; Data; Disease; Dysplasia; Engineering; Exhibits; Facies; Freezing; G Protein-Coupled Receptor Signaling; G-Protein-Coupled Receptors; GTP-Binding Proteins; Genes; Genetic Diseases; Genomics; Genotype; Goals; Harvest; Histology; Human; Immunohistochemistry; Knowledge; Lead; Lesion; Malignant Neoplasms; McCune-Albright Syndrome; Medical; Mesoderm; Modeling; Molecular; Morbidity - disease rate; Mosaicism; Mus; Mutation; Neural Crest; Organ; Osteitis Fibrosa Disseminata; Osteoblasts; Pain; Pathogenesis; Pathway interactions; Patients; Phenotype; Phosphorylation; Pituitary Neoplasms; Population; Precocious Puberty; Protocols documentation; Role; Sampling; Scientist; Serum Markers; Signal Pathway; Signal Transduction; Site; Skeleton; Solid; Somatic Mutation; System; Testing; Tissues; WNT Signaling Pathway; Work; activating transcription factor 1; beta catenin; bone; bone fragility; bone loss; bone turnover; career; causal variant; cell type; clinically significant; craniofacial; craniofacial bone; disability; effective therapy; face bone structure; facial disfigurement; hearing impairment; induced pluripotent stem cell; inhibitor; innovation; melanocyte; microCT; mosaic; mosaic nature; mouse model; osteogenic; preference; prevent; receptor; single cell technology; skeletal; skeletal disorder; skeletal dysplasia; skeletal stem cell; skin lesion; skull base; stem cells; therapeutic target; tissue mosaicism; transcriptomics; translational physician

PROJECT SUMMARY/ABSTRACT Craniofacial FD is one of the most common and debilitating skeletal dysplasias. The expansile, fibrous bony lesions that form in the facial bones and skull base cause significant disability, including dysmorphic facies, bone fragility, pain, and vision and hearing loss. There are no effective medical treatments for FD, making this an area of critical medical need. FD can develop in one bone (monoostotic) or multiple bones (polyostotic) and can occur in association with McCune-Albright Syndrome (MAS), which is a somatic mosaic genetic disease characterized by FD, precocious puberty, café-au-lait skin lesions, various endocrinopathies, and solid organ malignancies. FD is arguably the most clinically significant feature of MAS. FD can form at any site within the skeleton, but the most commonly involved sites are the neural crest-derived craniofacial bones. Despite this, little is known about the cellular and molecular mechanisms that drive craniofacial FD, and why there is a predilection for FD formation in neural crest-derived bone. In addition to craniofacial FD, patients with MAS also commonly have mosaic involvement of other neural crest-derived tissues, including melanocytic skin lesions and pituitary and adrenal tumors. The goal of this proposal is to better understand the cellular and molecular mechanisms that drive craniofacial FD so that we can work towards identifying potential treatment targets. We will pursue this goal through the following 3 aims: Aim 1: Test if WNT inhibition reverses craniofacial FD in Col1(2.3)+/Rs1+ mice. We have previously modeled FD in mice by activating Gs-GPCR signaling in osteoblastic cells via the engineered GPCR “Rs1.” Col1(2.3)+/Rs1+ mice develop a dramatic bone phenotype that resembles human FD. We have shown that stopping the abnormal Gs-GPCR signaling reverses the FD lesions, suggesting that FD can be reversed. We also found increased levels of WNT expression in bones harvested from these mice, suggesting that WNT signaling is important in FD. We will use this model to test if inhibition of WNT signaling can reverse craniofacial FD lesions. Aim 2: Determine the GNASR201H mutational burden in human craniofacial FD bone and identify the cell type(s) that harbor this mutation. Despite knowing the causative mutation in FD/MAS, we still do not know which cell type(s) carry the mutation in human FD bone and what level of mosaicism is needed in order to cause disease. We will use an innovative approach to isolate and genotype single cells from human craniofacial FD lesions. This will allow us to understand the mutational burden of human FD lesions and identify the cell type(s) that carry the mutation. Aim 3: Elucidate how the mosaic GNASR201H mutation affects cell multipotency and differentiation capacity using a human iPSC model of FD/MAS. We will use our newly-developed human iPSC model of MAS that contains the causative GNAS mutation in its endogenous locus to explore the effects of GNAS on early cell fate, particularly the neural crest and osteoblast lineages. Together, these aims seek to address the critical knowledge gaps that exist in our understanding of the pathogenesis of craniofacial FD.

项目摘要/摘要 颅面部功能障碍是最常见和最衰弱的骨骼发育不良之一。可膨胀的纤维状骨 在面骨和颅底形成的病变会导致严重的残疾，包括畸形相， 骨骼脆弱、疼痛、视力和听力丧失。目前还没有有效的药物治疗FD，使得 这是一个急需医疗服务的领域。FD可发生在单骨(单骨性)或多骨(多骨性)。 并可与McCune-Albright综合征(MAS)一起发生，MAS是一种体细胞花叶遗传病 以FD、性早熟、咖啡馆皮损、各种内分泌疾病和实质器官为特征 恶性肿瘤。FD可以说是MAS最重要的临床特征。Fd可以在 骨骼，但最常见的受累部位是神经脊来源的头面部骨骼。尽管如此， 关于驱动颅面部功能障碍的细胞和分子机制以及为什么会有一个 神经沟衍生骨中FD形成的偏好。除了头面部FD，MAS患者还 通常有其他神经脊源性组织的马赛克受累，包括黑色素细胞皮肤损害和 脑下垂体和肾上腺肿瘤。这项提议的目标是更好地理解细胞和分子 驱动颅面部功能障碍的机制，以便我们能够努力识别潜在的治疗靶点。我们 将通过以下3个目标实现这一目标：目标1：测试WNT抑制是否逆转颅面部功能障碍 Col1(2.3)+/Rs1+小鼠。我们以前通过在成骨细胞中激活Gs-GPCR信号来建立小鼠FD模型 细胞通过工程的gpr“rs1”。Col1(2.3)+/Rs1+小鼠发展出一种戏剧性的骨表型，类似于 人类功能性消化不良。我们已经证明，停止异常的Gs-GPCR信号可以逆转FD的损害，提示 这个FD是可以逆转的。我们还发现，从这些骨骼中提取的WNT表达水平增加 提示WNT信号在功能性消化不良中起重要作用。我们将使用这个模型来测试WNT的抑制作用 信号转导可以逆转颅面部FD病变。目的2：测定人类GNASR201H突变负荷 并鉴定含有该突变的细胞类型(S)。尽管知道起因 在FD/MAS中突变，我们仍然不知道在人FD骨中是哪种细胞类型(S)携带突变以及水平是什么 需要马赛克才能致病。我们将使用一种创新的方法来分离和分型 取自人颅面部FD病变的单个细胞。这将使我们能够理解人类的突变负担 FD损害并确定携带突变的细胞类型(S)。目的3：阐明镶嵌GNASR201H是如何 利用FD/MAS的人iPSC模型，突变影响细胞的多能性和分化能力。 我们将使用我们新开发的人类MAS IPSC模型，该模型在其 探索GNAS对早期细胞，特别是神经脊和成骨细胞命运的影响的内源性位点 血统。总而言之，这些目标试图解决我们在理解 颅面部FD的发病机制。