Genetic Studies of Human Craniofacial Diseases

人类颅面疾病的遗传学研究

• 批准号: 7967091
• 负责人: THOMAS C HART
• 金额: $ 170.36万
• 依托单位: NATIONAL INSTITUTE OF DENTAL & CRANIOFACIAL RESEARCH
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7967091
• 关键词: Affect; Alleles; Amelogenesis Imperfecta; Amino Acids; Animal Model; Area; Bone Density; Bone Marrow; Bone Resorption; Bone Surface; Caring; Cell Culture Techniques; Cell model; Cells; Cervical; Characteristics; Clinical; Clinical Research; Cobalamin; Collagen; Complex; Cyclophosphamide; Data; Defect; Deletion Mutation; Dental; Dental Clinics; Dental Enamel; Dental Papilla; Dentin; Dentin Dysplasia; Dentinogenesis Imperfecta; Development; Diagnosis; Diagnostic; Diagnostic tests; Disease; Distal; Enamel Formation; Etiology; Exhibits; Family; Family Study; Femur; Fluorochrome; Fostering; Gene Expression; Gene Mutation; Gene Proteins; Genes; Genetic; Genetic Polymorphism; Genets; Genotype; Gingiva; Gingival Fibromatosis; Gingival Overgrowth; Goals; Hair; Health Services Accessibility; Healthcare; Hereditary Disease; High Prevalence; Human; Image; Immunohistochemistry; In Vitro; Individual; Inherited; Interferon Type II; Investigation; Label; Laboratories; Mesenchymal; Metabolic Diseases; Minerals; Molecular Biology; Molecular Profiling; Mouth Diseases; Mus; Mutation; National Institute of Dental and Craniofacial Research; Nonsense Mutation; Odontoblasts; Oral; Oral health; Organ; Osteoblasts; Osteoclasts; Osteogenesis; Pathogenesis; Pathology; Patients; Phenotype; Physiology; Progeria; Proteins; Rare Diseases; Recruitment Activity; Research Personnel; Role; Salivary; Serum; Stromal Cells; Subgroup; Syndrome; Testing; Thick; Tissues; Tooth Diseases; Tooth structure; Transgenic Mice; Transgenic Model; Translating; United States National Institutes of Health; Work; Wound Healing; base; bone; cleft lip and palate; craniofacial; density; design; gene environment interaction; human disease; in vivo; insight; interest; methylmalonate; methylmalonic aciduria; monocyte; mutant; novel; orofacial; platelet-derived growth factor C; programs; promoter; research study; substantia spongiosa; tool; treatment strategy; tricho-dento-osseous syndrome

Our working paradigm is that there is a genetic basis to human disease and that understanding the genetic basis of disease will foster development of better diagnostic and treatment strategies. We study Mendelian diseases to identify the underlying gene defect and to understand how the product(s) of this/these gene mutation(s) result in abnormal development or disease. In some cases we have developed animal models (transgenic mice) and in vitro cell models to study disease pathogeneses. Dentinogenesis imperfecta (DI) (10% effort): Genetic mutations of the dentin sialophosphosphoprotein (DSSP) gene are responsible for most cases of DI and dentin dysplasia (DD). Dentin, the most abundant tissue in teeth, is produced by odontoblasts, which differentiate from mesenchymal cells of the dental papilla. Dentin defects are broadly classified into two major types: DIs, types I-III and dentin dysplasias (DDs, types I and II). To date, mutations in DSPP have been found to underlie DI types II and III and DD type II. With the elucidation of the underlying genetic mechanisms has come the realization that the clinical characteristics associated with DSPP mutations appear to represent a continuum of phenotypes. (Hart & Hart, Ortho Craniofac Res, 2009). Recent work (McKnight et al, Hum Mut, 2008) describing studies of 9 families segregating DI or DD has substantiated that the clinical characteristics associated with DSPP mutations appear to represent a continuum of phenotypes. Additionally, studies of Mendelian syndromes with dental findings have determined that mutations of other genes can cause dentin defects including GALNT3 mutations (Dumitrescu et al 2009) and DLX3 mutations. Understanding the genetic basis of diseases of dental importance provides an important first step to the development of diagnostic and treatment strategies to provide better care for affected individuals. To effectively translate clinical and research findings it will also be necessary to develop more effective strategies to educate dental health care clinicians in the field of genetics. (Johnson et al, J Dent Educ, 2008). Tricho-dento-osseous syndrome (TDO) (20 % effort). DLX3 mutations are responsible for TDO which is clinically characterized by anomalies of tooth, hair and bone. A cardinal feature of TDO is an increased thickness and density of bone. To characterize how mutant DLX3 contributes to alterations of bone we generated a transgenic model with mice carrying the 4bp DLX3 mutation driven by a collagen 1A1 promoter. Microcomputed tomographic analyses demonstrated markedly increased trabecular bone volume and bone mineral density in femora from TG mice. In ex vivo experiments, TG mice showed enhanced differentiation of bone marrow stromal cells to osteoblasts and increased expression levels of bone formation markers. However, TG mice did not show enhanced dynamic bone formation rates in in vivo fluorochrome double labeling experiments. Osteoclastic differentiation capacities of bone marrow monocytes were reduced in TG mice in the presence of osteoclastogenic factors and the numbers of TRAP(+) osteoclasts on distal metaphyseal trabecular bone surfaces were significantly decreased. TRACP 5b and CTX serum levels were significantly decreased in TG mice, while IFN-gamma levels were significantly increased. These data demonstrate that increased levels of IFN-gamma decrease osteoclast bone resorption activities, contributing to the enhanced trabecular bone volume and mineral density in these TG mice. These data suggest a novel role for this DLX-3 mutation in osteoclast differentiation and bone resorption. (Choi et al, Dev Biol. 2009). Amelogenesis Imperfecta (AI) (20%effort): We continue to study families segregating the AI phenotype. AI is caused by AMEL, ENAM, MMP20 and KLK4 gene mutations. Mice lacking expression of the AmelX, Enam and Mmp20 genes have been generated, providing tools to better understand enamel formation and AI pathogenesis. (Wright et al., Cells Tissues Organs, 2009). We have continued our linkage studies of families segregating AI to further characterize genotype-phenotype correlations for AI and increase the diagnostic potential for genetics to provide a definitive diagnosis for individuals with AI, a step towards integrating diagnostic testing with care and access to care. We have expanded on the identification of FAM83H gene mutations etiologic for autosomal dominant AI (ADAI) (Hart et al, Clin Genet, 2009). Additionally, our studies of families segregating ADAI have identified multiple novel FAM83H mutations were identified, including two 2-bp-deletion mutations, the first non-nonsense mutations identified. Craniofacial deviation from normal was more prevalent in the affected individuals. Affected individuals having truncating FAMH3H mutations of 677 or fewer amino acids presented a generalized ADHCAI phenotype, while those having mutations capable of producing a protein of at least 694 amino acids had a unique and previously unreported phenotype affecting primarily the cervical enamel. This investigation shows that unique phenotypes are associated with specific FAM83H mutations. (Wright et al, J Dent Res, 2009). Studies of families segregating AI have also provided direct evidence for the existence of at least one additional, currently unidentified gene locus etiologic for ADAI. (Becerik et al, AM J Med Genet, 2009). We have characterized dental/oral and craniofacial findings in a number of genetic syndromes (30% effort). These studies are performed based on genetic referrals to the NIDCR intramural dental clinic as well as with participation in the NIH Clinical Center Undiagnosed Diseases Program. Through our studies we have continued to characterize orofacial and dental findings in a number of rare diseases and syndromes including Hutchinson-Gilford progeria (Domingo et al, Oral Dis, 2009). Additionally, our studies of Mendelian syndromes continue to expand and more fully document dental findings of importance. We have characterized enamel defects in patients with methylmalonic acidemia (MMA) and cobalamin (cbl) metabolic disorders. (Bassim et al, Oral Diseases, 2009). Our studies have determined that enamel defects are significantly more prevalent in MMA affected individuals, across complementation types (P < 0.0001). The mut MMA subgroup show a significantly higher prevalence of severe enamel defects, and individuals with enamel defects exhibited higher serum methylmalonate levels. These findings suggest an association between enamel developmental pathology and disordered metabolism. Salivary methylmalonate levels were extremely elevated and were significantly higher in MMA affected individuals than controls, indicating that salivary testing may be an effective diagnostic strategy. Through our studies of human cleft lip/palate we have identified and validated a SNP in the PDGF-C gene that is associated with CL/P. The presence of the -986T allele in the PDGF-C promoter is associated with a significant decrease (up to 80%) of PDGF-C gene promoter activity. This functional polymorphism acting on a susceptible genetic background may represent a component of human CL/P etiology.(Choi et al, Eur J Hum Genet 2009) Hereditary Gingival Fibromatosis/Gingival overgrowth conditions (20%). We have continued our studies of gingival overgrowth conditions to identify novel genetic loci etiologic for syndromic forms of the condition. As part of these studies we are characterizing tissue and cell specific gene expression profiles to increase our understanding of the physiology of gingival tissues. The goal of these studies is to increase our ability to diagnose and treat these conditions as well as to understand unique qualities of gingival tissues such as their increased wound healing capabilities. (Hart & Hart, Ortho Craniofac Res, 2009).

我们的工作范式是，人类疾病有遗传基础，了解疾病的遗传基础将促进更好的诊断和治疗策略的发展。我们研究孟德尔疾病是为了识别潜在的基因缺陷，并了解该/这些基因突变的产物如何导致异常发育或疾病。在某些情况下，我们开发了动物模型（转基因小鼠）和体外细胞模型来研究疾病发病机制。 牙本质发育不全 (DI)（10% 努力）：牙本质唾液酸磷蛋白 (DSSP) 基因的基因突变是大多数 DI 和牙本质发育不良 (DD) 病例的原因。牙本质是牙齿中最丰富的组织，由成牙本质细胞产生，成牙本质细胞与牙乳头的间充质细胞分化。牙本质缺陷大致分为两种主要类型：DI，I-III 型和牙本质发育不良（DD，I 型和 II 型）。迄今为止，DSPP 突变已被发现是 DI II 型和 III 型以及 DD II 型的基础。随着对潜在遗传机制的阐明，人们认识到与 DSPP 突变相关的临床特征似乎代表了一系列表型。 （Hart 和 Hart，Ortho Craniofac Res，2009 年）。最近的工作（McKnight 等人，Hum Mut，2008）描述了分离 DI 或 DD 的 9 个家族的研究，证实与 DSPP 突变相关的临床特征似乎代表了表型的连续体。此外，对孟德尔综合征和牙科发现的研究已确定其他基因的突变可导致牙本质缺陷，包括 GALNT3 突变 (Dumitrescu et al 2009) 和 DLX3 突变。了解牙科疾病的遗传基础为制定诊断和治疗策略提供了重要的第一步，以便为受影响的个体提供更好的护理。为了有效地转化临床和研究结果，还需要制定更有效的策略来教育牙科保健临床医生在遗传学领域。 （Johnson 等人，J Dent Educ，2008）。 毛齿骨综合征 (TDO)（20% 努力）。 DLX3 突变导致 TDO，其临床特征是牙齿、头发和骨骼异常。 TDO 的一个主要特征是骨厚度和密度增加。为了表征突变体 DLX3 如何促进骨骼改变，我们用携带由胶原蛋白 1A1 启动子驱动的 4bp DLX3 突变的小鼠建立了转基因模型。显微计算机断层扫描分析表明，TG 小鼠股骨的骨小梁体积和骨矿物质密度显着增加。在离体实验中，TG 小鼠表现出骨髓基质细胞向成骨细胞的分化增强，并且骨形成标志物的表达水平增加。然而，TG小鼠在体内荧光染料双标记实验中并未表现出动态骨形成率增强。破骨细胞因子存在下，TG小鼠骨髓单核细胞破骨分化能力降低，远端干骺端骨小梁表面TRAP(+)破骨细胞数量显着减少。 TG 小鼠中 TRACP 5b 和 CTX 血清水平显着降低，而 IFN-γ 水平显着升高。这些数据表明，IFN-γ水平的增加会降低破骨细胞的骨吸收活性，从而有助于增加这些TG小鼠的骨小梁体积和矿物质密度。这些数据表明这种 DLX-3 突变在破骨细胞分化和骨吸收中具有新的作用。 （Choi 等人，Dev Biol.2009）。 釉质生成不完美 (AI)（20% 努力）：我们继续研究分离 AI 表型的家族。 AI 是由 AMEL、ENAM、MMP20 和 KLK4 基因突变引起的。缺乏 AmelX、Enam 和 Mmp20 基因表达的小鼠已经诞生，为更好地了解牙釉质形成和 AI 发病机制提供了工具。 （Wright 等人，细胞组织器官，2009）。 我们继续对分离人工智能的家庭进行连锁研究，以进一步表征人工智能的基因型-表型相关性，并提高遗传学的诊断潜力，为人工智能患者提供明确的诊断，这是将诊断测试与护理和获得护理相结合的一步。我们扩展了常染色体显性遗传性 AI (ADAI) 病因的 FAM83H 基因突变的鉴定（Hart 等人，Clin Genet，2009）。此外，我们对分离 ADAI 的家族的研究发现了多个新的 FAM83H 突变，包括两个 2 bp 缺失突变，这是第一个发现的非无义突变。在受影响的个体中，颅面偏离正常的情况更为普遍。具有 677 个或更少氨基酸截短 FAMH3H 突变的受影响个体呈现出普遍的 ADHCAI 表型，而那些具有能够产生至少 694 个氨基酸的蛋白质的突变的个体具有独特且先前未报道的表型，主要影响宫颈釉质。这项研究表明，独特的表型与特定的 FAM83H 突变相关。 （Wright 等人，J Dent Res，2009）。对隔离 AI 的家族的研究也提供了直接证据，证明存在至少一种目前尚未确定的 ADAI 病因基因位点。 （Becerik 等人，AM J Med Genet，2009）。 我们已经对多种遗传综合征的牙齿/口腔和颅面表现进行了表征（30% 的努力）。这些研究是根据 NIDCR 壁内牙科诊所的遗传转诊以及参与 NIH 临床中心未确诊疾病计划进行的。通过我们的研究，我们继续描述了许多罕见疾病和综合征的口面部和牙科表现，包括 Hutchinson-Gilford 早衰症（Domingo 等人，Oral Dis，2009）。此外，我们对孟德尔综合症的研究不断扩大，并更全面地记录重要的牙科发现。我们对甲基丙二酸血症 (MMA) 和钴胺素 (cbl) 代谢性疾病患者的牙釉质缺陷进行了表征。 （Bassim 等人，《口腔疾病》，2009 年）。 我们的研究已确定，在不同的互补类型中，受 MMA 影响的个体中牙釉质缺陷明显更为普遍 (P < 0.0001)。 mut MMA 亚组显示严重牙釉质缺陷的患病率显着较高，并且有牙釉质缺陷的个体表现出较高的血清甲基丙二酸水平。 这些发现表明牙釉质发育病理学与代谢紊乱之间存在关联。 MMA 受影响个体的唾液丙二酸甲酯水平显着升高，并且显着高于对照组，表明唾液检测可能是一种有效的诊断策略。通过对人类唇裂/腭裂的研究，我们鉴定并验证了 PDGF-C 基因中与 CL/P 相关的 SNP。 PDGF-C 启动子中 -986T 等位基因的存在与 PDGF-C 基因启动子活性的显着降低（高达 80%）相关。这种作用于易感遗传背景的功能多态性可能代表人类 CL/P 病因学的一个组成部分。（Choi 等人，Eur J Hum Genet 2009） 遗传性牙龈纤维瘤/牙龈过度生长（20%）。我们继续对牙龈过度生长状况进行研究，以确定该病症综合征形式的新遗传位点病因。作为这些研究的一部分，我们正在表征组织和细胞特异性基因表达谱，以增加我们对牙龈组织生理学的了解。这些研究的目的是提高我们诊断和治疗这些疾病的能力，以及了解牙龈组织的独特品质，例如增强的伤口愈合能力。 （Hart 和 Hart，Ortho Craniofac Res，2009 年）。