Clinical Research of Oral Connective Tissue Program

项目A.确定导致特发性牙根吸收的遗传易感性和免疫病理学机制 背景：以前，我们报道过BSP基因敲除小鼠表现出牙根吸收表型。基于这一发现，我们试图确定人类受试者表现出多发性特发性颈根吸收（MICRR），MICRR的家族模式与建议的遗传易感性。在获得底特律大学慈善牙科学院和NIH的IRB批准后，从表现出MICRR的亲属（4名受影响成员和4名未受影响成员）中收集牙科/病史、X射线、唾液样本和拔牙。使用唾液的全外显子组测序鉴定了IRF 8的c-末端区域中的突变，该突变负责破骨细胞功能过度活跃。结果发表在2019年JBMR上，该项目与Thumbigere-Math博士（K99/R 00接受者）一起转移到马里兰州大学。 正在进行中：我们继续与Thumbigere-Math博士合作，作为IRF 8项目的负责人，与其他NIH合作者，Ozato博士和Holland博士合作，并在Bone，2020（见下文）上发表文章。与其他临床医生建立了合作关系：Steve Russo博士（芝加哥，IL），托马斯施耐德博士（新泽西）和Janina Golob Deeb博士（VCU），他们见过快速MICRR的患者。 Golob Deeb博士注意到与MICRR和停用地舒单抗的潜在相关性，因此我们与Alison博伊斯和玛丽Kao-Hseih博士（NIDCR）合作，对服用地舒单抗治疗纤维性发育不良的患者进行牙科检查。 我们还与微生物分析专家Purnima Kumar博士（俄亥俄州州立大学）建立了合作关系，以确定Russos患者牙菌斑样本的微生物特征。此外，我们分析了从贝蒂·哈吉申加利斯博士（美国）那里获得的牙齿。Penn），来自一名患有听小骨缺失和特发性牙根吸收的患者，诊断为家族性膨胀性骨质溶解（10岁，编码RANK蛋白的TNFRSF 11基因突变，过度表达），并注意到牙骨质缺陷。 项目B。矿化障碍： 与NIDCR临床研究人员和其他IC临床医生合作，我们一直在检查患有矿化组织代谢障碍的个体的DOC复合物组织/细胞的改变。 1. Pi/PPi关键调控因子的突变。骨骼和牙齿的矿化受到细胞外无机磷酸盐（Pi）和焦磷酸盐（PPi）水平的严格调节。 三种控制Pi和PPi的细胞周围浓度的调节剂包括组织非特异性碱性磷酸酶（TNAP）、进行性强直蛋白（ANK）和外核苷酸焦磷酸酶/磷酸二酯酶1（ENPP 1）。 这些因素的失活导致影响牙齿及其支撑结构的矿化障碍。 我们研究了PPi降低对患有婴儿全身性动脉钙化（GACI）的人类受试者（4）牙齿发育和成熟的影响，这些受试者在ENPP 1中具有功能缺失突变。 受试者报告了咬合不全或乳牙过度保留、正畸牙齿移动不良的病史，表明矿物质代谢改变是一个影响因素。 对GACI受试者拔除的乳牙进行的Micro-CT分析显示，与年龄匹配的健康对照牙相比，颈部牙骨质厚度和密度显着增加。 GACI和对照牙的牙釉质和牙本质密度无差异。 组织学显示GACI牙颈部牙骨质显著扩张，包括牙骨质细胞样细胞和牙骨质吸收和修复的不寻常模式。 Enpp 1基因敲除小鼠磨牙的Micro-CT分析显示脱细胞牙骨质厚度和体积显著增加。 总的来说，这些研究结果报告了一种新的牙齿表型在GACI和进一步支持我们的假设，Pi/PPi调制是一个关键机制，调节跨物种的牙骨质形成。Thumbigere-Math V等人，JDR，2018年。 正在进行中：我们继续在NIH CRC检查Pi/PPi疾病患者，并分析Pi/PPi疾病患者的脱落或拔牙。 我们还纳入了ABCC 6（ATP结合盒亚家族C成员6）患者，该患者与某些GACI病例相关。 我们还检查了患有与骨骼紊乱相关的疾病的患者的牙齿，例如，人类家族性肿瘤性钙质沉着症（合作者：Michael柯林斯博士，2020年7月提交的手稿）、缺氧诱导因子2a突变患者（合作者：Pacak、Rosenblum和Zhuang博士）。 作为对照，我们与北卡罗来纳州州立大学和杜克大学合作，在新生儿表观遗传学研究中，从健康个体中获得脱落的乳牙。

Project A. Determine genetic susceptibility and immunopathological mechanisms contributing to idiopathic tooth root resorption Background: Previously, we reported that BSP KO mice exhibit a tooth root resorption phenotype. Based on this finding, we sought to identify human subjects exhibiting multiple idiopathic cervical root resorption (MICRR), a familial pattern of MICRR with suggested genetic susceptibility. Following IRB approval from the University of Detroit Mercy School of Dentistry and NIH, dental/medical histories, x-rays, saliva samples, and extracted teeth were collected from a kindred (4 affected and 4 unaffected members) exhibiting MICRR. Whole exome sequencing using saliva identified a mutation in the c-terminal region of IRF8 responsible for overactive osteoclast function. Results were published in JBMR, 2019 and this project transitioned to University Maryland with Dr. Thumbigere-Math (K99/R00 recipient). Ongoing: We continue to collaborate with Dr. Thumbigere-Math as the lead on the IRF8 project, with other NIH collaborators, Drs. Ozato and Holland, with a publication in Bone, 2020 (see below). Collaborations were established with other clinicians: Dr. Steve Russo (Chicago, IL), Dr. Thomas Schneider (New Jersey), and Dr. Janina Golob Deeb (VCU), who have seen patients with rapid MICRR. Dr. Golob Deeb noticed a potential association with MICRR and cessation of denosumab, leading us to collaborate with Drs. Alison Boyce and Marie Kao-Hseih (NIDCR) to conduct dental exams on patients taking denosumab for fibrous dysplasia. We have also set up a collaboration with an expert in microbial analyses, Dr. Purnima Kumar (Ohio State University), to determine the microbial profile of plaque samples from Russos patient. Further, we analyzed teeth obtained from Dr. Betty Hajishengallis (U. Penn), from a patient with missing ossicles and idiopathic root resorption, diagnosed with familial expansile osteolysis (age 10, mutations in the TNFRSF11 gene encoding RANK protein, over expression) and noted cementum defects. Project B. Disorders of mineralization: In collaboration with NIDCR clinical researchers and other IC clinicians, we have been examining individuals with mineralized tissue metabolism disorders for alterations in tissues/cells of the DOC complex. 1. Mutations in key regulators of Pi/PPi. Mineralization of skeleton and teeth is tightly regulated by levels of extracellular inorganic phosphate (Pi) and pyrophosphate (PPi). Three regulators that control pericellular concentrations of Pi and PPi include tissue-nonspecific alkaline phosphatase (TNAP), progressive ankylosis protein (ANK), and ectonucleotide pyrophosphatase/phosphodiesterase 1 (ENPP1). Inactivation of these factors results in mineralization disorders affecting teeth and their supporting structures. We examined the effect of decreased PPi on development and maturation of teeth in human subjects (4) with generalized arterial calcification of infancy (GACI), who harbor loss-of-function mutations in ENPP1. Subjects reported a history of infraocclusion or over-retained primary teeth,poor orthodontic tooth movement, suggesting altered mineral metabolism as a contributing factor. Micro-CT analyses of extracted primary teeth from GACI subjects revealed marked increase cervical cementum thickness and density vs. age-matched healthy control teeth. There were no differences in enamel and dentin densities between GACI and control teeth. Histology revealed dramatically expanded cervical cementum in GACI teeth, including cementocyte-like cells and unusual patterns of cementum resorption and repair. Micro-CT analysis of Enpp1 knock-out mouse molars revealed a marked increase in acellular cementum thickness and volume. Collectively, these findings report a novel dental phenotype in GACI and further support our hypothesis that Pi/PPi modulation is as a key mechanism for regulating cementogenesis across species. Thumbigere-Math V et al., JDR, 2018. Ongoing: We have continued to examine patients with Pi/PPi disorders at NIH CRC as well as analyzing exfoliated or extracted teeth from patients with Pi/PPi disorders. We have also included patients with ABCC6 (ATP Binding Cassette Subfamily C Member 6), which is associated with some cases of GACI. We have also been examining teeth from patients with disorders associated with skeletal disturbances, e.g., human familial tumoral calcinosis (collaborators: Dr. Michael Collins, manuscript submitted July 2020) , patients with Hypoxia-inducible factor 2a mutations (collaborators: Drs. Pacak, Rosenblum, and Zhuang). For controls, in collaboration with North Carolina State University and Duke under the Newborn Epigenetics Study, we are obtaining exfoliated primary teeth from healthy individuals.