Submitted by Rodrigo Martins Cruz (rodrigo.cruz@ufvjm.edu.br) on 2014-12-22T12:08:16Z No. of bitstreams: 2 mauricio_rocha_dourado.pdf: 2196583 bytes, checksum: ffa323001018e93a3c8d5bfd16675742 (MD5) license_rdf: 23898 bytes, checksum: e363e809996cf46ada20da1accfcd9c7 (MD5) / Approved for entry into archive by Rodrigo Martins Cruz (rodrigo.cruz@ufvjm.edu.br) on 2014-12-22T12:19:49Z (GMT) No. of bitstreams: 2 mauricio_rocha_dourado.pdf: 2196583 bytes, checksum: ffa323001018e93a3c8d5bfd16675742 (MD5) license_rdf: 23898 bytes, checksum: e363e809996cf46ada20da1accfcd9c7 (MD5) / Approved for entry into archive by Rodrigo Martins Cruz (rodrigo.cruz@ufvjm.edu.br) on 2014-12-22T12:20:06Z (GMT) No. of bitstreams: 2 mauricio_rocha_dourado.pdf: 2196583 bytes, checksum: ffa323001018e93a3c8d5bfd16675742 (MD5) license_rdf: 23898 bytes, checksum: e363e809996cf46ada20da1accfcd9c7 (MD5) / Made available in DSpace on 2014-12-22T12:20:06Z (GMT). No. of bitstreams: 2 mauricio_rocha_dourado.pdf: 2196583 bytes, checksum: ffa323001018e93a3c8d5bfd16675742 (MD5) license_rdf: 23898 bytes, checksum: e363e809996cf46ada20da1accfcd9c7 (MD5) Previous issue date: 2014 / Amelog?nese imperfeita (AI) representa um complexo grupo de condi??es herdadas que provocam malforma??es do esmalte dent?rio em quantidade ou qualidade. Nefrocalcinose (NC) ? a deposi??o de sais de c?lcio no par?nquima renal que pode, em longo prazo e na aus?ncia de tratamento adequado, levar ? insufici?ncia renal cr?nica e outros dist?rbios renais. Estas duas condi??es, simultaneamente, representam a s?ndrome da amelog?nese imperfeita e nefrocalcinose (SAINC), ou s?ndrome esmalte renal, uma desordem autoss?mica recessiva causada por muta??es no gene FAM20A, com poucos casos relatados na literatura. A proposta do presente estudo foi investigar as caracter?sticas da s?ndrome em indiv?duos de quatro fam?lias brasileiras. Os pacientes foram submetidos ao exame cl?nico, radiogr?fico, investiga??o renal e exames hematol?gicos e de marcadores bioqu?micos, sendo identificados nove pacientes portadores da s?ndrome. O exame cl?nico revelou dentes pequenos e de colora??o amarelada, desgaste oclusal/incisal, reten??o da denti??o dec?dua e hiperplasia gengival. As radiografias mostraram les?es pericoron?rias radiol?cidas envolvendo dentes permanentes impactados, aus?ncia de contraste entre esmalte e dentina e calcifica??es intrapulpares. Os exames bioqu?micos evidenciaram, em quatro pacientes, baixos n?veis de vitamina D 25-OH e altos n?veis de fosfatase alcalina e paratorm?nio. A ultrassonografia revelou nefrocalcinose bilateral, mas, os pacientes ainda, n?o demonstraram nenhuma manifesta??o de doen?a renal. Outras caracter?sticas menos comuns como defici?ncia mental e cistos renais foram encontradas, o que confirma o grande espectro de altera??es fenot?picas desta s?ndrome, j? descrita anteriormente. Estes resultados confirmam as varia??es fenot?picas da SAINC e refor?am a necessidade de investiga??o renal em pacientes com diagn?stico de AI, principalmente do tipo hipopl?sica. / Disserta??o (Mestrado) ? Programa de P?s-Gradua??o em Odontologia, Universidade Federal dos Vales do Jequitinhonha e Mucuri, 2014. / ABSTRACT Amelogenesis imperfecta (AI) is a complex group of conditions that cause inherited defects of dental enamel in quantity or quality. Nephrocalcinosis (NC) is the deposition of calcium salts at the renal parenchyma, which may in the long term and in the absence of proper treatment, lead to chronic renal failure and other kidney disorders. These two conditions simultaneously represent the amelogenesis imperfecta and nephrocalcinosis syndrome (AINCS), or enamel renal syndrome, an autosomal recessive disorder caused by mutations in FAM20A gene, with few cases reported in the literature. The purpose of this study was to investigate the characteristics of the syndrome in patients from four Brazilian families. Patients underwent clinical examination, radiographic, renal and haematological investigation and biochemical markers examinations, being identified nine patients with this syndrome. Clinical examination revealed small and yellowish teeth, occlusal/incisal wear, retention of deciduous dentition and gingival hyperplasia. The radiographs showed pericoronal radiolucencies involving impacted permanent teeth, absence of contrast between enamel and dentin and intrapulpal calcifications. Biochemical tests showed low levels of 25-OH vitamin D and high levels of alkaline phosphatase and parathyroid hormone in four patients. Ultrasonography revealed bilateral nephrocalcinosis, but patients still showed no manifestation of renal disease. Other characteristics as mental retardation and renal cysts were found, confirming the wide spectrum of phenotypic alterations of this syndrome previously described. These results confirm the phenotypic variations AINCS and reinforce the need for kidney research in patients with AI, especially the hypoplastic type.

Family with sequence similarity 20 (FAM20) consists of three members: FAM20A, FAM20B and FAM20C. Mutations in FAM20 family have been linked to developmental disorders involving bones, cartilage and teeth. FAM20A mutations in humans are associated with amelogenesis imperfecta with gingival fibromatosis and enamel renal syndrome. Fam20a knockout (KO) mouse showed growth retardation. The aim of this study was to characterize the skeletal and dental phenotypes using Fam20a KO mouse. Our results showed that body size and bone length of KO mice were smaller than those of WT. The microcomputed tomography (μCT) analyses of trabecular and cortical bones in KO displayed lower bone volume, thinner trabeculae and thinner bone cortex as compared to WT. Histological examination of KO growth plate demonstrated disorganized chondrocyte zones and extended hypertrophic zone. qRT-PCR results showed downregulation of several osteoblast differentiation markers in KO long bone. Immunohistochemical examination demonstrated reduced chondrocyte proliferation, apoptosis and increased collagen X expression in KO growth plate. Our data showed a lower number of osteoblasts and osteoclasts in KO as compared to WT. In vitro study, Fam20a KO showed a lower number of bone marrow stromal cells and osteoprogenitors. In vitro mineralization was impaired in KO osteoblasts. Fam20a KO had hypoplastic enamel, delayed tooth eruption and gingival overgrowth. The µCT results demonstrated that enamel in Fam20a KO was not fully mineralized and enamel matrix was detached from dentin. Scanning electron microscopy displayed absence of decussation patterns in Fam20a KO enamel. Histological examination of maxillary first molar at differentiation stage showed no difference between WT and KO. At the secretory stage, Fam20a KO ameloblasts were short and non-polarized as compared to WT. Immunohistochemical analysis showed diffuse staining pattern of amelogenin in Fam20a KO first molar compared to WT. Western blot analysis demonstrated that amelogenin proteolytic process was impaired in KO and showed slower migration pattern of MMP20. In conclusion, endochondral ossification defects and reduced number of osteoblasts and their precursors led to the bone phenotype in Fam20a KO. Amelogenin processing defects caused amelogenesis imperfecta phenotype in KO. Our study indicated that Fam20a plays a role in skeletal development and amelogenesis.