Objetivo: elaborar diretrizes para o tratamento. Modelo: Análise retrospectiva dos históricos de 236 crianças, durante o período de julho de 2003 a julho de 2006 e análise descritiva dos resultados. Ambiente: Unidade de Cuidados Especiais (UCE) do HRAC-USP. Participantes: 236 crianças com disfagia orofaríngea, com idade de 3 dias a 8 anos, 204 (86,4%) apresentavam fissura labiopalatina. A Seqüência de Robin foi diagnosticada em 139 crianças (58,9%), sendo que 95 (68,3%) tinham SRI e 44 (31,7%) síndrome associada. Variáveis: incidência de disfagia, síndromes ou malformações associadas, grau da disfagia, presença de desconforto respiratório, uso de sondas alimentadoras (SNG e gastrostomia), DRGE, complicações relacionadas às gastrostomias e ao tratamento cirúrgico da DRGE, reinternações e o tempo total de hospitalização. Resultados: Das 236 crianças, 110 (46,6%) tinham disfagia leve, 47 (19,9%) moderada e 79 (33,5%) grave. A disfagia leve foi maior nas crianças com fissuras de lábio e/ou palato isoladas (88,5%), com malformações congênitas (81,3%) e com SRI (56,8%). A disfagia grave foi mais freqüente em crianças com síndrome (61,6%). Do universo estudado, 195 crianças (82,6%) necessitaram do uso da SNG. O tempo de uso variou de 1 dia a 750 dias, com média de 113 dias e mediana de 60 dias. A gastrostomia foi indicada em 67 (28,4%). Conclusões: A disfagia orofaríngea é um dos principais problemas apresentados pelas crianças com malformações craniofaciais. O diagnóstico de outras malformações ou síndromes associadas e a classificação do grau da disfagia orofaríngea são determinantes na orientação do tratamento. / Objective: To draw up guidelines for the treatment. Model: Retrospective analysis of historical of 236 children, during the period July 2003 to July 2006 and descriptive analysis of the results. Environment: Special Care Unit by HRAC-USP. Participants: 236 children with oropharyngeal dysphagia, aged 3 days to 8 years, 204 (86.4%) had cleft lip, and / or palate. The sequence of Robin was diagnosed in 139 children (58.9%), whereas 95 (68.3%) had SRI and 44 (31.7%) syndrome associated. Variables: incidence of dysphagia, syndromes or associated malformations, degree of dysphagia, presence of respiratory discomfort, use of probes feeding (nasogastric tube and gastrostomy), GERD, complications related to gastrostomies and the surgical treatment of GERD, readmissions and the total time of hospitalization. Results: Of the 236 children, 110 (46.6%) had mild dysphagia, 47 (19.9%) moderate and 79 (33.5%) serious. The mild dysphagia was higher for children with cleft lip, and / or palate alone (88.5%) with congenital malformations (81.3%) and with SRI (56.8%). The severe dysphagia was more frequent in children with syndrome (61.6%). Of the study, 195 children (82.6%) required the use of nasogastric tube . The time of use ranged from 1 day to 750 days, with an average of 113 days and a median of 60 days. The gastrostomy was indicated in 67 (28.4%). Conclusions: The oropharyngeal dysphagia is one of the main problems presented by children with craniofacial deformities. The diagnosis of malformations or other syndromes associated and classification the degree of oropharyngeal dysphagia are crucial in guiding treatment.

Introdução: A craniossinostose é causada pela fusão prematura de uma ou mais suturas cranianas, levando à deformidade do crânio. Formas sindrômicas ocorrem quando a craniossinostose é associada a características dismorfológicas adicionais. A fusão precoce das suturas pode ser causada por fatores ambientais e genéticos. No presente trabalho, pretende-se reconhecer os diagnósticos clínicos e características fenotípicas da craniossinostose em pacientes atendidos nos ambulatórios de Genética Médica do Hospital de Clinicas de Porto Alegre no período de 2006 a 2016. O protocolo de investigação incluiu anamnese, exame dismorfológico e revisão de prontuário, incluindo exames de investigação realizados. Resultados: Entre 2006 e 2016, foram avaliados 133 indivíduos com craniossinostose, sendo que 121 reuniram critérios para inclusão neste estudo. A idade média de diagnóstico da craniossinostose foi de 38,4 meses. A sutura mais frequentemente acometida foi a sutura metópica. Houve maior proporção de casos sindrômicos (69,4%). Em 25 desses pacientes, foram identificadas as síndromes de Apert, Crouzon, Pfeiffer, Muenke, Craniofrontonasal ou Saethre-Chotzen. Síndromes não tipicamente relacionadas a craniossinostose foram também identificadas, como distrofia miotônica tipo 1 (n=2), síndrome de Gorlin, síndrome de Beckwith-Wiedemann e galactosemia. Os sinais clínicos não eram típicos de qualquer síndrome particular em 32 indivíduos (38,1% dos casos sindrômicos). Características fenotípicas frequentes incluíram malformações de extremidades (35,5%), do sistema nervoso central (32,1%), cardiovasculares (21,4%) e genito-urinárias (16,6%). Foram observadas malformações raras como espinha bífida (n=3), hérnia diafragmática congênita (n=3) e hipoplasia congênita de parede abdominal (n=2). Anormalidades citogenéticas ou moleculares foram identificadas em 18 indivíduos sindrômicos, sendo a síndrome de Muenke o diagnóstico mais frequente (n=7). Discussão: A maior proporção de casos sindrômicos em relação a outras séries é possivelmente relacionada ao fato de tratar-se de casos atendidos em um serviço de Genética clínica. Observou-se diagnóstico significativamente tardio na presente casuística, reforçando a necessidade de estratégias de saúde pública envolvendo treinamento de recursos humanos e otimização da referência aos centros terciários. O acometimento multissistêmico reforça a importância do acompanhamento multidisciplinar. Conclusão: O estudo demonstra uma amostra amplamente heterogênea em termos clínicos, genéticos e terapêuticos. É fundamental o desenvolvimento de estratégias de educação contínua não apenas dentro da equipe, mas também ao acessar pacientes e familiares, através do aconselhamento genético e de ferramentas de comunicação. Para isso, propõe-se uma cartilha informativa sobre craniossinostoses para pacientes e familiares. Faltam estudos em países em desenvolvimento para análise comparativa dos dados em contextos sociais semelhantes. / Introduction: Craniosynostosis is caused by premature fusion of one or more cranial sutures, leading to deformity of the skull. Syndromic forms occur when craniosynostosis is associated with additional dysmorphological features. Early suture fusion can be caused by environmental and genetic factors. In this study, it is intended to recognize the clinical diagnosis and phenotypic characteristics of craniosynostosis in patients attending Medical Genetics outpatient clinics of Hospital de Clínicas de Porto Alegre from 2006 to 2016. The research protocol included anamnesis, dysmorphological examination, review of medical records and investigations carried out. Results: Between 2006 and 2016, 133 individuals with craniosynostosis were evaluated, and 121 met inclusion criteria for this study. The mean age at diagnosis of craniosynostosis was 38.4 months. Metopic suture was the most commonly involved. There was a higher proportion of syndromic cases (69.4%). In 25 of these patients, Apert, Crouzon, Pfeiffer, Muenke, Craniofrontonasal or Saethre-Chotzen syndromes were identified. Syndromes not typically associated to craniosynostosis were also identified, such as myotonic dystrophy type 1 (n = 2), Gorlin syndrome, Beckwith-Wiedemann syndrome and galactosemia. Clinical signs were not typical of any particular syndrome in 32 individuals (38.1% of syndromic cases). Frequent phenotypic features included extremities (35.5%), central nervous system (32.1%), cardiovascular (21.4%) and genitourinary malformations (16.6%). Rare malformations such as spina bifida (n = 3), diaphragmatic hernia (n = 3) and congenital abdominal wall hypoplasia (n = 2) were observed. Cytogenetic or molecular abnormalities were identified in 18 syndromic patients, and Muenke syndrome was the most frequent diagnosis (n = 7). Discussion: The higher proportion of syndromic cases than in other series is possibly due to the fact that these cases are treated in a clinical genetics service. Significantly late diagnosis was observed in the present series, reinforcing the need for public health strategies involving training of human resources, optimization of referral to tertiary centers and active search strategies. Multisystemic involvement reinforces the importance of multidisciplinary follow-up. Conclusion: This study demonstrates a widely heterogeneous clinical, genetic and therapeutic sample. Strategies for continuous education within the team, patients and family members, through genetic counseling and communication tools are important, so it is proposed an information booklet for patients and families. There is a scarcity of case series from developing countries for comparative analysis in similar social contexts.

In this thesis, several techniques were combined to optimize, evaluate and characterize craniofacial development in Xenopus, with additional focus on understanding the alterations made during maturation in the craniofacial region and the cartilage. Three important techniques used were: confocal microscopy in conjunction with Acridine Orange (AO) labeling, Alcian Blue (AB) labeling, and geometric morphometric analysis. I found that facial width increased across all techniques used to evaluate it. Included within this focus was the study of the development of the ceratohyal (CH) cartilage, which supported the mouth and snout. This was also found to increase width wise, in unison with facial and orofacial growth. This data may suggest a link between the face, mouth and CH growth, in which the developing cartilage elongates and widens causing the increase seen in the width and distension of the mouth.

Introduction: The purpose of this study was to examine phenotypic expression of craniofacial form, shape, and size as it relates to the genotype of an individual. Shape analyses were completed on 3-D images of each subject's craniofacial structure by landmarking 45 points of interest on the cranial base, facial bones, and upper and lower jaws. A candidate gene analysis was undertaken focusing on specific genes in the Hippo Signaling Pathway to examine genotype-phenotype correlations that play a role in craniofacial development. This study is a continuation of a larger project aimed at the identification of candidate genes associated with human dento-skeletal bite problems led by Dr. Lina Moreno-Uribe. Methods: The sample size for our study included 166 individuals who had never been treated orthodontically at the time of records. Each individual was genotyped and a CBCT of the craniofacial complex was captured. Each CBCT image was landmarked by a single observer using 45 points to mark points on the cranial base and facial bones including the maxilla and mandible. General Procrustes superimposition was used to find correlations with phenotype and genotype. Size analysis was completed with average Euclidean Distances and ANOVA analysis. Results: 2 SNP's from the FOX03 gene had significant associations with size. The AA genotyped individuals appeared larger in overall size than AB genotyped individuals. 3 SNP's had statistically significant associations with facial form. The FOX06 SNPs had significant associations with increased anterior-posterior growth of the maxilla. The AJUBA SNP had significant associations with increased overall craniofacial breadth. Conclusion: Genes in the Hippo signaling pathway have specific roles in the development of facial form and size.

There are many craniofacial anomalies that exist in the oral mucosa, gingiva, lips, tongue, maxilla, mandible, floor of the mouth, palate, and teeth. These anomalies cause secondary issues such as airway obstruction, respiratory problems, feeding problems, ear disease, distal systemic issues, and speech and communication problems. Children that experience craniofacial anomalies and subsequent problems are often at a disadvantage with medical and dental related consequences, and especially speech, communication, and often present with psychosocial concerns. This paper explores such anomalies, consequential problems, and emphasizes the importance of having a multidisciplinary team when treating patients with craniofacial anomalies. Multidisciplinary teams consist of otolaryngologists, plastic surgeons, general dentists, prosthodontists, orthodontists, oral surgeons, pediatricians, neurologists, geneticists, social workers, psychologists, audiologists, and speech therapists. Based on past studies and data, multidisciplinary treatment has shown not only to provide the best options to correct an anomaly, but also to optimize the overall health and well-being of an individual as well. Multidisciplinary treatment of craniofacial anomalies outlines a coherent, inclusive, and revolutionized way on how to holistically treat a patient. This approach is present in the healthcare realm, but often underrated and not adopted by all healthcare professionals; this paper will demonstrate how such an approach will advance healthcare. Surgery removes, corrects, or improves a condition that exists in the oral cavity or oropharynx. Nevertheless, surgery often causes subsequent conditions and may even not be successful. Among the many disciplines exercised in treating patients with craniofacial anomalies and conditions, this paper highlights the importance of speech and language pathologists, psychologists, dentists, orthodontists, and geneticists to be included in the treatment plan. Studies demonstrated that each discipline’s responsibility, when implemented in a coordinated and timely fashion, can improve the outcomes, possibly prevent ensuing conditions, and therefore, optimize an individual’s health and quality of life.

New Zealand Maori and Pacific Island ethnic groups are marking up an increasingly larger proportion of New Zealand�s population. Intercanthal distance (ICD) and management of congenital and acquired deformities of the craniofacial complex. The ICD and IPD have been found to differ to establish these measurements for New zealand Maori and Samoan populations. For New Zealand Maori males the mean ICD was 32.1mm with a standard deviation (SD) of 2.6mm, and the mean IPD was 63.3mm, SD 3.8mm. For New Zealand Maori females the mean ICD was 30.7mm, SD2.7mm and the mean IPD was 60.1mm, SD2.8mm. For Samoan males the mean ICD was 33.9mm, SD2.5mm and the mean IPD 64.5mm, SD3.5mm. For Samoan females the mean ICD was 32.9mm, SD2.3mm and their mean IPD was 61.7mm, SD2.8mm. Consistent with other ethnicities New Zealand Maori and Samoan males have wider values for ICD and IPD than females respectively. New Zealand Maori measurements tend to lie within the normal values established for Caucasian populations, whilst Samoans have larger values.

The hedgehog morphogenic pathway is essential for the development of numerous organs and tissues in both vertebrates and invertebrates, and dysregulation of hedgehog signalling is also associated with a broad range of mammalian cancers. While a great deal of research has been dedicated to understanding the molecular interactions of the hedgehog signalling pathway itself, much work remains in understanding the downstream transcriptional output of the pathway, and how that output modulates cellular behaviour in target tissues to produce developmental outcomes. The hedgehog pathway is activated by hedgehog proteins and repressed by patched. Downstream of these regulators, the hedgehog signalling cascade involves modification and trafficking of a series of key proteins and ultimately leads to regulation of the GLI family of transcription factors, thereby modulating the transcriptional output of the pathway. This thesis builds on previous work investigating downstream targets of one GLI protein – GLI3 – in the mouse limb (McGlinn et al., 2005). This previous study identified genes that were dysregulated in the anterior limb of the Gli3-null, extra-toes strain of mice (Gli3Xt/Xt). Amongst the identified targets of GLI3 were a number of novel genes. However, further detailed analysis of these genes was not conducted, and therefore, this thesis investigates the embryonic expression or function of three of these novel downstream targets of GLI3, to clarify their regulation by the hedgehog pathway and identify their broader role throughout development. One published work and one paper submitted for publication are contained within this thesis, describing detailed expression of two novel SHH targets, Zinc finger protein 503 (Zfp503) and Pitrolysin metallopeptidase 1 (Pitrm1). Zfp503 belongs to a family of transcription factors that regulate aspects of development across a diversity of species. However, their role in mammals and avians has been poorly described. This manuscript presents a detailed description of Zfp503 expression in the mouse and chicken and examines regulation of Zfp503 in the limb by SHH and BMP signalling. My contribution to this paper was the analysis of WT Zfp503 expression in mouse and chick by section in situ hybridisation, and as such, I am listed as a middle author. Pitrm1 is a metallopeptidase with a broad range of predicted target molecules. Comparisons with family members in mammals and plants suggest Pitrm1 has mitochondrial function and is implicated in the pathology of Alzheimers disease. It is upregulated in response to hedgehog pathway activation in the anterior limb of two mouse models of hedgehog signalling– the Gli3Xt/Xt and Ptch1:Prx-Cre mouse line, which deletes patched1 in the developing limb. It is expressed in multiple developing tissues that are patterned by SHH, suggesting that Pitrm1 may be an important regulator of developmental processes downstream of SHH. For the Pitrm1 manuscript, I contributed the majority of the experimental data and prepared the manuscript, and therefore, I am the first listed author. A third downstream hedgehog target gene described in this thesis is Tmem26. Tmem26 is an entirely novel gene with unknown cellular function, although concurrent work in the Wicking laboratory suggests that Tmem26 regulates cell migration and morphology in cell culture. Tmem26 is negatively regulated by SHH in the anterior mouse limb at 11.5dpc, as shown by use of Gli3Xt/Xt and Ptch1:Prx-Cre mice. Tmem26 expression in wild-type mice is spatially restricted and strikingly evident in the facial prominences, particularly near the point of fusion of the developing lip and in the shelves of the secondary palate. This suggests that Tmem26 may be involved in lip and palate formation and possibly play a role in the common human birth disorders of cleft lip and cleft palate. Generation of a Tmem26 conditional knockout mouse line, followed by germline inactivation of Tmem26 using a ubiquitously expressed Cre line, did not reveal a craniofacial phenotype in embryos or adults. Knockout mice appear healthy and fertile with no obvious developmental defects. This does not preclude a role for Tmem26 in facial development however, as molecular redundancy may be able to compensate for Tmem26 loss in mice. Tmem26 is also expressed in cells and organs of the adult immune system and suggests an alternative possible role for Tmem26 in regulating immune function that could be further investigated using the Tmem26 conditional knockout mouse line.

Objective: To follow a cohort of children from age 4-6-12 with respect to sleep disordered breathing (SDB) and orofacial development. Questionnaires were completed about sleep, snoring, apneas, enuresis, sucking habits, and adenotonsillar surgery and, from age 12, about allergies, asthma, and general health. Children snoring regularly had an ENT- examinations including sleep studies (at ages 4 and 12) and an orthodontic evaluation. Development of biometric data in snoring children and not snoring controls was studied in relation to adenotonsillar surgery. Result: Of the original group of 615 children, 509 (83%) participated at age 6 and 393 (64%) at age 12. 27 snored regularly and 231 did not snore at age 12. Differences between groups were seen on all answers. From age 4–12 the prevalence of OSA decreased from 3.1% to 0.8%, and the minimum prevalence of snoring regularly from 5.3% to 4.2%. The odds for a child who snored regularly at four or six to be snoring regularly at age 12 was 3.7 times greater than for a not snoring child in spite of surgery (OR 3.7, 95% CI 2.4-5.7). 63 children were operated for snoring by age 12, of them 14 never snored and 17 snored regularly at age 12. Cross-bite was more common among snoring children at ages 4, 6 and 12 as was a narrower maxilla. In most cases, surgery cured the snoring temporarily, but the maxillar width was still smaller by age 12—even when nasal breathing was attained. Children snoring regularly at age 12, operated or not operated, showed long face anatomy and were oral breathers; the seven cases who were not operated and the five who were still snoring in spite of surgery, did not have reduced maxillary arch width. Conclusion: The prevalence of children snoring regularly is about the same from age four to twelve in a cohort where adenotonsillar surgery has been performed on obstructed cases, but the prevalence of OSA decreases considerably. The children snoring regularly have a more narrow maxilla compared to children not snoring—a condition that is not changed by adenotonsillar surgery regardless of symptom relief.

Craniofacial defects resulting from trauma and resection present many challenges to reconstruction due to the complex structure, combinations of tissues, and environment, with exposure to the oral, skin and nasal mucosal pathogens. Tissue engineering seeks to regenerate the tissues lost in these defects; however, the composite nature and proximity to colonizing bacteria remain difficult to overcome. Additionally, many tissue engineering approaches have further hurdles to overcome in the regulatory process to clinical translation. As such these studies investigated a two stage strategy employing an antibiotic-releasing porous polymethylmethacrylate space maintainer fabricated with materials currently part of products approved or cleared by the United States Food and Drug Administration, expediting the translation to the clinic. This porous space maintainer holds the bone defect open allowing soft tissue to heal around the defect. The space maintainer can then be removed and one regenerated in the defect. These studies investigated the individual components of this strategy. The porous space maintainer showed similar soft tissue healing and response to non-porous space maintainers in a rabbit composite tissue defect. In humans, the porous space maintainers were well tolerated and maintained a soft tissue envelope for closure after implantation of a bone regeneration technology. The antibiotic-releasing space maintainers showed release of antibiotics from 1-5 weeks, which could be controlled by loading and fabrication parameters. In vivo, space maintainers releasing a high dose of antibiotics for an extended period of time increased soft tissue healing over burst release space maintainers in an infected composite tissue defect model in a rabbit mandible. Finally, stabilization of bone defects and regeneration could be improved through scaffold structures and delivery of a bone forming growth factor. These studies illustrate the possibility of the two stage strategy for repair of composite tissue defects of the craniofacial complex.

Class III malocclusion in orthodontic patients typically results from midfacial retrognathia. However, the etiology of the midfacial retrognathia remains unclear. The cranial base is considered to play an important role in the emergence of midfacial morphology due to its location within the craniofacial region as well as its dramatic growth activity during the later prenatal and early postnatal periods. Previously, the nasal septum was considered a key cranial base component which functioned to pull the maxilla anteriorly during growth. Although this nasal septal theory of midfacial advancement is generally accepted, certain midfacial abnormalities occur in the presence of normal nasal septal morphology and growth indicating that additional craniofacial regions must contribute to the control of midfacial prognathism. The purpose of this study was to describe a mouse mutant which displays midfacial retrognathia and to delineate regions of cranial base malgrowth. Further, cellular growth mechanisms responsible for causing the abnormal cranial base growth trajectories were identified. Adult 3H1 Brachyrrhine (Br) male mice, displaying midfacial retrognathia with a characteristic Class III malocclusion, were bred to normal C3H females. Litters were examined to determine whether Br offspring could be distinguished from one another between Theiler stages 23 (E15) and 27 (E19) using qualitative and quantitative methods. Results showed that two distinct groups of offspring were derived: one with midfacial retrognathia and the other without. The cranial base of Br mutants displayed a malformed sphenoidal region while the nasal septum appeared much less affected as revealed by finite element morphometric analysis. In vivo autoradiographic analysis demonstrated the existence of temporal growth sites (TGS). TGS in the sphenoidal regions were deficient in the prenatal Br mouse. Using immunohistochemistry, insulin growth factor (IGF-I) and epidermal growth factor (EGF) as well as their receptors (IGF-IR and EGF-R) were found to be expressed in cranial base chondrocytes. In order to determine whether chondrocytic responses to these growth factors were deficient in the mutant, cell cultures were established, treated with IGF-I or EGF, and cellular proliferation and differentiation were measured using (³H) -thymidine and (³⁵S) -sulfate incorporation. Results from this analysis showed that chondrocytes from the Br posterior cranial base were less responsive to EGF compared to cells from normal posterior cranial bases. Data from this study suggest that the Br mouse displays midfacial retrognathia in a heritable fashion. Deficient growth of the Br sphenoid, particularly in the presphenoidal and sphenoethmoid region, is crucial for the establishment of midfacial retrognathia. In the Br mouse, the sphenoidal deficiency is associated with depressed proliferation in TGS. IGF-I and EGF, as well as their receptors, are expressed within the murine cranial base and the depressed level of cellular proliferation in the Br sphenoidal region results, at least in part, from a diminished response to EGF. Based on this study, I propose a "sphenoidal mechanism of midfacial advancement" whereby the sphenoid actively propels the midface forward in order to achieve proper maxillary prognathism during later prenatal and early postnatal period.