Topical treatment of several inner ear diseases through the round window membrane (RWM) might be feasible in the near future. Bacteria toxins, ototoxic drugs and noise trauma seem to harm the inner ear by a common pathway which involves, excessive outflow of the afferent neurotransmitter glutamate and formation of nitric oxide (NO), which can severely damage cells/nerve endings and lead to cell death. In this study we used 98 Sprague-Dawley rats and seven human temporal bones. Various substances were instilled into the middle ear of the rat, such as Pseudomonas Aeruginosa Exotoxin (PaExoA), gentamicin, NO-inhibitor N-Omega-Nitro-L-Arginine Methyl Ester (L-NAME), and glucocorticoids. The effects of the substances were studied by morphological analysis of RWM and the endolymphatic sac (ES) by light and electron microscopic. Hearing level was measured in the rats by ABR technique. The human temporal bones were studied immunomorphologically to search for glutamate. In the human inner ear, glutamate receptors and glutamine synthetase, were identified. In the rat, we found, following PaExoA exposure, reversible and permanent hearing loss and morphological changes in the RWM. The ES showed increased numbers of macrophages and thickening of the epithelia. When L-NAME was used as an otoprotector from gentamicin ototoxicity a therapeutic effect in the high frequency area was found. Hydrocortisone (but not dexamethasone) exposure of the RWM resulted in membrane thickening, and adjacent to the membrane, inflammatory cells. The importance of the RWM as a portal for toxic substances and topical treatment of inner ear diseases was highlighted in this study. The difficulties of applying drugs in the round window niche were exposed. The results of this study add important knowledge concerning certain mechanisms of inner ear injury and help us to understand possibilities and problems of local treatment of inner ear diseases in patients.

En els darrers anys s'ha produït un gran avenç en l'enteniment dels mecanismes implicats en la inducció de la placoda òtica. Tanmateix, poc es coneix encara de com s'estableix un domini amb competència neural i un altre no-neural i aquest ha estat l'objectiu d'aquesta tesi doctoral. Hem analitzat els mecanismes moleculars rellevants per la regionalització primarenca de la placoda òtica i hem explorat el paper de Sox3 i Lmx1b en l'establiment i manteniment d'un territori competent neural, emprant l'embrió de pollet com organisme model. Els resultats mostren que el gen Sox3, inicialment expressat en un territori extens, es regionalitza en un domini òtic i epibranquial proneural. La sobreexpressió de Sox3 a estadis preòtics, indueix la generació de precursors neuronals que expressen Sox2 i Delta1, però aquests no aconsegueixen progressar a estadis de major diferenciació. A la vegada, Sox3 és capaç de inhibir la expression de Lmx1b, un gen expressat en el domini no-neural, suggerint que el seu patró final depèn de l'activitat neurogènica de la oïda interna. Finalment, presento evidències que la senyalització mitjançada per BMP té un paper primerenc en l'establiment de l'expressió de Lmx1b en el territory òtic, però que ni l'activitat de BMP ni l'expressió de Lmx1b influencien el procés de determinació neural. En conclusió, els nostres resultats posen de relleu nova informacióour dels mecanismes moleculars que governen els primers passos de la competencia neural i regionalizació de la placoda òtica en un territori neural i un no-neural. / During the last years, a great progress has been made in understanding the mechanisms involved in otic induction but the mechanism behind otic patterning into neural and non-neural domains is still an open question and the major aim of this work was to address this question. We have analyzed the molecular mechanisms underling the early regionalization of the otic placode, and explored the role of Sox3 and Lmx1b in the establishment and maintenance of a neural competent domain in the otic placode by using the chick as a model system. The results show that Sox3 expression initially expressed in a broad domain gets regionalized in otic/epibranchial proneural domain. Overexpression of Sox3 at preotic stages can induce ectopic neuronal precursor cells expressing Sox2 and Delta1 but does not allow the ectopically developed neuronal precursor cells for further differentiation. Sox3, besides providing neural competence to the proneural domain, regulates the posterior non-neural gene Lmx1b suggesting that its final expression pattern depends on the neural activity. Finally, I present evidence that BMP signaling has an early role in inducing Lmx1b expression in the otic field but that neither BMP activity nor Lmx1b expression influence neural commitment. Taken together, our results provide new information and shed light on the molecular mechanisms that underlie the first steps of the neural competence and otic patterning in proneural and non-neural domain.

No description available.

Introdução: O 1º e 2º arcos faríngeos contribuem com o desenvolvimento craniofacial e interferências no seu desenvolvimento podem resultar em alterações de gravidade variável, envolvendo maxila, mandíbula e orelha. Das anomalias associadas à malformação de orelha externa, a hipoplasia mandibular, o dermóide epibulbar e a anomalia de coluna cervical são as mais freqüentes e, este conjunto de sinais tem sido denominado espectro oculoauriculovertebral. O EOAV é uma condição heterogênea e complexa e o espectro de anomalias inclui desde microtia isolada até outras anomalias cranianas e extracranianas. Objetivo: analisar e descrever a morfologia da mandíbula, com ênfase na articulação temporomandibular (ATM) e investigar orelha média e interna em indivíduos com anomalias de 1º e 2º arcos faríngeos cadastrados no Serviço de Genética Clinica do HRAC USP. Indivíduos estudados e métodos: avaliação genética-clínica e avaliação por imagem enfocando orelha média, interna e côndilo mandibular foram realizadas em 56 indivíduos, cadastrados no Serviço de Genética Clínica do Hospital de Reabilitação de Anomalias Craniofaciais USP. O critério mínimo utilizado foi a presença de microtia isolada ou de microssomia hemifacial com malformação auricular leve, como apêndices pré-auriculares. Resultados e conclusão: anomalias envolvendo côndilo mandibular e/ou mandíbula e anomalias de orelha média mostraram alta freqüência (87.5% e 96.1%, respectivamente) nos indivíduos da presente casuística. Relação preditiva entre a ocorrência e a gravidade das anomalias de côndilo mandibular e/ou mandíbula com o acometimento da orelha externa e, relação preditiva para a ocorrência do acometimento do côndilo mandibular com a ocorrência de anomalias estruturais de orelha média e orelha interna foram observadas. Para a avaliação da ATM, a tomografia computadorizada foi considerada o exame de eleição. A investigação das estruturas do côndilo mandibular, da orelha média e interna deve fazer parte do protocolo de avaliação dos indivíduos com anomalias de 1º e 2º arcos faríngeos. / Introduction: The first and second pharingeal arches contribute to craniofacial development and interferences in normal development of these structures can result in maxillary, mandibular, and ear abnormalities. Mandible hypoplasia, epibulbar dermoids, and spinal vertebral defects are the most frequent anomalies combined with ear anomalies and this group has been called oculoauriculovertebral spectrum. This is a heterogeneous and complex condition that includes isolated microtia until other cranial and extracranial anomalies. Purpose: To analyze and describe the morphology of the mandible, with special approach to the temporomandibular joint and to investigate the middle and inner ear in patients with first and second pharyngeal arches involvement. Methods: Clinical genetics evaluation, radiological and CT scan evaluation with main focus in middle and inner ear structures as well as in mandible. This study was performed in fifty six patients at the Hospital of the Rehabilitation of the craniofacial anomalies USP. Minimal diagnostic criteria were the presence of preauricular tags or mild external ear anomaly associated to facial asymmetry. Results and Conclusions: Condyle and/or mandible and middle ear anomalies showed high frequency (87.5% and 96.1%) for the patients of the present study. Predictive relation between the occurrence and the gravity for condyle anomalies and/or mandible with involvement of the external ear was observed. Other predictive relation was observed between the occurrence of condyle anomalies with involvement of structural anomalies of middle and inner ear. CT scan proved to be the most appropriated tool for temporomandibular joint evaluation. Evaluation of condyle structures, middle and inner ear should be included in protocols for evaluation of conditions with first and second pharingeal arches involvement.

Les traitements des pathologies de l’oreille interne par voie locale se développent en alternative aux traitements par voie générale peu efficaces et responsables de nombreux effets secondaires. Dans ce travail, nous avons développé une formulation originale constituée de liposomes dispersés au sein d’un gel d’acide hyaluronique pour la libération prolongée d’un corticoïde dans l’oreille interne après administration locale dans l’oreille moyenne. D'abord, une étude physicochimique approfondie a permis d'identifier les paramètres clés de formulation qui ont un impact sur les propriétés du gel de liposomes en termes de caractéristiques rhéologiques, de seringabilité, de stabilité, de microstructure et enfin de diffusion des liposomes dans le gel. Ensuite, le gel de liposomes contenant la dexamethsone phosphate a été évalué in vivo chez le cobaye. L'administration locale du gel de liposomes a abouti à une libération prolongée du corticoïde dans l'oreille interne, sans aucun effet négatif sur la fonction auditive. L’évaluation de l’efficacité thérapeutique de la formulation pour le traitement de la surdité secondaire à un traumatisme sonore et pour la préservation de l’audition durant l’implantation cochléaire a montré des résultats préliminaires très prometteurs. / Local, rather than systemic drug delivery is being developed to treat inner ear diseases. In this work, we developed an original drug delivery system based on liposomes dispersed within a hyaluronic acid gel for the sustained delivery of a corticoid to the inner ear after local administration in the middle ear. First, a thorough physicochemical study allowed to identify the key formulation parameters that impact the liposomal gel properties in terms of rheological behavior, syringeability, stability, microstructure and diffusion of liposomes within the gel. Then, the liposomal gel containing dexamethsone phosphate was evaluated in vivo in guinea pig. The local administration of the liposomal gel in the middle ear resulted in a sustained release of the corticoid in the inner ear without any negative effect on the hearing function. Promising preliminary data were obtained regarding the therapeutic efficacy of the formulation for hearing recovery after acoustic trauma and for hearing preservation during cochlear implantation.

The main functional cells of the inner ear are neurons and sensory cells that are formed from a common embryonic epithelial neurosensory domain. Discovering genes important for specification and differentiation of sensory cells and neurons in the inner ear is a crucial basis for understanding the pathophysiology of hearing loss. Some of these factors are necessary not only for the inner ear but also for the development of other neurosensory systems such as the visual and olfactory system. The aim of this work was to reveal functions of transcription factor SOX2 in inner ear development by using mouse models with different conditional deletions of Sox2 gene. Sox2 gene was deleted by cre-loxP recombination. In Isl1-cre, Sox2 CKO mutant, reduced number of hair cells differentiated only in some inner ear organs (utricle, saccule and cochlear base) and not in others (cristae and cochlear apex). Early forming inner ear neurons in the vestibular ganglion and neurons innervating the cochlear base developed in these mutants but died by apoptosis due to the lack of neurotrophic support from sensory cells. Late forming neurons in the cochlear apex never formed. In Foxg1-cre, Sox2 CKO mutant, only rudimental ear with no sensory cells was formed. The initial formation of vestibular ganglion with peripheral and...

No description available.

L’espèce humaine est caractérisée par une forme de locomotion bipède qui lui est propre. Toumaï, l’holotype de l’espèce Sahelanthropus tchadensis découverte au Tchad, est le plus ancien représentant connu du rameau humain. Daté de 7 millions d’années, il est très proche de la dichotomie entre ce rameau et celui des chimpanzés. C’est donc un témoin précieux des premières phases de l’évolution de la locomotion dans ce groupe. Le but de ce travail est de reconstituer le comportement locomoteur de Sahelanthropus tchadensis à partir de l’étude de la partie osseuse de son labyrinthe, organe de l’équilibre.Pour cela, les relations entre locomotion et morphologie du labyrinthe osseux sont recherchées chez les hominoïdes actuels. Les covariations entre le labyrinthe et le crâne sont étudiées chez Homo sapiens afin de déterminer l’influence sur la morphologie du labyrinthe de son intégration dans le crâne. Ces résultats sont enfin appliqués au labyrinthe de Toumaï, comparé à d’autres hominines fossiles.La morphométrie géométrique discrimine l’ensemble des espèces actuelles par l’orientation, la taille et la forme relatives de chacune des parties du labyrinthe. Cependant, la torsion et l’orientation relative des canaux semi-circulaires sont majoritairement liées à l’intégration du labyrinthe dans le crâne (orientation des pyramides pétreuses et de la face par rapport à la ligne médio-sagittale du basicrâne). Au contraire, l’excentricité des canaux et leur taille sont uniquement liées à la locomotion.Chez Sahelanthropus tchadensis, la taille des canaux semi-circulaires indique une espèce relativement plus agile que les grands singes. En revanche, il ne présente pas les rotations de canaux caractéristiques du genre Homo. La forme de ses canaux constitue une combinaison unique de caractères : canal postérieur rond et canal antérieur légèrement étiré, comme les humains et les australopithèques, mais canal latéral légèrement projeté postéro-latéralement, comme les gorilles et les babouins. Ceci indiquerait un comportement locomoteur associant le grimper vertical dans les arbres à des déplacements au sol bipèdes mais aussi quadrupèdes. / Humans are characterised by a specific form of bipedal locomotion. Toumaï, holotype specimen of Sahelanthropus tchadensis discovered in Chad, is the oldest known member of the human branch. Dated from 7 million years ago, it is very close to the dichotomy between this branch and the branch of chimpanzees. Therefore, it gives valuable information on the first stages of the evolution of locomotion is this group. The purpose of this work is to reconstruct the locomotor behaviour of Sahelanthropus tchadensis based on the study of the osseous part of its labyrinth, the balance organ.To this aim, the relationships between the locomotion and the bony labyrinth morphology are investigated in extent hominoids. The covariations between the labyrinth and the skull are studied in Homo sapiens in order to determine the influence of cranial integration on labyrinth morphology. Finally, these results are applied to the bony labyrinth of Toumaï, compared to other hominin fossils.Geometric morphometrics discriminates all extant species by the relative orientation, size and shape of each part of the labyrinth. However, the torsion and the relative orientation of the semicircular canals are mainly due to the integration of the labyrinth in the skull (petrous pyramids and face orientations relative to the midline of the basicranium). On the contrary, canal eccentricity and size are linked only to locomotion.In Sahelanthropus tchadensis, the canal size indicates that this species was more agile than great apes. However, it does not have the semicircular canal rotations characterising the genus Homo. The shape of its canals constitutes a unique combination of features: a round posterior canal and a slightly stretched anterior canal, as in humans and australopithecines, but a slightly posterolaterally projected lateral canal, as in gorillas and baboons. This is the sign of a locomotor behaviour associating vertical climbing in trees to both bipedalism and quadrupedalism on the ground.

The inner ear, the sensory organ responsible for hearing and balance, contains specialized sensory and non-sensory epithelia arranged in a highly complex threedimensional structure. To achieve this complexity, a tight coordination between morphogenesis and cell fate specification is essential during otic development. Tisúes surrounding the otic primordium, and more particularly the adjacent segmented hindbrain, have been implicated in specifying structures along the anteroposterior and dorsoventral axes of the inner ear. In this work we have first characterized the generation and axial specification of the otic neurogenic domain, and second, we have investigated the effects of the mutation of kreisler/MafB -a gene transiently expressed in the rhombomeres 5 and 6 of the developing hindbrain- in early otic patterning and cell specification. We show that kr/kr embryos display an expansion of the otic neurogenic domain, due to defects in otic patterning. Although many reports have pointed to the role of FGF3 in otic regionalization, we provide evidence that FGF3 is not sufficient to govern this process. Neither Krox20 nor Fgf3 null mutant embryos, in which Fgf3 is either downregulated or absent in r5 and r6, present ectopic otic neuroblasts in the otic primordium. However, Fgf3-/-Fgf10-/- double mutants show a phenotype very similar to kr/kr embryos: they present ectopic neuroblasts along the AP and DV otic axes. Finally, and remarkably, partial rescue of the kr/kr phenotype is obtained when Fgf3 or Fgf10 are ectopically expressed in the hindbrain of kr/kr embryos. These results highlight a compensatory mechanism between FGFs, and the importance of hindbrain-derived signals in instructing otic patterning and the establishment of the neurogenic domain.

No description available.