Introdução: A patogênese da apneia obstrutiva do sono (AOS) é complexa e pode variar de acordo com a etnia. O componente anatômico que predispõe à AOS é resultado da interação entre a estrutura óssea e partes moles da via aérea superior (VAS), e pode ser acessado através da pressão crítica de fechamento da faringe (Pcrit). Hipotetizamos que os descendentes de Japoneses e Caucasianos apresentam diferentes preditores para a colapsabilidade da VAS, sugerindo diferentes vias que levam ao desenvolvimento da AOS nestes dois grupos étnicos. Métodos: Homens descendentes de Japoneses (n=39) e Caucasianos (n=39), pareados para idade e gravidade da AOS, foram avaliados através de polissonografia (PSG), Pcrit e tomografia computadorizada (TC) da VAS e abdome para estudo da anatomia da via aérea e gordura abdominal, respectivamente. Resultados: Pcrit foi similar entre descendentes de Japoneses e Caucasianos (-1.0 ± 3.3 vs -0.4 ± 3.1 cmH20). Descendentes de Japoneses apresentaram menores dimensões ósseas craniofaciais (comprimento da base do crânio, maxila e mandíbula), enquanto que os Caucasianos apresentaram maior tamanho das partes moles da VAS (comprimento e volume da língua) e maior desbalanço entre o volume da língua e da mandíbula (razão entre o volume da língua e o volume da mandíbula). O ângulo da base do crânio apresentou associação com a Pcrit somente entre os descendentes de Japoneses (r=-0.535, p < 0.01). A razão volume da língua/volume mandibular se associou com a Pcrit somente nos Caucasianos (r=0.460, p < 0.01). Variáveis relacionadas à obesidade (IMC, circunferências cervical e abdominal, gordura visceral) mostraram correlação semelhante com a Pcrit em ambos os grupos. Conclusões: Descendentes de Japoneses e Caucasianos apresentam diferentes preditores da colapsabilidade da VAS. Enquanto a restrição óssea craniofacial foi determinante para a Pcrit somente entre os descendentes de Japoneses, o desbalanço anatômico entre o volume da língua e da mandíbula foi importante para a Pcrit nos Caucasianos. Estes achados podem ter implicações terapêuticas no tocante à melhora da predisposição anatômica para a AOS entre as etnias / Introduction: Obstructive sleep apnea (OSA) pathogenesis is complex and may vary according to ethnicity. The anatomical component predisposing to OSA is the result of the interaction between bony structure and upper airway soft tissues and can be assessed using passive critical closing pressure (Pcrit). We hypothesized that Japanese-Brazilians and Caucasians present different predictors to upper airway collapsibility, suggesting different causal pathways to develop OSA in these two groups. Methods: Male Japanese-Brazilians (n=39) and Caucasians (n=39) well matched for age and OSA severity were evaluated by full polysomnography, Pcrit and upper airway plus abdomen CT scans for determination of upper airway anatomy and abdominal fat, respectively. Results: Pcrit was similar between Japanese-Brazilians and Caucasians (-1.0 ± 3.3 vs -0.4 ± 3.1 cmH20). Japanese-Brazilians presented smaller upper airway bony dimensions (cranial base, maxillary and mandibular length) while Caucasians presented larger upper airway soft tissue (tongue length and volume) and greater imbalance between tongue and mandible (tongue/mandibular volume ratio). Cranial base angle was associated with Pcrit only among Japanese-Brazilians (r=-0.535, p < 0.01). Tongue/mandibular volume ratio was associated with Pcrit only among Caucasians (r=0.460, p < 0.01). Obesity-related variables (visceral fat, BMI, neck and waist circumferences) showed similar correlation with Pcrit in Japanese-Brazilians and Caucasians. Conclusions: Japanese-Brazilians and Caucasians present different predictors of upper airway collapsibility. While craniofacial bony restriction was determinant to Pcrit only in the Japanese-Brazilians, anatomical imbalance between tongue and mandible volume was important to Pcrit among Caucasians. These findings may have therapeutic implications regarding how to improve anatomical predisposition to OSA across ethnicities

INTRODUÇÃO: A apneia obstrutiva do sono (AOS) é comum na população geral e é caracterizada pelo colapso recorrente da via aérea superior. Há um interesse crescente no desenvolvimento de métodos para melhor entendimento da fisiopatologia da AOS. A técnica da pressão negativa expiratória (NEP) é um método relativamente simples que avalia a colapsabilidade da via aérea superior durante a vigília. Porém, a metodologia varia muito e a maioria dos estudos utilizou o bocal, que pode não retratar de forma adequada o comportamento da nasofaringe e pode interferir na posição da língua. Adicionalmente, não existem estudos que avaliaram a associação da NEP com variáveis anatômicas da via aérea superior. A pressão crítica de fechamento da faringe (Pcrit) é um método bem estabelecido que reflete o componente anatômico da AOS, porém é realizada durante o sono e envolve metodologia complexa. OBJETIVOS: Realizamos 2 estudos em indivíduos normais e portadores de AOS com o objetivo de: Estudo 1) Determinar a influência da interface e posição sobre a medida da colapsabilidade da via aérea superior durante a vigília avaliada pela NEP. Estudo 2) Avaliar a associação entre a colapsabilidade da via aérea superior durante a vigília medida pela NEP com máscara nasal na posição supina e durante o sono medida pela Pcrit com variáveis anatômicas da via aérea superior avaliadas pela tomografia computadorizada (TC). MÉTODOS: Foram recrutados indivíduos com idade entre 18 e 65 anos com suspeita de AOS referidos do Laboratório do Sono do InCor. Os indivíduos foram submetidos a prova de função pulmonar, polissonografia e NEP em 4 situações: posição sentada e supina utilizando tanto bocal como máscara nasal. A NEP foi avaliada pelo parâmetro V0,2SB/V0,2NEP (relação entre o volume expirado a 0,2 s durante a respiração espontânea (3 expirações precedentes à aplicação da NEP) sobre o volume expirado a 0,2 s durante a aplicação da NEP). Um subgrupo dos indivíduos realizou o exame de Pcrit e TC de via aérea superior. RESULTADOS: Estudo 1) Foram estudados um total de 86 indivíduos (72 homens, idade: 46 ± 12 anos, índice de massa corpórea (IMC): 30,2 ± 4,4 kg/m2, índice de apneia/hipopneia (IAH): 32,9 ± 26,4 eventos/hora). Encontramos uma interação entre interface e posição sobre a colapsabilidade da via aérea superior na análise multivariada (p=0,007), sendo que a via aérea superior foi mais colapsável com bocal do que com máscara nasal na posição sentada. A colapsabilidade da via aérea superior foi maior na posição supina do que sentada quando a NEP foi realizada com máscara nasal. Em contraste, a NEP não foi influenciada pela posição quando avaliada com bocal. A resistência expiratória foi significativamente maior e independente da posição com bocal do que máscara nasal (20,7 cmH2O/L.s-1 vs 8,6 cmH2O/L.s-1 respectivamente, p=0,018). Estudo 2) Vinte e oito indivíduos realizaram a NEP com máscara nasal na posição supina, Pcrit e TC da via aérea superior (idade: 45 ± 13 anos, IMC: 29.4±4.9 kg/m2 e IAH: 30 ± 26 eventos/hora). A NEP e a Pcrit se associaram de maneira semelhante com a área da língua (r=0,646 e r=0,585), volume da língua (r=0,565 e r=0,613), comprimento da faringe (r=0,580 e r=0,611) e IAH (r=0,490 e r=0,531), respectivamente (p < 0,05 para todas as correlações). A NEP e a Pcrit foram significativamente piores em pacientes com AOS grave do que no restante da população (p < 0,05). CONCLUSÕES: Estudo 1) A interface e a posição influenciam a colapsabilidade da via aérea superior medida pela NEP. Propomos que a NEP seja realizada com máscara nasal na posição supina em estudos futuros de avaliação da colapsabilidade da via aérea superior em pacientes sob investigação de AOS. Estudo 2) A NEP avaliada com máscara nasal na posição supina é um método simples e promissor que reflete o componente anatômico da colapsabilidade da via aérea superior de forma similar a Pcrit / INTRODUCTION: Obstructive sleep apnea (OSA) is common in the general population and is characterized by recurrent collapse of the upper airway. There is a growing interest in developing methods for better understanding of OSA pathophysiology. Negative expiratory pressure (NEP) technique is a simple method that evaluates upper airway collapsibility during wakefulness. However, the method of NEP determination varies among published studies and is mostly evaluated with a mouthpiece, which could inadequately reflect the behavior of nasopharynx and also interfere on the tongue position. In addition, there are no studies evaluating the association between NEP and upper airway anatomy. Pharyngeal critical closing pressure (Pcrit) is a well established technique that reflects the anatomical component of OSA, however, it is performed during sleep and requires a complex methodology. OBJECTIVES: We performed 2 studies in OSA and normal subjects with the objectives of: Study 1) To determine the influence of interface and position on the measurement of upper airway collapsibility while awake evaluated by NEP. Study 2) To evaluate the association among upper airway collapsibility while awake evaluated by NEP with nasal mask in supine position and during sleep evaluated by Pcrit with upper airway anatomy evaluated objectively by upper airway computed tomography (CT) scan. METHODS: We recruited subjects with age between 18 and 65 years with suspect OSA referred to the outpatient sleep clinic at the Heart Institute, University of São Paulo. Subjects underwent pulmonary function test, polysomnography and NEP evaluations in four conditions: sitting and supine position either with mouthpiece or with nasal mask. NEP was evaluated by the parameter V0.2SB/V0.2NEP (ratio between the volume exhaled at 0.2 s during stable breathing (3 expirations prior to NEP application) over the volume exhaled at 0.2 s during NEP application). A subgroup of subjects performed Pcrit and upper airway CT evaluations. RESULTS: Study 1) We studied a total of 86 subjects (72 male, age: 46 ± 12 years, body mass index (BMI): 30.2 ± 4.4 kg/m2, apnea/hypopnea index (AHI): 32.9 ± 26.4 events/hour). We found an interaction between interface and position on upper airway collapsibility in multivariate analysis (p=0.007), with the upper airway being more collapsible with mouthpiece than with nasal mask in sitting position. Upper airway collapsibility was higher in supine than in sitting position when NEP was performed with nasal mask. In contrast, NEP was not influenced by position when evaluated with mouthpiece. Expiratory resistance was significantly higher and independent of position with mouthpiece than with nasal mask (20.7 cmH2O/L.s-1 vs 8.6 cmH2O/L.s-1 respectively, p=0.018). Study 2) Twenty-eight subjects performed NEP with nasal mask in supine position, Pcrit and upper airway CT scan (age: 45±13 years, BMI: 29.4 ± 4.9 kg/m2, and AHI: 30 ± 26 events/h). NEP evaluated with nasal mask in supine position and Pcrit were similarly associated with tongue area (r=0.646 and r=0.585), tongue volume (r=0.565 and r=0.613), pharyngeal length (r=0.580 and r=0.611), and AHI (r=0.490 and r=0.531) respectively (p < 0.05 for all comparisons). NEP and Pcrit were significantly worse in patients with severe OSA than the remaining population (p < 0.05). CONCLUSIONS: Study 1) Interface and position influence upper airway collapsibility measured by NEP. We propose NEP to be performed with nasal mask in supine position in future studies of upper airway collapsibility evaluation in patients under investigation for OSA. Study 2) NEP evaluated with nasal mask in supine position is a simple and promising method that is associated with the anatomical component of upper airway collapsibility similarly to Pcrit

L’embranchement Hemichordata regroupe les classes Enteropneusta et Pterobranchia. Hemichordata constitue, avec l’embranchement Echinodermata, le groupe-frère des chordés. Les entéropneustes sont des organismes vermiformes solitaires qui vivent sous ou à la surface du substrat et s’alimentent généralement par déposivorie, alors que les ptérobranches sont des organismes coloniaux filtreurs habitant dans un réseau de tubes appelé coenecium. Ce mémoire présente trois études dont le point commun est l’utilisation des hémichordés actuels pour répondre à des questions concernant l’évolution des hémichordés, des chordés, et du super-embranchement qui les regroupe, Deuterostomia. Notre première étude démontre que les fentes pharyngiennes, l’organe pré-oral cilié (POCO) et le pharynx de l’entéropneuste Protoglossus graveolens sont utilisés pour l’alimentation par filtration. Le système de filtration de P. graveolens permet la capture de particules jusqu’à 1.3 um, à un débit de 4.05 mm.s-1, pour une demande énergétique de 0.009 uW. Les similarités structurales et fonctionnelles avec le système de filtration des céphalochordés suggèrent que la filtration pharyngienne est ancestrale aux deutérostomes. Lors de notre deuxième étude, nous avons exploré l’hypothèse selon laquelle le POCO des entéropneustes, une structure ciliée pré-buccale au rôle possiblement chémorécepteur, serait homologue au « wheel organ » des céphalochordés et à l’adénohypophyse des vertébrés. Pour cela, nous avons déterminé par immunohistochimie l’expression de Pit-1, un facteur de transcription spécifique à ces deux structures, chez l’entéropneuste Saccoglossus pusillus. Pit-1 est exprimé dans des cellules sensorielles du POCO, mais aussi dans des cellules épithéliales distribuées dans le proboscis, collet et tronc. Ce patron d’expression ne permet pas de confirmer ou rejeter l’homologie du POCO et de l’adénohypophyse des vertébrés. Lors de notre troisième étude, nous avons caractérisé l’ultrastructure du coenecium des ptérobranches Cephalodiscus hodgsoni, Cephalodiscus nigrescens et Cephalodiscus densus par microscopie électronique à transmisison et à balayage. Cephalodiscus est le groupe frère de Graptolithina, un groupe qui inclut les graptolithes éteints ainsi que les ptérobranches du genre Rhabdopleura. Nous avons décrit les types de fibrilles de collagène présents, leur taille et leur organisation, ainsi que l’organisation globale du coenecium. Nous avons ainsi démontré la présence chez Cephalodiscus d’une organisation similaire au paracortex, pseudocortex et eucortex des graptolithes. La présence chez Cephalodiscus de ce type d’organisation suggère que le cortex est ancestral à la classe Pterobranchia. Ces trois études illustrent plusieurs axes importants de la recherche sur les hémichordés, qui en intégrant des données morphologiques, fonctionnelles et moléculaires permet de reconstruire certains évènements clés de l’évolution des deutérostomes. / The phylum Hemichordata comprises the classes Enteropneusta and Pterobranchia. Together with echinoderms, hemichordates are the sister-group to chordates. Enteropneusts are worm-shaped solitary deposit feeders. Pterobranchs are colonial filter feeders that live in a secreted collagenous domicile called a coenecium. In this thesis, three studies are presented. These studies are based on observations of extant hemichordates, and adress a variety of issues relating to the evolution of hemichordates, chordates, and the super-phylum to which they belong: Deuterostomia. Our first study demonstrates that the gill slits, pre-oral ciliary organ (POCO), and lining of the pharynx of the enteropneust Protoglossus graveolens are used in filter feeding. The filter-feeding system of P. graveolens enables particle capture down to 1.3 um, at a rate up to 4.05 mm.s-1, with a power consumption of 0.009 uW. Structural and functional similarities with the cephalochordate filter-feeding system suggest that pharyngeal filter-feeding is ancestral to the deuterostomes. In our second study, we address the hypothesis that the enteropneust POCO, a putative chemosensory structure located anterior to the mouth, is homologous to the cephalochordate wheel organ and vertebrate adenohypophysis. We characterized the expression pattern of the adenohypophysis-specific transcription factor Pit-1 in the adult enteropneust Saccoglossus pusillus with immunohistochemistry. Pit-1 is expressed in sensory cells of the POCO and in scattered epithelial cells of the proboscis, collar and trunk. This expression pattern does not allow to confirm or reject the homology of the POCO with the vertebrate adenohypophysis. In our third study, we characterized the ultrastructure of the coenecium of the pterobranchs Cephalodiscus hodgsoni, Cephalodiscus nigrescens and Cephalodiscus densus using transmission and scanning electron microscopy. Cephalodiscus is the sister-group to the Graptolithina, which includes the extinct graptolites and the extant pterobranch genus Rhabdopleura. We described the fibril types, size and organization, as well as the general organization of the coenecium. We demonstrated that the coenecium of Cephalodiscus shows similarities with the graptolite eucortex, paracortex and pseudocortex. The cortical-like organization of the coenecium of Cephalodiscus suggests that the cortex is ancestral to the Pterobranchia. Together, these three studies illustrate different axes of hemichordate research, and show how integrating morphological, functional and molecular data allows us toinfer key events in the evolution of deuterostomes.

L’embranchement Hemichordata regroupe les classes Enteropneusta et Pterobranchia. Hemichordata constitue, avec l’embranchement Echinodermata, le groupe-frère des chordés. Les entéropneustes sont des organismes vermiformes solitaires qui vivent sous ou à la surface du substrat et s’alimentent généralement par déposivorie, alors que les ptérobranches sont des organismes coloniaux filtreurs habitant dans un réseau de tubes appelé coenecium. Ce mémoire présente trois études dont le point commun est l’utilisation des hémichordés actuels pour répondre à des questions concernant l’évolution des hémichordés, des chordés, et du super-embranchement qui les regroupe, Deuterostomia. Notre première étude démontre que les fentes pharyngiennes, l’organe pré-oral cilié (POCO) et le pharynx de l’entéropneuste Protoglossus graveolens sont utilisés pour l’alimentation par filtration. Le système de filtration de P. graveolens permet la capture de particules jusqu’à 1.3 um, à un débit de 4.05 mm.s-1, pour une demande énergétique de 0.009 uW. Les similarités structurales et fonctionnelles avec le système de filtration des céphalochordés suggèrent que la filtration pharyngienne est ancestrale aux deutérostomes. Lors de notre deuxième étude, nous avons exploré l’hypothèse selon laquelle le POCO des entéropneustes, une structure ciliée pré-buccale au rôle possiblement chémorécepteur, serait homologue au « wheel organ » des céphalochordés et à l’adénohypophyse des vertébrés. Pour cela, nous avons déterminé par immunohistochimie l’expression de Pit-1, un facteur de transcription spécifique à ces deux structures, chez l’entéropneuste Saccoglossus pusillus. Pit-1 est exprimé dans des cellules sensorielles du POCO, mais aussi dans des cellules épithéliales distribuées dans le proboscis, collet et tronc. Ce patron d’expression ne permet pas de confirmer ou rejeter l’homologie du POCO et de l’adénohypophyse des vertébrés. Lors de notre troisième étude, nous avons caractérisé l’ultrastructure du coenecium des ptérobranches Cephalodiscus hodgsoni, Cephalodiscus nigrescens et Cephalodiscus densus par microscopie électronique à transmisison et à balayage. Cephalodiscus est le groupe frère de Graptolithina, un groupe qui inclut les graptolithes éteints ainsi que les ptérobranches du genre Rhabdopleura. Nous avons décrit les types de fibrilles de collagène présents, leur taille et leur organisation, ainsi que l’organisation globale du coenecium. Nous avons ainsi démontré la présence chez Cephalodiscus d’une organisation similaire au paracortex, pseudocortex et eucortex des graptolithes. La présence chez Cephalodiscus de ce type d’organisation suggère que le cortex est ancestral à la classe Pterobranchia. Ces trois études illustrent plusieurs axes importants de la recherche sur les hémichordés, qui en intégrant des données morphologiques, fonctionnelles et moléculaires permet de reconstruire certains évènements clés de l’évolution des deutérostomes. / The phylum Hemichordata comprises the classes Enteropneusta and Pterobranchia. Together with echinoderms, hemichordates are the sister-group to chordates. Enteropneusts are worm-shaped solitary deposit feeders. Pterobranchs are colonial filter feeders that live in a secreted collagenous domicile called a coenecium. In this thesis, three studies are presented. These studies are based on observations of extant hemichordates, and adress a variety of issues relating to the evolution of hemichordates, chordates, and the super-phylum to which they belong: Deuterostomia. Our first study demonstrates that the gill slits, pre-oral ciliary organ (POCO), and lining of the pharynx of the enteropneust Protoglossus graveolens are used in filter feeding. The filter-feeding system of P. graveolens enables particle capture down to 1.3 um, at a rate up to 4.05 mm.s-1, with a power consumption of 0.009 uW. Structural and functional similarities with the cephalochordate filter-feeding system suggest that pharyngeal filter-feeding is ancestral to the deuterostomes. In our second study, we address the hypothesis that the enteropneust POCO, a putative chemosensory structure located anterior to the mouth, is homologous to the cephalochordate wheel organ and vertebrate adenohypophysis. We characterized the expression pattern of the adenohypophysis-specific transcription factor Pit-1 in the adult enteropneust Saccoglossus pusillus with immunohistochemistry. Pit-1 is expressed in sensory cells of the POCO and in scattered epithelial cells of the proboscis, collar and trunk. This expression pattern does not allow to confirm or reject the homology of the POCO with the vertebrate adenohypophysis. In our third study, we characterized the ultrastructure of the coenecium of the pterobranchs Cephalodiscus hodgsoni, Cephalodiscus nigrescens and Cephalodiscus densus using transmission and scanning electron microscopy. Cephalodiscus is the sister-group to the Graptolithina, which includes the extinct graptolites and the extant pterobranch genus Rhabdopleura. We described the fibril types, size and organization, as well as the general organization of the coenecium. We demonstrated that the coenecium of Cephalodiscus shows similarities with the graptolite eucortex, paracortex and pseudocortex. The cortical-like organization of the coenecium of Cephalodiscus suggests that the cortex is ancestral to the Pterobranchia. Together, these three studies illustrate different axes of hemichordate research, and show how integrating morphological, functional and molecular data allows us toinfer key events in the evolution of deuterostomes.

Diese Arbeit untersucht die Auswirkungen von Cytochrom P450 (CYP)-produzierten Eicosanoiden auf das Pumpen des Pharynx im Nematoden Caenorhabditis elegans und deren funktionellen Vernetzung mit dem Wirken von Neurohormonen. Bei der Erforschung der CYP-Eicosanoid-Signalübertragung konzentrierte sich diese Arbeit hauptsächlich auf die Identifizierung und Charakterisierung von Schlüsselkomponenten, welche an der Regulation der pharyngalen Aktivität des Wurms beteiligt sind, insbesondere Regio- und Stereometaboliten von EPA und AA, Neurohormone (Serotonin und Octopamin) sowie relevante GPCR. Zunächst wurde der Einfluss einer Kurzzeitbehandlung mit CYP-Eicosanoiden und Neurohormonen auf die Bildung von freien CYP-Eicosanoiden im Wildtyp und in verschiedenen Mutantenstämmen analysiert. Die Ergebnisse zeigten, dass 17,18-EEQ die stimulierende Wirkung von Serotonin zeigt, während 20-HETE mit der hemmenden Wirkung von Octopamin überlappt. Darüber hinaus erhöhte Serotonin den freien 17,18-EEQ-Spiegel, während Octopamin selektiv die Synthese von Hydroxymetaboliten induzierte. Zweitens wurde die Stereodiskriminierung von 17,18-EEQ getestet, um das wirksame Enantiomer in der stimulierenden Funktion auf die Pharynxmuskelzellen zu identifizieren. Es wurde festgestellt, dass nur 17(R),18(S)-EEQ Beeinträchtigungen des pharyngalen Pumpens beheben konnte. Darüber hinaus wurden drei Mutantenstämme, die in verschiedenen Serotonin-GPCRs defekt sind, ser-1(ok345), ser-7(tm1325) und ser-7(tm1325)ser-1(ok345), ausgewählt, um zu entschlüsseln, welcher Rezeptor am CYP-Eicosanoid-Signaltransduktionsweg beteiligt sein könnte. Tatsächlich scheint SER-7 für die 17-18-EEQ-Effekte auf die Regulierung der pharyngalen Pumpaktivität von C. elegans erforderlich zu sein. Drittens wurde mit der Identifizierung eines GPCR des vorgeschlagenen Signaltransduktionsweges stromabwärts der CYP-Eicosanoid-Produktion begonnen. NMUR-2 wurde als potenzielles Kandidaten-GPCR-Gen ausgewählt. / This thesis investigated the effects of cytochrome P450 (CYP)-produced eicosanoids on the pharyngeal pumping in the nematode Caenorhabditis elegans and their cross-link with neurohormones. Within the framework of exploring CYP-eicosanoid signaling in C. elegans this work mainly focused on identifying and characterizing key components involved in worm’s pharyngeal activity, in particular regio- and stereoisomeric metabolites of EPA (eicosapentaenoic acid, 20:5 n-3) and AA (arachidonic acid, 20:4 n-6), neurohormones, (serotonin and octopamine) as well as the involvement of relevant GPCR (G protein-coupled receptors). Firstly, the impact of short-term treatment with CYP-eicosanoids and neurohormones on CYP-eicosanoid formation in wildtype and different mutant strains was analyzed. The results showed that 17,18-EEQ (17,18-epoxyeicosatetraenoic acid) mimics the stimulatory effect of serotonin while 20-HETE (20-hydroxeicosatetraenoic acid) overlapped with the inhibitory effect of octopamine. Moreover, serotonin increased free 17,18- EEQ levels, whereas octopamine selectively induced the synthesis of hydroxy-metabolites. Secondly, stereo-discrimination of 17,18-EEQ was tested to identify the effective enantiomer in the stimulatory function on pharyngeal muscle cells. Only 17(R),18(S)-EEQ was found to rescue impairments of pharyngeal pumping. Moreover, three mutant strains defective in different serotonin GPCRs, ser-1(ok345), ser-7(tm1325) and ser-7(tm1325) ser-1(ok345), were selected to decipher which receptor might be involved in the CYP-eicosanoid signaling transduction pathway. In fact, SER-7 seems to be required for the 17-18-EEQ effects on regulation of pharyngeal pumping activity in C. elegans. Third, the identification of a GPCR of the proposed signal transduction pathway down-stream of CYP-eicosanoid production was started. NMUR-2 was selected as a potential candidate GPCR gene.

Introdução: A síndrome da apnéia-hipopnéia obstrutiva do sono (SAHOS) é caracterizada por episódios recorrentes de colapso parcial ou completo da faringe responsáveis por roncos e eventos de apnéia ou hipopnéia, com queda da saturação de oxigênio (SaO2) e despertares freqüentes durante a noite de sono. A nasofaringoscopia com a manobra de Muller (MM) é utilizada rotineiramente na avaliação pré-operatória destes pacientes para diagnóstico do sítio da obstrução retropalatal (RP) e retrolingual (RL) e planejamento terapêutico, porém existem controvérsias a respeito de seus benefícios. Os estudos que mostram sensibilidade do sono induzido (SI) neste sentido utilizaram métodos subjetivos de avaliação e doses não controladas do indutor do sono. Objetivos: O principal objetivo do estudo foi comparar o colapso da faringe observado pelos dois métodos diagnósticos, a MM e o SI, através da análise quantitativa da obstrução documentada pela vídeo-nasofaringoscopia, e estudar as correlações deste colapso com o índice de apnéia-hipopnéia (IAH) e a SaO2 mínina noturna. Métodos: Foram estudados prospectivamente 24 pacientes com SAHOS (IAH > 10) e 15 controles saudáveis (IAH < 5) por meio da vídeo-nasofaringoscopia com MM e SI. O video-broncoscópio Olympus com canal de trabalho de 2,0 mm foi utilizado durante o procedimento. Todos os pacientes foram avaliados em decúbito dorsal horizontal e com a cabeça vi em posição neutra. Durante a MM, a pressão negativa intrafaríngea foi controlada por meio de um manovacuômetro conectado ao canal de trabalho do video-broncoscópio. Baixa dose e infusão lenta de midazolam foram utilizadas na indução do sono, e o controle da dose foi realizado por meio da polissonografia (PSG) em tempo real ao exame endoscópico. As imagens das variações de áreas e diâmetros, laterolateral e ântero-posterior obtidas, durante a MM e o SI, foram capturadas e posteriormente analisadas no programa de computador OSIRIS (Osiris Medical Imaging Software - University Hospital of Geneva). Resultados: O colapso faríngeo foi mais significativo em indivíduos com SAHOS durante o SI. Na região RP o colapso das paredes da faringe à MM foi semelhante àquele do SI. Na região RL o colapso à MM foi significativamente inferior ao observado no SI. Diferente do que ocorreu na MM, o colapso RL durante o SI apresentou boa correlação com o IAH e a SaO2 mínima noturna. A dose total de midazolam necessária à indução do sono foi significativamente menor no grupo SAHOS. Conclusão: A vídeo-nasofaringoscopia com SI e análise quantitativa do colapso se mostrou um método simples, superior à MM para documentar a obstrução funcional da faringe, apresentando boa correlação com os índices da PSG noturna devendo ser realizada na prática clínica. / Introduction: Obstructive sleep apnea syndrome (OSAHS) is characterized by partial or total collapse of the retropalatal (RP) or retroglosal (RG) pharynx during sleep. It is responsible for episodes of apnea-hypopnea, snore and sleep fragmentation. Nasopharyngoscopy with Muller Maneuver (MM) has been used to evaluate the site of obstruction in awaking patients. Literature has not definitively considered as the best technique for evaluation. Some authors suggest that induced sleep (IS) is better than MM, but the method has been criticized because of subjective analysis of collapse of and for not being an ideal level of sedation. Some protocols compared the two methods using subjective analysis. Objectives: The aim of this study was to compare quantitatively RP and RL collapse of normal and apneic subjects during MM and IS video-nasopharyngoscopy, determine the best correlations to the apnea-hypopnea index (AHI) and minimal oxygen saturation (SaO2) in nocturnal polissomnography (PSG). Methods: We prospectively evaluated 24 OSAHS patients (AHI >10) and 15 controls (AHI< 5), identified by nocturnal PSG, using video-nasopharyngoscopy with MM and IS. An Olympus-video-bronchoscope with 2.0 mm channel was used during the procedure. All patients were supine with the head in neutral position. The negative pharyngeal pressure was measured during MM by an aneroid manometer connected to the bronchoscope channel. Low dose and slow infusion of midazolam was necessary to IS and the dosage was controlled by PSG in real time during the procedure. The images of the airway obstruction were captured and analyzed by computed software. Variations between cross sectional areas, lateral dimension and antero-posterior dimension of the RP and RG pharynx during MM and IS was analyzed by OSIRIS program (Osiris Medical Imaging Software - University Hospital of Geneva). Results: the collapse of the pharynx was greater in OSAHS than in controls during IS. Retropalatal obstruction was similar between the two methods. In contrast, RG collapse was significantly lower during MM than IS. This study has also shown that RG collapse during IS correlated to AHI and nocturnal SaO2. The total dose of midazolam was lower in OSAHS patients than in controls. Conclusion: Video-nasopharyngoscopy with IS is better than MM in understanding the functional site of pharyngeal obstruction when performed by a controlled technique and quantitative analysis. It should be considered in clinical practice.

Introdução: A síndrome da apnéia-hipopnéia obstrutiva do sono (SAHOS) é caracterizada por episódios recorrentes de colapso parcial ou completo da faringe responsáveis por roncos e eventos de apnéia ou hipopnéia, com queda da saturação de oxigênio (SaO2) e despertares freqüentes durante a noite de sono. A nasofaringoscopia com a manobra de Muller (MM) é utilizada rotineiramente na avaliação pré-operatória destes pacientes para diagnóstico do sítio da obstrução retropalatal (RP) e retrolingual (RL) e planejamento terapêutico, porém existem controvérsias a respeito de seus benefícios. Os estudos que mostram sensibilidade do sono induzido (SI) neste sentido utilizaram métodos subjetivos de avaliação e doses não controladas do indutor do sono. Objetivos: O principal objetivo do estudo foi comparar o colapso da faringe observado pelos dois métodos diagnósticos, a MM e o SI, através da análise quantitativa da obstrução documentada pela vídeo-nasofaringoscopia, e estudar as correlações deste colapso com o índice de apnéia-hipopnéia (IAH) e a SaO2 mínina noturna. Métodos: Foram estudados prospectivamente 24 pacientes com SAHOS (IAH > 10) e 15 controles saudáveis (IAH < 5) por meio da vídeo-nasofaringoscopia com MM e SI. O video-broncoscópio Olympus com canal de trabalho de 2,0 mm foi utilizado durante o procedimento. Todos os pacientes foram avaliados em decúbito dorsal horizontal e com a cabeça vi em posição neutra. Durante a MM, a pressão negativa intrafaríngea foi controlada por meio de um manovacuômetro conectado ao canal de trabalho do video-broncoscópio. Baixa dose e infusão lenta de midazolam foram utilizadas na indução do sono, e o controle da dose foi realizado por meio da polissonografia (PSG) em tempo real ao exame endoscópico. As imagens das variações de áreas e diâmetros, laterolateral e ântero-posterior obtidas, durante a MM e o SI, foram capturadas e posteriormente analisadas no programa de computador OSIRIS (Osiris Medical Imaging Software - University Hospital of Geneva). Resultados: O colapso faríngeo foi mais significativo em indivíduos com SAHOS durante o SI. Na região RP o colapso das paredes da faringe à MM foi semelhante àquele do SI. Na região RL o colapso à MM foi significativamente inferior ao observado no SI. Diferente do que ocorreu na MM, o colapso RL durante o SI apresentou boa correlação com o IAH e a SaO2 mínima noturna. A dose total de midazolam necessária à indução do sono foi significativamente menor no grupo SAHOS. Conclusão: A vídeo-nasofaringoscopia com SI e análise quantitativa do colapso se mostrou um método simples, superior à MM para documentar a obstrução funcional da faringe, apresentando boa correlação com os índices da PSG noturna devendo ser realizada na prática clínica. / Introduction: Obstructive sleep apnea syndrome (OSAHS) is characterized by partial or total collapse of the retropalatal (RP) or retroglosal (RG) pharynx during sleep. It is responsible for episodes of apnea-hypopnea, snore and sleep fragmentation. Nasopharyngoscopy with Muller Maneuver (MM) has been used to evaluate the site of obstruction in awaking patients. Literature has not definitively considered as the best technique for evaluation. Some authors suggest that induced sleep (IS) is better than MM, but the method has been criticized because of subjective analysis of collapse of and for not being an ideal level of sedation. Some protocols compared the two methods using subjective analysis. Objectives: The aim of this study was to compare quantitatively RP and RL collapse of normal and apneic subjects during MM and IS video-nasopharyngoscopy, determine the best correlations to the apnea-hypopnea index (AHI) and minimal oxygen saturation (SaO2) in nocturnal polissomnography (PSG). Methods: We prospectively evaluated 24 OSAHS patients (AHI >10) and 15 controls (AHI< 5), identified by nocturnal PSG, using video-nasopharyngoscopy with MM and IS. An Olympus-video-bronchoscope with 2.0 mm channel was used during the procedure. All patients were supine with the head in neutral position. The negative pharyngeal pressure was measured during MM by an aneroid manometer connected to the bronchoscope channel. Low dose and slow infusion of midazolam was necessary to IS and the dosage was controlled by PSG in real time during the procedure. The images of the airway obstruction were captured and analyzed by computed software. Variations between cross sectional areas, lateral dimension and antero-posterior dimension of the RP and RG pharynx during MM and IS was analyzed by OSIRIS program (Osiris Medical Imaging Software - University Hospital of Geneva). Results: the collapse of the pharynx was greater in OSAHS than in controls during IS. Retropalatal obstruction was similar between the two methods. In contrast, RG collapse was significantly lower during MM than IS. This study has also shown that RG collapse during IS correlated to AHI and nocturnal SaO2. The total dose of midazolam was lower in OSAHS patients than in controls. Conclusion: Video-nasopharyngoscopy with IS is better than MM in understanding the functional site of pharyngeal obstruction when performed by a controlled technique and quantitative analysis. It should be considered in clinical practice.

A Parede lateral da faringe parece ser a estrutura central envolvida no colapso da faringe nos pacientes com síndrome da apnéia obstrutiva do sono (SAOS). Os pacientes com roncos e apnéia têm uma faringe mais colapsável associada a um espessamento da musculatura da parede lateral da faringe, em grande parte formada pelo músculo constritor superior da faringe. O endomísio da matriz extracelular do músculo esquelético tem uma íntima relação com as células musculares, influenciando suas propriedades mecânicas e biológicas, podendo modificar seu comportamento e função. As fibras colágenas e elásticas formam o arcabouço do tecido conectivo, enquanto os proteoglicanos e as glicoproteínas estruturais (fibronectina) têm importante papel na propriedade de adesão e hidratação dos tecidos e as metaloproteínas são responsáveis pela degradação dos componentes da matriz extracelular. Os fatores determinantes da complacência excessiva da musculatura da parede lateral são desconhecidos. É possível que a MEC tenha um papel relevante neste aspecto. O objetivo deste estudo é descrever e comparar a densidade dos componentes da matriz extracelular do endomísio da musculatura da parede lateral da faringe em controles, roncadores e apnéicos. Neste estudo prospectivo foram avaliados 61 pacientes maiores de 18 anos e não obesos do Departamento de Otorrinolaringologia do Hospital das Clínicas da Faculdade de medicina da Universidade de São Paulo, com indicação de cirurgias faríngeas no período de 2005 e 2006. Os pacientes foram divididos em três grupos: controles (14 pacientes sem roncos ou SAOS), 18 roncadores primários e 29 pacientes com SAOS. Os espécimes foram obtidos da musculatura da parede lateral da faringe (constritor superior da faringe) durante cirurgias faríngeas. Por meio de análise histoquímica e imunohistoquímica foi determinada a área proprocional do colágeno total, fibras elásticas, colágeno tipos I e III, metaloproteínas 1 e 2, versican e fibronectina, no endomísio destas amostras e comparada entre os grupos. Os resultados foram ainda correlacionados com dados antropométricos. O colágeno tipo I foi o componente mais abundante da matriz extracelular no constritor superior da faringe. Observamos uma correlação positiva entre o colágeno I e a idade (r=0,42, p=0,01) e inversa com o colágeno III (r=-0,28, p=0.027). O índice de massa corpórea também mostrou uma correlação inversa com o colágeno III (r=-0,311, p=0,017). Não houve diferenças estatisticamente significantes para os componentes da MEC entre os grupos. Em resume, a análise histológica da musculatura da parede lateral da faringe de um largo grupo de pacientes com distúrbios do sono, revelaram que a composição da matriz extracelular é relacionada à idade e o peso, e não pode ser envolvida nos mecanismos que levam a colapsibilidade e espessamento da parede lateral da faringe. / The lateral muscular pharyngeal wall appears to be the predominant structure involved in the collapse of the pharynx in obstructive sleep apnea (OSA). OSA patients and snorers have a narrower pharynx which is more susceptible to collapse and is associated with muscle thickening in the lateral pharyngeal wall, largely formed by superior pharyngeal constrictor. The endomysial extracellular matrix of skeletal muscle has an intimate relationship with the muscle cells and has an important influence on its mechanical and biological properties and can modify their behavior and function. Collagen and elastic fibers form the scaffolding of the connective tissue and proteoglycans and structural glycoproteins (fibronectin) have important roles in hydration, resiliency and adhesive properties of the tissues and the matrix metalloproteinases are involved in extracellular matrix components degradation. The determining factors in the compliance of lateral muscular pharyngeal wall are also unknown. It is possible that the ECM has a relevant role in this aspect. Our objective was to determine and compare the density of the components of the extracellular matrix in the deep, structural lateral pharyngeal muscular walls in snorers, OSA and control patients and correlate with age and weight. In this prospective study we evaluated 61 patients older than 18 and nonobese with indications for pharyngeal surgery at the department of Otolaryngology of the Hospital das Clínicas of the São Paulo University, between 2005 and 2006. The patients were divided in three groups: controls (14 patients), 18 primary snorers and 29 OSA patients. We obtained specimens from the lateral pharyngeal muscular wall (superior constrictor muscle) during pharyngeal surgeries. Using histochemical and immunohistochemical analyses, we determined the fractional area of total collagen, elastic fibers, collagen I and III, matrix metalloproteinase 1 and 2, versican and fibronectin in the endomysium of these samples, and compared among groups. Results were further correlated with anthropometric data. Collagen type I was the most abundant component of the ECM within the pharyngeal muscle. There was a positive correlation between collagen I and age (r=0.419, p=0.01) and an inverse one with type III collagen (r=-0.284, p=0.027). Body mass index inversely correlated with collagen type III (r=-0,311, p=0,017). There were no statistically significant differences for any of the components of the ECM among groups. In summary, histological analysis of the lateral muscular pharyngeal wall of a large group of patients with sleep disturbances revealed that its extracellular matrix composition is age and weight related but may not be involved in the mechanisms leading to the collapsibility of the lateral wall of the pharynx.

Contexte : L'amiante est maintenant une cause avérée de cancer du larynx, mais le rôle des expositions professionnelles dans la survenue des cancers des voies aéro-digestives supérieures (VADS) reste largement méconnu. Bien que plusieurs études aient rapporté des associations entre les expositions professionnelles et ces cancers, il est difficile de synthétiser les résultats et d'en tirer des conclusions définitives. Objectifs : (1) Le premier objectif est de synthétiser les données épidémiologiques disponibles sur les associations entre les cancers de la cavité buccale, du pharynx et du larynx (les cancers des VADS les plus fréquents) et les expositions professionnelles ; (2) le second objectif est, à partir des données d'une large étude cas-témoins, l'étude Icare, d'identifier les professions ou industries présentant des risques élevés de cancer des VADS, puis d'étudier le rôle de certaines expositions professionnelles suspectées (amiante, laines minérales (LM), poussières de ciment, silice). Matériel et méthodes : (1) Une recherche bibliographique et des séries de méta-analyses ont été réalisées pour certaines expositions professionnelles suspectées. (2) L'étude Icare est une étude cas-témoins en population générale française incluant 2415 cas de cancer des VADS et 3555 témoins. L'historique professionnel complet des sujets a été recueilli, avec une description détaillée de chaque emploi exercé. L'évaluation des expositions à l'amiante, aux LM, aux poussières de ciment et à la silice sur l'ensemble de la vie professionnelle a été réalisée à l'aide des matrices emplois-expositions développées dans le cadre du programme Matgéné (Institut de Veille Sanitaire). Résultats : Des méta-risques relatifs (méta-RR) significativement augmentés ont été observés pour le cancer du larynx et les expositions aux hydrocarbures aromatiques polycycliques (HAP), aux gaz d'échappement de moteurs, au travail dans l'industrie textile et dans l'industrie du caoutchouc, et pour le cancer de la cavité buccale et du pharynx et les expositions à l'amiante, aux HAP et aux gaz d'échappement de moteurs. Des augmentations de risque significatives ont été observées pour plusieurs professions et secteurs d'activité exposant notamment aux nuisances présentant des méta-RR augmentés. Les résultats des analyses par nuisance confirment l'association entre l'amiante et le cancer du larynx et suggèrent également une association avec les cancers de la cavité buccale et du pharynx. Une association avec les poussières de ciment est également suggérée. Les résultats ne sont pas en faveur d'un rôle de l'exposition aux LM et à la silice.Conclusion : Ce travail renforce l'hypothèse d'un rôle des expositions professionnelles dans la survenue des cancers des VADS. Dans l'ensemble, nos résultats suggèrent des associations entre le risque de cancer des VADS et les expositions à l'amiante, aux HAP, aux poussières de ciment, et le travail dans l'industrie du caoutchouc.

Thesis (Ph. D.) -- University of Texas Southwestern Medical Center at Dallas, 2004. / Vita. Bibliography: 148-158.