The airway epithelium demonstates the ability to quickly repair following physical injury. The morphologic features of this dynamic repair process have been well characterised at the anatomic and cellular level using a number of animal model systems and these studies have provided a solid foundation upon which our understanding of normal repair is build. With the advent of molecular and bioinformatic tools and resources the opportunity exists to extend the value of these models in defining the molecular pathways and interactions that underlay the normal repair process. This thesis represents a realisation of this opportunity. A large animal model was developed in which selected areas of airway epithelium were subjected to bronchial brush biopsy as part of routine bronchoscopic examination prcedures in anaethetised sheep. The process resulted in a physical perturbation of the normal pseudostratified structure of the sheep airwway epithelium at specific locations. By careful experimental design it was possible, within the same animals. to identify and sample from sites undergoing repair at different intervals subsequent in injury. To supplement the histological evaluation of the repair process and align findings with extablished small animal models of airway epithelial repair proliferative cell labelling strategies were implemented in order to study the location and extent of cellular proliferation occurring duringthe repair process. Molecular approaches towards defining the transcriptional response to physical injury comprised application of microarray technology using a commercially sources array platform. Such approach demanted preliminary effort directed towards optimising RNA integrity and yield from airway samples. Following preliminary studies directed towards optimising the model conditions patterns of airway epithelial repair following bronchial brush biopsy were studies in eight sheep at degined time points (6 hours, 1,3, & 7 days) post-injury. Bronchial brush biopsy resulted in the acute removal of the epithelial cell layer and components of the underlying structures. repair processes were rapidly implemented through initial epithelial dedifferentiation, proliferatino and migration at the wound margins and subsequent time-depentend changes in the proportion of subepithelial structures, including smooth muscle and blood vessels, as the epithelial surface moved towards repair. Transcriptional analysis revewaled that over 13,000 probes showed evidence of differential expession at some point during the repair process (p<0.05), whilst of these, 1491 probes had in excess of a two-fold change in expression. array results were validated against conventional semi-quantitative RT-PCR for selected genes. Differentially expressed genes with previously characterised roles in epithelial migration, prolifereation and differentiation were identified during the repair process. The relative emphasis of gene products with particular functional roles varied during the course of repair. Indeed gene ontology (GO) terms identified included those associated with the inflammatory response, cellular migration, extracellular matrix activities, differentiation, proliferation, cellular development, cell cycle activities, cellular adhesion, apoptosis and mitosis. In addition the Kyoto Encyclopedia of Genes and Gneomes (KEGG) databases were queried and such process indicated the involvement of cell communication, 053 and complement and coagulation cascade pathways throughout the repair process, initial (6h) Toll-like receptor and cytokine-cytoine receptor interaction pathways, and the progressive involement of cell cycle, focal adhesion and extracellulaar matrix (ECM)-receptor, and cytokine interaction pathways as the epithelium repaired. The model of airway epithelial injury developed in this thesis generated features broadly consistent with those previosly described in relation to various small animal model systems. Importantly, and in addition, this thesis defines the molecular features associated with repair in this model system and provides a useful resource with which to assess the comparative fetures of the airway transcriptional response to physical injury, It is through such comparison, using analogous methodology, that the fundamental pathways and interactions that underlay normal repair and regeneration can be identified and therafter extended towards inderstanding the basis for variation associated with natural and experimental disease.

Patients with an anterior mediastinal mass pose significant risk for cardiorespiratory compromise during surgical procedures and general anesthesia. Several techniques have been described to reverse airway obstruction in these patients. In extreme circumstances, patients may require cardiac bypass or extracorporeal membrane oxygenation (ECMO) until definitive treatment of the mass and patient stabilization is achieved. We present a case in which the RulTract (R) system was used for emergency sternal elevation as a bridge to ECMO in acute respiratory collapse in an 11-year-old female with a minimally symptomatic anterior mediastinal mass. (C) 2016 The Authors. Published by Elsevier Inc. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).

published_or_final_version / Dentistry / Master / Master of Dental Surgery

Safe and efficient swallowing requires integrity of both motor and sensory systems. Prior studies have established that motor impairment in individuals with PD frequently manifests as abnormalities in swallowing biomechanics. In contrast, very few studies have investigated the contribution of sensory impairment towards pharyngeal biomechanics and airway protection in this patient cohort. This area should be addressed in light of evidence that the severity of limb motor dysfunction in PD does not reliably predict severity of dysphagia. Emerging data suggests that dysphagia in PD cannot be solely attributed to motor impairment, but may also be influenced by deficits in sensory aspects of airway protection. As an example, silent aspiration in up to 100% has been reported in individuals with PD due to laryngopharyngeal sensory deficits have. Even so, current research lacks information on the integration of both motor and sensory components that make up the swallowing process. The aim of this study was to document changes in airway protection with age, in PD and across severity levels of PD. The project was comprised of two parts. In part one, three parallel studies were conducted to assess a series of both motor and sensory airway mechanism (Chapters 4 to 9). In the first study, 16 young (8 males, age range 21.3 - 32.4) and 16 elder adults (8 males, age range 61.5 - 84.7), were assessed to investigate changes in airway protection that accompany ageing. In the second study, data from individuals diagnosed with PD across severity levels (Hoehn-Yahr 1 - 4, age range 64.2 - 84.5) were age and gender-matched to 16 healthy elders in order to examine the effects of PD on airway protection. In the third, the impact of disease severity was studied with data from 16 individuals in the earlier stages (Hoehn-Yahr ≤ 2, 13 males, age range 51.3 - 82.5, ) compared to 16 individuals in the later stages (Hoehn-Yahr ≥ 2.5, 10 males, age range 61.5 - 78.9). In part two of this project, two smaller, pilot studies were completed to probe the influence of pharmacologic and behavioural treatments on airway protection mechanisms. In the first pilot study, the effect of pharmacotherapy on airway protection was investigated in 10 patients 'on' and 'off' levodopa (Chapter 10). In the second study, 5 patients were assessed before and after completing the Lee Silverman Voice Treatment (LSVT) to document effects of speech rehabilitation on airway protection (Chapter 11). Multimodality assessment elicited data from all participants on both motor and sensory components of airway protection (Chapter 3). Specifically, breathing-swallowing coordination (BSC) and swallowing apnoea (SA) were captured using simultaneous directional nasal airflow and surface electromyography (sEMG). Standard, closed-loop spirometry was used to assess pulmonary function. Swallowing biomechanics were screened using a validated timed test of swallowing efficiency and further evaluated using fibreoptic endoscopic evaluation of swallowing (FEES). Finally, chemo-sensation of the laryngopharynx was determined with the administration of the inhalation cough challenge while mechanosensation was examined using FEES. Results suggest that motor control for airway protection is reasonably robust in PD, although sensory response is impaired. The predominant pattern for swallowing respiratory coordination was mid-expiration for all participants regardless of age and disease severity (Chapter 4). Individuals with PD demonstrated a reduction in average time and volume per swallow, leading to an overall decrease in swallowing capacity (Chapter 5). No difference was found for swallowing efficiency between those in early and later stages of PD. Pulmonary function measures were not significantly different as a function of age, PD or PD severity (Chapter 6). In summary, results from motor assessments contributing to airway protection support the robustness of breathing-swallowing coordination (BSC) and pulmonary function across research groups, but identify a reduction in overall swallowing efficiency in PD. Results from sensory assessments contributing to airway protection revealed that chemosensation was not different between age groups but base of tongue mechano-sensation was diminished in individuals with PD. Natural cough thresholds did not differ between young adults and elders but when asked to stifle coughing, elders were less able to do so compared to young adults (Chapter 7). For the first time, a reduction in mechano-reception at the base of tongue was recorded in individuals with PD (Chapter 8). These patients also demonstrated increased post swallow residual (Chapter 5), which offers an explanation for the complaint of globus in this population. These assessments highlight some compromise to sensory aspects of airway protection in PD. Overall, dysphagia had a negative impact on the quality of life of individuals with PD and even more as disease severity progresses (Chapter 9). Results from part two of the study looking at the effects of therapeutic interventions on airway protection revealed some unexpected findings. In chapter 10, results showed a reduction in pulmonary function when 'on' levodopa, but no differences in swallowing efficiency, BSC, or laryngopharyngeal chemo- and mechano-reception were observed. These results suggested a reduction in pulmonary function with levodopa without any increase in risk of airway protection compromise1. Unexpectedly and documented for the first time, the percentage of post swallow inspiration increased after LSVT (Chapter 11) but as with the levodopa study, this was also not accompanied by any apparent increase in aspiration risk. An increase in submental surface electromyography (sEMG) amplitude across all 5 participants may serve as a proxy measure of improvement in hyolaryngeal excursion. Finally, participants reported an overall improvement in social functioning and communication after LSVT. In conclusion, this study provided evidence that mechano-sensory aspect of airway protection is diminished in individuals with PD, possibly compromising airway protection. Patients not only demonstrated increased residue but the lack of sensation may prevent clearing or spontaneous multiple swallows. Overall, airway protection is maintained in ageing but swallowing efficiency declines in the presence of PD. This study contributes significantly to current research efforts in PD by expanding on existing reports regarding motor aspects of airway protection. Specifically, BSC, swallowing efficiency and evaluation of biomechanics using FEES research have never before been investigated exclusively in the PD population. Finally, the chemo- and mechano-sensation evaluated in this study are an important addition to the limited evidence that sensory impairment in individuals with PD potentially compromises airway protection. Results of the present study will serve as a platform upon which future studies may compare and expand.

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Orientador: Eliana Marisa Ganem / Banca: Norma Suely Pinheiro Módulo / Banca: Tânia Mara Vilela Abud / Resumo: A dificuldade na intubação traqueal é causa de mortalidade em anestesiologia e pode estar relacionada à obesidade. Reconhecer o paciente com intubação difícil contribui para o sucesso da abordagem da via aérea, contudo os parâmetros preditores de intubação difícil não estão bem estabelecidos. A classificação de Mallampati, a distância interincisivos, a circunferência do pescoço, a distância tireomentoniana, a conformação da face e a presença da síndrome da apnéia obstrutiva do sono são parâmetros que podem indicar a intubação difícil. A manobra de BURP tem por objetivo melhorar a visualização da via aérea durante a laringoscopia e variações da mesma também podem ser efetivas. Assim, o objetivo desta pesquisa foi correlacionar os parâmetros acima descritos com a dificuldade de intubação traqueal observada durante a laringoscopia, avaliada por meio da escala de Cormack e Lehane (ECL), sem e com manipulação de laringe (variação da manobra de BURP). Cento e trinta e quatro pacientes de ambos os sexos, foram avaliados no período pré-operatório, quanto ao índice massa corpórea, a classificação de Mallampati, a circunferência do pescoço, a distância interincisivos, a distância tireomentoniana e o grau da síndrome da apnéia obstrutiva do sono. Após a monitorização, o posicionamento adequado e a indução anestésica os pacientes foram submetidos à laringoscopia direta, sendo avaliado o grau da escala de Cormack e Lehane. Nos pacientes com graus superiores a um, realizou-se a manipulação da laringe, reavaliando-se o grau da mesma. A manobra de manipulação da laringe é variação da de BURP e consiste no deslocamento da laringe pelo anestesiologista que esta Introdução e Objetivo 9 realizando a laringoscopia, sem um direcionamento pré-estabelecido. Foram registrados os graus da... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: The difficulty in tracheal intubation is a cause of mortality in anesthesiology and can be associated with obesity. To be able to recognize a patient of difficult intubation may add to the success of the airway approach; however, parameters used to predict difficult intubations are not well established. Mallampati's classification, the distance between incisive teeth, neck circumference, thyroid-chin distance, facial shape and the presence of sleep apnea syndrome are parameters that can point towards a challenging intubation. BURP's maneuver aims at improving airway visualization during laryngoscopy and its variations may also be effective. Thus, the goal of the present study is to correlate the aforementioned parameters with the difficulties associated with tracheal intubation observed during laryngoscopy, evaluated by means of Cormack and Lehane's scale, with and without laryngeal handling (BURP's maneuver variation). One hundred and thirty four patients from both genders were assessed in the pre-operative time as to body mass index, Mallampati's classification, neck circumference, distance between incisive teeth, thyroid-chin distance and the level of obstructive sleep apnea. After monitoring, proper positioning and anesthetic induction, the patients were submitted to direct laryngoscopy, and we assessed their classification according to Cormack and Lehane's scale. In those patients classified above level one, we manipulated their larynx and reassessed their classification. Laryngeal manipulation maneuver is a variation of BURP's maneuver and is based on laryngeal shifting by the anesthesiologist who is doing the laryngoscopy, with no pre-established direction. We recorded the patient's classifications according to Introdução e Objetivo 11 Cormack and Lehane's scale ...(Complete abstract click electronic access below) / Mestre

The mechanisms by which epithelial progenitor cells integrate local signals to balance proliferation with differentiation and regulate patterning during lung organogenesis are still poorly understood. The Hippo pathway and its transcription co-activator Yap have recently emerged as major regulators of progenitor cell expansion and differentiation in development and cancer. Here we investigated the role of Yap signaling in the cellular and molecular events associated with lung epithelial morphogenesis and differentiation. We provide evidence that when airway epithelial tubules are forming and branching, a nuclear to cytoplasmic shift of Yap marks the boundary between the progenitors of the distal lung and the airway compartment. At this transition zone, Yap specifies a transcriptional program that controls the expression of Sox2, restricting distal gene expression and initiating an airway progenitor cell program key to generate the airway epithelium and its branched tubular structures. In Yap deficient mice, epithelial progenitors are unable to properly respond to local Tgf beta-induced cues to control levels and distribution of Sox2, resulting in expansion of the distal epithelial compartment and inability to form airways. Moreover, we show that Yap levels and phosphorylation status play a major role in regulating differentiation of airway progenitors later in development and in adult life. Analysis of YAP-interacting partners in adult airway progenitors by Mass Spectroscopy suggests phosphorylated Yap interactions with ciliome proteins. Our study reveals a crucial role for Yap in specification and differentiation of airway progenitors likely to be also relevant in regeneration-repair of the adult airway epithelium.

Tobacco smoke remains a major public health concern and a factor contributing to the development and progression of various lung diseases world-wide. Smoking cessation can significantly reduce the risk of developing smoking-related diseases, although some smokers remain at an elevated risk despite quitting. Here, I used high-throughput genomic technologies to pave the way for understanding the transcriptomic response in airway epithelium to tobacco exposure, smoking cessation and potentially reduced exposure products (PREP). First, using a longitudinal dataset of nasal airway epithelial cells obtained from active smokers enrolled in smoking cessation programs over 24 weeks, I demonstrated that tobacco-related alterations in the airway gene expression are rapidly reversed within 4 to 8 weeks following smoking cessation. Genes with different biological functions revert towards baseline with different dynamics following smoking cessation. These findings suggest that the nasal-epithelium can serve as a minimally-invasive site to measure the reversible impact of smoking. Secondly, using a dataset of nasal airway epithelial cells from active smokers who switched to potentially reduced exposure products (PREP) for 6 weeks, I showed that gene expression differences induced by switching to PREP may only constitute a partially reduced exposure relative to smoking cessation. My results demonstrate that the nasal-epithelium can also serve as a minimally-invasive site to measure the responses to PREP and may ultimately yield biomarkers to evaluate the potential disease risks associated with these products. Lastly, using small RNA-seq data from bronchial epithelial cells of smokers, I found alterations in airway micro-RNA expression that associate with time since quitting in former smokers. These studies have provided novel insights into the physiologic responses of the airway epithelium to tobacco smoke and PREP and may ultimately serve as a useful approach for evaluating the disease risks associated with changes in smoking behavior. This work sets the stage for additional studies aimed at identifying prevention strategies that decrease the persistent risk of smoking-related lung disease in former smokers and identify biomarkers to assess lung disease risk in former smokers. / 2016-12-31T00:00:00Z