Submitted by Maria Aparecida (cidazen@gmail.com) on 2017-07-31T13:41:46Z No. of bitstreams: 1 Mariane de Amarante Souza.pdf: 1596658 bytes, checksum: 44d5d39f8f8a41833027151f10b44e37 (MD5) / Made available in DSpace on 2017-07-31T13:41:46Z (GMT). No. of bitstreams: 1 Mariane de Amarante Souza.pdf: 1596658 bytes, checksum: 44d5d39f8f8a41833027151f10b44e37 (MD5) Previous issue date: 2017-05-25 / Introduction: Hepatitis C virus (HCV) is an RNA virus with six different genotypes (1 to 6). This virus is mainly transmitted through parenteral route. An estimated 170 million individuals are HCV carriers worldwide. HCV infection is considered as the main cause of liver cirrhosis in the West. The natural history of HCV is related to viral and host factors. Among the latter, type 2 diabetes mellitus (T2DM) has been related with more rapid progression of liver fibrosis. Aim: To evaluate HCV carriers and identify factors associated with more advanced degrees of liver fibrosis. Methods: A cross-sectional study with chronic HCV carriers. The study was performed at the Center for Liver Study Outpatient Clinic of the Presidente Dutra University Hospital, Federal University of Maranhão, São Luís, Maranhão, northeast Brazil. The patients had their medical records analysed. The subjects with the following features were not enrolled for this study: incomplete data, HIV or HBV coinfection, end-stage chronic renal disease on dialysis and individuals who underwent kidney or liver transplantation. Demographic data (sex and age), alcohol intake (yes or no), T2DM (presence or absence), HCV genotype and degrees of liver fibrosis were retrieved from the patients’ medical records. The patients were classified as less or more advanced fibrosis. Both groups were compared to demographic variables, viral genotypes, T2DM presence and alcohol intake. We used the chi-square test and student’s t-test for nominal and numerical variables respectively. Multivariate logistic regression analysis was performed to identify factors independently associated with more advanced degrees of liver fibrosis. We used the SPSS version 23.0 for statistical analyses. Results: A total of 235 patients participated of this study. These patients had complete data on their medical records and met all the inclusion and exclusion criteria. Most of them were male (138/235; 59%) ranging from 18 to 78 years of age (53 ± 10). They were associated with more advanced degrees of fibrosis: Age (OR=1.061, 95% CI: 1.025- 1.098, p<0.001), presence of T2DM (OR 2.227, 95%CI: 1.059-4.142, p = 0.035) and alcohol intake (OR 1.921, 95%CI: 1.129-3.269, p=0.036). Conclusion: T2DM was associated with more advanced degrees of liver fibrosis among the HCV carriers. Thus, diagnosis and treatment of HCV carriers with diabetes are of great importance for interrupting the progression of liver cirrhosis. / Introdução: O vírus da Hepatite C (HCV) é um RNA vírus, que apresenta seis genótipos diferentes (1 a 6) e é transmitido por via predominantemente parenteral. Existem cerca de 170 milhões de indivíduos portadores do HCV em todo o mundo. É a principal causa de cirrose hepática no mundo ocidental. A história natural da infecção pelo HCV é modulada pela interação de fatores virais e do hospedeiro. Entre os fatores associados ao hospedeiro, Diabetes mellitus tipo 2 (T2DM) tem sido associada a maior progressão da doença hepática. Objetivo: avaliar portadores do HCV e identificar fatores associados a graus mais avançados de fibrose hepática. Metodologia: Estudo transversal com portadores da infecção crônica pelo HCV. Foram analisados dados dos prontuários de pacientes atendidos no ambulatório do Núcleo de Estudo do Fígado do Hospital Universitário da Universidade Federal do Maranhão, excluídos portadores de co-infecção com vírus da imunodeficiência humana (HIV) ou vírus da hepatite B (HBV), portadores de doença renal crônica terminal em diálise e transplantados de fígado ou rim. Resgatados dados demográficos (idade e gênero), graus de fibrose hepática, genótipo viral, história de ingestão alcoólica e diagnóstico de T2DM. Os portadores foram categorizados em graus mais e menos avançados de fibrose e comparados quanto aos dados demográficos (idade e sexo), genótipo viral, presença ou não de T2DM e ingestão alcoólica. Diferenças entre variáveis numéricas foram calculadas pelo teste t de Student e entre nominais pelo Quiquadrado. Foi realizada regressão logística multivariada para identificar fatores independentemente associados com graus mais avançados de fibrose. O programa SPSS versão 23.0 foi utilizado. Resultados: Incluídos 225 pacientes, a maioria do sexo masculino (138/235, 59%), com média de idade 53 ± 10 anos. Foram associados a graus mais avançados de fibrose: idade (OR=1,061 (IC 95% 1.025-1,098) P<0.001), presença de T2DM (OR 2,227 (IC 95% 1,059-4,142) P= 0,035) e ingestão alcoólica (OR 1,921(IC 95% 1,129-3,269) P=0,036). Conclusão: Entre portadores crônicos do HCV no Maranhão, a presença de T2DM esteve associada a graus mais avançados de fibrose hepática, sugerindo que é importante o diagnóstico da infecção crônica pelo HCV entre diabéticos, para que o tratamento da infecção seja feito, prevenindo progressão para cirrose hepática.

Pulmonary fibrosis is a common consequence of lung inflammation, leading to organ dysfunction and significant morbidity and mortality. Macrophages, through their diverse functions associated with polarisation status, play a role in lung homeostasis and alternatively activated (M2) macrophages have been associated with lung fibrosis. Prolyl hydroxylases (PHDs) are the main oxygen sensors and regulators of hypoxia inducible factors (HIFs). The PHD/HIF pathway is known to play a role in tissue inflammation and fibrosis, but their role in macrophage polarisation is not fully understood. Aim To study the role of the PHD/HIF pathway in macrophage polarisation and lung fibrosis, and specifically in Idiopathic Pulmonary Fibrosis (IPF). Hypothesis It was hypothesised that pan hydroxylase inhibition alters macrophage polarisation and modulates lung inflammation and fibrosis. Methods A combination of pharmacological (pan hydroxylase inhibitors DMOG and FG41) and genetic (HIF and PHD-null) tools were used to manipulate the PHD/HIF pathway. The bleomycin induced lung fibrosis model was used to define the effect of pan hydroxylase inhibition during the early, inflammatory or the late, fibrotic phase of this model. Murine bone marrow derived macrophages (BMDM), human monocyte derived macrophages and alveolar macrophages obtained from patients with lung fibrosis were used to study the effect of pan hydroxylase inhibition on macrophage polarisation. Bronchoalveolar lavage fluid (BALF) from patients was used to define the association between lung CCL18, an M2 associated chemokine, and disease progression in IPF. Results DMOG therapy during the early phase of the bleomycin model significantly reduced lung fibrosis at day 24. In contrast, late phase pan hydroxylase inhibition enhanced lung fibrosis at day 24. In both instances there was evidence of enhanced alveolar macrophage M2-like polarisation following pan hydroxylase inhibition. Reduced fibrosis after early pan hydroxylase inhibition was not a consequence of reduced acute lung inflammation or direct inhibition of collagen synthesis. In BMDM, pan hydroxylase inhibition resulted in an ‘augmented M2-like’ macrophage. Using LysM-Cre HIF-1α, HIF-2α and PHD-3 KO mice as well as chetomin, a potent inhibitor of HIF-1α and HIF-2α mediated gene expression, the HIF-dependent and HIF-independent polarisation markers were defined. PHD-3 deficiency was not sufficient to enhance M2 skewing. In contrast to murine BMDM, in human monocyte derived macrophages and alveolar macrophages from healthy volunteers and patients with interstitial lung disease including IPF, pan hydroxylase inhibition did not augment M2 polarisation and indeed significantly inhibited macrophage CCL18 expression. CCL18 studies in clinical BALF samples confirmed that CCL18 was elevated in the lungs of patients with IPF and other ILDs compared to controls. However, baseline BALF CCL18 concentrations did not correlate with disease severity or with disease progression, suggesting this is not a useful biomarker in IPF. Further, a unique study of serial BAL in IPF patients showed no association between 12-month change in CCL18 and disease progression over the same period. Indeed CCL18 concentrations mostly fell over 12 months in patients that did progress, strongly suggesting that CCL18 does not play a major pathogenic role in IPF. Concluding, it was shown that in both BMDM and murine lung pan hydroxylase inhibition promoted an ‘augmented M2-like’ polarisation. Pharmacological pan hydroxylase inhibition during the late fibrotic phase of injury enhanced fibrosis but it is not known if there was a causal association between M2 macrophages and lung fibrosis. Similarly, the functional relevance of finding enhanced M2 polarisation observed during early DMOG therapy, which subsequently resulted in attenuated fibrosis, is not known. In human macrophages, pan hydroxylase inhibition unexpectedly attenuated CCL18 production, a chemokine associated with an M2-like phenotype in man whilst other M2 markers were unchanged. However, there was no evidence to support a pathogenic role for CCL18 in IPF, and therefore there is little potential for using pan hydroxylase inhibition to target CCL18 and treat IPF.

Introdução: O tratamento dos pacientes com fibrose cística envolve o uso de medicamentos ototóxicos, sendo que os mais frequentemente utilizados são os antibióticos aminoglicosídeos. Devido ao uso frequente deste tipo de medicamento, os pacientes com fibrose cística apresentam risco de desenvolver perda auditiva. Objetivo: o objetivo deste estudo foi avaliar a audição dos pacientes com fibrose cística pela audiometria de altas frequências (AAF) e emissões otoacústicas por produto de distorção (EOAPD). Pacientes e métodos: estudo transversal retrospectivo e prospectivo, incluindo 75 indivíduos, sendo 39 do grupo de estudo e 36 do grupo controle. Foram realizados os exames de AAF (de 250 a 16.000 Hz) e EOAPD. Resultados: o grupo de estudo apresentou limiares na AAF significativamente mais elevados em 250, 1.000, 8.000, 9.000, 10.000, 12.500 e 16.000 Hz. (p=0,004) e maior prevalência de alterações nas EOAPD em 1.000 e 6.000 Hz (p=0,001), com amplitudes significativamente mais baixas em 1.000, 1.400 e 6.000 Hz. Houve associação significativa entre as alterações dos limiares auditivos na AAF com o número de cursos de aminoglicosídeos realizados (p=0,005). Oitenta e três por cento dos pacientes que realizaram mais de 10 cursos de aminoglicosídeos apresentaram perda auditiva na AAF. Conclusão: Um número expressivo de pacientes com fibrose cística que receberam repetidos cursos de aminoglicosídeos apresentou alterações na AAF e EOAPD. realização de 10 ou mais cursos de aminoglicosídeos esteve associada às alterações na AAF. / Introduction: the treatment of patients with cystic fibrosis involves the use of ototoxic drugs, and the most frequently used are the aminoglycoside antibiotics. Due to the frequent use of this drug, cystic fibrosis patients are at risk to develop hearing loss. Objective: the aim of this study was to evaluate the hearing of patients with cystic fibrosis by high frequency audiometry (HFA) and distortion product otoacoustic emissions (DPOAE). Patients and methods: retrospective and prospective crosssectional study including 75 individuals, 39 of the study group and 36 in the control group. HFA (250 – 16,000 Hz) and DPOAE tests were conducted. Results: the study group had thresholds significantly higher in the HFA in 250, 1,000, 8,000, 9,000, 10,000, 12,500 and 16,000 Hz (p=0.004) and higher prevalence of abnormal DPOAE at 1,000 and 6,000 Hz (p=0.001), with significantly lower amplitudes of 1,000, 1,400 and 6,000 Hz. There was a significant association between changes in hearing thresholds in HFA with the number of courses of aminoglycosides performed (p=0.005). Eighty-three percent of patients who completed more than 10 courses of aminoglycosides had hearing loss in HFA. Conclusion: a significant number of patients with cystic fibrosis who received repeated courses of aminoglycosides showed alterations in HFA and DPOAE.

Despite affecting many organ systems, the leading cause of morbidity and mortality in the cystic fibrosis (CF) population is lung disease. For the current studies we investigated elements of CF lung disease in a porcine model of CF and in people with CF. Our primary analysis tool was chest computed tomography (CT). To investigate early CF lung disease we examined three week old CF and non-CF pigs. We found three week old CF pigs to have large, irregular tracheal smooth muscle bundles, airways of reduced size, airways of irregular shape, and airways of abnormal distensibility. Three week old CF pig lung parenchyma was more heterogenous in density than three week non-CF pigs, especially in the right cephalad lung. The degree of lung tissue heterogeneity in CF pigs correlated with the degree of lung infection. Three week old CF pigs also had significantly more air trapping upon exhalation, evidence of airflow obstruction, than non-CF pigs. The degree of air trapping correlated with the degree of mucus accumulation in the airways. These data show that CF pigs spontaneously develop hallmark features of CF lung disease within weeks of birth, and that abnormal airway growth and development in CF may contribute to lung disease. This study helped set the foundation for future comparative studies involving CF therapeutics, for example, antibiotics and mucolytics. In adults with CF we performed a before drug, after drug study. The drug was ivacaftor, and it restores the basic underlying defect in a subset of people with CF: impaired function of a particular anion channel. We hypothesized that abnormal airway smooth muscle behavior in people with CF, known as “CF asthma,” is, in part, a primary pathogenic mechanism of CF lung disease. We tested our hypothesis by assaying smooth muscle tone before and after administration of ivacaftor. We limited the time duration to two days. We reasoned two days was long enough for ivacaftor to become effective, but not long enough to reverse long standing lung infection and inflammation which could affect smooth muscle function independently. The implication being, that observed changes would be directly due to restoration of the CF defect. We found evidence suggesting relaxation of airway and vascular smooth muscle tone. And, the change in airway smooth muscle tone correlated with the change in vascular smooth muscle tone. These data suggest that impaired smooth muscle function is a primary element of CF lung disease. Many of the people in our two day ivacaftor study returned for follow up after one year of ivacaftor therapy. We hypothesized that radiographic features of lung disease would improve following one year of ivacaftor therapy. We observed no change in lung volume upon inspiration, but a reduction in expiratory lung volume, approximately half of which occurred within two days. Our airway measurements were confounded by errors in scan reconstruction, however, other published studies report airway wall thinning over long term ivacaftor administration. Taken together, these studies of pigs with CF and people with CF, help us understand this disease.

A major challenge in drug development is ensuring that each new candidate drug is delivered to the appropriate location, in a timely manner and at an optimal concentration. Low drug solubility, drug instability, drug degradation, drug toxicity, or rapid clearance from the body can reduce the effectiveness of an otherwise promising drug candidate. Formulations such as nano/microparticles and melt extruded pellets made with synthetic and natural polymers are effective solutions for the advancement of drug delivery technology. These polymeric formulations can provide controlled release of therapeutic agents by delivering constant doses over long periods, cyclic dosages, and tunable release of both hydrophilic and hydrophobic drugs in order to improve the bioavailability and bioactivity of a drug. PLGA-based nanoparticles formed by emulsion or nanoprecipitation techniques can be designed to have a range of degradation times. Particle degradation and drug release kinetics can be controlled by the physiochemical properties of the polymer, such as molecular weight, hydrophobicity, and polydispersity. This study is focused on developing polymeric-based delivery systems for small and large molecules as treatment strategies for arthrofibrosis and bone tissue engineering. In developing arthrofibrotic treatments, several mechanosignaling and biochemical pathways were targeted using small molecule therapeutics such as blebbistatin (a myosin II ATPase inhibitor), paclitaxel (a microtubule stabilizer), sulfasalazine (a kappa B suppressor), beta-aminopropionitrile (a lysyl oxidase inhibitor) and cis-hydroxyproline (inhibits the formation of stable triple helix structure of collagen). The aforementioned drugs were delivered either via PLGA micro/nanoparticles or via pellets formed by melt extrusion. From the studies performed, it was found that blebbistatin delivered by PLGA nanoparticles could reversibly inhibit fibroblast contractile activity and could significantly inhibit collagen synthesis. These findings lay the foundations for further optimization of drug dosing and potentially enabling a new drug delivery technology for treating arthrofibrosis. Sulfasalazine delivered by melt extruded PLGA pellets significantly inhibited myofibroblast numbers as deduced from α-SMA expression and col1A1 gene expression results and thus can be considered a potential treatment for arthrofibrosis. For bone tissue engineering, plasmids encoding differentiation promoting factors or growth factors such as BMP-2 (pBMP-2), FGF-2 (pFGF-2), PDGF (pPDGF) and VEGF (pVEGF) were delivered via polyethylenimine (PEI), a cationic carrier that interacts electrostatically with negatively charged DNA. The formed nanoplexes were either tested directly or by coating them onto biocompatible titanium metal implants and cultured with human bone marrow derived mesenchymal stem cells (hBMSCs). We found that the combinatorial delivery of pBMP-2 and pFGF-2 significantly enhanced bone regeneration as deduced from Runx-2, alkaline phosphatase and osteocalcin gene expression results as well as from data yielded from alizarin red staining assays and atomic absorption spectroscopy where calcium ion levels were measured. It was also found that pBMP-2 nanoplex-coated titanium discs could significantly enhance bone regenerative gene expression for osteocalcin, Runx-2, and alkaline phosphatase as well as enhance calcium ion expression in human adipose derived mesenchymal stem cells (hADMSCs). Thus, it can be concluded that pFGF-2 and pBMP-2 nanoplexes have osteogenic potential and our studies demonstrate a new methodology with the potential to modify titanium disc implant surfaces for the purposes of enhancing osseointegration.

Objetivo: Probar si existe asociación entre la administración de vacuna DPT y el diagnóstico de Neumonitis Intersticial, como hallazgo histopatológico en niños fallecidos, de dos a seis meses de edad. Materiales y métodos: Realizamos un estudio observacional, descriptivo y analítico, de casos y controles no pareado. Los casos fueron lactantes de 2 a 6 meses de edad, fallecidos en los años 2001 y 2002 con estudios histopatológicos realizados en la Morgue Central de Lima y diagnóstico de neumonitis intersticial. Los controles fueron niños de la misma edad, fallecidos en el mismo periodo, con diagnóstico diferente a neumonitis intersticial. Se utilizó el Odds Ratio para analizar la fuerza de asociación y la regresión logística para realizar un análisis multivariado. / Objective: To prove if there is association between immunization with vaccine DTP and Interstitial Pneumonitis, in children, between two and six months of age, died. Materials and methods: We made observacional, descriptive and analytical a case control study. The cases were children of 2 to 6 months old, died in 2001 and 2002 with histopathology studies made in the forensic office of Lima and diagnosis of interstitial pneumonitis. The controls were children of the same age, died in the same period, with diagnosis different from interstitial pneumonitis. Odds Ratio was used to analyze the force of association and the logistic regression to make a multivaried analysis. / Tesis

Cystic fibrosis (CF) is an inherited disease leading to disrupted function of the cystic fibrosis transmembrane conductance regulator (CFTR) anion channel. CF affects many organ systems including the pancreas, liver, intestine, sweat glands, and gallbladder. The leading cause of morbidity and mortality, however, is lung disease. A porcine model of CF was developed, and over time it develops lung disease that recapitulates many of the characteristics observed in humans with CF including airway remodeling, mucus accumulation, infection, and inflammation. At birth, and despite the absence of inflammation and infection, the CF pig airways exhibit a host of abnormalities including tracheal cartilage ring defects, abnormal appearing smooth muscle bundles, reduced trachea diameter, and reduced mainstem bronchi diameter. The primary objectives of this study were to construct an experimental method that allowed for the attainment of airway size information at multiple inflation pressures, to assess the extent of airway narrowing in the newborn CF porcine lung at 20 cmH2O, to determine the tracheal lobe volume for CF and non-CF, and to perform morphologic assessment of the parenchymal airspaces for CF and non-CF newborn pigs. Micro-computed tomography (micro-CT) was selected as the primary analysis tool. The volumetric, high resolution data sets of micro-CT provided a means to virtually track airways through the three dimensional space of the lung, and to image airways as small as 250 microns in diameter. Due to experimental constraints, only one lobe was analyzed: the tracheal lobe; it is the porcine equivalent of the human right upper lobe. Each excised tracheal lobe was cannulated and micro-CT scanned five times. Each lobe was scanned at multiple inflation pressures ranging from 0 to 20 cmH2O. The airways were segmented with a custom designed, substantially-automated computer algorithm. Quantitative analysis of airway size was done with the Pulmonary Workstation 2 software package. At a pressure of 20 cmH2O, the CF airway narrowing was most pronounced in the large airways of the tracheal lobe, and the percent difference in airway cross sectional area between CF and non-CF lessened for airways of smaller size. The volume of the newborn CF pig tracheal lobe was approximately twenty percent smaller than non-CF, but no differences were observed in tracheal lobe airspace histology between the groups. Airway size deviations at birth imply developmental abnormalities in utero that are dependent upon CFTR function. Additionally, the observation that reduced airway caliber exists only in relatively large airways suggests a time-dependent role of CFTR on airway development, as the large airways develop before the small ones in utero. These findings may provide insight to the early pathogenesis of CF lung disease.

The most common cystic fibrosis transmembrane conductance regulator (CFTR) gene mutation is δF508 and this causes cystic fibrosis (CF). Animal models that recapitulate the human disease phenotype are critical to understanding pathophysiologic mechanisms in CF and developing therapies. New CF models in the pig and ferret have been generated that develop lung, pancreatic, liver, and intestinal pathologies that reflect disease in CF patients. Species-specific biology in the processing of CFTR has demonstrated that pig and mouse δF508-CFTR proteins are more effectively processed to the apical membrane of airway epithelia than human δF508-CFTR. The processing behavior of ferret wild-type (WT) and δF508-CFTR proteins remain unknown and such information is important to predicting the utility of a δF508-CFTR ferret. To this end, we sought to compare processing, membrane stability, and function of human and ferret WT- and δF508-CFTR proteins in a heterologous expression system using HT1080, HEK293T, BHK21, and Cos7 cells, as well as human and ferret CF polarized airway epithelia. Analysis of the protein processing and stability by metabolic pulse-chase and surface On-Cell Western blots revealed that WT-fCFTR half-life and membrane stability were increased relative to WT-hCFTR. Furthermore, in BHK21, Cos7, and CuFi cells, human and ferret δF508-CFTR processing was negligible, while low levels of processing of δF508-fCFTR could be seen in HT1080 and HEK293T cells. Only the WT-fCFTR, but not δF508-fCFTR, produced functional cAMP-inducible chloride currents in both CF human and ferret airway epithelia. Further elucidation of the mechanism responsible for elevated fCFTR protein stability may lead to new therapeutic approaches to augment CFTR function. These findings also suggest that generation of a ferret CFTRδF508/δF508 animal model may be useful. Furthermore, in the CFTR and CFTR+/+ ferret model we have characterized abnormalities in the bioelectric properties of the trachea, stomach, intestine and gallbladder of newborn CF ferrets. Short circuit current (ISC) analysis of CF and WT tracheas revealed the following similarities and differences: 1) amiloride sensitive sodium currents were similar between genotypes, 2) responses to 4,4'-diisothiocyano-2,2'-stilbene disulphonic acid (DIDS) were ~4-fold greater in CF animals, suggesting elevated baseline chloride transport through non-CFTR channels, and 3) as expected, there was a lack of IBMX/forskolin-stimulated and GlyH-101-inhibited currents in CF animals due to the lack of CFTR. CFTR mRNA and protein was present throughout all levels of the WT ferret and IBMX/forskolin-inducible ISC was only observed in WT animals. Interestingly, IBMX/forskolin-inducible intestinal ISC in WT animals was not inhibited by the CFTR inhibitor GlyH-101 or bumetanide. The luminal pH of the CF ferret stomach was significantly decreased relative to the controls, while both genotypes maintained near neutral pH along the length of the intestine. The WT stomach and gallbladder exhibited significantly enhanced IBMX/forskolin ISC responses and inhibition by GlyH-101 relative to CF samples. These findings demonstrate that multiple organs affected by disease in the CF ferret have bioelectric abnormalities consistent with the lack of cAMP-mediated chloride transport.

Cystic fibrosis (CF) is a lethal genetic disease that affects 30,000 people in the United States alone. While the disease affects organs throughout the body, it is the lung disease that is the primary cause of morbidity and mortality for people with the disease. CF lung disease is characterized by thick and sticky mucus that obstructs the airways, acute and chronic bacterial infections, and chronic inflammation and remodeling. Thanks to the creation of the CF pig, it is now possible to study the manifestations of CF lung disease at birth. The CF pig develops spontaneous lung disease, similar to that found in humans with CF, making it the ideal model for our studies. One of the critical findings that revealed in studies of the CF pig is that airway surface liquid (ASL) bactericidal activity is impaired in CF at birth, and this activity is pH dependent. Because infants and children with CF tend to suffer greater morbidity from respiratory viruses than non-CF infants and children, we sought to determine if ASL has antiviral activity and if that activity is reduced in newborn CF pigs. We found that pre-incubating either tracheal or nasal ASL from wild-type pigs reduced the infectivity of various recombinant viruses expressing an eGFP or GFP reporter gene. Those viruses include Sendai virus (SeV-eGFP), respiratory syncytial virus (RSV-GFP), the PR8 strain of influenza virus A (PR8-eGFP), and adenovirus (Ad-eGFP), indicating ASL has broad-spectrum antiviral activity. Nasal secretions from newborn CF pigs had strikingly reduced antiviral activity against SeV-eGFP and Ad-eGFP compared to nasal secretions from WT littermates. Unlike what was observed for ASL antibacterial activity, nasal secretion antiviral activity was not affected by pH, nor was it affected by bicarbonate concentration, one of the molecules that drives pH in the airways. However, when we mixed CF and WT nasal secretions at different ratios, we found the antiviral activity to follow a linear trend, with antiviral activity increasing as the percentage of WT nasal secretions increased. This suggests that one or more components of nasal secretions are found less abundantly in CF nasal secretions compared to WT nasal secretions, leading to reduced antiviral activity in CF. The CF pig has facilitated a much greater understanding of the early stages of CF lung disease. This model will allow us to determine what antiviral components are lacking in the CF airways and why they are reduced in CF.

Background: Cystic fibrosis (CF) is a genetically inherited disease which adversely affects the respiratory and gastrointestinal systems. Malnutrition is a major clinical problem in individuals with the disease. Nutritional interventions are warranted as improvements in nutritional status could improve the rates of morbidity and mortality associated with the disease. The review of the literature indicated the need to develop a behavioural-based nutrition prevention program in order for children to achieve CF dietary requirements and appropriate pancreatic enzyme replacement therapy.Methods: The intervention program, Go and Grow with CF, and nutrition and pancreatic enzyme knowledge and self-management questionnaires were developed for children with CF and their carers as part of this thesis. Social learning theory constructs which particularly assist children in achieving desirable behaviours were applied during the development of the Go and Grow with CF program. The program consisted of workshops and a home-based course.Fifty eight children with cystic fibrosis, aged 2 to 11 years, and their carers participated in a clinical trial that was designed to assess the effects of the Go and Grow with CF pilot program on knowledge, self-management, behaviour, dietary intake and body composition, using anthropometry. Process evaluation was conducted on the pilot program and on the clinic-wide implementation of the revised Go and Grow with CF program. The revised program included the Australian Pancreatic Enzyme Replacement Therapy Guidelines and the effects of fat-based dosing were assessed with a cohort of 29 children with CF-related pancreatic insufficiency aged 1 to 13 years.Results: Similar to the process evaluation of the pilot program, 100% of carers who completed the revised home-based course indicated that they would recommend Go and Grow with CF to other families ++ / with a child who has CF. The 'objective assessment of knowledge indicated a significant m improvement in' children's knowledge in the short-term. There were no statistically significant improvements in any of the other parameters assessed. The lack of significant improvements in self-management, behaviour, dietary intake and anthropometry may have been because the program had no effect, the parameters assessed or the instruments used (particularly the questionnaires) were not sufficiently sensitive, the sample size (which was determined by the CF population available) was too small or the duration of the intervention and follow-up was too short.Conclusion: Carers' unanimous recommendation of Go and Grow with CF, together with high levels of perceived learning, reported increase in confidence and improvement in children's knowledge in the short-term, indicate the benefits of the program.Although there was no statistically significant improvement in the anthropometric measurements after the intervention, 'the extensive data obtained during this study suggest that measurements of height and weight may underestimate the presence of poor nutritional status. It is likely that comprehensive assessments of body composition of children with CF would be useful in detecting mild degrees of malnutrition and in providing information about the effects of nutritional status on morbidity and mortality associated with the disease.Fat-based pancreatic enzyme replacement therapy dosing warrants further investigation given that parents had a strong preference for this method and that fat absorption remains abnormal in the majority of individuals who have pancreatic insufficiency. Evaluation of all pancreatic enzyme replacement therapy dosing methods are needed and this research suggests that dose should be assessed on a meal and snack basis, rather than just on daily intake, in order ++ / for levels of adherence to be examined.The apparent absence of a long-term effect of a single exposure to the program on knowledge suggests that regular, ongoing education and counselling is required by families to reinforce aspects related to the child's current stage of development and disease status.