L’œdème cérébral est une complication associée à l’encéphalopathie hépatique (EH) lors d’une insuffisance hépatique chronique (cirrhose du foie). Présentement, l’origine de sa pathogenèse, vasogénique (rupture de la barrière hémato-encéphalique (BHE)) ou cytotoxique (prise anormale d’ions), n’a pas encore été déterminée. Il a été démontré que le co-transporteur Na-K-Cl (NKCC1) du côté luminal des microvaisseaux sanguins cérébraux (CMV) joue un rôle dans le développement de l’œdème cérébral dans des modèles d’ischémie où la bumetanide, un inhibiteur de NKCC, atténue l’œdème cérébral. Deux modèles d’EH ont été utilisés pour cette étude i) la ligature de la voie biliaire (BDL) qui présente l’hyperammoniémie chronique, l’œdème cérébral et le stress oxydatif systémique ; ii) l’anastomose portocave (PCA) qui présente de l’hyperammoniémie chronique seulement. Les buts du projet étaient de: i) définir l’origine du développement de l’œdème chez les rats BDL en étudiant l’extravasation de macromolécules, les jonctions serrées et l’activation des métalloprotéinases matricielles de la BHE; ii) observer les effets de l’hyperammoniémie chronique indépendamment sur la BHE chez les rats PCA; iii) évaluer le rôle de l’hyperammoniémie et du stress oxydatif et iv) étudier le rôle du NKCC1 dans les CMV dans la pathogenèse de l’œdème cérébral. Les résultats du projet démontrent que l’œdème est d’origine cytotoxique chez les rats BDL et que l’intégrité de la BHE est conservée chez les rats PCA malgré l’hyperammoniémie. L’expression génique du NKCC1 est associée à l’œdème mais pas son expression protéique et sa phosphorylation. Enfin, l’étude démontre que l’hyperammoniémie et le stress oxydatif indépendant ne jouent pas un rôle dans la pathogenèse de l’œdème mais suggère qu’ils y aient un effet synergique. / Brain edema is a complication associated with hepatic encephalopathy (HE) due to chronic liver failure (cirrhosis). It is unclear whether brain edema is of vasogenic (blood brain barrier (BBB) breakdown) or cytotoxic (abnormal cellular uptake of ions) origin. It has been demonstrated that the Na-K-Cl cotransporter (NKCC1) located on the luminal side of the cerebral microvessels (CMV) is implicated in the pathogenesis of brain edema in animal models of ischemia and that the administration of bumetanide, an inhibitor of NKCC, attenuates brain water increase. Two distinct animal models of chronic liver failure and HE are used in the present study; 1) bile duct ligation (BDL) where brain edema, chronic hyperammonemia and systemic oxidative stress are observed; 2) portacaval anastomosis (PCA) where only chronic hyperammonemia is observed. The aims of the study were to: i) determine the origin of brain edema in BDL rats measuring brain extravasation, tight junctions expression and matrix metalloproteinase activation; ii) observe the effects of chronic hyperammonemia on the BBB in PCA rats; iii) study the role of oxidative stress and hyperammonemia; iv) evaluate the role of NKCC in CMV in the pathogenesis of brain edema. The results of the study determined that brain edema in BDL rats is of cytotoxic origin and chronic hyperammonemia independently has no effect on the BBB. An increase of NKCC1 mRNA is associated with brain edema but protein expression and phosphorylation are not. Furthermore, hyperammonemia and oxidative stress independently are not implicated in the development of brain edema however a synergistic effect between the two pathogenic factors in BDL rats remains a possibility.

L’encéphalopathie hépatique (EH) est un syndrome neuropsychiatrique découlant des complications de l'insuffisance hépatique. Les patients souffrant d'une insuffisance hépatique chronique (IHC) présentent fréquemment une EH minimale (EHM) caractérisée par des dysfonctions cognitives subtiles qui affectent leur qualité de vie. L'insuffisance hépatique entraîne une hyperammoniémie, le facteur central dans la pathogenèse de l'EH. Pourtant, les taux d'ammoniaque sérique ne sont pas corrélés avec la sévérité de l'EH lors d'une IHC, suggérant que d'autres facteurs y contribuent. L'oedème cérébral est une caractéristique neuropathologique décrite chez les patients souffrant d'une EHM et plusieurs facteurs dont le stress oxydatif, les altérations du métabolisme énergétique et l'augmentation de la glutamine cérébrale pourraient contribuer à la pathogenèse de l'oedème cérébral lors d'une EHM induite par une IHC. Les mécanismes sous-jacents exacts ainsi que les relations entre ces facteurs et l'ammoniaque ne sont pas connus. Présentement, le seul traitement efficace de l'IHC est la transplantation hépatique, une option thérapeutique très limitée. Le but de cette thèse est de contribuer à l'avancement des connaissances sur les mécanismes sous-jacents liés au rôle du stress oxydatif, de la glutamine et du lactate dans la pathogenèse de l'oedème cérébral lors d'une EHM induite par une IHC afin d'envisager de nouvelles options thérapeutiques. Les objectifs précis étaient: 1. Établir le rôle de l’ammoniaque et sa relation avec le stress oxydatif dans la pathogenèse de l'oedème cérébral lors d'une EHM induite par une IHC. 2. Établir le rôle du stress oxydatif dans la pathogenèse de l'oedème cérébral, sa relation avec l'ammoniaque et l'effet du traitement avec des antioxydants. 3. Confirmer l'effet synergique entre l'ammoniaque et le stress oxydatif dans la pathogenèse de l'oedème cérébral. 4. Établir le rôle du lactate et de la glutamine dans la pathogenèse de l'oedème cérébral et leur relation avec l’ammoniaque. Pour atteindre ces objectifs, 2 modèles animaux d'EHM obtenus par microchirurgie chez le rat ont été utilisés: 1) la ligature de voie biliaire, un modèle d'IHC et 2) l'anastomose porto-cave, un modèle d'hyperammoniémie induite par la dérivation portosystémique. Nos résultats démontrent que l'ammoniaque et le stress oxydatif indépendamment n'induisent pas l'oedème cérébral lors d'une EHM. Pourtant, lorsque les 2 facteurs agissent ensemble ils présentent ii un effet synergique qui entraîne le développement de l'oedème cérébral, le stress oxydatif étant une première insulte, qui est suivie par l'hyperammoniémie comme deuxième insulte. En plus, le stress oxydatif a été mis en évidence seulement au niveau systémique, et non au niveau central dans notre modèle d'IHC en association avec l'oedème cérébral, suggérant que le stress oxydatif systémique est une conséquence de la dysfonction hépatique et que l'hyperammoniémie n’induit pas le stress oxydatif ni systémique ni central. Nous avons démontré qu’une augmentation du lactate cérébral est une conséquence directe de l'hyperammoniémie et joue un rôle important dans la pathogenèse de l'oedème cérébral lors d'une EHM induite par une IHC, tandis qu’une augmentation de la glutamine au niveau cérébral n'est pas un facteur clé. La compréhension de ces mécanismes a entraîné la proposition de 3 nouvelles stratégies thérapeutiques potentielles pour l'EHM. Elles ciblent la diminution de l'ammoniaque sérique, la réduction du stress oxydatif et l'inhibition de la synthèse du lactate. / Hepatic encephalopathy (HE) is a metabolic neuropsychiatric syndrome which occurs as a complication of liver failure/disease. Patients with chronic liver disease (CLD) present often with minimal HE (MHE) characterized by subtle cognitive dysfunction which impairs their quality of life. Impaired liver function leads to hyperammonemia which is a central factor in the pathogenesis of HE. However, ammonia alone is poorly correlated with the severity of HE during CLD, strongly suggesting other factors may contribute. Brain edema is a neuropathological feature described in MHE patients and several factors such as oxidative stress, energy metabolism alterations and an increase in glutamine may to contribute to the pathogenesis of brain edema during HE related to CLD. However the exact underlying mechanisms and the relationships between these factors and ammonia are poorly understood. To date, the only effective treatment of CLD remains liver transplantation, a limited therapeutic option. The aim of this thesis is to advance the knowledge into the mechanisms underlying the role of oxidative stress, glutamine and lactate in the pathogenesis of brain edema during MHE associated with CLD in order to uncover new therapeutic options. The study objectives were: 1. Define the role of ammonia and its relationship with oxidative stress in the pathogenesis of brain edema in CLD. 2. Define the role of oxidative stress in the pathogenesis of brain edema, its relationship with ammonia as well as the effect of antioxidant treatment. 3. Confirm a synergistic role of ammonia and oxidative stress in the pathogenesis of brain edema. 4. Define the role of lactate and glutamine in the pathogenesis of brain edema and their relationship with ammonia. To achieve these objectives, we used 2 microsurgical rat models: 1) bile-duct ligation, a cirrhosis model and 2) portacaval anastomosis, a hyperammonemia model following portal-systemic shunting. Our findings demonstrate that ammonia and systemic oxidative stress independently do not induce brain edema in MHE related to CLD. However, when both factors are present, they exert a synergistic effect leading to the development of brain edema with oxidative stress presenting as a “first hit”, followed by hyperammonemia as a “second hit”. Moreover, solely systemic and not central oxidative stress was observed in our CLD rat model in relation to brain edema implying that systemic oxidative stress is a consequence of liver dysfunction and that central oxidative stress is not a direct iv effect of hyperammonemia in the setting of CLD. Moreover, we revealed that increased cerebral lactate is a direct consequence of hyperammonemia and also plays an important role in the pathogenesis of brain edema, while increased cerebral glutamine does not. The understanding of these mechanisms led to the proposal of three different strategies as potential HE therapies. These are directed towards lowering ammonia, reducing oxidative stress and inhibiting lactate synthesis.

Trichorhinophalangeal syndrome is a malformation syndrome characterized by craniofacial and skeletal abnormalities and is inherited in an autosomal dominant manner. We distinguish free subtypes on clinical and molecular level - TRPS I, TRPS II, TRPS III. All TRPS patients have sparse hair, a pear-shaped nose, a long flat philtrum, a thin upper lip and protruding ears. Skeletal abnormalities include cone-shaped epiphyses at the phalanges, hip malformations and short stature are present. The subgroups TRPS I and TRPS III are result of the mutated TRPS1 gene, which is maped into the 8q24 region. This gene is situated proximal of the EXT1 gene, both genes are affected in a subgroup of patients with TRPS II. These patients suffer more from multiple (cartilaginous) exostoses and mental retardation. In this work we performed molecular genetic analysis of a sample of 16 patients, 8 probands showed a TRPS phenotype and 8 probands had only isolated exostoses. The peripheral venous blood of patients was used to gain purified DNA, which was subsequently used to investigate the chromosome 8q24 region using MLPA ("multiplex ligation-dependent probe amplification"). This analysis revealed a deletion in 1 TRPS patient and 1 patient with exostoses. Sequencing of the TRPS1 gene coding exons in remaining 7 TRPS...

INTRODUÇÃO: Apesar do índice de sucesso da cirurgia da ligadura ou cauterização da artéria esfenopalatina, para o tratamento da epistaxe severa, ser maior que 95%, a falha pode variar de 2% a 10%. Algumas variações anatômicas na parede lateral do nariz são relatadas, sendo referentes à localização do forame esfenopalatino (FEP), à presença de um forame acessório, à ramificação das artérias e à dimensão e morfologia do FEP. A variação anatômica dessa região assim como a escassez de estudos endoscópicos mostrando pontos de reparo para o encontro da artéria esfenopalatina e seus ramos podem justificar a falha cirúrgica em alguns casos, assim como a dificuldade técnica encontrada por alguns autores. OBJETIVO: Descrever a anatomia da região do FEP na parede lateral do nariz e as possíveis variações anatômicas, durante a dissecção endoscópica em cadáveres, e observar as possíveis diferenças entre os achados anatômicos, o gênero (masculino/feminino) e o grupo étnico/racial, assim como a simetria entre as fossas nasais. CASUÍSTICA E MÉTODOS: Estudo anatômico prospectivo realizado de setembro de 2006 a janeiro de 2007. A região do FEP de 61 cadáveres frescos (122 fossas nasais) foi cuidadosamente dissecada, sob visibilização endoscópica. Prevaleceram os cadáveres do sexo masculino (75%) e grupo étnico/racial pardo, seguidos de negros e brancos. Foram observados a presença da crista etmoidal da lâmina perpendicular do osso palatino, a localização dos forames esfenopalatino e acessório, o número de ramos arteriais emergentes pelos forames e a distância dos mesmos à espinha nasal anterior. Os dados foram analisados em relação ao gênero, grupo étnico/racial e simetria entre as fossas nasais do mesmo cadáver. Foi, ainda, avaliada a predição da presença do forame acessório em relação ao número de ramos arteriais emergentes através do FEP, à localização do FEP e à distância do FEP à espinha nasal anterior. RESULTADOS: A crista etmoidal esteve presente em 100% dos cadáveres, sendo anterior ao mesmo em 98,4% das vezes. A localização mais freqüente do FEP foi a região de transição do meato médio e meato superior (86,9%). A distância média do FEP e do forame acessório à espinha nasal anterior foi respectivamente de 66mm e 67mm, com desvio padrão de 5,3 e 4,7mm. O forame acessório esteve presente em 9,83% dos casos. Um único tronco arterial emergia através do FEP em 67,2% das vezes e em 100% dos forames acessórios. A análise da prevalência das variáveis estudadas em relação ao gênero e grupo étnico/racial não mostrou diferenças estatisticamente significantes (p<0,05). A análise da simetria mostrou concordância boa a excelente em relação à localização do FEP (índice Kappa 0,71/ p<0,001); concordância pobre em relação ao número de ramos arteriais emergentes através do FEP (índice Kappa 0,22/p=0,03) e ausência de concordância estatisticamente significante em relação à presença de forame acessório (p=0,53). Nenhuma das variáveis de interesse apresentou associação estatisticamente significante (p>0,05) que permita predizer a presença do forame acessório. CONCLUSÕES: Existem variações anatômicas na parede lateral do nariz que devem ser levadas em consideração para o sucesso do tratamento cirúrgico endoscópico da epistaxe severa. / INTRODUCTION: Even though the success rate of sphenopalatine ligation is greater than 95%, some authors have reported some difficulties in isolating those arteries during endoscopic surgical procedure. The failure rate of the sphenopalatine artery ligation or cauterization may vary from 2% to 10%. Some anatomical variations on the nose lateral wall are reported, with reference to the sphenopalatine foramen (SPF) location, the presence of an accessory foramen, arteries ramification and SPF dimension and morphology. Anatomical variation of the region, as well as scarcity of endoscopic studies showing landmarks to find the sphenopalatine artery and its branches may justify surgical failure. OBJECTIVE: The purpose of this study was to describe the anatomy of SPF region and possible anatomical variations, during the endoscopic cadaver dissection and to observe the symmetry between nasal sides and the relationship to gender and racial group. CASUISTICS AND METHODS: It is a prospective anatomical study developed from September, 2006 to January, 2007. The SPF of 61 fresh cadavers (122 nasal fossae) was carefully endoscopic dissected. Male (75%) and mixed race cadavers prevailed. Presence of ethmoidal crest, location of sphenopalatine and accessory foramens, number of arterial branches emerging through foramens and distances from the foramens to anterior nasal spine were observed. Data were analyzed in relation to gender, racial group and symmetry of the same cadaver. Prediction of the presence of accessory foramen was evaluated in relation to number of arterial branches emerging through SPF, SPF location and distance from the SPF to the anterior nasal spine. RESULTS: Ethmoidal crest was present in 100% of cadavers, being anterior to the SPF in 98.4% of times. The most frequent SPF location was the transition region of middle and superior meatus (86.9%). Mean distance from SPF and accessory foramen to anterior nasal spine was 6.6cm and 6.7cm, respectively. Accessory foramen was present in 9.83% of cases. A single arterial stem emerged through the SPF in 67.2% of times, and 100% through accessory foramens. The prevalence analyses showed no differences statistically significant (p>0,05) between gender and racial group. The symmetry analyses showed a strong conformity (Kappa index 0,71/p<0,01) between nasal fossae in relation to the SPF location; and a poor conformity (Kappa index 0,22/p=0,03) in relation to the number of arterial branches emerging through the SPF. There was no statistically significant conformity between nasal fossae and the presence of accessory foramen (p = 0,53). None of the variables of interest presents any statistically significant (p>0,05) association with the presence of the accessory foramen. CONCLUSIONS: Anatomical variations in the lateral nose wall exist, and should be taken into account, for a well-succeeded endoscopic surgical treatment of severe epistaxis.

L’œdème cérébral est une complication associée à l’encéphalopathie hépatique (EH) lors d’une insuffisance hépatique chronique (cirrhose du foie). Présentement, l’origine de sa pathogenèse, vasogénique (rupture de la barrière hémato-encéphalique (BHE)) ou cytotoxique (prise anormale d’ions), n’a pas encore été déterminée. Il a été démontré que le co-transporteur Na-K-Cl (NKCC1) du côté luminal des microvaisseaux sanguins cérébraux (CMV) joue un rôle dans le développement de l’œdème cérébral dans des modèles d’ischémie où la bumetanide, un inhibiteur de NKCC, atténue l’œdème cérébral. Deux modèles d’EH ont été utilisés pour cette étude i) la ligature de la voie biliaire (BDL) qui présente l’hyperammoniémie chronique, l’œdème cérébral et le stress oxydatif systémique ; ii) l’anastomose portocave (PCA) qui présente de l’hyperammoniémie chronique seulement. Les buts du projet étaient de: i) définir l’origine du développement de l’œdème chez les rats BDL en étudiant l’extravasation de macromolécules, les jonctions serrées et l’activation des métalloprotéinases matricielles de la BHE; ii) observer les effets de l’hyperammoniémie chronique indépendamment sur la BHE chez les rats PCA; iii) évaluer le rôle de l’hyperammoniémie et du stress oxydatif et iv) étudier le rôle du NKCC1 dans les CMV dans la pathogenèse de l’œdème cérébral. Les résultats du projet démontrent que l’œdème est d’origine cytotoxique chez les rats BDL et que l’intégrité de la BHE est conservée chez les rats PCA malgré l’hyperammoniémie. L’expression génique du NKCC1 est associée à l’œdème mais pas son expression protéique et sa phosphorylation. Enfin, l’étude démontre que l’hyperammoniémie et le stress oxydatif indépendant ne jouent pas un rôle dans la pathogenèse de l’œdème mais suggère qu’ils y aient un effet synergique. / Brain edema is a complication associated with hepatic encephalopathy (HE) due to chronic liver failure (cirrhosis). It is unclear whether brain edema is of vasogenic (blood brain barrier (BBB) breakdown) or cytotoxic (abnormal cellular uptake of ions) origin. It has been demonstrated that the Na-K-Cl cotransporter (NKCC1) located on the luminal side of the cerebral microvessels (CMV) is implicated in the pathogenesis of brain edema in animal models of ischemia and that the administration of bumetanide, an inhibitor of NKCC, attenuates brain water increase. Two distinct animal models of chronic liver failure and HE are used in the present study; 1) bile duct ligation (BDL) where brain edema, chronic hyperammonemia and systemic oxidative stress are observed; 2) portacaval anastomosis (PCA) where only chronic hyperammonemia is observed. The aims of the study were to: i) determine the origin of brain edema in BDL rats measuring brain extravasation, tight junctions expression and matrix metalloproteinase activation; ii) observe the effects of chronic hyperammonemia on the BBB in PCA rats; iii) study the role of oxidative stress and hyperammonemia; iv) evaluate the role of NKCC in CMV in the pathogenesis of brain edema. The results of the study determined that brain edema in BDL rats is of cytotoxic origin and chronic hyperammonemia independently has no effect on the BBB. An increase of NKCC1 mRNA is associated with brain edema but protein expression and phosphorylation are not. Furthermore, hyperammonemia and oxidative stress independently are not implicated in the development of brain edema however a synergistic effect between the two pathogenic factors in BDL rats remains a possibility.

INTRODUÇÃO: Apesar do índice de sucesso da cirurgia da ligadura ou cauterização da artéria esfenopalatina, para o tratamento da epistaxe severa, ser maior que 95%, a falha pode variar de 2% a 10%. Algumas variações anatômicas na parede lateral do nariz são relatadas, sendo referentes à localização do forame esfenopalatino (FEP), à presença de um forame acessório, à ramificação das artérias e à dimensão e morfologia do FEP. A variação anatômica dessa região assim como a escassez de estudos endoscópicos mostrando pontos de reparo para o encontro da artéria esfenopalatina e seus ramos podem justificar a falha cirúrgica em alguns casos, assim como a dificuldade técnica encontrada por alguns autores. OBJETIVO: Descrever a anatomia da região do FEP na parede lateral do nariz e as possíveis variações anatômicas, durante a dissecção endoscópica em cadáveres, e observar as possíveis diferenças entre os achados anatômicos, o gênero (masculino/feminino) e o grupo étnico/racial, assim como a simetria entre as fossas nasais. CASUÍSTICA E MÉTODOS: Estudo anatômico prospectivo realizado de setembro de 2006 a janeiro de 2007. A região do FEP de 61 cadáveres frescos (122 fossas nasais) foi cuidadosamente dissecada, sob visibilização endoscópica. Prevaleceram os cadáveres do sexo masculino (75%) e grupo étnico/racial pardo, seguidos de negros e brancos. Foram observados a presença da crista etmoidal da lâmina perpendicular do osso palatino, a localização dos forames esfenopalatino e acessório, o número de ramos arteriais emergentes pelos forames e a distância dos mesmos à espinha nasal anterior. Os dados foram analisados em relação ao gênero, grupo étnico/racial e simetria entre as fossas nasais do mesmo cadáver. Foi, ainda, avaliada a predição da presença do forame acessório em relação ao número de ramos arteriais emergentes através do FEP, à localização do FEP e à distância do FEP à espinha nasal anterior. RESULTADOS: A crista etmoidal esteve presente em 100% dos cadáveres, sendo anterior ao mesmo em 98,4% das vezes. A localização mais freqüente do FEP foi a região de transição do meato médio e meato superior (86,9%). A distância média do FEP e do forame acessório à espinha nasal anterior foi respectivamente de 66mm e 67mm, com desvio padrão de 5,3 e 4,7mm. O forame acessório esteve presente em 9,83% dos casos. Um único tronco arterial emergia através do FEP em 67,2% das vezes e em 100% dos forames acessórios. A análise da prevalência das variáveis estudadas em relação ao gênero e grupo étnico/racial não mostrou diferenças estatisticamente significantes (p<0,05). A análise da simetria mostrou concordância boa a excelente em relação à localização do FEP (índice Kappa 0,71/ p<0,001); concordância pobre em relação ao número de ramos arteriais emergentes através do FEP (índice Kappa 0,22/p=0,03) e ausência de concordância estatisticamente significante em relação à presença de forame acessório (p=0,53). Nenhuma das variáveis de interesse apresentou associação estatisticamente significante (p>0,05) que permita predizer a presença do forame acessório. CONCLUSÕES: Existem variações anatômicas na parede lateral do nariz que devem ser levadas em consideração para o sucesso do tratamento cirúrgico endoscópico da epistaxe severa. / INTRODUCTION: Even though the success rate of sphenopalatine ligation is greater than 95%, some authors have reported some difficulties in isolating those arteries during endoscopic surgical procedure. The failure rate of the sphenopalatine artery ligation or cauterization may vary from 2% to 10%. Some anatomical variations on the nose lateral wall are reported, with reference to the sphenopalatine foramen (SPF) location, the presence of an accessory foramen, arteries ramification and SPF dimension and morphology. Anatomical variation of the region, as well as scarcity of endoscopic studies showing landmarks to find the sphenopalatine artery and its branches may justify surgical failure. OBJECTIVE: The purpose of this study was to describe the anatomy of SPF region and possible anatomical variations, during the endoscopic cadaver dissection and to observe the symmetry between nasal sides and the relationship to gender and racial group. CASUISTICS AND METHODS: It is a prospective anatomical study developed from September, 2006 to January, 2007. The SPF of 61 fresh cadavers (122 nasal fossae) was carefully endoscopic dissected. Male (75%) and mixed race cadavers prevailed. Presence of ethmoidal crest, location of sphenopalatine and accessory foramens, number of arterial branches emerging through foramens and distances from the foramens to anterior nasal spine were observed. Data were analyzed in relation to gender, racial group and symmetry of the same cadaver. Prediction of the presence of accessory foramen was evaluated in relation to number of arterial branches emerging through SPF, SPF location and distance from the SPF to the anterior nasal spine. RESULTS: Ethmoidal crest was present in 100% of cadavers, being anterior to the SPF in 98.4% of times. The most frequent SPF location was the transition region of middle and superior meatus (86.9%). Mean distance from SPF and accessory foramen to anterior nasal spine was 6.6cm and 6.7cm, respectively. Accessory foramen was present in 9.83% of cases. A single arterial stem emerged through the SPF in 67.2% of times, and 100% through accessory foramens. The prevalence analyses showed no differences statistically significant (p>0,05) between gender and racial group. The symmetry analyses showed a strong conformity (Kappa index 0,71/p<0,01) between nasal fossae in relation to the SPF location; and a poor conformity (Kappa index 0,22/p=0,03) in relation to the number of arterial branches emerging through the SPF. There was no statistically significant conformity between nasal fossae and the presence of accessory foramen (p = 0,53). None of the variables of interest presents any statistically significant (p>0,05) association with the presence of the accessory foramen. CONCLUSIONS: Anatomical variations in the lateral nose wall exist, and should be taken into account, for a well-succeeded endoscopic surgical treatment of severe epistaxis.

Along with proteins, a myriad of different molecular biomarkers, such as post-translational modifications and autoantibodies, could be used in an attempt to improve disease detection and progression. In this thesis, I build on several iterations of the proximity ligation assay to develop and apply new adaptable methods to facilitate detection of proteins, autoantibodies and post-translational modifications. In paper I, we present an adaptation of the solid-phase proximity ligation assay (SP-PLA) for the detection of post-translational modification of proteins (PTMs). The assay was adapted for the detection of two of the most commons PTMs present in proteins, glycosylation and phosphorylation, offering the encouraging prospect of using detection of PTMs in a diagnostic or prognostic capacity.  In paper II, we developed a variant of the proximity ligation assay using micro titer plate for detection and quantification of protein using optical density as readout in the fluorometer, termed PLARCA. With a detection limit considerably lower than ELISA, PLARCA detected femtomolar levels of these proteins in patient samples. In paper III, we aim to compare detection values of samples collected from earlobe capillary, venous plasma, as well as capillary plasma stored in dried plasma spots (DPS) assessed with a 92-plex inflammation panel using multiplex proximity extension assay (PEA). Despite the high variability in protein measurements between the three sample sources, we were able to conclude that earlobe capillary sampling is a suitable less invasive alternative, to venipuncture. In paper IV, we describe the application of PLARCA and proximity extension assay (PEA) for the detection of GAD65 autoantibodies (GADA). Thus, offering highly sensitive and specific autoimmunity detection.

Autosomal recessive polycystic kidney disease (ARPKD) is a rare severe inherited disease manifested by cystic renal disease, congenital hepatic fibrosis and dilatatation of bile ducts. The spectrum of clinical manifestations is very wide and variable, depends on the age at which the disease was manifested. In severe forms of the disease, it is possible to detect the first symptoms prenatally around the 20th week of pregnancy due to increased echogenic kidneys and the presence of oligohydramnios. The causal gene of this disease is thePKHD1 gene with protein product fibrocystin that is most likely contributing on maintaining the intracellular concentration of Ca2+ cations. The exact phatophysiology mechanism of ARPKD remains unknown. Phenotypic manifestations of this disease may overlap with mutations associated with other genes. One of the genes mimicking the ARPKD phenotype is the HNF1B gene. Mutations associated with HNF1B gene are the most common monogenic cause of developmental kidney abnormalities. HNF1B is a tissue-specific transcription factor that regulates the expression of PKHD1. In experimental part I worked on genetic analysis of the HNF1B gene in 28 patients who have not been confirmed ARPKD diagnosis by detection of 2 PKHD1 mutations. For the purposes of mutational screening, I used...

Chronic pain affects 20 % of the global population, causes suffering, is difficult to treat, and constitutes a large economic burden for society. So far, the characterization of molecular mechanisms of chronic pain-like behaviors in animal models has not translated into effective treatments. In this thesis, consisting of five studies, pain patient biofluids were analyzed with modern proteomic methods to identify biomarker candidates that can be used to improve our understanding of the pathophysiology chronic pain and lead to more effective treatments. Paper I is a proof of concept study, where a multiplex solid phase-proximity ligation assay (SP-PLA) was applied to cerebrospinal fluid (CSF) for the first time. CSF reference protein levels and four biomarker candidates for ALS were presented. The investigated proteins were not altered by spinal cord stimulation (SCS) treatment for neuropathic pain. In Paper II, patient CSF was explored by dimethyl and label-free mass spectrometric (MS) proteomic methods. Twelve proteins, known for their roles in neuroprotection, nociceptive signaling, immune regulation, and synaptic plasticity, were identified to be associated with SCS treatment of neuropathic pain. In Paper III, proximity extension assay (PEA) was used to analyze levels of 92 proteins in serum from patients one year after painful disc herniation. Patients with residual pain had significantly higher serum levels of 41 inflammatory proteins. In Paper IV, levels of 55 proteins were analyzed by a 100-plex antibody suspension bead array (ASBA) in CSF samples from two neuropathic pain patient cohorts, one cohort of fibromyalgia patients and two control cohorts. CSF protein profiles consisting of levels of apolipoprotein C1, ectonucleotide pyrophosphatase/phosphodiesterase family member 2, angiotensinogen, prostaglandin-H2 D-isomerase, neurexin-1, superoxide dismutases 1 and 3 were found to be associated with neuropathic pain and fibromyalgia. In Paper V, higher CSF levels of five chemokines and LAPTGF-beta-1were detected in two patient cohorts with neuropathic pain compared with healthy controls. In conclusion, we demonstrate that combining MS proteomic and multiplex antibody-based methods for analysis of patient biofluid samples is a viable approach for discovery of biomarker candidates for the pathophysiology and treatment of chronic pain. Several biomarker candidates possibly reflecting systemic inflammation, lipid metabolism, and neuroinflammation in different pain conditions were identified for further investigation. / Uppsala Berzelii Technology Centre for Neurodiagnostics