Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / CHAPTER II: The stability of the erythrocyte membrane, which is essential for maintenance of cell functions, occurs in a critical region of fluidity, which depends largely on its composition and the composition and characteristics of the medium. As the composition of the erythrocyte membrane is influenced by several blood variables, the stability of the erythrocyte membrane must have relations with them. The present study aimed to evaluate, by bivariate and multivariate statistical analyses, the correlations and causal relationships between hematological and biochemical variables and the stability of the erythrocyte membrane against the chaotropic action of ethanol. The validity of this type of analysis depends on the homogeneity of the population and on the variability of the studied parameters, conditions that can be filled by patients who undergo bariatric surgery by the technique of Roux-en-Y gastric bypass, since they will suffer feeding restrictions that have great impact on their blood composition. Pathway analysis revealed that an increase in hemoglobin leads to decreased stability of the cell, probably through a process mediated by an increase in MCV. Furthermore, an increase in the MCH leads to an increase in the erythrocyte membrane stability, probably because higher values of MCH are associated to smaller quantities of RBC and larger contact area between the cell membrane and ethanol present in the medium. CHAPTER III: The need to treat obesity, a growing worldwide public health problem, has led to an increase in performing bariatric surgery, particularly the Roux-en-Y gastric bypass. The sudden change in eating habits, resulting from this type of surgery, leads to abrupt changes in the body. This study analyzed the correlation between the osmotic stability of erythrocytes and various biochemical and hematological indices in a population consisting of 24 female volunteers, before and at four different times after surgery, distributed along eight weeks, what allowed the generation of 120 sampling points. The osmotic stability of erythrocytes proved to be of great importance for understanding the meaning of the redcell distribution width (RDW), because the stability variables (1/H50 and dX) were positively correlated with this hematological index. However, the stability variables and RDW seem to suffer different influences from other variables, as the LDL-cholesterol (LDL-C), because only RDW has increased throughout time. Indeed, the stability of variable 1/H50 showed positive correlation with the blood levels of LDL-C, which declined throughout time. Path analysis showed that the body mass index (BMI) has an indirect effect, mediated by RDW, on the osmotic stability of erythrocytes. The correlations that the osmotic stability variables presented with RDW may help to understand the origin of the predictive ability of this hematological index in relation to various pathological conditions. / CAPÍTULO II: A estabilidade de membrana de eritrócitos, que é essencial para a manutenção da função dessas células, ocorre em uma região crítica de fluidez, que depende largamente de sua composição e das características do meio. Como a composição da membrana do eritrócito é influenciada por muitas variáveis sanguíneas, a estabilidade de membrana do eritrócito deve ter relações com elas. O presente estudo objetivou avaliar, por análises estatísticas bivariadas e multivariadas, as correlações e relações causais entre variáveis hematológicas e bioquímicas e a estabilidade de membrana de eritrócitos conta a ação caotrópica do etanol. A validade deste tipo de análise depende da homogeneidade da população e da variabilidade dos parâmetros estudados, condições que podem ser satisfeitas por pacientes que sofrem cirurgia bariátrica pela técnica do desvio gástrico em Y-de-Roux, uma vez que eles passam por restrições alimentares que têm grande impacto sobre a composição sanguínea deles. A análise de caminho revelou que um aumento na concentração de hemoglobina leva a uma diminuição da estabilidade da célula, provavelmente através de um processo mediado por um aumento no volume corpuscular médio (MCV). Além disso, um aumento na hemoglobina corpuscular media (MCH) leva a um aumento na estabilidade de membrana do eritrócito, provavelmente porque valores elevados de MCH são associados a menores quantidades de células vermelhas (RBC) e maiores áreas de contato entre a membrana da célula e o etanol presente no meio. CAPÍTULO III: A necessidade de tratar a obesidade, um crescente problema de saúde pública em todo o mundo, tem levado a um aumento na execução de cirurgia bariátrica, particularmente o desvio gástrico pelo Y-de-Roux. A súbita mudança nos hábitos alimentares, resultante deste tipo de cirurgia, leva a mudanças abruptas no corpo. Este estudo analisou a correlação entre a estabilidade osmótica de eritrócitos e vários índices hematológicos e bioquímicos em uma população constituída de 24 participantes do sexo feminino, antes e em quatro diferentes momentos após a cirurgia, distribuídos ao longo de oito semanas, o que permitiu a geração de 120 pontos amostrais. A estabilidade osmótica de eritrócitos mostrou ser de grande importância para a compreensão do significado da distribuição de volumes das células vermelhas do sangue (RDW), porque as variáveis de estabilidade (1/H50 and dX) foram positivamente correlacionadas com este índice hematológico. Entretanto, as variáveis de estabilidade e o RDW parecem sofrer diferentes influências de outras variáveis, como o LDLcolesterol (LDL-C), porque somente o RDW aumentou ao longo do tempo após a cirurgia. Realmente, a variável de estabilidade 1/H50 apresentou correlação positiva com os níveis sanguíneos de LDL-C, os quais diminuíram ao longo do tempo. A análise de caminho mostrou que o índice de massa corporal (BMI) tem um efeito indireto, mediado pelo RDW, sobre a estabilidade osmótica de eritrócitos. As correlações que as variáveis de estabilidade osmótica apresentaram com RDW podem ajudar na compreensão da origem da habilidade preditiva que este índice hematológico tem em relação a várias condições patológicas. / Doutor em Genética e Bioquímica

The cellular prion protein (PrPc) is essential for pathogenesis of fatal neurodegenerative prion diseases. Recently reported four cases of vCJD transmission by blood transfusion raise concerns about the safety of blood products. Proper understanding of PrPc in blood is necessary for development of currently unavailable blood screening tests for prion diseases. Flow cytometry is an attractive method for prion detection, however, the reports on the quantity of PrPc on human blood cells are contradictory. We showed that the majority of PrPc in resting platelets is present in the intracellular pool and is localized in α-granules. We demostrated that both, human platelets and red blood cells (RBC) express significant amount of PrPc and thus may play an important role in the transmission of prions by blood transfusion. Our results suggest a unique modification of PrPc on human RBC. Such modification of pathological prion protein could distort the results of blood screening tests for prions. Further we showed that the storage of blood prior to analysis and the choice of anti-prion antibody greatly affect the detection of PrPc by flow cytometry and we identified platelet satellitism as a factor contributing to the heterogeneity of PrPc detection in blood cells. Moreover, we demonstrated existence of...

Abstract Coronary artery bypass grafting (CABG) is the treatment of choice for patients with three-vessel disease or left main stenosis. However, it is associated with considerable risk of perioperative complications such as myocardial infarction, stroke, infections, and mortality to which excessive bleeding is a contributing factor. This thesis aims to determine the factors involved in and clinical implications of bleeding after CABG. The 1st study evaluated the effects of preoperative ASA discontinuation on the patient’s outcome after CABG. The results showed that late or no discontinuation of low-dose ASA before CABG may decrease the risk of postoperative stroke without increasing the risk of postoperative bleeding. In the 2nd study the use of warfarin was found to be a safe during CABG with no excess bleeding nor other major complications. The 3rd study estimated the impact of surgeons´ performances on blood loss and need for re-exploration after CABG. With 2001 study patients, this study clearly demonstrated that an individual surgeon is a powerful determinant of postoperative bleeding and need for re-exploration after CABG. Using systematic review and meta-analysis, we estimated the risk of complications related to re-exploration for bleeding after CABG. In literature search in 2011, 8 articles with 557 923 patients fulfilled the inclusion criteria. Re-exploration for bleeding after cardiac surgery carries a significantly increased risk of postoperative mortality and morbidity, and thus has a major impact on the patient’s immediate postoperative outcome. We also studied the impact of blood transfusion on the development of post-operative stroke after CABG. Of the study population of 2 226 CABG patients, stroke occurred postoperatively in 53 patients (2.4%). The statistical analysis showed that transfusion of blood products after CABG has a strong, dose-dependent association with the risk of stroke. The use of Octaplas® and platelet transfusions seem to have an even larger impact on the development of stroke than red blood cell transfusions. The 6th study investigated the impact of transfusion of blood products on intermediate outcome after CABG in 2001 patients. The findings indicated that transfusion of any blood product is associated with a significant risk of all-cause and cardiac mortality after CABG. / Tiivistelmä Sepelvaltimotauti on yleisin kuolinsyy ja sepelvaltimoiden ohitusleikkaus hyvine pitkäaikaistuloksineen on todettu parhaaksi hoidoksi potilailla, joilla on monen suonen tai vasemman päärungon tauti. Ohitusleikkaukseen liittyy kuitenkin verenvuodon sekä näihin kytkeytyvien komplikaatioiden riski. Tämän väitöskirjan tavoitteena oli määrittää verenvuodon riskitekijöitä sekä verituotteiden siirtojen vaikutusta ohitusleikkauspotilaiden ennusteeseen. Verenhyytymistä estävien lääkkeiden tiedetään lisäävän verenvuotoja. Ensimmäinen tutkimus osoitti, että ASA:n jatkaminen keskeytyksettä ohitusleikkauksissa vähentää aivoinfarktien riskiä lisäämättä silti verenvuodon riskiä. Toisessa tutkimuksessa pitkäaikainen warfariinihoito osoittautui turvalliseksi ohitusleikkauksen aikana eikä sen käyttö lisännyt verenvuotoja eikä muita komplikaatioita. Kolmas tutkimus osoitti kirurgin taidon merkityksen verenvuotojen ja uusintaleikkausten määrään 2001 potilaalla. Verenvuotojen vuoksi tehtävien uusintaleikkausten negatiivinen vaikutus postoperatiiviseen mortaliteettiin sekä morbiditeettiin on todettu yksiselitteisesti useissa tutkimuksissa. Vuonna 2011 tehdyllä systemaattisella kirjallisuuskatsauksella ja meta-analyysillä selvitimme yhteensä 557 923 ohitusleikkauspotilaan aineistosta, että verenvuodon jälkeisiin uusintaleikkauksiin liittyy huomattava kuoleman ja komplikaatioiden riski. Verenvuotoja hoidetaan yleisesti verensiirroilla, vaikkakin useat tutkimukset ovat osoittaneet verituotteiden annon lisäävän mortaliteettia sekä komplikaatioriskiä. Viides tutkimus selvitteli sepelvaltimoleikkauksissa potilaalle annettujen verituotteiden ja leikkauksen yhteydessä sairastettujen aivoinfarktien välistä yhteyttä. Osoittautui, että verituotteiden käyttöön liittyy annosriippuvaisesti lisääntynyt riski saada aivoinfarkti leikkauksen yhteydessä. Varsinkin verihiutale- ja jääplasmasiirtoihin on todettu liittyvän vielä suurempi aivoinfarktin riski kuin punasolusiirtoihin. Kuudes tutkimus selvitteli sepelvaltimoleikkauksien yhteydessä annettujen verituotteiden vaikutusta 2001 potilaan keskipitkään ennusteeseen. Tutkimus osoitti, että minkä tahansa verituotteen antoon sepelvaltimoleikkauksissa liittyy lisääntynyt kuoleman ja sydänkuoleman riski.

No description available.

The cellular prion protein (PrPc) is essential for pathogenesis of fatal neurodegenerative prion diseases. Recently reported four cases of vCJD transmission by blood transfusion raise concerns about the safety of blood products. Proper understanding of PrPc in blood is necessary for development of currently unavailable blood screening tests for prion diseases. Flow cytometry is an attractive method for prion detection, however, the reports on the quantity of PrPc on human blood cells are contradictory. We showed that the majority of PrPc in resting platelets is present in the intracellular pool and is localized in α-granules. We demostrated that both, human platelets and red blood cells (RBC) express significant amount of PrPc and thus may play an important role in the transmission of prions by blood transfusion. Our results suggest a unique modification of PrPc on human RBC. Such modification of pathological prion protein could distort the results of blood screening tests for prions. Further we showed that the storage of blood prior to analysis and the choice of anti-prion antibody greatly affect the detection of PrPc by flow cytometry and we identified platelet satellitism as a factor contributing to the heterogeneity of PrPc detection in blood cells. Moreover, we demonstrated existence of...

No description available.

Les plaquettes sont essentielles au développement lymphatique dès l’embryogenèse et maintiennent la séparation lympho-sanguine au cours de la vie. Elles participent directement à la lymphangiogenèse et à la réparation des vaisseaux en plus d’améliorer l’intégrité lymphatique in vitro. En présence d’apolipoprotéine A-I (apoA-I), les plaquettes consolident les connexions intercellulaires en adhérant à l’endothélium lymphatique. Toutefois, les plaquettes sont absentes de la lymphe, contrairement à leurs vésicules extracellulaires (pEVs) exprimant un protéome similaire. De plus, leur concentration dans la lymphe s’accentue en condition d’inflammation chronique, par exemple lors de l’athérosclérose. En ayant caractérisé les effets de ces vésicules sur l’intégrité lymphatique durant ma maîtrise, nous montrons que les pEVs sont rapidement internalisées par les LECs et aident à préserver l’intégrité lymphatique des effets délétères des constituants de la lymphe tels que les EVs dérivées des globules rouges (rbEVs). In vitro, des concentrations physiologiques de pEVs limitent la production d’espèces réactives d’oxygène (ROS) et diminuent la nécrose des cellules. Comme les pEVs injectées dans le derme de la peau sont prises en charge par les cellules endothéliales des vaisseaux lymphatiques collecteurs et qu’elles voyagent jusqu’aux ganglions afférents, nous croyons qu’une partie des pEVs circulantes dans la lymphe serait cruciale pour le maintien d’une fonction lymphatique adéquate lors de maladies chroniques inflammatoires telles que l’athérosclérose. / Platelets have a protective role in lymphatic function both at the embryogenic stage and throughout life by maintaining blood-lymphatic separation. In addition, platelets enhance the integrity of lymphatic endothelial cells (LECs) in vitro and appear to exert a shielding effect on the lymphatic endothelium by consolidating the connexion between lymphatic endothelial cells when incubated with apolipoprotein A-I (apoA-I). Whereas platelets are absent from lymph, platelet extracellular vesicles (pEVs) are abundantly circulating within lymphatic vessels, and their level increases during chronic inflammation such as atherosclerosis. Having characterized their effect on lymphatic integrity during my Master internship, we show that pEVs are rapidly internalized by LECs, which in turn help preserve the integrity of the lymphatic endothelium when harmful blood constituent such as red blood cell EVs (rbEVs) are infiltrating lymph. In vitro, physiological concentrations of pEVs are limiting the production of reactive oxygen species (ROS) by lymphatic endothelial cells and decreasing their necrosis rate. Furthermore, pEVs injected in the skin interstitium travel through the collecting lymphatics and are rapidly internalized by LECs, which in turn might help preserve the integrity of the lymphatic endothelium. Lymph pEVs might be critical for the maintenance of a proper lymphatic function during chronic inflammatory settings such as atherosclerosis.

L’utilisation de stratégies transfusionnelles restrictives (pas de transfusion de globules rouges si hémoglobine > 70 g/L) a été démontrée comme étant sécuritaire en aiguë et celles-ci sont maintenant largement utilisées aux soins intensifs pédiatriques (SIP). En conséquence, plus de patients quittent les SIP en étant anémiques. L’anémie chronique, plus spécifiquement l’anémie ferriprive, est associée à des issues neurodéveloppementales défavorables. Peu d’études sont disponibles pour caractériser la prévalence de l’anémie au congé des SIP. Les études présentées dans ce mémoire s’intéressent à cette question en examinant d’abord le problème de l’anémie aux SIP et son traitement (revue narrative), puis sa prévalence au congé chez tous les patients ayant survécu aux SIP sur une période de 5 ans et pour finir, la prévalence de ce problème chez les nourrissons admis pour une chirurgie cardiaque (via 2 études de cohorte rétrospectives). Nous avons démontré que 50,9% des patients survivant à un séjour aux SIP sont anémiques au congé. Cette proportion varie en fonction du type d’admission (44,8% médicale, 70,3% chirurgicale non-cardiaque et 53,3% chirurgicale cardiaque). Le fait d’être anémique à l’admission aux SIP est le facteur de risque le plus souvent associé à l’anémie au congé des SIP (rapport des cotes, RC : 14,41). Chez les nourrissons ayant subi une chirurgie cardiaque, la prévalence d’anémie au congé est de 27,7%. La proportion de patients quittant les SIP anémiques est alarmante : il est primordial de déterminer la trajectoire et l’impact de ce problème chez les patients survivant à une maladie critique. / It is now clear that restrictive transfusion strategies (no red blood cell transfusion if the hemoglobin level is > 70 g/L) are safe in acute settings. Therefore, these strategies are widely being used in Pediatric Intensive Care Unit (PICU) and as a consequence, numerous children leave the PICU anemic. Prolonged anemia, especially iron-deficiency anemia, is associated with worst neurocognitive outcomes. Data regarding the true prevalence of anemia at PICU discharge is lacking. This thesis addresses this question. Etiology and management of anemia in the PICU is explored through a narrative review. Prevalence of anemia at PICU discharge is examined through two retrospective cohort studies: the first describes the prevalence of this complication in PICU survivors over a five-year period and the second one, the prevalence of anemia in infants admitted to the PICU after cardiac surgery. Through our studies, we established that the prevalence of anemia at PICU discharge was 50.9% in PICU survivors. This proportion varied depending on the type of admission (44.8%, 70.3%, and 53.3% for medical, non-cardiac surgery, and cardiac surgery respectively). Anemia at PICU admission was the most important risk factor associated with anemia at PICU discharge (odds ratio, OR 14.41). In newborns less than 6 weeks old who underwent cardiac surgery, the prevalence of anemia at PICU discharge was 27.7%. Given the alarming proportion of PICU survivors that are anemic at discharge, it is paramount that we determined the trajectory and impact of this problem in critically ill survivors.

Les cellules sanguines jouent un rôle fondamental dans le bon fonctionnement du corps et sont régulées de manière à répondre à ses besoins immédiats. Malgré leurs fonctions distinctes, plusieurs études suggèrent que les processus de régulations des globules rouges et des plaquettes sanguines interagissent entre eux, notamment par l'intervention d'hormones. On s'intéresse ici aux interactions conceptuelles entre ces deux familles de cellules sanguines à l'aide d'un modèle de deux équations différentielles à délais couplées. L'analyse de la distribution des valeurs propres de l'équation caractéristique du modèle linéarisé permet de dresser un portrait de stabilité de l'équilibre du système dans un plan de paramètre approprié. On s'intéresse notamment aux possibilités de déstabilisation et de restabilisation par le couplage des deux équations. L'analyse des diagrammes de stabilité pour ces différents cas de figure permet de mettre en évidence des dynamiques intéressantes comme des alternances de stabilité par l'action des délais et différents types de bifurcations déstabilisatrices de l'équilibre. / Blood cells play a fundamental role in the proper functioning of the body and are regulated to respond to its immediate needs. Despite their distinct functions, several studies suggest that the regulatory processes of red blood cells and blood platelets interact with each other, notably through the intervention of hormones. We aim to study the conceptual interactions between these two families of blood cells using a coupled two-delay differential equation model. The analysis of the distribution of eigenvalues of the characteristic equation of the linearized model allows us to outline a stability portrait of the system in a suitable parameter plane. We are particularly interested in exploring the possibilities of destabilization and restabilization through the coupling of the two equations. The analysis of stability diagrams for these different scenarios highlights interesting dynamics such as stability switches due to the influence of delays and various types of destabilizing bifurcations of the equilibrium.

Einleitung: Eisenmangel ist ein häufiges Problem bei Ausdauersportlern, da dieser die Leistungsfähigkeit einschränken und zu einer Blutanämie führen kann. Von den klassischen Eisenstoffwechselparametern sind nur wenige geeignet, den tatsächlichen Eisenstatus anzuzeigen, da sie durch sportliche Aktivität per se beeinflusst werden können. In der Studie wird die Wertigkeit moderner Eisenstoffwechselparameter im Ausdauersport untersucht. Ergebnisse und Diskussion: Der Hämoglobingehalt der Retikulozyten (CHr) und der prozentuale Anteil der hypochromen Erythrozyten (% HYPO) bleiben als moderne Parameter der Eisenstoffwechseldiagnostik auch nach regenerativen, extensiven sowie intensiven Ausdauerbelastungen konstant. Sie zeigen somit unbeeinflusst von der (Ausdauer-)Sportart als auch von der Belastungsdauer und –intensität die aktuelle Eisenversorgung der Erythropoese im Knochenmark an. Als Monitoringparameter im Rahmen einer Eisensubstitution sind der CHr sowie der prozentuale Anteil hypochromer Retikulozyten sensible Parameter, mit denen sich schnell und zuverlässig die Effektivität einer oralen Eisensubstitution nachweisen lässt. Der Eisenstatus kann durch die alleinige Bestimmung des CHr bzw. des % HYPO nicht exakt ermittelt werden. Im Gegensatz zum Einsatz bei klinischen Fragestellungen können sie zu diesem Zeitpunkt in der Sportmedizin allenfalls nur ein ergänzender Parameter in der Differentialdiagnostik eines Eisenmangels sein. Die für Ausdauersportler ermittelten Referenzwerte liegen bei 28,8-35,9 pg für den CHr und bei 0-0,5 % für die hypochromen Erythrozyten. Ein belastungsbedingter Anstieg des Eisen-regulierenden Hormons Hepcidin könnte auf Grund seiner inhibitorischen Wirkung auf die intestinale Eisenabsorption sowie auf die Freisetzung von Eisen aus den Makrophagen zur Entwicklung von Eisenmangel-zuständen führen. Die Entwicklung von Hepcidin-Antagonisten könnte in Zukunft therapeutische Anwendungen finden. / Introduction: Athletes are commonly diagnosed with iron deficiency, particularly those involved in endurance sports. It often not only decreases athletic performance but also contributes to the development of anaemia. Many biochemical markers are used to evaluate body iron stores but some of them are affected by physical exercises. This study was designed to investigate new haematological parameters in endurance athletes. Results and discussion: Reticulocyte hemoglobin content (CHr) and percentage of hypochromic red cells (% HYPO) were highly stable in athletes subjected to varying physical loads. Furthermore, they were neither affected by different forms of endurance sports nor by their duration and intensity. These modern indices are able to reflect the availability of iron during erythropoiesis at all times. In the early prediction of response to oral iron supplementation, the reticulocyte indices CHr and % HYPOr (percentage of hypochromic reticulocytes) are the most sensitive parameters. However, it is not possible to assess the iron status in athletes by CHr and % HYPO alone. In contrast to other clinical cases they can only be an additional parameter in the differential diagnosis of iron deficiency. The calculated reference range for endurance athletes are 28,8 - 35,9 pg for CHr and 0-0,5 % for % HYPO. As the iron-regulating hormone hepcidin is inhibiting the absorption of iron from the diet at the site of the duodenal enterocytes and blocking the release of iron from macrophages that have collected senescent erythrocytes, an exercise induced up-regulation of hepcidin activity might potentially be a new mechanism causing iron deficiency in athletes. Therefore, the development of hepcidin antagonists could prove to be helpful with regard to therapeutic utilization.