L'hypertension est l'une des maladies humaines les plus courantes. Aujourd'hui, de nombreuses études confirment l'hypothèse selon laquelle l'hypertension implique nécessairement un transport anormal de sel (NaCl) par les reins.Le néphron, l'unité fonctionnelle du rein, est composé de plusieurs segments présentant différents systèmes de transport de NaCl. Notre équipe a récemment démontré que les cellules intercalaires du canal collecteur (CD) étaient capables de réabsorber du NaCl. Ce transport est possible grâce au couplage fonctionnel de deux échangeurs : la pendrine et NDCBE. L'objectif de cette thèse est de caractériser ce transport et de montrer que les cellules intercalaires ( IC) jouent un rôle important dans le maintien de la balance sodée et de la pression artérielle. Grâce à plusieurs modèles de souris transgéniques nous avons pu montrer que la délétion de NDCBE provoque une hypotension et la mise en place d'une compensation par un autre transporteur du néphron : le cotransporteur NCC. A partir d'un autre modèle murin knock-out pour la pompe à proton H+-ATPase spécifiquement dans les cellules intercalaires et nécessaire à la réabsorption de NaCl par les ICs, nous avons montré que les ICs étaient capables de moduler le transport de sodium des cellules principales du CD par la sécrétion de facteurs paracrines. Enfin, chez des souris possédant une mutation de kinase WNK4, connue pour provoquer le syndrôme de Gordon, une maladie caractérisée entre autre par une hypertension artérielle et une hyperkaliémie, le transport de NaCl par les ICs est suractivée et cette suractivation contribue probablement à la mise en place de la maladie. / Hypertension is one of the most common human diseases. Today, many studies support the hypothesis that hypertension necessarily involve abnormal transport of salt (NaCl) by the kidneys. The nephron, the functional unit of the kidney, is composed of several segments with different NaCl transport systems. Our team has recently demonstrated that β-intercalated cells of the collecting duct (CD) were able to reabsorb NaCl. This transport is possible through functional coupling of two exchangers : pendrin and NDCBE. The objective of this thesis is to characterize the transport and show that β-intercalated cells (β-IC) play an important role in maintaining of sodium balance and blood pressure. Through several transgenic mouse models, we have shown that deletion of NDCBE causes hypotension and induces a compensation by another transporter of NaCl : the NCC cotransporter. With another mouse model, knockout for the proton pump H+- ATPase specifically in the intercalated cells and necessary for the reabsorption of NaCI in the β-ICs, we have shown that the β-ICs were capable of modulating the sodium transport of the principal cell of the CD through the secretion of paracrine factors.Otherwise, in mice with a mutated form of the WNK4 kinase, known to cause the Gordon syndrome, a disease characterized by hypertension and hyperkalemia, the NaCl transport by β-ICs is overactive and this overactivation probably contributes to the establishment of the disease.

No description available.

Sudden cardiac death is largely attributable to sudden onset ventricular arrhythmias. Alterations in cardiac conduction, particularly the slowing of conduction velocity is one major factor in arrhythmogenesis. By understanding the mechanisms and factors that modulate cardiac conduction velocity, we can assess and perhaps mitigate the risk of arrhythmia in patients for whom slowed conduction is a arrhythmogenic substrate. Cardiac conduction has traditionally been described by cable theory, which predicts an inverse relationship between extracellular resistance and conduction velocity (CV). However, in studies that reduce extracellular resistance by inducing interstitial edema, there are conflicting results, with some labs showing increased CV when edema is induced with one agent, and others showing reduced CV when edema is induced with a different agent. In the first part of this dissertation, we present experimental data in support of ephaptic coupling (EpC), a theorized mechanism of conduction that resolves these apparent contradictions. In the later part of this dissertation, we address how changes in sodium concentration can alter conduction, despite conventional wisdom suggesting that it should not. We show that when sodium channels are impaired, such as by genetic mutation or pharmacologic blockade, that conduction is sensitized to changes in sodium concentrations that would not otherwise induce changes in CV. We go on to explore the mechanisms that modulate this sensitivity and present data that show it is a function of both EpC and outward potassium currents. Taken together, these data expand our understanding of the mechanics behind cardiac conduction and demonstrate that EpC has a clinically relevant impact on conduction and represents a new pathway to explore in regard to the treatment and management of arrhythmogenic and conduction disorders. / Doctor of Philosophy / In all large animals, life is sustained by the regular coordinated beating of the heart to pump blood throughout the body. Throughout this continuous activity, and even with minute-to-minute changes in heart rate, this electrically driven activity continues without major disruption. Until it doesn't. Major arrhythmias can occur suddenly, and without warning. Over the last century, we have begun to understand some of the reasons why heart, even an injured one, will work normally for hundreds of thousands of beats, and on the next fall into a life-threatening new pattern, and one of the most important of these reasons is the speed of conduction: the spread of electrical activation throughout the heart tissue. Understanding the mechanisms of conduction provides a way to assess and mitigate the risk of arrhythmias and may open up new avenues for treatment and prevention. This dissertation presents evidence for a previously theoretical mechanism of conduction called ephaptic coupling. We show that this electric field mediated form of conduction can be modulated with clinically used osmotic agents, and that it has a physiologically relevant impact on conduction. We also show that hyponatremia (i.e. low sodium), a condition that is traditionally thought to have minimal impact on cardiac conduction, because a significant modulator of conduction when sodium channel functions are impaired. As a great many drugs block sodium channels, this sensitization to hyponatremia and the factors that mediate it are underappreciated concerns that are relevant to a wide array of patients. The new findings presented in this dissertation advance our collective understanding of the mechanisms of cardiac conduction and provide evidence for new avenues of exploration in the prevention and management of arrhythmias and conduction disorders.

No / Background Anecdotally, it has long been felt by academic staff that students on the Bradford 5-year sandwich degree programme (intercalated pre-registration training) performed differently on return to university from those on the continuous 4-year programme. Direct comparisons between cohorts have been difficult to undertake as the two groups were taught separately in their final stage. In 2016-17, a cohort of returning sandwich students was taught alongside a comparable group of continuous students in a final stage module. This study compares the results from these two student cohorts. Method The Pharmacy Special Studies module offered a very broad range of opportunities across laboratory research, systematic and scoping review, product development and care-orientated topics including audit, and analysis of clinical cases, organised in 9 separate “strands.” Students from the sandwich (n=99) and continuous (n=89) courses were offered the same selection of learning experiences. Assessment was by oral presentation and discussion (slides or poster) and written report. The Level 7 marking schemes used were designed so that the highest marks were only available to reward student demonstration of the higher-level critical, analytical and interpretative skills. Results Student performance across all 9 strands of the module was comparable. When module results were split according to cohort, a strong divergence was observed. Sandwich student results displayed an approximately bell-shaped distribution with a mean mark 73.2% (SD 8.1). In contrast, the continuous student results had a lower mean 67.3% (SD 8.5, p<10-5); moreover, the distribution of these marks was distorted with a ‘cliff edge’ in the low 70s and a long tail. Conclusion This analysis shows that students who had completed 6 months pre-registration training achieved, on average, higher grades. Moreover, they demonstrated improved higher-level skills of interpretation and critical analysis compared with the continuous group. Although this is a one-year “snapshot” observation it appears to show that following 6 months preregistration training, students are better able to critically evaluate and interpret data and draw evidence-based conclusions. Such a result could provide evidence for the benefits of intercalated placements and indicate the optimal location of professional training within the academic course. Any impact on pre-registration training itself remains to be determined. / Abstract, presented at the Pharmacy Education Conference 2017, Manchester, United Kingdom.

Bilayers formed by molecules that possess long alkyl hydrophobic tails are ubiquitous in the natural world manifesting both in biological systems as well as in chemistry. The lipid bilayer is an integral feature of cell membranes of living systems with functions that are of critical importance to the life of the cell. Long chain amphiphilic surfactant molecules can be introduced within the interlamellar region of layered inorganic host lattices to form anchored alkyl chainbilayerswithinthegalleries.Theintercalatedbilayerbearsastriking resemblance to lipid bilayers. However, unlike lipid bilayers where individual molecules can undergo lateral diffusion and also flip-flop between layers the anchored bilayer is characterized by the total absence of translational mobility. The degrees of freedom of the alkyl chains of the anchored bilayer are restricted to changes in conformation. This thesis describes a detailed investigation of the anchored bilayer formed by the intercalation of the anionic surfactant dodecyl sulphate (DDS) in a layered solid, Mg-Al Layered Double Hydroxide(Mg-AlLDH) using both experimental measurements and Molecular Dynamics (MD) simulations (Chapter 2). The thesis is organized as two parts. The first (Chapters 2 -4) deals with the anchored bilayer in the condensed phase -the conditions for the formation of the bilayer arrangement of the intercalated surfactant chains and the conformation and dynamics of the alkyl chains of the surfactant in the galleries of the layered solid. The surfactant intercalated Mg-AlLDH-DDS may be delaminated in nonpolar solvents to give colloidal dispersions of individual Mg-Al LDH sheets with the DDS surfactant chains remaining tethered to the inorganic sheets(Scheme 1).The second part of thesis(Chapters 5 -9)describe studies on the dispersions of the Mg-AlLDH-DDS in toluene. A summary of the results of the of the investigations of the anchored bilayer, formed by the intercalation of DDS ions in Mg-Al LDH, in the condensed and the dispersed phases is presented in the concluding chapter(Chapter10). Layered Double Hydroxides(LDH) are insulating lamellar solids of the general chemical formula[M’(1-x) Mx(OH)2], where M’ is a divalent metal ion and M a trivalent ion. Their structure may be derived from that of Brucite, Mg(OH)2, by isomorphous substitution of apart of the Mg2+ by trivalent ions like Al3+ with electrical neutrality maintained by interlamellar exchangeable ions. The studies reported in this thesis are on an Mg-Al LDH,Mg(1−x)Alx(OH)2, x ranging from 0.17 to 0.37. Dodecyl sulphate surfactant ions have been ion-exchange intercalated in Mg-AlLDH (Chapter 3). By varying the Mg-Al ratio, differing packing densities of the surfactant chains in the interlamellar space of the Mg-Al LDH-DDS are realized. At high packing densities the alkyl chains of the intercalated dodecyl sulphate ions anchored on opposing Mg-Al LDH sheets are arranged as bilayers while at lower packing densities the surfactant chains form a monolayer with the chains oriented flat in the galleries. This composition driven monolayer to bilayer transformation in the surfactant intercalated Mg-AlLDH-DDS is also reproduced by MD simulations. The simulations also indicate that there are profound differences in the factors that decide the arrangement of the surfactant chains. In the bilayer arrangement it is dispersive van der Waals interactions between the chains in opposing layers that is responsible for the cohesive energy of the solid whereas at lower packing densities, where a monolayer arrangement is favored, Coulomb interactions between the positively charged Mg-Al LDH sheets and the negatively charged head-group of the DDS anion dominate. The conformation and dynamics of the alkyl chains of the intercalated surfactant chains in both the monolayer and bilayer arrangements as well as the effect of packing density on these parameters is reported in Chapter 4. The conformation was studied using spectroscopic techniques, infra-red, Raman and 13C Nuclear Magnetic Resonance (NMR) while the dynamics by Variable Contact Time Cross Polarization Magic Angle Spinning(VCT -CPMAS) and2DWidelineSeparation(2DWiSe)NMR techniques. The results showed the expected trends; the concentration of gauche defects and the dynamics of the chains increase with decreasing packing density. There is, however a sharp increase in the gauche concentration and conformational mobilities of the intercalated surfactant chains associated with the bilayer to monolayer transformation. The results of the MD simulations, too, reflect these trends. The second part of thesis describes the delamination of the intercalated anchored bilayer (Mg-AlLDH-DDS) in non-polar solvents. Delamination results in a colloidal dispersion Of the anchored bilayer, isolated Mg-AlLDH sheets with the DDS chains tethered to them, as neutral nanosheets of nanometer thickness and micron size. With increasing concentration of the anchored bilayers in the solvent a gel state is realized. The sol to gel transformation of the dispersions of the anchored bilayer in toluene has been investi-gated. Frequency dependent rheology measurements (Chapter6) were used to investigate the visco-elastic properties of the dispersions and Small Angle X-ray Scattering (SAXS) measurements(Chapter 7) to understand the structure and shape of the nanosheets. The rheology experiments showed that the dispersions irrespective of their concentrations showed shear thinning. The SAXS results indicate a tactoid structure of the dispersions as well as in the gel phase. At higher concentrations, the X-ray scattering curves indicated that the layers stack loosely with an interlamellar space of ~ 39 Å , a value much larger than the interlayer lattice spacing of solid Mg-AlLDH-DDS( ~ 27 Å). The nature of interactions between solvent molecules and the anchored DDS chains were probed by 1Hand 2H NMR measurements(Chapter8). A clear association between the toluene molecules and the alkyl chains of the anchored surfactant was observed. 2D NOESY experiments established that there are toluene molecules in close proximity that interact with the methyl tail of the anchored surfactant. NMR measurements were also able to distinguish two types of solvent molecules based on their widely differing mobilities. MD simulations(Chapter9)of the dispersed anchored bilayer are able to reproduce the essential features of the experimental observations including the formation of a loosely bound lamellar structure. It also provides an explanation on how the spectroscopic observation of motional heterogeneity gives rise to the viscoelastic properties of the dispersed anchored bilayer.

No description available.

A sépsis e o choque séptico representam uma síndrome complexa de intensa resposta inflamatória sistêmica, com múltiplas anormalidades fisiológicas e imunológicas, comumente causadas por infecção bacteriana. A principal conseqüência dessa resposta é o comprometimento de muitos órgãos e tecidos. A disfunção cardíaca, decorrente de um prejuízo na contratilidade miocárdica, tem sido reconhecida como um fator importante que contribui para os altos índices de mortalidade observados na sépsis. Dados recentes do nosso laboratório indicam que alterações estruturais no miocárdio podem ser responsáveis pela disfunção cardíaca observada na sépsis. Considerando que a maquinaria contrátil interna das miofibras deve permanecer intimamente conectada com a membrana e a matriz extracelular, o presente estudo foi proposto para avaliar alterações nas comunicações intercelulares e acoplagem mecânica entre os cardiomiócitos vizinhos e avaliar a expressão de proteínas do arcabouço celular e da matriz extracelular (especificamente a laminina-?2) durante a sépsis grave. Nossos resultados mostraram que há uma diminuição na expressão das proteínas envolvidas na formação das gap junctions (conexina43) e junções aderentes (N-caderina), o que resultaria na perda da integridade estrutural dos discos intercalares, alterando o acoplamento mecânico e eletro-químico entre os cardiomiócitos vizinhos. Além disso, demonstramos que há redução na expressão de distrofina e das proteínas que constituem o complexo de glicoproteínas associadas a distrofina (CGD) durante a sépsis experimental. A redução ou perda da expressão de distrofina é o evento primário que ocorre seguido pela degeneração miofilamentar, caracterizada pela lise dos filamentos de actina e miosina. A diminuição na expressão das glicoproteínas associadas à distrofina: -distroglicana e laminina foram considerados eventos secundários. Os resultados sugerem que durante a sépsis induzida por ligação e perfuração do ceco (CLP), há perda de proteínas importantes envolvidas tanto no remodelamento do disco intercalar quanto na expressão de glicoproteínas envolvidas na ligação mecânica entre o citoesqueleto intracelular e a matriz extracelular. Embora estudos funcionais sejam necessários para determinar o efeito direto dessas alterações sobre o miocárdio podemos sugerir que as alterações estruturais são parcialmente responsáveis pela depressão miocárdica observada na sépsis. / Sepsis and septic shock represent a complex syndrome of systemic inflammatory response, with multiple physiological and immunological abnormalities, commonly caused by bacterial infection. The most important consequence of the response is the involvement of many organs and tissues. Cardiac dysfunction, caused by impairment in myocardial contractility, has been recognized as an important factor that contributes to the high mortality observed in sepsis. Evidence from our laboratory indicates that myocardial structural changes could be responsible for sepsis-induced myocardial dysfunction. Taking into account that the contractile machinery inside the myofibers must remain intimately connected with the membrane and extracellular matrix, the present investigation sought to evaluate changes in intercellular communications and mechanical coupling between the neighbor cardiomyocytes and the expression of the cell scaffold protein and extracellular matrix (specifically merosin laminin-2 chain) during the severe sepsis. Our results showed a decrease in the expression of proteins involved in formation of gap junctions (connexin-43) and adherens junctions (N-cadherin). These alterations may result in the loss of intercalated disc structural integrity, changing the mechanical and electrical-chemical coupling between neighboring cardiomyocytes. Additionally, we demonstrated the decrease of dystrophin and dystrophin-glycoprotein complex (DGC) components resulting from severe septic injury. The reduction or loss of dystrophin is the primary event that occurs followed by miofilamentar degeneration characterized by actin and myosin lysis. The decrease of glycoproteins associated with dystrophin: -dystroglican and laminin were considered secondary events. The results suggest that during experimental severe sepsis induced by cecal ligation and puncture (CLP), there is loss of important proteins involved in both the remodeling of the intercalated disc and the glycoproteins expression implicated in the mechanical link between the intracellular cytoskeleton and extracellular matrix. Although the functional studies are needed to determine the direct effect of these alterations on myocardium, we can suggest that myocardial structural changes may be partly responsible for sepsis-induced cardiac depression.

A sépsis e o choque séptico representam uma síndrome complexa de intensa resposta inflamatória sistêmica, com múltiplas anormalidades fisiológicas e imunológicas, comumente causadas por infecção bacteriana. A principal conseqüência dessa resposta é o comprometimento de muitos órgãos e tecidos. A disfunção cardíaca, decorrente de um prejuízo na contratilidade miocárdica, tem sido reconhecida como um fator importante que contribui para os altos índices de mortalidade observados na sépsis. Dados recentes do nosso laboratório indicam que alterações estruturais no miocárdio podem ser responsáveis pela disfunção cardíaca observada na sépsis. Considerando que a maquinaria contrátil interna das miofibras deve permanecer intimamente conectada com a membrana e a matriz extracelular, o presente estudo foi proposto para avaliar alterações nas comunicações intercelulares e acoplagem mecânica entre os cardiomiócitos vizinhos e avaliar a expressão de proteínas do arcabouço celular e da matriz extracelular (especificamente a laminina-?2) durante a sépsis grave. Nossos resultados mostraram que há uma diminuição na expressão das proteínas envolvidas na formação das gap junctions (conexina43) e junções aderentes (N-caderina), o que resultaria na perda da integridade estrutural dos discos intercalares, alterando o acoplamento mecânico e eletro-químico entre os cardiomiócitos vizinhos. Além disso, demonstramos que há redução na expressão de distrofina e das proteínas que constituem o complexo de glicoproteínas associadas a distrofina (CGD) durante a sépsis experimental. A redução ou perda da expressão de distrofina é o evento primário que ocorre seguido pela degeneração miofilamentar, caracterizada pela lise dos filamentos de actina e miosina. A diminuição na expressão das glicoproteínas associadas à distrofina: -distroglicana e laminina foram considerados eventos secundários. Os resultados sugerem que durante a sépsis induzida por ligação e perfuração do ceco (CLP), há perda de proteínas importantes envolvidas tanto no remodelamento do disco intercalar quanto na expressão de glicoproteínas envolvidas na ligação mecânica entre o citoesqueleto intracelular e a matriz extracelular. Embora estudos funcionais sejam necessários para determinar o efeito direto dessas alterações sobre o miocárdio podemos sugerir que as alterações estruturais são parcialmente responsáveis pela depressão miocárdica observada na sépsis. / Sepsis and septic shock represent a complex syndrome of systemic inflammatory response, with multiple physiological and immunological abnormalities, commonly caused by bacterial infection. The most important consequence of the response is the involvement of many organs and tissues. Cardiac dysfunction, caused by impairment in myocardial contractility, has been recognized as an important factor that contributes to the high mortality observed in sepsis. Evidence from our laboratory indicates that myocardial structural changes could be responsible for sepsis-induced myocardial dysfunction. Taking into account that the contractile machinery inside the myofibers must remain intimately connected with the membrane and extracellular matrix, the present investigation sought to evaluate changes in intercellular communications and mechanical coupling between the neighbor cardiomyocytes and the expression of the cell scaffold protein and extracellular matrix (specifically merosin laminin-2 chain) during the severe sepsis. Our results showed a decrease in the expression of proteins involved in formation of gap junctions (connexin-43) and adherens junctions (N-cadherin). These alterations may result in the loss of intercalated disc structural integrity, changing the mechanical and electrical-chemical coupling between neighboring cardiomyocytes. Additionally, we demonstrated the decrease of dystrophin and dystrophin-glycoprotein complex (DGC) components resulting from severe septic injury. The reduction or loss of dystrophin is the primary event that occurs followed by miofilamentar degeneration characterized by actin and myosin lysis. The decrease of glycoproteins associated with dystrophin: -dystroglican and laminin were considered secondary events. The results suggest that during experimental severe sepsis induced by cecal ligation and puncture (CLP), there is loss of important proteins involved in both the remodeling of the intercalated disc and the glycoproteins expression implicated in the mechanical link between the intracellular cytoskeleton and extracellular matrix. Although the functional studies are needed to determine the direct effect of these alterations on myocardium, we can suggest that myocardial structural changes may be partly responsible for sepsis-induced cardiac depression.

No description available.