Regenerative potential of corneal endothelium from patients with fuchs endothelial corneal dystrophy

La dystrophie cornéenne endothéliale de Fuchs (FECD, pour l’abréviation du terme anglais « Fuchs endothelial corneal dystrophy ») est une maladie de l'endothélium cornéen. Sa pathogenèse est mal connue. Aucun traitement médical n’est efficace. Le seul traitement existant est chirurgical et consiste dans le remplacement de l’endothélium pathologique par un endothélium sain provenant de cornées de la Banque des yeux. Le traitement chirurgical, en revanche, comporte 10% de rejet immunologique. Des modèles expérimentaux sont donc nécessaires afin de mieux comprendre cette maladie ainsi que pour le développement de traitements alternatifs. Le but général de cette thèse est de développer un modèle expérimental de la FECD en utilisant le génie tissulaire. Ceci a été réalisé en trois étapes. 1) Tout d'abord, l'endothélium cornéen a été reconstruit par génie tissulaire en utilisant des cellules endothéliales en culture, provenant de patients atteints de FECD. Ce modèle a ensuite été caractérisé in vitro. Brièvement, les cellules endothéliales cornéennes FECD ont été isolées à partir de membranes de Descemet prélevées lors de greffes de cornée. Les cellules au deuxième ou troisième passages ont ensuite été ensemencées sur une cornée humaine préalablement décellularisée. Suivant 2 semaines de culture, les endothélia cornéens reconstruits FECD (n = 6) ont été évalués à l'aide d'histologie, de microscopie électronique à transmission et d’immunomarquages de différentes protéines. Les endothélia cornéens reconstruits FECD ont formé une monocouche de cellules polygonales bien adhérées à la membrane de Descemet. Les immunomarquages ont démontré la présence des protéines importantes pour la fonctionnalité de l’endothélium cornéen telles que Na+-K+/ATPase α1 et Na+/HCO3-, ainsi qu’une expression faible et uniforme de la protéine clusterine. 2) Deux techniques chirurgicales (DSAEK ; pour « Descemet stripping automated endothelial keratoplasty » et la kératoplastie pénétrante) ont été comparées pour la transplantation cornéenne dans le modèle animal félin. Les paramètres comparés incluaient les défis chirurgicaux et les résultats cliniques. La technique « DSAEK » a été difficile à effectuer dans le modèle félin. Une formation rapide de fibrine a été observée dans tous les cas DSAEK (n = 5). 3) Finalement, la fonctionnalité in vivo des endothélia cornéens reconstruits FECD a été évaluée (n = 7). Les évaluations in vivo comprenaient la transparence, la pachymétrie et la tomographie par cohérence optique. Les évaluations post-mortem incluaient la morphométrie des cellules endothéliales, la microscopie électronique à transmission et des immunomarquage de protéines liées à la fonctionnalité. Après la transplantation, la pachymétrie a progressivement diminué et la transparence a progressivement augmenté. Sept jours après la transplantation, 6 des 7 greffes étaient claires. La microscopie électronique à transmission a montré la présence de matériel fibrillaire sous-endothélial dans toutes les greffes d’endothelia reconstruits FECD. Les endothélia reconstruits exprimaient aussi des protéines Na+-K+/ATPase et Na+/HCO3-. En résumé, cette thèse démontre que les cellules endothéliales de la cornée à un stade avancé FECD peuvent être utilisées pour reconstruire un endothélium cornéen par génie tissulaire. La kératoplastie pénétrante a été démontrée comme étant la procédure la plus appropriée pour transplanter ces tissus reconstruits dans l’œil du modèle animal félin. La restauration de l'épaisseur cornéenne et de la transparence démontrent que les greffons reconstruits FECD sont fonctionnels in vivo. Ces nouveaux modèles FECD démontrent une réhabilitation des cellules FECD, permettant d’utiliser le génie tissulaire pour reconstruire des endothelia fonctionnels à partir de cellules dystrophiques. Les applications potentielles sont nombreuses, y compris des études physiopathologiques et pharmacologiques.Fuchs endothelial corneal dystrophy (FECD) is a primary disease of the corneal endothelium. Its pathogenesis is poorly understood. No medical treatment is effective. Surgical treatment (the only available treatment) carries 10% of immunogenic rejection. Experimental models are needed in order to better understand the disease and to investigate potential autologous treatments (to prevent immunogenic rejection). The overall goal of this thesis is to develop an experimental model for FECD using tissue engineering. This was achieved in three steps. 1) An in vitro tissue-engineered FECD model was created and characterized. Briefly, Descemet’s membranes from patients with late-stage FECD undergoing Descemet’s Stripping Automated Endothelial Keratoplasty (DSAEK) were used to isolate and culture FECD endothelial cells. Second or third-passaged FECD endothelial cells were seeded on a previously decellularized human cornea. After 2 weeks in culture, TE-FECD corneas (n=6) were assessed using histology, transmission electron microscopy (TEM) and immunofluorescence labeling of various proteins. TE-FECD endothelium yielded a monolayer of polygonal cells well adhered to Descemet’s membrane. The TE-FECD corneal endothelium expressed the function-related proteins Na+-K+/ATPase α1 and Na+/HCO3-. Clusterin expression was faint and uniform. 2) In order to determine the best surgical procedure to transplant the TE-FECD corneas in the feline model, a DSAEK procedure was evaluated and compared to penetrating keratoplasty technique. DSAEK assessments included surgical challenges and clinical outcomes. DSAEK technique was challenging to perform in the feline model. Rapid fibrin formation was observed in all DSAEK cases (n=5). 3) The in vivo functionality of the TE-FECD corneas was assessed. TE-FECD corneas were grafted in the feline model (n=7) using penetrating keratoplasty procedure and observed for seven days. In vivo assessments included transparency, pachymetry, optical coherence tomography, endothelial cell morphometry, TEM and immunostaining of function-related proteins. After transplantation, pachymetry gradually decreased and transparency gradually increased. Seven days after transplantation, 6 out of 7 grafts were clear. Post-mortem TEM showed subendothelial loose fibrillar material deposition in all TE-FECD grafts. The TE grafted endothelium expressed Na+-K+/ATPase and Na+/HCO3-. This thesis demonstrates that endothelial cells from late-stage FECD corneas can be used to engineer a corneal endothelium. Compared to DSEAK, penetrating keratoplasty is a more appropriate procedure for corneal transplantation in the feline model, since the DSAEK procedure in the feline model presently yields inconsistent clinical results. Restoration of corneal thickness and transparency demonstrates that the TE-FECD grafts are functional in vivo. This novel FECD living model suggests a potential role of tissue engineering for FECD cell rehabilitation. Potential applications are numerous, including pathophysiological and pharmacological studies

Semileptonic form factors D → \rightarrow π \pi , K and B → \rightarrow π \pi , K from a fine lattice

We extract the form factors relevant for semileptonic decays of D and B mesons from a relativistic computation on a fine lattice in the quenched approximation. The lattice spacing is a = 0.04 fm (corresponding to a -1 = 4.97 GeV), which allows us to run very close to the physical B meson mass, and to reduce the systematic errors associated with the extrapolation in terms of a heavy-quark expansion. For decays of D and Ds mesons, our results for the physical form factors at \ensuremath q^2 = 0 are as follows: \ensuremath f_+^{D\rightarrow\pi}(0) = 0.74(6)(4) , \ensuremath f_+^{D \rightarrow K}(0) = 0.78(5)(4) and \ensuremath f_+^{D_s \rightarrow K} (0) = 0.68(4)(3) . Similarly, for B and Bs we find \ensuremath f_+^{B\rightarrow\pi}(0) = 0.27(7)(5) , \ensuremath f_+^{B\rightarrow K} (0) = 0.32(6)(6) and \ensuremath f_+^{B_s\rightarrow K}(0) = 0.23(5)(4) . We compare our results with other quenched and unquenched lattice calculations, as well as with light-cone sum rule predictions, finding good agreemen

Theoretical and Phenomenological Constraints on Form Factors for Radiative and Semi-Leptonic B-Meson Decays

We study transition form factors for radiative and rare semi-leptonic B-meson decays into light pseudoscalar or vector mesons, combining theoretical constraints and phenomenological information from Lattice QCD, light-cone sum rules, and dispersive bounds. We pay particular attention to form factor parameterisations which are based on the so-called series expansion, and study the related systematic uncertainties on a quantitative level. In this context, we also provide the NLO corrections to the correlation function between two flavour-changing tensor currents, which enters the unitarity constraints for the coefficients in the series expansion.Comment: 52 pages; v2: normalization error in (29ff.) corrected, conclusion about relevance of unitarity bounds modified; form factor fits unaffected; references added; v3: discussion on truncation of series expansion added, matches version to be published in JHEP; v4: corrected typos in Tables 5 and

Bayesian Fit of Exclusive b→sℓˉℓb \to s \bar\ell\ell Decays: The Standard Model Operator Basis

We perform a model-independent fit of the short-distance couplings $C_{7,9,10}$ within the Standard Model set of $b\to s\gamma$ and $b\to s\bar\ell\ell$ operators. Our analysis of $B \to K^* \gamma$, $B \to K^{(*)} \bar\ell\ell$ and $B_s \to \bar\mu\mu$ decays is the first to harness the full power of the Bayesian approach: all major sources of theory uncertainty explicitly enter as nuisance parameters. Exploiting the latest measurements, the fit reveals a flipped-sign solution in addition to a Standard-Model-like solution for the couplings $C_i$. Each solution contains about half of the posterior probability, and both have nearly equal goodness of fit. The Standard Model prediction is close to the best-fit point. No New Physics contributions are necessary to describe the current data. Benefitting from the improved posterior knowledge of the nuisance parameters, we predict ranges for currently unmeasured, optimized observables in the angular distributions of $B\to K^*(\to K\pi)\,\bar\ell\ell$.Comment: 42 pages, 8 figures; v2: Using new lattice input for f_Bs, considering Bs-mixing effects in BR[B_s->ll]. Main results and conclusion unchanged, matches journal versio

Two-loop Corrections to the B to pi Form Factor from QCD Sum Rules on the Light-Cone and |V(ub)|

We calculate the leading-twist O(alphas^2 beta0) corrections to the B to pi transition form factor f+(0) in light-cone sum rules. We find that, as expected, there is a cancellation between the O(alphas^2 beta0) corrections to fB f+(0) and the large corresponding corrections to fB, calculated in QCD sum rules. This suggests the insensitivity of the form factors calculated in the light-cone sum rules approach to this source of radiative corrections. We further obtain an improved determination of the CKM matrix element |V(ub)|, using latest results from BaBar and Belle for f+(0)|V(ub)|.Comment: 18 pages, 3 figure

Implications of unitarity and analyticity for the D\pi form factors

We consider the vector and scalar form factors of the charm-changing current responsible for the semileptonic decay D\rightarrow \pi l \nu. Using as input dispersion relations and unitarity for the moments of suitable heavy-light correlators evaluated with Operator Product Expansions, including O(\alpha_s^2) terms in perturbative QCD, we constrain the shape parameters of the form factors and find exclusion regions for zeros on the real axis and in the complex plane. For the scalar form factor, a low energy theorem and phase information on the unitarity cut are also implemented to further constrain the shape parameters. We finally propose new analytic expressions for the $D\pi$ form factors, derive constraints on the relevant coefficients from unitarity and analyticity, and briefly discuss the usefulness of the new parametrizations for describing semileptonic data.Comment: 10 pages, 7 figures, uses EPJ style files: expanded version of v1 with extended discussion, additional analysis, explanation, figure and references; corresponds to EPJA versio

Matrix elements of heavy-light mesons from a fine lattice

We present results for the first two moments of the distribution amplitudes of pseudoscalar mesons. Using two flavors of non-perturbatively improved clover fermions and non-perturbative renormalization of the matrix elements we perform both chiral and continuum extrapolations and compare with recent results from models and experiments

Influence of Dietary Aspirin on Growth Performance, Antioxidant Status, and Mortality due to Ascites in Broiler Chickens

This study was conducted to investigate the effects of dietary influence of aspirin on growth performance, antioxidant status and mortality in the broilers subjected to cold-induced ascites. A total six hundred 1-d old male broilers (Ross, 308) were randomly allotted to four treatment groups, with five replicate pens per treatment and 30 birds each. The experimental groups were kept in a cold chamber to induce ascites and fed the basal diet supplemented with 0, 20, 40 or 80 mg of aspirin /kg diet. Results showed, aspirin had a significant effect (P< 0.05) on broiler performance so that, 80 mg group of aspirin had greater body weight gain and lower feed conversion ratio. Compared to other groups, 20 mg of aspirin significantly reduced malondialdehyde concentration and increased total antioxidant capacity with higher glutathione peroxidase activity in plasma (