Orientador: Daniela Ponzoni / Banca: Mariza Akemi Matsumoto / Banca: André Luís da Silva Fabris / Resumo: Objetivo: Avaliar por meio da análise histomorfométrica e histológica o potencial osteocondutor dos biomateriais Bio-Oss® (Geistilich, Suécia), GenOx Inorgânico® (Baumer S.A, São Paulo, Brasil) e GenOx Inorgânico 700 (Baumer S.A, São Paulo, Brasil) no reparo ósseo de defeitos críticos em calvária de ratos. Métodos: Foram utilizados 48 ratos divididos em quatro grupos de acordo com o material de preenchimento. Grupo controle (GC) com o defeito preenchido somente com coágulo sanguíneo (n=12), GBO preenchido com Bio-Oss® (n=12), G300 preenchido com GenOx Inorgânico® (n=12) e o G700 com GenOx Inorgânico 700 (n=12). Na calvária de cada animal foi utilizada uma broca trefina de 5 mm de diâmetro interno para confecção de defeito central com 6 mm de diâmetro e a após a inserção dos biomateriais correspondentes. Sobre todos os defeitos foi sobreposta a membrana bovina Gen Derm®. Posteriormente, submetidos à eutanásia aos 30 dias e 60 dias pós-cirúrgicos. Após 30 e 60 dias, tais defeitos foram encaminhados para análise histológica e análise histométrica para avaliação das seguintes variáveis: neoformação óssea, presença de biomaterial, leucócitos mononucleares e polimorfonucleares e outros tecidos encontrados (de granulação e medular). Resultados: O grupo mais representativo para neoformação óssea foi GC. Aos 30 dias apresentou maior média de tecido ósseo maduro (75,8). Aos 60 dias não houve diferença estatística entre os grupos GC (64,9), GBO (32,9), G300 (45,3) e G700 (26,6). O GBO a... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Abstract :Objective:The aim of this study was to evaluate the osteoconductive potential of Bio-Oss® (Geistilich, Sweden), GenOx Inorganic® (Baumer SA, São Paulo, Brazil) and GenOx Inorganic 700 (Baumer SA, São Paulo, Brazil) in the bone repair of critical defects in rat calvaria. Methods: 48 rats were divided into four groups, submitted to euthanasia at 30 days and 60 postoperative days: the control group (CG) with the defect filled only with blood clot (n = 12), GBO filled with Bio-Oss® (n = 12), G300 filled with GenOx Inorganic® (n = 12) and G700 with GenOx Inorganic 700 (n = 12). In the calvaria of each animal a 5mm trephine drill was used to make a central defect with 6 mm diameter and after the insertion of the corresponding biomaterials. Gen Derm® membrane was superimposed over all defects. Subsequently, these cavities were sent to histological and histometric analysis to evaluate the following variables: bone neoformation, presence of biomaterial, mononuclear and polymorphonuclear leukocytes and other tissues found (granulation and medullary). Results:The most representative group for bone neoformation was GC. At 30 days presented a higher mean of mature bone tissue (75.8). At 60 days there was no statistical difference between the GC (64.9), GBO (32.9), G300 (45.3) and G700 (26.6) groups. GBO had a higher amount of biomaterial after 30 days (115.9) and 60 days (118.5). Conclusion:All bovine biomaterials tested showed to be biocompatible and osteoconductive,Among the b... (Complete abstract click electronic access below) / Mestre

1 / Sarah Elizabeth Galvez

acase@tulane.edu

Coral reefs are threatened in Florida and worldwide. Successful resource management requires rapid identification of anthropogenic sources of stress before they affect the reef community. I tested a multi-scale approach for assessing reef condition at seven reefs within the Florida Keys National Marine Sanctuary and Biscayne National Park between 2001 and 2003. I examined multiple environmental parameters to identify potential sources of stress. I utilized the Atlantic and Gulf Rapid Reef Assessment Biotic Reef Index to assess benthic community structure and an indicator species of Foraminifera (Amphistegina gibbosa) to determine if environmental conditions were suitable for calcareous organisms that host algal endosymbionts. Small tissue samples were extracted from colonies of Montastraea annularis species complex to assay a suite of cellular biomarkers to elucidate possible mechanisms of the coral stress response. I monitored regeneration rates of the resultant lesions to determine if the coral colonies were capable of recovering from damage. Multivariate data analyses indicated that corals at all study sites were experiencing stress with different degrees of response and decline. On reefs with coarse grain sediments that are adjacent to an intact mangrove shoreline, the Cellular Diagnostic System indicated that corals were responding to a xenobiotic stress but appeared to be compensating as evidenced by consistently high lesion regeneration rates, a high percentage of healed lesions, low coral mortality and high abundances of A. gibbosa. On reefs with silt-sized sediments adjacent to developed coastlines, corals also were responding to xenobiotic stresses, but were negatively affected as evidenced by low regeneration rates, a low percentage of healed lesions, high coral mortality, and low abundances of A. gibbosa. Corals at an 18 m offshore site exhibited abnormally low biomarker levels and some died during the study, indicating that sampled colonies were incapable of upregulating necessary protective proteins. Further research will be required to determine stressor sources. This study demonstrates that a multiple-indicator approach, spanning scales from cellular to community, can provide marine resource managers with data linking decline of coral populations to specific environmental conditions and events, thereby providing potential for early detection of stressors allowing for preventive management.

Traumatic injuries to peripheral nerves often leave gaps that cannot be repaired by direct suture methods. In such instances, repair with a tubular nerve guide, allows connection of the nerve ends, provides directional guidance, and concentrates endogenous trophic factors for regenerating axons. We hypothesized that collagen nerve guides containing longitudinally oriented channels would further improve the outcome of nerve repair by increasing the surface area available for cell migration. We restored the continuity of a 10mm peripheral nerve gap (rat sciatic nerve) by suturing the nerve stumps into a type I collagen nerve guide (1.5 mm ID), which contained longitudinal channels. Two different channel designs were tested. They were compared to empty nerve guides and autografts. One channel design contained five longitudinally-oriented collagen microtubes (0.4 mm ID) and the other contained 32 longitudinally-oriented collagen filaments (90 micro m diameter). Nerve regeneration was examined at 6 weeks and 12 weeks post repair by a determination of the number and diameter of myelinated axons in the middle sections of the nerve guides. Sciatic function Indices were calculated from walking tracks and static stance images, and electrophysiological assessments were performed. Compound muscle action potentials of the gastrocnemius and intrinsic muscles of the foot were recorded from animals in each group at 12 weeks, indicating that axons regenerated through the nerve repair site, into the distal nerve stump, and successfully reinnervated peripheral targets. At 6 weeks, there was no significant difference between the mean number of myelinated axons with the mid sections of the 3 types of nerve guides (P = 0.488). At 12 weeks, the nerve guide that contained 5 microtubes within its lumen had significantly more axons than the nerve guide that contained 32 filaments in its lumen (P = 0.008). The mean myelinated axon number in the microtube group is larger than the empty nerve guide group but this difference was not statistically significance (P < 0.05). Autografts at both 6 and 12 weeks had significantly more myelinated axons in the mid section of the repair site than either of the nerve guide repairs at the respective time points (P < 0.05).

Orientador: Leonardo Perez Faverani / Coorientador Roberta Okamoto / Banca: Letícia Helena Theodoro / Banca: Ana Paula Farnezi Bassi / Banca: Claudio Maldonado Pastori / Banca: Cassio Edvard Sverzut / Resumo: Este trabalho teve como objetivo avaliar o efeito da biomodulação e bioatividade no reparo ósseo de ratas ovariectomizadas submetidas à simulação de fraturas femurais. Sessenta e quatro ratas Wistar (Rattus novergicus), fêmeas, com 6 meses de idade, em que metade dos animais foram submetidos à ovariectomia bilateral (OVX) e a outra metade à cirurgia fictícia de ovariectomia (SHAM), e um período de 3 meses foi acompanhado para a verificar se a osteoporose estava presente. As ratas foram submetidas à simulação de fratura em ambos os fêmures e a fratura foi fixada com miniplaca e parafusos do sistema 1,5 mm. A metade das amostras tiveram miniplacas com texturização de superfície tratadas por oxidação com plasma eletrolítico (PEO), que ficou em contato com o "gap" reparacional e, a outra metade com miniplacas convencionais, com superfície (CONV). Estes grupos experimentais não foram submetidos à biomodulação (CO). Para o mesmo desenho experimental dos grupos, no ato operatório, após a fixação das fraturas, os demais grupos experimentais foram submetidos à fotobiomodulação por meio da irradiação do laser de baixa intensidade (LASER), durante 5 minutos. Aos 14 e 42 dias de pós-operatório, foi administrada calceína e alizarina, respectivamente, pela via intramuscular, na dose de 20 mg/kg de peso. A eutanásia de todos os animais foi realizada aos 60 dias de pós-operatório. Os grupos experimentais (SHAM/LASER/CONV; SHAM/CO/CONV; SHAM/LASER/PEO; SHAM/CO/PEO; OVX/LASER/CONV; OVX/CO/CONV... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: The aim of this study was to evaluate the effect of biomodulation and bioactivity on the bone repair of ovariectomized rats submitted to simulation of femoral fractures. 64 female adults Wistar rats (Rattus novergicus), which half of the animals was submitted to bilateral ovariectomy (OVX) and the other half to the dummy ovariectomy surgery (SHAM), and after 3-month the osteoporosis was induced. The rats were submitted to fracture simulation in both of the femurs and the fracture was fixed with miniplate and screws of the system 1.5 mm. Half of the samples had surface-textured miniplates treated by with plasma electrolytic oxidation (PEO), which was in contact with the repair gap, and the other half with conventional miniplates with surface (CONV). These experimental groups were not submitted to biomodulation (CO). For the same experimental design of the groups, after fracture fixation, the other experimental groups were submitted to photobiomodulation through low level laser (laser), for 5 minutes. At 14 and 42 postoperative days, calcein and alizarin, respectively, were administered intramuscularly at a dose of 20 mg / kg body weight. The animals were euthanized at 60 postoperative days. The experimental groups (SHAM/LASER/CONV, SHAM/CO/CONV, SHAM/LASER/PEO, SHAM/CO/PEO, OVX/LASER/CONV, OVX/CO/CONV, OVX/LASER/PEO), after euthanasia, had the regions of interest, initially scanned in computerized microtomography to evaluate the bone volume parameters [BV / TV] and bone qualit... (Complete abstract click electronic access below) / Doutor

This thesis concerned the design and examination of a scaffold for tissue engineering applications. The template for the presented scaffold came from nature itself: the intercellular space in tissues that provides structure and support to the cells of the respective tissue, known as extracellular matrix (ECM). Fibres are a predominant characteristic feature of ECM, providing adhesion sites for cell-matrix interactions. In this dissertation a fibrous mesh was generated using the electrospinning technique to mimic the fibrous structure of the ECM. Two base polymers were explored: a biodegradable polyester, poly(D,L-lactide-co-glycolide); and a functional PEG-based star polymer, NCO-sP(EO-stat-PO). This topic was described in three major parts: the first part was materials based, concerning the chemical design and characterisation of the polymer scaffolds; the focus was then shifted to the cellular response to this fibrous scaffold; and finally the in vivo performance of the material was preliminarily assessed. The first steps towards an electrospun mesh started with adjusting the spinning parameters for the generation of homogeneous fibres. As reported in Chapter 3 a suitable setup configuration was on the one hand comprised of a spinning solution that consisted of 28.5 w/v% PLGA RG 504 and 6 w/v% NCO-sP(EO-stat-PO) in 450 µL acetone, 50 µL DMSO and 10 µL of an aqueous trifluoroacetic acid solution. On the other hand an ideal spinning behaviour was achieved at process parameters such as a flow rate of 0.5 mL/h, spinneret to collector distance of 12-16 cm and a voltage of 13 kV. The NCO-sP(EO-stat-PO) containing fibres proved to be highly hydrophilic as the functional additive was present on the fibre surface. Furthermore, the fibres featured a bulk degradation pattern as a consequence of the proportion of PLGA. Besides the morphologic similarity to ECM fibres, the functionality of the electrospun fibres is also decisive for a successful ECM mimicry. In Chapter 4, the passive as well as active functionality of the fibres was investigated. The fibres were required to be protein repellent to prevent an unspecific cell adhesion. This was proven as even 6.5 % sP(EO-stat-PO) in the PLGA fibres reduced any unspecific protein adsorption of bovine serum albumin and foetal calf serum to less than 1 %. However, avidin based proteins attached to the fibres. This adhesion process was avoided by an additional fibre surface treatment with glycidol. The active functionalisation of NCO-sP(EO-stat-PO)/PLGA fibres was investigated with two fluorescent dyes and biocytin. A threefold, chemically orthogonal, fibre modification was achieved with these dyes. The chapters about the chemical and mechanical properties laid the basis for the in vitro chapters where a specific fibre functionalisation with peptides was conducted to analyse the cell adhesion and biochemical expressions. Beginning with fibroblasts in Chapter 5 the focus was on the specific cell adhesion on the electrospun fibres. While NCO-sP(EO-stat-PO)/PLGA fibres without peptides did not allow any adhesion of fibroblasts, a fibre modification with GRGDS (an adhesion mediating peptide sequence) induced the adhesion and spreading of human dermal fibroblasts on the fibrous scaffolds. The control sequence GRGES that has no adhesion mediating qualities did not lead to any cell adhesion as observed on fibres without modifications. While the experiments of Chapter 5 were a proof-of-concept, in Chapter 6 a possible application in cartilage tissue engineering was examined. Therefore, primary human chondrocytes were seeded on fibrous scaffolds with various peptide sequences. Though the chondrocytes exhibited high viability on all scaffolds, an active interaction of cells and fibres was only found for the decorin derived sequence CGKLER. Live-cell-imaging revealed both cell attachment and migration within CGKLER-modified meshes. As chondrocytes undergo a de-differentiation towards a fibroblast-like phenotype, the chondrogenic re-differentiation on these scaffolds was investigated in a long term cell culture experiment of 28 days. Therefore, the glycosaminoglycan production was analysed as well as the mRNA expression of genes coding for collagen I and II, aggrecan and proteoglycan 4. In general only low amounts of the chondrogenic markers were measured, suggesting no chondrogenic differentiation. For conclusive evidence follow-up experiments are required that support or reject the findings. The success of an implant for tissue engineering relies not only on the response of the targeted cell type but also on the immune reaction caused by leukocytes. Hence, Chapter 7 dealt with primary human macrophages and their behaviour and phenotype on two-dimensional (2D) surfaces compared to three-dimensional (3D) fibrous substrates. It was found that the general non-adhesiveness of NCO-sP(EO-stat-PO) surfaces and fibres does not apply to macrophages. The cells aligned along the fibres on surfaces or resided in the pores of the meshes. On flat surfaces without 3D structure the macrophages showed a retarded adhesion kinetic accompanied with a high migratory activity indicating their search for a topographical feature to adhere to. Moreover, a detailed investigation of cell surface markers and chemokine signalling revealed that macrophages on 2D surfaces exhibited surface markers indicating a healing phenotype while the chemokine release suggested a pro-inflammatory phenotype. Interestingly, the opposite situation was found on 3D fibrous substrates with pro-inflammatory surface markers and pro-angiogenic cytokine release. As the immune response largely depends on cellular communication, it was concluded that the NCO-sP(EO-stat-PO)/PLGA fibres induce an adequate immune response with promising prospects to be used in a scaffold for tissue engineering. The final chapter of this thesis reports on a first in vivo study conducted with the presented electrospun fibres. Here, the fibres were combined with a polypropylene mesh for the treatment of diaphragmatic hernias in a rabbit model. Two scaffold series were described that differed in the overall surface morphology: while the fibres of Series A were incorporated into a thick gel of NCO-sP(EO-stat-PO), the scaffolds of Series B featured only a thin hydrogel layer so that the overall fibrous structure could be retained. After four months in vivo the treated defects of the diaphragm were significantly smaller and filled mainly with scar tissue. Thick granulomas occurred on scaffolds of Series A while the implants of Series B did not induce any granuloma formation. As a consequence of the generally positive outcome of this study, the constructs were enhanced with a drug release system in a follow-up project. The incorporated drug was the MMP-inhibitor Ilomastat which is intended to reduce the formation of scar tissue. In conclusion, the simple and straight forward fabrication, the threefold functionalisation possibility and general versatile applicability makes the meshes of NCO-sP(EO-stat-PO)/PLGA fibres a promising candidate to be applied in tissue engineering scaffolds in the future. / Diese Dissertation beschäftigte sich mit der Entwicklung und Untersuchung eines Gerüsts zur Geweberegeneration. Der interzelluläre Raum, der in Geweben für die Gewebestruktur verantwortlich ist, wurde als Vorbild aus der Natur für das entwickelte Gerüst verwendet. Fasern sind in dieser extrazellulären Matrix (EZM) ein charakteristischer Bestandteil, die Adhäsionssequenzen für Zell-Matrix-Interaktionen enthalten und zur strukturellen Organisation der Gewebe beitragen. In der vorliegenden Arbeit wurden Faservliese mit Hilfe des elektrostatischen Verspinnens hergestellt, um die natürlichen Fasern der EZM zu imitieren. Zwei Polymere bildeten die chemische Grundlage für diese Fasern: Ein bioabbaubarer Polyester, Poly(D,L-Laktid-co-Glykolid) (PLGA) und ein funktionales auf Polyethylenglykol basierendes, sternförmiges Polymer, NCO-sP(EO-stat-PO). Der erste Teil des in drei Hauptteile untergliederten Themas beschäftigte sich mit dem chemischen Design und der Fasercharakterisierung im Sinne der Materialeigenschaften. Der zweite Teil betrachtet die Auswirkungen der Fasern auf zellulärer Ebene, während der dritte Teil einen ersten Eindruck über die in vivo Reaktion auf die Materialien vermittelt. Die ersten Schritte in Richtung eines elektrostatisch gesponnenen Vlieses begannen mit der Erforschung geeigneter Einstellungen für eine homogene Faserproduktion. Kapitel 3 thematisiert geeignete Spinnparameter, zu denen auf der einen Seite eine spinnfähige Lösung gehörte, die aus 28.5 w/v% PLGA RG 504 und 6 w/v% NCO-sP(EO-stat-PO) in 450 µL Aceton, 50 µL DMSO und 10 µL trifluoressigsaurer wässriger Lösung besteht. Auf der anderen Seite wurden Prozessparameter gefunden, wie zum Beispiel eine Flussrate von 0.5 mL/h und ein Kollektor-Abstand von 12-16 cm, die bei einer Potentialdifferenz von 13 kV ein stabiles Spinnverhalten garantierten. Fasern mit dem Additiv NCO-sP(EO-stat-PO) zeigten eine äußerst starke Hydrophilie, da das Additiv während des Spinnprozesses an die Faseroberfläche segregierte. Des Weiteren sind die Fasern dank des PLGA-Anteils nach einem Volumenabbaumechanismus unter physiologischen Bedingungen degradierbar. Neben der morphologischen Ähnlichkeit zwischen natürlichen Fasern der EZM und elektrogesponnenen Fasern ist die Funktionalität der synthetischen Fasern entscheidend für eine erfolgreiche Imitation der EZM. Kapitel 4 betrachtet deswegen sowohl die passive als auch aktive Funktionalität der Fasern. Unter passive Funktionalität fällt das proteinabweisende Verhalten, welches eine unspezifische Zelladhäsion verhindert. Es wurde gezeigt, dass ein Anteil von 6.5 % sP(EO-stat-PO) in den PLGA-Fasern ausreicht, um die unspezifische Adhäsion von Albumin aus Rinderserum und fötalem Kälberserum auf weniger als 1 % zu senken. Dennoch adhärierten avidinbasierte Proteine auf den Fasern, was jedoch durch eine Behandlung mit Glycidol unterbunden werden konnte. Die aktive Funktionalisierung wurde exemplarisch mit zwei Fluoreszenzfarbstoffen und Biocytin untersucht. Mit diesen Modellmolekülen wurde eine dreifache, chemisch orthogonale Fasermodifizierung erreicht. Die Kapitel über die chemischen und mechanischen Eigenschaften haben die Grundlage für in vitro Zellversuche gelegt, bei denen eine Faserfunktionalisierung mit Peptidsequenzen durchgeführt wurde, um eine spezifische Zelladhäsion zu erreichen und die biochemische Reaktion der Zellen zu untersuchen. In Kapitel 5 lag der Fokus auf der spezifischen Adhäsion von humanen dermalen Fibroblasten an den elektrogesponnenen Fasern. Während NCO-sP(EO-stat-PO)/PLGA Fasern ohne Peptide keine Zelladhäsion zuließen, induzierte eine Fasermodifikation mit GRGDS, einer adhäsionsvermittelnden Peptidsequenz, sowohl die Adhäsion als auch Ausbreitung der Fibroblasten auf den Fasern. Eine Kontrollsequenz ohne adhäsionsvermittelnde Eigenschaften (GRGES), führte, wie auch Fasern ohne Peptide, zu keiner Zelladhäsion. Die Experimente von Kapitel 6 gingen über das reine Machbarkeitskonzept von Kapitel 5 hinaus, indem eine mögliche Anwendung im Bereich der Knorpelregeneration untersucht wurde. Daher wurden primäre humane Chondrozyten auf Faservliesen ausgesät, die mit unterschiedlichen Peptiden modifiziert wurden. Trotz einer allgemein sehr guten Vitalität der Zellen auf allen Fasertypen, zeigten die Chondrozyten nur auf Vliesen mit der aus Decorin abgeleiteten CGKLER-Sequenz eine aktive Interaktion. Diese konnte mit dem Live-Cell-Imaging-Verfahren anhand der Zelladhäsion und Zellmigration beobachtet werden. Da Chondrozyten in der 2D-Expansionszellkultur einer Dedifferenzierung in Richtung eines Fibroblasten ähnlichen Zelltypen unterliegen, wurde eine 28-tägige Studie durchgeführt, um das Redifferenzierungsverhalten auf den Fasergerüsten zu untersuchen. Dazu wurde sowohl die Glykosaminoglykanproduktion analysiert als auch die mRNA Expression der Gene, die die Kollagen I und II, Aggrecan und Proteoglykan 4 Produktion regulieren. Die chondrogenen Marker wurden in diesen Versuchen nur geringfügig ausgeschüttet, was in Anbetracht der großen Varianzen in den Messwerten auf keine Redifferenzierung schließen lässt. Für eine abschließende Beurteilung werden Folgeexperimente empfohlen, die die gemachten Beobachtungen bestärken oder widerlegen. Der Erfolg eines Implantats zur Geweberegeneration beruht nicht nur auf der gewünschten Reaktion des Zielzelltyps, sondern auch auf der Immunreaktion des Organismus, welche durch Leukozyten gesteuert wird. Folglich beschäftigte sich Kapitel 7 mit dem Verhalten und den Phänotypen primärer humaner Makrophagen auf dreidimensionalen Fasergerüsten und zweidimensionalen Oberflächen im Vergleich zueinander. Bei den Versuchen zeigte sich, dass die generelle Nicht-Adhäsivität von NCO-sP(EO-stat-PO) Oberflächen für Makrophagen nicht zutrifft. Die Zellen richteten sich an den Fasern auf den Oberflächen aus oder saßen in den Poren der Vliese. Auf flachen Oberflächen ohne dreidimensionale Struktur wiesen die Makrophagen ein verzögertes Adhäsionsverhalten auf und migrierten stark über die Oberfläche auf der Suche nach topographischen Unebenheiten, um dort adhärent zu werden. Des Weiteren zeigte eine detaillierte Untersuchung der Oberflächenmarker und der Zytokinausschüttung, dass Makrophagen auf 2D-Oberflächen gemäß der Oberflächenmarker einen entzündungs-hemmenden Phänotypen aufwiesen, während die Zytokinausschüttung einen entzündungs-fördernden Phänotypen suggerierte. Interessanterweise bot sich das entgegengesetzte Bild auf 3D-Faseroberflächen. Hier wurde die Erkenntnis gewonnen, dass die Morphologie einen größeren Einfluss auf die Zellreaktion hat als die Oberflächenchemie. Da die Immunantwort eines Organismus auf ein Implantat stark von der interzellularen Kommunikation abhängt, wurde gefolgert, dass die NCO-sP(EO-stat-PO)/PLGA Fasern eine adäquate Immunantwort hervorrufen mit vielversprechenden Aussichten, die Fasergerüste im Bereich der Geweberegeneration einzusetzen. Das letzte Kapitel der Dissertation berichtet über eine erste in vivo Studie der hier vorgestellten Fasern. Mit den Fasern wurde ein bestehendes Behandlungskonzept für Hernien des Zwerchfells erweitert und die Leistungsfähigkeit in einem Kaninchenmodell überprüft. Zwei Gerüsttypen wurden untersucht, die sich in der Oberflächenmorphologie maßgeblich unterschieden: In Serie A wurden die elektrogesponnenen Fasern in ein Gel aus NCO-sP(EO-stat-PO) gebettet, während die Fasern in Serie B nur mit einer dünnen Gelschicht bedeckt wurden, so dass die topografische Faserstruktur erhalten blieb. Nach 4 Monaten in vivo waren die behandelten Zwerchfelldefekte signifikant kleiner und überwiegend mit Narbengewebe gefüllt. Die ausgeprägte Granulombildung bei Fasergerüsten der Serie A konnte in der darauffolgenden Studie (Serie B) minimiert werden. Das gute Abschneiden dieser Studien wurde zum Anlass genommen, die Vliese weiterzuentwickeln und eine Medikamentenfreisetzung (Ilomastat) zu integrieren, um die Narbenbildung zu minimieren. Zusammenfassend beschreibt diese Dissertation einen einfachen und direkten Weg, Fasern für eine gezielte Geweberegeneration zu erzeugen, die dreifach funktionalisierbar und vielseitig anwendbar sind. Dies macht Fasergerüste auf der Basis von NCO-sP(EO-stat-PO)/PLGA zu einem vielversprechenden Kandidaten um in der Geweberegeneration eingesetzt zu werden.

Le pancréas est impliqué dans la digestion alimentaire et le contrôle glycémique. Cette diversité fonctionnelle repose sur des facteurs transcriptionnels et épigénétiques qui assurent une régulation coordonnée de l’identité et de la fonction des cellules. Ma thèse vise à caractériser le rôle de certains de ces facteurs. Nous avons précédemment montré le rôle de Hnf1b dans le développement du pancréas. Nous montrons que son inactivation post-natale chez la souris induit de graves défauts canalaires ainsi qu’une pancréatite associée à des néoplasies intra-épithéliales. La pathologie MODY5 est liée à des mutations de HNF1B. Nous avons analysé un modèle murin du MODY5 généré par insertion d’une mutation humaine chez la souris. Notre étude montre que ces souris constituent un bon modèle de la maladie. Elles sont intolérantes au glucose et présentent une réduction du volume total des gros ilots et des cellules bêta. Sur le plan exocrine, elles développent une pancréatite. Nos analyses suggèrent une étiologie embryonnaire. Antérieurement, notre équipe a montré le rôle des HDACs dans le développement du pancréas. Basés sur l’analogie des voies qui régulent développement et régénération du pancréas, nous avons étudié le rôle des HDACs dans la régénération pancréatique. Nous montrons que l’inactivation de HDAC3 sensibilise le tissu acinaire à l’inflammation et abroge sa régénération. Nos études dévoilent l’implication de nouveaux mécanismes moléculaires dans la survenue de la pancréatite, du cancer du pancréas et du diabète. Ces données pourront contribuer à l’identification de nouvelles cibles thérapeutiques pour le traitement ou la prévention des pathologies du pancréas. / The pancreas is crucial for food digestion and glycemic control. Its functional diversity is based on transcriptional and epigenetic factors which ensure the regulation of cellular identity and function. My thesis aims to characterize the role of some of these factors. We previously showed that Hnf1b is required for pancreas development. We show here that Hnf1b post-natal inactivation in mouse leads to strong ductal defects and pancreatitis associated with intraepithelial neoplasia. In human, HNF1B heterozygous mutations cause the MODY5 pathology. We analyzed a unique MODY5 mouse model generated by insertion of human mutation in mouse. We show that these mice constitute a valuable model of the disease. They are glucose intolerant and display reduced total volume of large islets and beta-cells. The exocrine compartment is also altered as mice develop a pancreatitis phenotype. Our analyses suggest an embryonic etiology. Previously, our team revealed HDAC roles in pancreas development. Pancreas regeneration and pancreas development sharing common molecular pathways, we investigated the role of HDACs in pancreas regeneration. We found that HDAC3 inactivation sensitizes acinar tissue to inflammation and prevents its regeneration. Our studies reveal new molecular mechanisms implied in the development of pancreatitis, pancreatic cancer and diabetes. These data could contribute to the identification of new therapeutic targets for the treatment of pancreatic pathologies.

No description available.

Exclosure plots were monitored to investigate the impact of browsing on seedling recruitment by Trichosurus vulpecula, Oryctolagus cuniculus and Rattus rattus on seedlings under Leptospermum scoparium and Kunzea ericoides plantings in two Auckland Regional Council Parks (ARC), Shakespear and Wenderholm. The number of woody seedlings that established over a 17-month period was recorded. Gaps within the same Leptospermum scoparium and Kunzea ericoides canopy were created to investigate the influence of the canopy on seedling recruitment. Soil samples were taken to investigate the existing seed bank beneath the same Leptospermum scoparium and Kunzea ericoides canopies.At Wenderholm, net change in seedling density differed among treatments (P=0.014). Seedling density increased within the plots that excluded Trichosurus vulpecula and Oryctolagus cuniculus and within plots that additionally excluded Rattus rattus, but declined in the control plots. In contrast at Shakespear, although seedling density increased more within both the exclosure plots than in the control plots, this result was not statistically significant (P=0.728). At Wenderholm, the average seedling height increased within both types of exclosure plots, but declined in the control plots. However, these differences among treatments were not statistically significant (P=0.204). At Shakespear, seedlings increased in height within the Trichosurus vulpecula, Oryctolagus cuniculus and Rattus rattus exclosures and declined marginally in the other two treatments. Again, differences in height change among treatments were not statistically significant (P=0.202).At both regional parks, the greatest cause of mortality within the exclosures excluding Trichosurus vulpecula and Oryctolagus cuniculus was desiccation. All of the mortalities within the Trichosurus vulpecula, Oryctolagus cuniculus and Rattus rattus exclosures was unidentifiable. However, within the control plots, at Wenderholm, the greatest identified cause of mortality was browsing and at Shakespear, the only cause of mortality within the control plots was browsing.Seedbanks at Wenderholm and Shakespear under the Leptospermum scoparium and Kunzea ericoides plantings were dominated by forb species. A total of 1308 seedlings germinated from soil taken from Wenderholm, with exotic species making up 99.4% of germinations, with exotic species making up 97.9% of germinations. Similarly a total of 801 seedlings germinated from soil samples taken from Shakespear.At Wenderholm, the number of native seedling germinations within the gaps created in the Leptospermum scoparium and Kunzea ericoides canopy, was more than twice the number that germinated under the closed canopy. However, this difference was marginally non-significant (P=0.065). At Shakespear, the number of native seedling germinations within gaps created in the Leptospermum scoparium and Kunzea ericoides canopy was similar to the number that germinated under the closed canopy (P=0.2603).The results suggest that at Wenderholm, despite ongoing predator control, Trichosurus vulpecula and/or Oryctolagus cuniculus have had an adverse effect on the survival and growth of seedlings. The results also suggest that at Shakespear, Rattus rattus have had an adverse effect on the survival and growth of seedlings under the Leptospermum scoparium and Kunzea ericoides canopy. The distance from mature forest may also have had an impact on the dispersal of native seeds within the Leptospermum scoparium and Kunzea ericoides canopy. The implication of these results for the future management of restoration plantings in regional parks is discussed.