A complex network of transcription factors, which are regulated by signalling molecules, is responsible in coordinating the formation of differentiated skeletal and cardiac myocytes from undifferentiated stem cells. The present study aims to understand and compare the transcriptional regulation of skeletal and/or cardiac muscle development in the absence of Sox7 or in the presence of a collagen-based matrix in P19 embyonal carcinoma (EC) and mouse embryonic stem (ES) cells. First, knock-down of Sox7 , by shRNA, in muscle inducing conditions (+DMSO) and in the absence of RA (-RA), decreased muscle progenitor transcription factor and myogenic regulatory factor (MRF) levels, suggesting that Sox7 is necessary for myogenesis. However, knock-down of Sox7 in the presence of RA (+RA) and DMSO increased expression of muscle progenitor markers and MRFs, suggesting that Sox7 is inhibitory for myogenesis +RA. Furthermore, Sox7 overexpression enhanced myogenesis -RA, but inhibited myogenesis and enhanced neurogenesis +RA. These results suggest an important interplay between RA signalling and Sox7 function during P19 differentiation. Second, Q-PCR analysis showed that compared to the mouse ES cells differentiated on the regular TC plates, differentiation on the collagen matrices had a higher expression of skeletal and cardiac precursors, MRFs and terminal differentiation markers. Collagen alone enhanced myotube formation. The enhanced collagen matrix, containing the oligosaccharide sialyl LewisX (sLeX), specifically enhanced cardiomyogenesis. These studies have added to our understanding of the transcriptional regulation of premyogenic mesoderm factors and the role of Sox7 in this process. In addition these studies provide a vision for possible use of biomaterials in directed differentiation of stem cells for the purpose of cell therapy.

The ultimate goal of this research was to develop a “self-fitting” shape memory polymer (SMP) scaffold for the repair of craniomaxillofacial (CMF) bone defects. CMF defects may be caused by trauma, tumor removal or congenital abnormalities and represent a major class of bone defects. Their repair with autografts is limited by availability, donor site morbidity and complex surgical procedures. In addition, shaping and positioning of these rigid grafts into irregular defects is difficult. Herein, we have developed SMP scaffolds which soften at T > ~56 °C, allowing them to conformally fit into a bone defect. Upon cooling to body temperature, the scaffold becomes rigid and mechanically locks in place. This research was comprised of four major studies. In the first study, photocrosslinkable acrylated (AcO) SMP macromers containing a poly(ε-caprolactone) (PCL) segment and polydimethylsiloxane (PDMS) segments were synthesized with the general formula: AcO-PCL40-block-PDMSm-block-PCL40-OAc. By varying the PDMS segment length (m), solid SMPs with highly tunable mechanical properties and excellent shape memory abilities were prepared. In the second study, porous SMP scaffolds were fabricated based on AcO-PCL40-block-PDMS37-block-PCL40-OAc via a revised solvent casting particulate leaching (SCPL) method. By tailoring scaffold parameters including salt fusion, macromer concentration and salt size, scaffold properties (e.g. pore features, compressive modulus and shape memory behavior) were tuned. In the third study, porous SMP scaffolds were produced from macromers with variable PDMS segment lengths (m = 0 – 130) via an optimized SCPL method. The impact on pore features, thermal, mechanical, and shape memory properties as well as degradation rates were investigated. In the final study, a bioactive polydopamine coating was applied onto pore surfaces of the SMP scaffold prepared from PCL diacrylate. The thin coating did not affect intrinsic bulk properties of the scaffold. However, the coating significantly increased its bioactivity, giving rise to the formation of “bone-bonding” hydroxyapatite (HAp) when exposed to simulated body fluid (SBF). It was also shown that the coating largely enhanced the scaffold’s capacities to support osteoblasts adhesion, proliferation and osteogenesis. Thus, the polydopamine coating should enhance the performance of the “self-fitting” SMP scaffolds for the repair of bone defects.

Diamond-like carbon (DLC) thin films were produced by pulsed laser deposition (PLD) on silicon, fused silica, and silicon nitride substrates. The films produced were either undoped, made using a pure graphite target, or doped, using multi-component targets made from a combination of graphite and silicon, silicon nitride, titanium dioxide, or silicon monoxide. These films were evaluated for their potential use in biomedical applications, including coatings for artificial joints, heart stents, and bronchoscopes. The films were characterized by Raman spectroscopy, atomic force microscopy, ball-on-flat tribometry, contact angle measurements, and spectrophotometry. Film thickness was determined by optical profilometry. Film adhesion was checked by soaking the films in simulated body fluid (SBF) and monitoring the quality of the film surface at varying time intervals using an optical microscope. DLC coatings were produced with a root mean square surface roughness of less than 1 nm and a 0.08 lubricated coefficient of friction. Contact angles of water on the undoped films varied with deposition conditions, ranging from 65 to 88 degrees. Contact angles as low as 25 degrees were achieved by incorporating silicon monoxide dopant. DLC coatings were produced on fused silica having high transparency and showing no delamination after forty-three weeks of immersion in SBF. These results indicate that these films have potential as biomedical coatings.

Ο σκοπός της παρούσας διπλωματικής εργασίας ήταν η δημιουργία ενός νανοσύνθετου υλικού με αυξημένες μηχανικές ιδιότητες και βιοσυμβατότητα. Η χιτοζάνη είναι ένας γραμμικός πολυσακχαρίτης και είναι παράγωγο της χιτίνης. Είναι ιδιαίτερα βιοσυμβατή και βιοδιασπώμενη, μπορεί να πραγματοποιήσει αδιάλυτες ενώσεις και μπορεί εύκολα να χρησιμοποιηθεί σε ογκώδη, πορώδη ικριώματα και φιλμς, Έχει ευρεία χρήση σε βιοφαρμακευτικές εφαρμογές σε τομείς όπως η μηχανική ιστών, τα φάρμακα, η θεραπεία πληγών και τα υποκατάστατα οστών. Μελέτες που προηγήθηκαν έδειξαν πως η χιτοζάνη αυξάνει τον αριθμό των κυττάρων που έχουν προσκολληθεί σε σύγκριση με τις επιφάνειες ελέγχου. Οι νανοσωλήνες άνθρακα αποτελούνται από άτομα άνθρακα με κατάλληλους δεσμούς μεταξύ τους ώστε να σχηματίζουν έναν κυλινδρικό σωλήνα του οποίου το μήκος είναι πολλές τάξεις μεγέθους μεγαλύτερο από τη διάμετρο του, περίπου 103 με 104. Οι νανοσωλήνες άνθρακα διακρίνονται σε μονού και πολλαπλού τοιχώματος. Οι πολλαπλού τοιχώματος αποτελούνται από πολλές στρώσεις ομόκεντρων νανοσωλήνων άνθρακα ενώ οι μονού, από μία. Οι νανοσωλήνες άνθρακα έχουν εξαιρετικές μηχανικές ιδιότητες, όπως εφελκυστική αντοχή μερικά GPa και μέτρο ελαστικότητας μερικά TPa, και, σαν έγκλεισμα, βελτιώνουν τις μηχανικές ιδιότητες της μήτρας, δίνοντας ένα νανοσύνθετο υλικό με πολύ καλύτερες μηχανικές ιδιότητες από αυτές που έχει το υλικό της μήτρας. Λαμβάνοντας υπ’ όψη τα δύο παραπάνω, παρασκευάστηκαν φιλμς από χιτοζάνη (CS) και δύο ειδών φιλμς από χιτοζάνη με διαφορετικές περιεκτικότητες σε νανοσωλήνες άνθρακα 0,001% w/v και 0,002% w/v (CS-CNTs). Κατόπιν κόπηκαν κατάλληλα δοκίμια CS και CS-CNTs καταπονήθηκαν σε εφελκυσμό. Η εφελκυστική μηχανή που χρησιμοποιήθηκε ήταν ο μικρογραφικός ελεγκτής υλικών MiniMat 2000. Τα προς εξέταση δείγματα που εφελκύστηκαν ήταν: CS, CS-CNTs 0,001% w/v, CS-CNTs 0,002% w/v. Τα πειράματα επαναλήφθηκαν περίπου 15 φορές για κάθε περίπτωση και με χρήση του MiniMat2000 προέκυψαν οι γραφικές παραστάσεις τάσης – παραμόρφωσης και με τους κατάλληλους υπολογισμούς βρέθηκε το μέτρο ελαστικότητας κάθε υλικού. Μετά τα πειράματα της μηχανικής καταπόνησης παρατηρήθηκε ότι το μέτρο ελαστικότητας του CS-CNTs 0,001% w/v εμφάνισε αύξηση της τάξεως 44% σε σύγκριση με το μέτρο ελαστικότητας της CS ενώ του CS-CNTs 0,002% w/v εμφάνισε αύξηση της τάξεως 36%. Από αυτό συμπεραίνουμε πως θα υπάρχει μια μέση τιμή πρόσμιξης CNTs για την οποία μεγιστοποιείται το μέτρο ελαστικότητας. Περαιτέρω πειράματα κρίνονται αναγκαία για την μελέτη του υλικού. Ο έλεγχος της βιοσυμβατότητας των υλικών CS-CNTs και MWCNTs έγινε με καλλιέργεια πρόβειων μυοϊνοβλαστικών κυττάρων. Τα πειράματα που πραγματοποιήθηκαν ήταν ο έλεγχος της κυτταροτοξικότητας, του πολλαπλασιασμού και της απόπτωσης για μυοϊνοβλαστικά κύτταρα, τα οποία εξετάστηκαν για 1, 3 και 8 ημέρες. Τα αποτελέσματα του ελέγχου της κυτταροτοξικότητας έδειξαν ότι και τα δυο MWCNTs και CS-CNTs παρήγαγαν πολύ χαμηλό επίπεδο τοξικότητας. Τα αποτελέσματα του πειράματος του πολλαπλασιασμού έδειξαν ότι στην περίπτωση των μυοϊνοβλαστικών κυττάρων υπήρξε αύξηση του πολλαπλασιασμού μετά από 8 ημέρες κυτταροκαλλιέργειας και για τα MWCNTs και τη CS-CNTs. Η μεγαλύτερη αύξηση παρατηρήθηκε στη CS-CNTs. Επιπλέον, δεν υπήρξε απόπτωση σε καμία επιφάνεια, όσο και για τα MWCNTs και για τη CS-CNTs. Συνοψίζοντας, καταλήγουμε ότι οι νανοσωλήνες άνθρακα αυξάνουν τις μηχανικές ιδιότητες του νανοσύνθετου υλικού CS-CNTs, δεν επηρεάζουν αξιοσημείωτα την κυτταροτοξικότητα, δεν δημιουργούν απόπτωση και αυξάνουν τον πολλαπλασιασμό των πρόβειων μυοϊνοβλαστικών κυττάτων. / The aim of this thesis was to create a nanocomposite material with increased mechanical properties and biocompatibility. Chitosan is a linear polysaccharide and is a derivative of chitin. It is highly biocompatible and biodegradable, can form ​​insoluble compounds and can easily be used in massive, porous scaffolds and films. It has been widely used in biomedical applications in areas such as tissue engineering, pharmaceuticals, wound healing and bone substitutes. Prior studies have shown that chitosan increases the number of adherent cells compared with control surfaces. Carbon nanotubes (CNTs) consist of carbon atoms with appropriate links between them to form a cylindrical tube whose length is several orders of magnitude larger than the diameter of around 103 to 104. CNTs can be either into single or multi walled. The multi-walled CNTs are composed of several concentric layers of carbon nanotubes while the single, one layer. CNTs have excellent mechanical properties such as tensile strength of the order of several GPa and young modulus of the order of TPa, and, as a filler improves the mechanical properties of the matrix, resulting in a nanocomposite material with much better mechanical properties than those of the material matrix. Taking into account the two above films prepared from chitosan (CS) and two types of chitosan films with different contents of carbon nanotubes 0,001% w / v and 0,002% w / v (CS-CNTs) were and the results were compared. A mold was designed and built in AutoCAD for the preparation of the into a suitable shape.The mechanical tests were performed with a tensile machine the miniature materials tester MiniMat 2000. The samples that were tested were: CS, CS-CNTs 0,001% w / v, CS-CNTs 0,002% w / v. The experiments were repeated about 15 times for each case and using the resulting MiniMat2000 graphs stress - strain and the young's modulus of each material was calculated. The results showed that the young's modulus of CS-CNTs 0,001% w / v was increased at 44% compared to the tensile modulus of the CS and that of CS-CNTs 0,002% w / v was increased at 36% . This suggests that there will be an optimum content of CNTs for which the young's modulus is maximized. Further experiments are necessary to study the material. Myofibroblasts were used, in order to evaluate the biocompatibility of the materials CS-CNTs and MWCNTs. In particular cytotoxicity, proliferation and apoptosis of cells, were evaluated after for 1, 3 and 8 days of culture. Test results showed that both MWCNTs and CS-CNTs produced very low toxicity myofibroblasts. The proliferation of Myofibroblasts in the case of cell proliferation was increased after 8 days of cell culture both on MWCNTs and on CS-CNTs. The largest increase was observed on CS-CNTs. Moreover, there was no apoptosis on any of the surfaces, MWCNTs or CS-CNTs. In summary, we conclude that carbon nanotubes enhance the mechanical properties of the nanocomposite material CS-CNTs, are not cytoxic, do not generate apoptosis and increase the proliferation of sheep Myofibroblasts cells.

Drugs can be made up of nucleic acids, sugars, small organic and inorganic compounds, peptides, and large macromolecules. Drug therapy can be optimized by controlled delivery systems that release an appropriate dose to the site of action, extend the duration of delivery, reduce administration sessions, and can target a precise site of activity. An advanced method of controlled drug delivery is through injectable polymeric biomaterial microparticles that entrap drugs within their matrix for slow release (1-6 months). Surface morphology of polymer microparticles is known to affect drug release; however, it is often reported in qualitative terms only. In this thesis, a mastery over the controlled fabrication of biodegradable poly (ε-caprolactone) (PCL) microspheres is shown, as well as their characterization using different imaging conditions/techniques of the scanning electron microscope (SEM). Retinoic acid (RA), a morphogenic molecule, is encapsulated to create RA/PCL microspheres that are used to successfully deliver drug to human induced pluripotent stem cell aggregates. Furthermore, this works reports the creation of variable surface morphology PCL microspheres and their characterization via size analysis and stereo-microscopy. A rough morphology candidate is identified and selected for 3D SEM surface model reconstruction via a computer vision technique. Surface studies via SEM have a lot of potential to advance the development of these particles. The 3D model first reported here serves as foundation for quantitative surface morphology measurements. / Graduate

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Submitted by Jarede Carvalho Pereira null (dr.jarede@hotmail.com) on 2017-08-30T17:29:26Z No. of bitstreams: 1 Tese Járede.pdf: 2356458 bytes, checksum: 4637812d3b83b5f71fecba5d9a1af034 (MD5) / Approved for entry into archive by Luiz Galeffi (luizgaleffi@gmail.com) on 2017-08-30T18:09:15Z (GMT) No. of bitstreams: 1 pereira_jc_dr_araca.pdf: 2356458 bytes, checksum: 4637812d3b83b5f71fecba5d9a1af034 (MD5) / Made available in DSpace on 2017-08-30T18:09:15Z (GMT). No. of bitstreams: 1 pereira_jc_dr_araca.pdf: 2356458 bytes, checksum: 4637812d3b83b5f71fecba5d9a1af034 (MD5) Previous issue date: 2017-08-07 / Este estudo objetivou avaliar a eficácia da membrana de celulose bacteriana (Nanoskin®) no reparo ósseo de defeitos críticos de 8mm em calota de ratos machos adultos quando comparada à membrana colágena Bio-Gide®. Foram utilizados 72 ratos machos adultos divididos em três grupos experimentais: grupo coágulo (GC), grupo Bio-Gide® (BG) e grupo chá verde (CV). Os períodos de estudo foram de 7, 14 30 e 60 dias após a criação do defeito cirúrgico e aposição da membrana. Foram realizadas a análise histológica, histométrica e imunoistoquímica. Foi possível observar que aos 30 e 60 dias de reparo, o grupo BG apresentou melhor preenchimento do defeito cirúrgico, com maior quantidade de tecido ósseo. Já o grupo CV apresentou preenchimento do defeito com grande quantidade de tecido conjuntivo maduro. Os dados obtidos na análise histométrica revelam que aos 30 e 60 dias a área de osso neoformado teve grande discrepância para o grupo BG em relação ao CV e GC (Teste de Tukey - P<0,001). Por outro lado, os resultados referentes à análise por imunistoquímica evidenciam marcações moderada e intensa para as proteínas osteocalcina (OC) e osteopontina (OP) tanto para o grupo BG quanto para o CV aos 60 dias de reparo. Pode-se concluir que apesar de a membrana de chá verde apresentar aspectos promissores no reparo de tecido mole, neste estudo não permitiu o reparo ósseo em calotas de ratos, quando comparada à Bio-Gide®. / The present study aimed to evaluate the efficiency of bacterial cellulose membrane (Nanoskin® ) in the bone repair of 8 mm critical-size defects in male adult rat calvaria compared to collagen membrane BioGide® . It was used 72 adult male rats divided into three experimental groups: Clot group (CG), Bio-Gide® group (BG) and bacterial cellulose group (BC). The study period were seven, 15, 30 and 60 days after the surgical defect and position of the membrane. It were performed the histologic, histometric and imunohistochemistry analysis and possible to observe that at 30 and 60 days, BG group showed higher filling of the surgical wound with higher amount of new formed bone. The BC group showed a mature connective tissue filling the defect. Data from histometric analysis showed that at 30 and 60 days the new bone formed area was discrepant for BG group compared with BC and CG group (Tukey test – P<0,001). On the other hands, immunohistochemistry analysis showed a moderate and intense immunolabeling of osteocalcin and osteopontin at 60 days of bone repair for BG and BC group. It was possible to conclude that despite the BC membrane showing promising aspects in the soft tissue repair, did not allow bone repair in rats calvaria, compared to BioGide®.

Orientador: Enilson Antônio Sallum / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Odontologia de Piracicaba / Made available in DSpace on 2018-08-17T16:35:38Z (GMT). No. of bitstreams: 1 Queiroz_LucasAraujo_M.pdf: 23764982 bytes, checksum: 9d01ff7e493a0eb69a8166673d96fe1b (MD5) Previous issue date: 2011 / Resumo: O objetivo do presente estudo foi avaliar clinicamente, a utilização das proteínas derivadas da matriz do esmalte (EMD) e do Beta-tricálciofosfato/ hidroxiapatita (?TCP/HA), isolado ou em associação, no tratamento de lesões de bifurcações classe II mandibulares. Foram utilizados neste estudo 25 pacientes adultos, que apresentavam lesões de bifurcações classe II livres (HAMP et al, 1975). Os pacientes foram aleatoriamente divididos em dois grupos: grupo um (n=13) - acesso cirúrgico para raspagem e alisamento radicular e ?TCP/HA e grupo dois (n=12) - acesso cirúrgico para raspagem e alisamento radicular e associação EMD ?TCP/HA. Os seguintes parâmetros clínicos: Índice de Placa (IP), Sangramento à Sondagem (SS); Posição da Margem Gengival Relativa (PMGR), Profundidade de Sondagem (PS), Nível de Inserção Clínico Vertical Relativo (NICVR), Nível de Inserção Clínico Horizontal Relativo (NICHR) foram avaliados antes do procedimento cirúrgico e após 6 meses. Após o período de acompanhamento, o ganho médio no nível clínico de inserção vertical foi de 2,33 mm para o grupo um e de 1,58 mm para o grupo dois. Com relação ao nível de inserção clínico horizontal as médias de ganho foram de 2,58 mm e 2,67 mm para o grupo um e dois respectivamente. Ambos os grupos não apresentaram diferença clínica estatisticamente significante entre eles. Pode-se concluir que, os dois procedimentos são eficazes no tratamento de lesões de bifurcação classe II mandibulares / Abstract: The aim of the present clinical trial is to evaluate the associated use of enamel matrix derivative proteins with a graft material including beta-tricalcium phosphate and hydroxyapatite for the treatment of class II furcation defects. Twenty five patients presenting mandibular class II furcation involvements were selected (HAMP et al, 1975). The patients were randomly assigned to: Group one (n=13) - Open Flap Debridement + beta-tricalcium phosphate in association with hydroxiapatyte (? TCP-HA); Group two (n=12) - Open Flap Debridement (OFD) + Enamel matrix derivative proteins (EMD) + beta-tricalcium phosphate in association with hydroxiapatyte (? TCP-HA); Plaque Index (PI), Bleeding on Probing (BOP), Probing Depth (PD), Relative Gingival Margin Position (GMPR), Relative Vertical and Horizontal Clinical Attachment Level (RVCAL and RHCAL), were evaluated at baseline and 6 months after the surgeries. After the following period the mean vertical attachment level was 2,33 mm for the group one and 1,58 mm for the group two. For the horizontal attachment level the mean gain was 2,58 mm and 2,67 mm for the group one and two respectively. Both groups did not show any statistic difference between them. It can be concluded that both procedures are capable in the treatment of mandibular class II furcation defects / Mestrado / Periodontia / Mestre em Clínica Odontológica