



@article {2880,
	title = {Endothelialization of chitosan porous conduits via immobilization of a recombinant fibronectin fragment (rhFNIII7-10)},
	journal = {Acta Biomaterialia},
	volume = {9},
	year = {2013},
	note = {Export Date: 12 December 2013Source: Scopus},
	month = {2013///},
	pages = {5643 - 5652},
	url = {http://www.scopus.com/inward/record.url?eid=2-s2.0-84873199566\&partnerID=40\&md5=f548695261ada2fc57ece26bff1f45d0},
	author = {Amaral, I.F. and Neiva, I. and Ferreira da Silva, F. and Sousa, S.R. and Piloto, A.M. and Lopes, C.D.F. and Barbosa, M.A. and Kirkpatrick, C.J. and P{\^e}go, A.P.}
}
@article {2830,
	title = {Macrophage polarization following chitosan implantation},
	journal = {Biomaterials},
	volume = {34},
	year = {2013},
	note = {Export Date: 12 December 2013Source: Scopus},
	month = {2013///},
	pages = {9952 - 9959},
	url = {http://www.scopus.com/inward/record.url?eid=2-s2.0-84885383515\&partnerID=40\&md5=a152c1c59bf410d725b5e4ce4eeed18f},
	author = {Vasconcelos, D.P. and Fonseca, A.C. and Costa, M. and Amaral, I.F. and Barbosa, M.A. and {\'A}guas, A.P. and Barbosa, J.N.}
}
@article {2847,
	title = {Modulation of stability and mucoadhesive properties of chitosan microspheres for therapeutic gastric application},
	journal = {International Journal of Pharmaceutics},
	volume = {454},
	year = {2013},
	note = {Cited By (since 1996):1Export Date: 12 December 2013Source: Scopus},
	month = {2013///},
	pages = {116 - 124},
	url = {http://www.scopus.com/inward/record.url?eid=2-s2.0-84884163785\&partnerID=40\&md5=1d040067777b023c63bb769260b66dcd},
	author = {Fernandes, M. and Gon{\c c}alves, I.C. and Nardecchia, S. and Amaral, I.F. and Barbosa, M.A. and Martins, M.C.L.}
}
@booklet {2179,
	title = {Endothelialization of chitosan porous conduits via immobilization of a recombinant fibronectin fragment (rhFNIII                    7-10)
},
	howpublished = {Acta BiomaterialiaActa Biomater.},
	year = {2012},
	note = {Export Date: 21 November 2012Source: ScopusArticle in Pressdoi: 10.1016/j.actbio.2012.10.029Language of Original Document: EnglishCorrespondence Address: Amaral, I.F.; INEB - Instituto de Engenharia Biom{\'e}dica, Universidade do Porto, Rua do Campo Alegre, 823, 41email: iamaral@ineb.up.pt},
	month = {2012///},
	abstract = {The present study aimed to develop a pre-endothelialized chitosan (CH) porous hollowed scaffold for application in spinal cord regenerative therapies. CH conduits with different degrees of acetylation (DA; 4\% and 15\%) were prepared, characterized (microstructure, porosity and water uptake) and functionalized with a recombinant fragment of human fibronectin (rhFNIII                        7-10). Immobilized rhFNIII 
                        7-10 was characterized in terms of amount ( 
                        125I-radiolabelling), exposure of cell-binding domains (immunofluorescence) and ability to mediate endothelial cell (EC) adhesion and cytoskeletal rearrangement. Functionalized conduits revealed a linear increase in immobilized rhFNIII 
                        7-10 with rhFNIII 
                        7-10 concentration, and, for the same concentration, higher amounts of rhFNIII 
                        7-10 on DA 4\% compared with DA 15\%. Moreover, rhFNIII 
                        7-10 concentrations as low as 5 and 20 μg ml 
},
	keywords = {Protein conformation, Protein radiolabelling, Spinal cord injury, Surface grafting, Three-dimensional scaffolds},
	isbn = {17427061 (ISSN)},
	url = {http://www.scopus.com/inward/record.url?eid=2-s2.0-84868680582\&partnerID=40\&md5=6599fd2565f4511b94e944978afc8073},
	author = {Amaral, I.F. and Neiva, I. and Ferreira da Silva, F. and Sousa, S.R. and Piloto, A.M. and Lopes, C.D.F. and Barbosa, M.A. and Kirkpatrick, C.J. and P{\^e}go, A.P.}
}
@article {2280,
	title = {Evaluation of the effect of the degree of acetylation on the inflammatory response to 3D porous chitosan scaffolds},
	journal = {Journal of Biomedical Materials Research - Part AJ. Biomed. Mater. Res. Part A},
	volume = {93},
	year = {2010},
	note = {Cited By (since 1996): 3Export Date: 21 November 2012Source: ScopusCODEN: JBMRCdoi: 10.1002/jbm.a.32499PubMed ID: 19484769Language of Original Document: EnglishCorrespondence Address: Barbosa, J. N.; INEB - Instituto de Engenharia Biom{\'e}dica, Divis{\~a}o de Biomateriais, Universidade do Porto, Rua do Campo Alegre 823, 4150-180 Porto, Portugal; email: judite@ineb.up.ptChemicals/CAS: chitosan, 9012-76-4; Chitosan, 9012-76-4References: Sefton, M.V., Babensee, J.E., Woodhouse, K.A., Innate and adaptative immune responses in tissue engineering (2008) Semin Immunol, 20, pp. 83-85;VandeVord, P.J., Matthew, H.W.T., DeSilva, S.P., Mayton, L., Wu, B., Wooley, P.H., Evaluation of the biocompatibility of a chitosan scaffold in mice (2002) J Biomed Mater Res, 59, pp. 585-590; 
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},
	month = {2010///},
	pages = {20 - 28},
	abstract = {The effect of the degree of acetylation (DA) of 3D chitosan (Ch) scaffolds on the inflammatory reaction was investigated. Chitosan porous scaffolds with DAs of 4 and 15\% were implanted using a subcutaneous air-pouch model of inflammation. The initial acute inflammatory response was evaluated 24 and 48 h after implantation. To characterize the initial response, the recruitment and adhesion of inflammatory cells to the implant site was studied. The fibrous capsule formation and the infiltration of inflammatory cells within the scaffolds were evaluated for longer implantation times (2 and 4 weeks). Chitosan with DA 15\% attracted the highest number of leukocytes to the implant site. High numbers of adherent inflammatory cells were also observed in this material. For longer implantation periods Ch scaffolds with a DA of 15\% induced the formation of a thick fibrous capsule and a high infiltration of inflammatory cells within the scaffold. Our results indicate that the biological response to implanted Ch scaffolds was influenced by the DA. Chitosan with a DA of 15\% induce a more intense inflammatory response when compared with DA 4\% Ch. Because inflammation and healing are interrelated, this result may provide clues for the relative importance of acetyl and amine functional groups in tissue repair and regeneration. {\textcopyright} 2009 Wiley Periodicals, Inc.},
	keywords = {acetic acid derivative, Acetylation, Acute inflammatory response, amine, Amine functional groups, animal cell, animal experiment, animal tissue, Animals, article, Biological materials, Biological response, Biomaterials, cell adhesion, cell infiltration, Chitin, chitosan, Chitosan scaffold, Degree of acetylation, Fibrous capsule, Fibrous capsule formation, freeze drying, Functional groups, Hydrogels, implant, implantation, Implantation periods, Implants, Experimental, Inflammation, Inflammatory cells, Inflammatory reaction, Inflammatory response, leukocyte count, Leukocytes, male, Mice, Mice, Inbred BALB C, Microscopy, Electron, Scanning, mouse, nonhuman, Pathology, porosity, Porous scaffold, Prosthesis Implantation, Relative importance, Scaffolds, Subcutaneous Tissue, Surface Properties, Three dimensional, Tissue regeneration, tissue repair, Tissue response, Tissue responses, Tissue Scaffolds},
	isbn = {15493296 (ISSN)},
	url = {http://www.scopus.com/inward/record.url?eid=2-s2.0-77649219383\&partnerID=40\&md5=58781af29b82f3108ded649c4776e428},
	author = {Barbosa, J.N. and Amaral, I.F. and {\'A}guas, A.P. and Barbosa, M.A.}
}
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Oliveira, S.M., Turner, G., Mijares, D., Amaral, I.F., Barbosa, M.A., Teixeira, C.C., (2007) Engineering endochondral bone: In vivo studies
},
	month = {2009///},
	pages = {625 - 634},
	abstract = {Chitosan scaffolds have been shown to possess biological and mechanical properties suitable for tissue engineering and clinical applications. In the present work, chitosan sponges were evaluated regarding their ability to support cartilage cell proliferation and maturation, which are the first steps in endochondral bone formation. Chitosan sponges were seeded with chondrocytes isolated from chicken embryo sterna. Chondrocyte/chitosan constructs were cultured for 20 days, and treated with retinoic acid (RA) to induce chondrocyte maturation and matrix synthesis. At different time points, samples were collected for microscopic, histological, biochemical, and mechanical analyses. Results show chondrocyte attachment, proliferation, and abundant matrix synthesis, completely obliterating the pores of the sponges. RA treatment caused chondrocyte hypertrophy, characterized by the presence of type X collagen in the extracellular matrix and increased alkaline phosphatase activity. In addition, hypertrophy markedly changed the mechanical properties of the chondrocyte/chitosan constructs. In conclusion, we have developed chitosan sponges with adequate pore structure and mechanical properties to serve as a support for hypertrophic chondrocytes. In parallel studies, we have evaluated the ability of this mature cartilage scaffold to induce endochondral ossification. {\textcopyright} Copyright 2009, Mary Ann Liebert, Inc. 2009.},
	keywords = {alkaline phosphatase, Alkaline phosphatase activity, animal, animal cell, animal tissue, Animals, article, biomechanics, Body fluids, Bone, Bone and Bones, Bone formation, Cartilage, cartilage cell, Cell culture, cell damage, cell hypertrophy, cell isolation, cell maturation, Cell membranes, cell proliferation, Cells, Cultured, chick embryo, Chicken embryo, Chitin, chitosan, Chitosan scaffold, Chondrocyte, Chondrocyte maturation, Chondrocytes, Clinical application, collagen type 10, Collagen Type X, controlled study, cytology, DNA, drug effect, embryo, enchondral ossification, Endochondral ossification, enzyme activity, enzymology, Extracellular matrices, extracellular matrix, gene expression profiling, Hypertrophic chondrocytes, in vitro study, In-vitro, Ligaments, Matrix synthesis, Mechanical analysis, Mechanical properties, metabolism, methodology, nonhuman, Pathology, physiology, Porifera, porosity, priority journal, retinoic acid, Retinoic acids, Scaffolds, sponge (Porifera), Sterna, sternum, synthesis, Time points, Tissue engineering, tissue scaffold, Tissue Scaffolds, Tretinoin, Type X collagen},
	isbn = {19373341 (ISSN)},
	url = {http://www.scopus.com/inward/record.url?eid=2-s2.0-64549143844\&partnerID=40\&md5=310f8e62d4dd51ccb9399db171440148},
	author = {Oliveira, S.M. and Amaral, I.F. and Barbosa, M.A. and Teixeira, C.C.}
}
@article {2317,
	title = {Engineering endochondral bone: In vivo studies},
	journal = {Tissue Engineering - Part ATissue Eng. Part A},
	volume = {15},
	year = {2009},
	note = {Cited By (since 1996): 14Export Date: 21 November 2012Source: Scopusdoi: 10.1089/ten.tea.2008.0052Language of Original Document: EnglishCorrespondence Address: Teixeira, C. C.; Department of Basic Science and Craniofacial Biology, College of Dentistry, New York University, 345 East. 24th St., New York, NY 10010; email: ct40@nyu.eduChemicals/CAS: chitosan, 9012-76-4; Chitosan, 9012-76-4; MineralsReferences: Yelin, E., Callahan, L.F., The economic cost and social and psychological impact of musculoskeletal conditions. National Arthritis Data Work Groups (1995) Arthritis Rheum, 38, p. 1351;Service, R.F. Tissue engineers build new bone. Science 289,1498, 2000Dupraz, A., Delecrin, J., Moreau, A., Pilet, P., Passuti, N., Long-term bone response to particulate injectable ceramic (1998) J Biomed Mater Res, 42, p. 368; 
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},
	month = {2009///},
	pages = {635 - 643},
	abstract = {The use of biomaterials to replace lost bone has been a common practice for decades. More recently, the demands for bone repair and regeneration have pushed research into the use of cultured cells and growth factors in association with these materials. Here we report a novel approach to engineer new bone using a transient cartilage scaffold to induce endochondral ossification. Chondrocyte/chitosan scaffolds (both a transient cartilage scaffold- experimental-and a permanent cartilage scaffold-control) were prepared and implanted subcutaneously in nude mice. Bone formation was evaluated over a period of 5 months. Mineralization was assessed by Faxitron, micro computed tomography, backscatter electrons, and Fourier transform infrared spectroscopy analyses. Histological analysis provided further information on tissue changes in and around the implanted scaffolds. The deposition of ectopic bone was detected in the surface of the experimental implants as early as 1 month after implantation. After 3 months, bone trabeculae and bone marrow cavities were formed inside the scaffolds. The bone deposited was similar to the bone of the mice vertebra. Interestingly, no bone formation was observed in control implants. In conclusion, an engineered transient cartilage template carries all the signals necessary to induce endochondral bone formation in vivo. {\textcopyright} Copyright 2009, Mary Ann Liebert, Inc. 2009.},
	keywords = {animal, animal experiment, animal tissue, Animals, article, Biological materials, Biomaterials, Bone, Bone and Bones, Bone formation, bone marrow, bone mineralization, Bone repair, bone transplantation, Calcification, Physiologic, Cartilage, cartilage cell, chick embryo, chitosan, Chondrocytes, Computerized tomography, controlled study, Cultured cell, cytology, drug effect, Ectopic bones, ectopic tissue, electron, enchondral ossification, Endochondral ossification, Fourier transform infrared spectroscopy, Growth (materials), Growth factor, Histological analysis, histology, implantation, in vivo study, In-control, In-vivo, infrared spectroscopy, Ligaments, male, metabolism, methodology, Mice, Mice, Nude, micro-computed tomography, Microcomputed tomography, Microscopy, Electron, Scanning, mineral, Minerals, mouse, Mus, Mus musculus, nonhuman, Nude mice, nude mouse, ossification, physiology, priority journal, Prosthesis Implantation, Scaffolds, Scanning electron microscopy, Spectroscopic analysis, Spectroscopy, Spectroscopy, Fourier Transform Infrared, Tissue changes, Tissue engineering, tissue scaffold, Tissue Scaffolds, Tomography, trabecular bone, vertebra},
	isbn = {19373341 (ISSN)},
	url = {http://www.scopus.com/inward/record.url?eid=2-s2.0-64549096039\&partnerID=40\&md5=d389cb8b10be8945007a5b6996b8896f},
	author = {Oliveira, S.M. and Mijares, D.Q. and Turner, G. and Amaral, I.F. and Barbosa, M.A. and Teixeira, C.C.}
}
@article {2297,
	title = {Fibronectin-mediated endothelialisation of chitosan porous matrices},
	journal = {BiomaterialsBiomaterials},
	volume = {30},
	year = {2009},
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},
	month = {2009///},
	pages = {5465 - 5475},
	abstract = {Chitosan (Ch) porous matrices were investigated regarding their ability to be colonized by human microvascular endothelial cells (HPMEC-ST1.6R cell line) and macrovascular endothelial cells namely HUVECs. Specifically we assessed if previous incubation of Ch in a fibronectin (FN) solution was effective in promoting endothelial cell (EC) adhesion to Ch matrices with different degrees of acetylation (DAs). Upon FN physiadsorption, marked differences were found between the two DAs investigated, namely DA 4\% and 15\%. While cell adhesion was impaired on Ch with DA 15\%, ECs were able to not only adhere to Ch with DA 4\%, but also to spread and colonize the scaffolds, with retention of the EC phenotype and angiogenic potential. To explain the observed differences between the two DAs, protein adsorption studies using                        125I-FN and immunofluorescent labelling of FN cell-binding domains were carried out. In agreement with the higher cell numbers found, scaffolds with DA 4\% revealed a higher number of exposed FN cell-binding domains as well as greater ability to adsorb FN and to retain and exchange adsorbed FN in the presence of competitive proteins. These findings suggest that the DA is a key parameter modulating EC adhesion to FN-coated Ch by influencing the adsorbed protein layer. {\textcopyright} 2009 Elsevier Ltd. All rights reserved.
},
	keywords = {Acetylation, Adhesion, Adsorption, angiogenesis, article, Biocompatible Materials, Blood vessel prostheses, cell adhesion, Cell culture, cell invasion, Cell Line, Cell lines, Cell membranes, Cell number, cell proliferation, Cell size, cell spreading, cell survival, Cell-binding domain, Chitin, Chitin/chitosan, chitosan, colony formation, controlled study, Degrees of acetylation, Electric conductivity measurement, Electric currents, Endothelial cell, Endothelial Cells, Endothelialisation, endothelium, endothelium cell, epithelization, Fibronectin, Fibronectins, human, human cell, Human microvascular endothelial cells, Humans, immunofluorescence test, Key parameters, Labels, materials testing, Matrix algebra, Neovascularization, Physiologic, phenotype, porosity, Porous matrixs, priority journal, Protein adsorption, Protein layers, Scaffold, Scaffolds, Self assembly, Tissue engineering, tissue scaffold, umbilical vein},
	isbn = {01429612 (ISSN)},
	url = {http://www.scopus.com/inward/record.url?eid=2-s2.0-68549107574\&partnerID=40\&md5=9babb57c1a4f2ae6eb7e7e0643bffe48},
	author = {Amaral, I.F. and Unger, R.E. and Fuchs, S. and Mendon{\c c}a, A.M. and Sousa, S.R. and Barbosa, M.A. and P{\^e}go, A.P. and Kirkpatrick, C.J.}
}
@article {2381,
	title = {Attachment, spreading and short-term proliferation of human osteoblastic cells cultured on chitosan films with different degrees of acetylation},
	journal = {Journal of Biomaterials Science, Polymer EditionJ. Biomater. Sci. Polym. Ed.},
	volume = {18},
	year = {2007},
	note = {Cited By (since 1996): 22Export Date: 21 November 2012Source: ScopusCODEN: JBSEEdoi: 10.1163/156856207780425068Language of Original Document: EnglishCorrespondence Address: Amaral, I.F.; INEB - Instituto de Engenharia Biom{\'e}dica, Laborat{\'o}rio de Biomateriais, Rua do Campo Alegre, 823, 4150-180 Porto, Portugal; email: iamaral@ineb.up.ptChemicals/CAS: chitosan, 9012-76-4; Biocompatible Materials; Chitosan, 9012-76-4References: Vacanti, C.A., Bonassar, L.J., Vacanti, J.P., (2000) Principles of Tissue Engineering, p. 671. , R. P. Lanza, R. Langer and J. Vacanti Eds, p, Academic Press, San Diego, CA;Bruder, S.P., Caplan, A.I., (2000) Principles of Tissue Engineering, p. 683. , R. P. Lanza, R. Langer and J. Vacanti Eds, p, Academic Press, San Diego, CA; 
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},
	month = {2007///},
	pages = {469 - 485},
	abstract = {Chitosan (Ch) is being actively investigated as a non-protein template for the growth of an increasing number of anchorage-dependent cells, including chondrocytes and bone cells. In the present work, Ch films with degrees of N-acetylation (DAs) in the range of 4 to 49\% were evaluated with respect to the attachment, spreading and short-term proliferation of osteoblasts, using human osteoblastic MG-63 cells. The films were characterized in terms of surface morphology and surface charge by atomic force microscopy and streaming potential measurements, respectively. Cell attachment was assessed after 3 and 24 h of cell culture. After 24 h of incubation, cell attachment was found to be dependent on the DA, lower DAs favouring cell adhesion. With time, cell spreading and cytoskeleton organization were only attained for DAs <= 13\%. Regarding cell proliferation, cells grown on films with the lowest DA (4\%) revealed a higher specific growth rate as compared to those grown on films with higher DAs. Films with a DA of 49\% failed to sustain cell proliferation. In addition, a longer lag-phase was observed on Ch, as compared to TCPS, in accordance to an observed delay of cell spreading. The present findings revealed that differences in the DA as small as 9\% may be critical in terms of the osteoblast response to two-dimensional Ch-based matrices. {\textcopyright} 2007 VSP.},
	keywords = {Acetylation, AFM, article, Atomic force microscopy, Biocompatible Materials, biofilm, Biofilms, biomaterial, bone cell, cartilage cell, cell adhesion, cell anchorage, Cell culture, cell growth, Cell Line, Tumor, cell proliferation, Cell Shape, cell spreading, cell survival, Chemical structure, chemistry, Chitin, chitosan, comparative study, controlled study, cytoskeleton, Degree of acetylation, drug effect, Growth kinetics, human, human cell, Humans, incubation time, metabolism, Microscopy, Atomic Force, osteoblast, Osteoblast-like cells, Osteoblasts, Pathology, priority journal, Surface charge, Surface Properties, surface property, Surface treatment, tumor cell line, Zeta potential},
	isbn = {09205063 (ISSN)},
	url = {http://www.scopus.com/inward/record.url?eid=2-s2.0-34347375992\&partnerID=40\&md5=437b9ed270e859425b76aa3655c611be},
	author = {Amaral, I.F. and Cordeiro, A.L. and Sampaio, P. and Barbosa, M.A.}
}
@article {2399,
	title = {Functionalization of chitosan membranes through phosphorylation: Atomic force microscopy, wettability, and cytotoxicity studies},
	journal = {Journal of Applied Polymer ScienceJ. Appl. Polym. Sci.},
	volume = {102},
	year = {2006},
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},
	month = {2006///},
	pages = {276 - 284},
	abstract = {Grafting negatively charged groups such as phosphates is a well-known strategy for inducing the deposition of apatite-like layers under simulated physiological conditions. In this investigation, chitosan was phosphorylated in an attempt to enhance its osteoconduction. Chitosan membranes were phosphorylated at 30{\textdegree}C with the H                        3PO
                        4/ Et 
                        3PO
                        4/P
                        2O
                        5/butanol reaction system for periods up to 48 h. This method is an alternative to the phosphoric acid/ urea/dimethylformamide phosphorylation method, which involves the use of much higher temperatures. In this study, the effects of the phosphorylation reaction time on the surface morphology and surface free energy of phosphorylated membranes were investigated with atomic force microscopy and static-contact-angle measurements, respectively. In addition, the modified membranes were evaluated with respect to their cytotoxiciry toward bone cells through the incubation of human osteoblastic cells with extracts of the materials for two different periods: 24 and 120 h. The results revealed a reduction of the average surface roughness at the nanometer scale with increasing phosphorylation reaction time. Wettability studies showed an increase in the polar component of the surface free energy with increasing reaction time as a result of the increase in the phosphate surface concentration. Cytotoxicity studies revealed no cytotoxic effect of phosphorylated membranes on osteoblastic cells, regardless of the incubation period. {\textcopyright} 2006 Wiley Periodicals, Inc.
},
	keywords = {Atomic force microscopy, Atomic force microscopy (AFM), Biocompatibility, Biological applications, Biological applications of polymers, Biomaterials, Chitin, chitosan, Chitosan membranes, grafting, Grafting (chemical), Osteoblastic cells, Phosphorylation, Polymeric membranes, polysaccharide, Polysaccharides, surface property, toxicity, Urea},
	isbn = {00218995 (ISSN)},
	url = {http://www.scopus.com/inward/record.url?eid=2-s2.0-33749478403\&partnerID=40\&md5=c365a50e3e767b1da90ad4ccd331440b},
	author = {Amaral, I.F. and Granja, P.L. and Melo, L.V. and Saramago, B. and Barbosa, M.A.}
}
@article {2421,
	title = {Three-dimensional culture of human osteoblastic cells in chitosan sponges: The effect of the degree of acetylation},
	journal = {Journal of Biomedical Materials Research - Part AJ. Biomed. Mater. Res. Part A},
	volume = {76},
	year = {2006},
	note = {Cited By (since 1996): 19Export Date: 21 November 2012Source: ScopusCODEN: JBMRCdoi: 10.1002/jbm.a.30522PubMed ID: 16270345Language of Original Document: EnglishCorrespondence Address: Amaral, I.F.; INEB-Instituto de Engenharia Biom{\'e}dica, Laborat{\'o}rio de Biomateriais, Rua do Campo Alegre, 823, 4150-180 Porto, Portugal; email: iamaral@ineb.up.ptChemicals/CAS: alkaline phosphatase, 9001-78-9; chitosan, 9012-76-4; collagen, 9007-34-5; Alkaline Phosphatase, EC 3.1.3.1; Chitosan, 9012-76-4; Collagen, 9007-34-5References: Bruder, S.P., Caplan, A.I., Bone regeneration through cellular engineering (2000) Principles of Tissue Engineering, pp. 683-696. , Lanza RP, Langer R, Vacanti J, editors. San Diego: Academic Press;Anselme, K., Osteoblast adhesion on biomaterials (2000) Biomaterials, 21, pp. 667-681; 
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},
	month = {2006///},
	pages = {335 - 346},
	abstract = {In this investigation, the effect of the degree of acetylation (DA) of chitosan on the behavior of human osteoblastic MG-63 cells cultured in three-dimensional chitosan matrices was assessed. Chitosan sponges with DAs in the range of 4 to 49\% were prepared and characterized in terms of microstructure, porosity, and pore size. Collagen sponges were used as 3D control. Cell proliferation was determined using the MTT assay while the retention of the osteoblastic phenotype was monitored by assaying alkaline phosphatase activity. Cell morphology, cytoskeletal organization, and viability were assessed using different microscopy techniques. Chitosan sponges showed a similar microstructure regardless the DA, except for the highest DA used, where a more heterogeneous pore distribution was observed. In terms of cell proliferation, alkaline phosphatase activity and cell viability, cells cultured in chitosan scaffolds performed as well as in the 3D control regardless the DA, except for the highest DA used, where an inhibitory effect on cell proliferation was found. However, while in sponges with DAs <= 13\% cells attached and spread displaying long cell filopodia and numerous cell-to-cell contacts, in sponges with higher DAs cells tended to remain spherical and grow into spheroid-like cellular aggregates. In the present study, the DA played a key role in determining the affinity of osteoblastic cells towards the substrates, possibly by influencing the nature of the initial adsorbed protein layer. {\textcopyright} 2005 Wiley Periodicals, Inc.},
	keywords = {Acetylation, Aggregates, alkaline phosphatase, article, Bone Regeneration, bone tissue, Bone tissue engineering, Cell culture, Cell Culture Techniques, Cell filopodia, cell growth, cell proliferation, Cell Shape, cell structure, cell survival, cell viability, chitosan, Collagen, confocal microscopy, controlled study, Cytoskeletal organization, cytoskeleton, Degree of acetylation (DA), enzyme activity, gel permeation chromatography, human, human cell, Humans, infrared spectroscopy, Microstructure, morphology, osteoblast, Osteoblastic cells, Osteoblasts, Phosphatase activity, porosity, Proteins, Scaffold, Scaffolds, Scanning electron microscopy, Substrates, Tissue engineering},
	isbn = {00219304 (ISSN)},
	url = {http://www.scopus.com/inward/record.url?eid=2-s2.0-33645101773\&partnerID=40\&md5=5adc6336963ec1c8a83d8ec89943159d},
	author = {Amaral, I.F. and Sampaio, P. and Barbosa, M.A.}
}
@article {2438,
	title = {Chemical modification of chitosan by phosphorylation: An XPS, FT-IR and SEM study},
	journal = {Journal of Biomaterials Science, Polymer EditionJ. Biomater. Sci. Polym. Ed.},
	volume = {16},
	year = {2005},
	note = {Cited By (since 1996): 50Export Date: 21 November 2012Source: ScopusCODEN: JBSEEdoi: 10.1163/156856205774576736Language of Original Document: EnglishCorrespondence Address: Amaral, I.F.; Laborat{\'o}rio de Biomateriais, INEB - Instituto de Engenharia Biom{\'e}dica, Rua do Campo Alegre, 823, 4150-180 Porto, Portugal; email: iamaral@ineb.up.ptChemicals/CAS: chitosan, 9012-76-4; phosphate, 14066-19-4, 14265-44-2; apatite, 64476-38-6; butanol, 35296-72-1, 71-36-3; calcium ion, 14127-61-8; Chitosan, 9012-76-4; PhosphatesReferences: Khor, E., (2001) Chitin: Fulfilling a Biomaterials Promise, , Elsevier Science, Oxford;Vande Vord, P.J., Matthew, H.W.T., DeSilva, S.P., Mayton, L., Wu, B., Wooley, P.H., (2002) J. Biomed. Mater. Res., 59, p. 585; 
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},
	month = {2005///},
	pages = {1575 - 1593},
	abstract = {In the present work, the surface of chitosan membranes was modified using a phosphorylation method carried out at room temperature. Phosphorylation may be of particular interest in materials for orthopaedic applications, due to the cation-exchange properties of phosphate functionalities. Phosphate groups chelate calcium ions, thus inducing the deposition of an apatite-like layer known to improve the osteoconduction of polymer-based implants. Additionally, the negatively charged phosphate functionalities, together with the positively charged amine groups from chitosan, are expected to provide chitosan with an amphoteric character, which may be useful as a combinatorial therapeutic strategy, by simultaneously allowing the immobilization of signalling molecules like growth factors. Phosphorylation was carried out at room temperature using the H                        3 PO
                        4/Et
                        3 PO
                        4/P
                        2O
                        5/butanol method. Surface characterization was performed by XPS, ATR-FT-IR, and SEM. Cross-sections were analyzed by SEM fitted with EDS. The phosphate content increased with the reaction time, as shown by XPS and ATR-FT-IR, a P/N atomic ratio of 0.73 being obtained after 48 h of treatment. High-resolution XPS spectra regarding C
                        1s, O
                        1s, N
                        1s and P
                        2p are discussed. The introduction of a neutralization step led to a reduction of P content, which pointed out to the presence of phosphates ionically bound to protonated amines, in addition to phosphate esters. EDS analysis of cross-sections revealed a gradual P reduction up to 50\% towards the inner part of the membrane. {\textcopyright} VSP 2005.
},
	keywords = {Acetylation, amine, animal, Animals, Apatite, article, bone conduction, butanol, calcium ion, cation exchange, cephalopod, chelate, Chemical modification, chemistry, Chitin, chitosan, comparative study, controlled study, Decapodiformes, Fourier transform infrared spectroscopy, Fourier transform infrared spectroscopy (FT-IR), Growth factor, immobilization, Implants (surgical), infrared spectroscopy, isolation and purification, Microscopy, Electron, Scanning, molecular weight, organophosphate, phosphate, Phosphate groups, Phosphates, Phosphorylation, Phosphorylation method, Plastics applications, priority journal, reflectometry, room temperature, Scanning electron microscopy, SEM-EDS, signal transduction, spectrometry, Spectrometry, X-Ray Emission, Spectroscopy, Fourier Transform Infrared, Surface chemical reaction, Surface Properties, surface property, temperature, time, Time Factors, viscosity, X ray photoelectron spectroscopy, X-rayphotoelectron spectroscopy (XPS)},
	isbn = {09205063 (ISSN)},
	url = {http://www.scopus.com/inward/record.url?eid=2-s2.0-33644875542\&partnerID=40\&md5=cb42a0fba6d2000d93b90af9724a5bc1},
	author = {Amaral, I.F. and Granja, P.L. and Barbosa, M.A.}
}
@article {2454,
	title = {Polysaccharides as scaffolds for bone regeneration},
	journal = {ITBM-RBMITBM-RBM},
	volume = {26},
	year = {2005},
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},
	month = {2005///},
	pages = {212 - 217},
	abstract = {This review is concerned with the use of polysaccharides for bone regeneration. Cellulose, alginate and chitosan have been chosen as examples of the formidable potential of this class of materials. Many others could be cited but our experience and research interests dictated the choice. As in many areas of research we know how they started but we cannot see how and if they will end. Modification of cellulose was our entrance to the world of polysaccharides and a lot of what we know we owe to Charles Baquey, to whom we would like to express our immense gratitude. This chapter is dedicated to him, for his cooperation, unlimited enthusiasm and friendship. {\textcopyright} 2005 Elsevier SAS. All rights reserved.},
	keywords = {Alginate, alginic acid, Amino Acid Sequence, arginylglycylaspartic acid, biomaterial, biotechnology, bone cell, bone prosthesis, Bone Regeneration, Cellulose, chitosan, enzyme degradation, polysaccharide, Polysaccharides, review, Tissue engineering},
	isbn = {12979562 (ISSN)},
	url = {http://www.scopus.com/inward/record.url?eid=2-s2.0-20744434609\&partnerID=40\&md5=43d83907f18b3e44035dfe14ff13a24f},
	author = {Barbosa, M.A. and Granja, P.L. and Barrias, C.C. and Amaral, I.F.}
}
@article {2444,
	title = {Rat bone marrow stromal cell osteogenic differentiation and fibronectin adsorption on chitosan membranes: The effect of the degree of acetylation},
	journal = {Journal of Biomedical Materials Research - Part AJ. Biomed. Mater. Res. Part A},
	volume = {75},
	year = {2005},
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},
	month = {2005///},
	pages = {387 - 397},
	abstract = {Cell adhesion, migration, and proliferation of a few anchorage-dependent cells cultured on chitosan (Ch) matrices are influenced by the degree of N-acetylation (DA) of Ch. In the present work, we examined the influence of the DA on the attachment, spreading, proliferation, and osteogenic differentiation of rat bone marrow stromal cells (rBM-SCs). Ch membranes were characterized in terms of surface morphology, roughness, and wettability, and in terms of adsorption of an adhesive protein, fibronectin (Fn). Chs with DAs in the range of 4 to 49\% were used. Among the Ch samples, the DA of 4\% led to the highest Fn surface concentration, both from single protein solution and from diluted serum. Furthermore, the levels of Fn adsorbed from serum found for this DA were threefold higher than for the tissue culture polystyrene control, indicating that in the presence of competitive proteins Ch is more specific toward Fn adsorption than tissue culture polystyrene. rBMSCs cultured on Ch carrying a DA of 4\% were able to spread, proliferate, and differentiate, reaching a higher level of osteogenic differentiation than on the control, despite the lower cell attachment observed for all Ch samples. Because the Ch sample with a DA of 4\% showed the highest Fn adsorption from serum, we suggest that cell adhesion, spreading, and osteogenic differentiation of rBMSCs on Ch may be mediated by the adsorbed layer of Fn. {\textcopyright} 2005 Wiley Periodicals, Inc.},
	keywords = {Acetylation, Adhesion, Adsorption, animal cell, Animals, article, artificial membrane, binding affinity, binding competition, Bone, bone development, bone marrow cell, Bone Marrow Cells, Bone marrow stomal cells, Bone marrow stromal cells, cell adhesion, cell anchorage, Cell culture, cell differentiation, Cell membranes, cell migration, cell proliferation, Cell Shape, cell spreading, chitosan, Coated Materials, Biocompatible, controlled study, cytoskeleton, Degree of acetylation, Fibronectin, Fibronectins, male, materials testing, morphology, nonhuman, polystyrene, Proteins, rat, Rats, Rats, Wistar, roughness, stroma cell, Stromal Cells, Surface Properties, surface property, Tissue culture, wettability},
	isbn = {00219304 (ISSN)},
	url = {http://www.scopus.com/inward/record.url?eid=2-s2.0-26844551535\&partnerID=40\&md5=fa91e3e29d946053ae7bb231b6a2640e},
	author = {Amaral, I.F. and Lamghari, M. and Sousa, S.R. and Sampaio, P. and Barbosa, M.A.}
}
