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|Title: ||Biopolymer based scaffolds for tissue engineering|
|Authors: ||Bellomo, Francesca|
|Supervisor/Tutor: ||Paoletti, Sergio|
|Co-Advisor: ||Biasotto, Matteo|
|Issue Date: ||8-Apr-2011|
|Publisher: ||Università degli studi di Trieste|
|Abstract: ||Tissue engineering in the orthopedic field is mainly focused on the development and design of material to serve as a temporary extracellular matrix or scaffold to overcame the limitation of actual treatments. In fact, current treatments are based on autologous or autogenous bone grafts and as an alternative to these, metals and ceramics. The limitation of this kind of implants is related to shortage of autograft that can be obtained and to donor site morbidity; the possibility of immune rejection and of pathogen transmission from donor to host in the case of allograft; the poor overall integration with the tissue at the implantation site for the metal and ceramic.
The aim of study is the development of bio-composite scaffolds for orthopedic applications, based on the combination of polysaccharides (mainly alginate, Alg) engineered with bioactive molecules and nano-hydroxyapatite (nHap). The first step was the synthesis and characterization of the different components of the scaffold. The nHap was synthesized with sol-gel method and the obtained crystals have been chemically and morphologically characterized by mean of Raman spectroscopy, X-Ray Diffraction and Transmission Electron Microscopy. All the collected results confirm the chemical composition of nHap and information about the average dimension of crystals (120nm). The synthesized nHap was used to produce composite hydrogels of Alg and nHap. The porous structure was achieved through freeze-casting. The obtained scaffold was characterized with µCT (µComputed tomography) and confocal microscopy; the histomorphometric data were compared with the parameters of native tissue with an high level of similarity between trabecular bone and scaffolds structure. Subsequently, the scaffolds have been investigated for their cytocompatibility and the results shows high rate of proliferation of cells seeded into the scaffold.
The modification with specific proteins or peptides of the backbone of a polymer can be an effective strategy to tailor and control cell attachment, migration, proliferation and differentiation. Incorporation of peptide motifs containing sequences that are recognized by integrin receptors, such as arginine-glycine-aspartic acid (RGD)-based sequences, are now a common strategy to enhance the biological properties as well as differentiation and proliferation of a variety of cells, including osteoblasts. Along the same line, reducing bacterial adhesion is important since microorganism surface attachment is the ﬁrst critical step in the development of implant-associated infections.
To introduce bioactive molecules and antimicrobial agents, enhancing in this way the biological property of the scaffolds, Chitlac was exploited as vector. Chitlac is a lactitolated-derivative of chitosan that can be modified by chemical grafting of bioactive peptides like RGD and that can be use to produce and stabilize silver nanoparticles (nAg) with antimicrobial activity. By adsorbing the Chitlac on the scaffold’s surface we spread the bio-signal inside the structure to induce a specific cell reaction. Alg/nHap scaffolds were coated with Chitlac-RGD and Chitlac-nAg, respectively, and on those, biological and antimicrobial in vivo tests were performed. For both scaffolds we observed good cytocompatibility and, in the case of scaffold with RGD, an improved cell proliferation; moreover, the scaffold with nAg showed a high level of antimicrobial activity.
According the results obtained from the previous cytocompatibility tests, preliminary cytocompatibility in vivo tests were performed. The animal model was New Zealand rabbit and the produced scaffolds were inserted into bone defects on femur. Post-operatively, three fluorochromes were administered sequentially every week. After 5 weeks the rabbits were sacrificed and all the implanted bones were analyzed using µ-CT and light and confocal microscopy. We observed a high level of osteointegration of the scaffolds and ingrowth of newly deposited structured lamellar bone inside them, indicating good osteoconductive properties.
In conclusion, the developed scaffolds have suitable biological properties both bioactive and antimicrobial. The in vitro results shows a high level of cytocompatibility for all the scaffolds studied and that the presence of Chitlac-nAg does not compromise the compatibility of scaffolds. In the case of scaffolds modified with RGD the results confirm that the presence of bioactive molecule is able to enhance the healing process also with respect to the BAG (BioActive Glass) control.
The findings of the present study revealed that the structures here developed could serve as promising filler for orthopedic application.|
|PhD cycle: ||XXIII Ciclo|
|PhD programme: ||SCUOLA DI DOTTORATO DI RICERCA IN NANOTECNOLOGIE|
|Keywords: ||tissue engineering, bone, polysaccharides, scaffold,bioactivity|
|Main language of document: ||en|
|Type: ||Tesi di dottorato|
|Scientific-educational field: ||BIO/10 BIOCHIMICA|
|Appears in Collections:||Scienze biologiche|
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