Co-cultures and cell sheet engineering as relevant tools to improve the outcome of bone tissue engineering strategies

Taking into consideration the complex biology of bone tissue it is quite clear that the understanding of the cellular interactions that regulate the homeostasis and regeneration of this remarkable tissue is essential for a successful Tissue Engineering strategy. The in vitro study of these cellular interactions relies on co-culture systems, a tremendously useful methodology where two or more cell types are cultured at the same time. Such strategy increases the complexity of typical monoculture systems, allowing the in vitro settings to closely mimic the in vivo environment. Moreover, 2D coculture systems have been extensively used by cell biologists to study cell interactions as an attempt to understand specific cellular mechanisms and signalling pathways. The interaction between osteoblasts/ osteoprogenitor cells and different cell types relevant within the bone Tissue Engineering context, namely mononuclear cells from peripheral blood and umbilical cord blood and fibroblasts, has been addressed in the first part of this thesis. The different co-cultures showed that mononuclear cells from peripheral blood were capable of accelerating the osteogenic differentiation of human bone marrow stromal cells by producing BMP-2. On the other hand, osteoblasts cultured on carrageenan membranes were capable of supporting the culture of endothelial progenitors cells present in the mononuclear fraction of umbilical cord blood that contributed to the in vivo angiogenesis after implantation in an inflammatory setting. Furthermore, fibroblasts, which are key players in the formation of fibrotic tissue after a biomaterial implantation, were shown to decrease the osteogenic activity of osteoblasts through gap junctional communication. A serious limitation of the paradigmatic use of scaffolds for bone Tissue Engineering is the lack of oxygen and nutrient supply to the cells in the core of the engineered construct leading to cell necrosis at the bulk of the constructs. Furthermore, foreign body response to the implanted biomaterial is a frequent reaction of the host and has as a consequence the formation of fibrotic tissue surrounding the implant. The use of cell sheet engineering for bone Tissue Engineering can potentially avoid those shortcomings. One of the explored strategies comprised the production of osteogenic cell sheets using this technology. Its potential for in vivo bone formation was analyzed and the formation of vascularized bone tissue with a marrow was originally demonstrated by implanting a single osteogenic cell sheet in a nude mice model. Furthermore, in order to promote vascularization, co-cultured osteogenic cell sheets with endothelial cells were also created. Endothelial cells, stacked in between osteogenic cell sheets, were proven to contribute to new vessel formation and increased bone formation in vivo This thesis demonstrates that monocytes/macrophages from peripheral blood can accelerate the osteogenic differentiation of osteoprogenitor cells while fibroblasts, have a deleterious effect on the osteogenic phenotype of osteoblasts. In addition, within an inflammatory host reaction, cells from the mononuclear fraction of umbilical cord blood were capable of contributing to new blood vessel formation after co-culture with osteoblasts. Moreover, when using the cell sheet technology to fabricate a bone tissue engineering construct, endothelial cells were also shown to improve in vivo bone formation.

Author dc:creator

• Pirraco, Rogério

Advisors dc:contributor.advisor

• Reis, R. L.
• Marques, Alexandra P.

Statement dc:rights

• openAccess

Language dc:language.iso

eng

Handle dc:identifier.uri

https://hdl.handle.net/1822/19686

OAI record oai:identifier

http://repositorium.sdum.uminho.pt/oai/request?verb=GetRecord&metadataPrefix=dim&identifier=oai%3Arepositorium.uminho.pt%3A1822%2F19686