Ana M. Frias

Carrageenan-Based Hydrogels for the Controlled Delivery of PDGF-BB in Bone Tissue Engineering Applications

One of the major drawbacks found in most bone tissue engineering approaches developed so far consists in the lack of strategies to promote vascularisation. Some studies have addressed different issues that may enhance vascularisation in tissue engineered constructs, most of them involving the use of growth factors (GFs) that are involved in the restitution of the vascularity in a damaged zone. The use of sustained delivery systems might also play an important role in the re-establishment of angiogenesis. In this study, kappa-carrageenan, a naturally occurring polymer, was used to develop hydrogel beads with the ability to incorporate GFs with the purpose of establishing an effective angiogenesis mechanism. Some processing parameters were studied and their influence on the final bead properties was evaluated. Platelet derived growth factor (PDGF-BB) was selected as the angiogenic factor to incorporate in the developed beads, and the results demonstrate the achievement of an efficient encapsulation and controlled release profile matching those usually required for the development of a fully functional vascular network. In general, the obtained results demonstrate the potential of these systems for bone tissue engineering applications.

Human Adipose Stem Cells Cell Sheet Constructs Impact Epidermal Morphogenesis in Full-Thickness Excisional Wounds

Among the wide range of strategies to target skin repair/regeneration, tissue engineering (TE) with stem cells at the forefront, remains as the most promising route. Cell sheet (CS) engineering is herein proposed, taking advantage of particular cell-cell and cell-extracellular matrix (ECM) interactions and subsequent cellular milieu, to create 3D TE constructs to promote full-thickness skin wound regeneration. Human adipose derived stem cells (hASCs) CS were obtained within five days using both thermoresponsive and standard cell culture surfaces. hASCs-based constructs were then built by superimposing three CS and transplanted into full-thickness excisional mice skin wounds with delayed healing. Constructs obtained using thermoresponsive surfaces were more stable than the ones from standard cell culture surfaces due to the natural adhesive character of the respective CS. Both CS-generating strategies lead to prolonged hASCs engraftment, although no transdifferentiation phenomena were observed. Moreover, our findings suggest that the transplanted hASCs might be promoting neotissue vascularization and extensively influencing epidermal morphogenesis, mainly through paracrine actions with the resident cells. The thicker epidermis, with a higher degree of maturation characterized by the presence of rete ridges-like structures, as well as a significant number of hair follicles observed after transplantation of the constructs combining the CS obtained from the thermoresponsive surfaces, reinforced the assumptions of the influence of the transplanted hASCs and the importance of the higher stability of these constructs promoted by cohesive cell-cell and cell-ECM interactions. Overall, this study confirmed the potential of hASCs CS-based constructs to treat full-thickness excisional skin wounds and that their fabrication conditions impact different aspects of skin regeneration, such as neovascularisation, but mainly epidermal morphogenesis.

Osteogenic differentiation of two distinct subpopulations of human adipose-derived stem cells : an in vitro and in vivo study

The first stem cells considered for the reconstruction of bone were bone marrow mesenchymal stem cells (BMSCs). Subsequently, cells with similar marker expression panel and differentiation potential were found in new sources of cells, such as adipose tissue. This source of stem cells has a promising future in tissue‐engineering applications, considering the abundance of this tissue in the human body, the easy harvesting and the high number of stem cells that are available from such a small amount of tissue. The isolation of the adipose stem cells is generally performed by means of enzymatic digestion of the tissues, followed by a natural selection of the stem cells based on their capacity to adhere to the culture flasks, leading to a quite heterogeneous population. This constitutes a major drawback for the use of these cells, since the heterogeneity of the cell culture obtained can compromise their proliferation and differentiation potential. In the present study we have analysed the in vitro and in vivo behaviour of two selected subpopulations with high osteogenic potential. For this purpose, ASCs

Perivascular-like cells contribute to the stability of the vascular network of osteogenic tissue formed from cell sheet-based constructs

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In vivo biodistribution of carboxymethylchitosan/poly(amidoamine) dendrimer nanoparticles in rats

and ASCs

Synthesis and functionalization of superparamagnetic poly-ɛ-caprolactone microparticles for the selective isolation of subpopulations of human adipose-derived stem cells

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Sulfonic groups induce formation of filopodia in mesenchymal stem cells

subpopulations were isolated and in vitro cultured onto a biodegradable polymeric scaffold, using osteogenic medium, before implantation in a nude mice model. The biodegradable polymeric scaffold used is a fibre‐mesh structure based on a blend of starch and polycaprolatone (SPCL) that has been successfully used in several bone tissue‐engineering studies. The implanted ASCs–scaffold constructs promoted the formation of new bone tissue in nude mice. However, the results obtained show differences in the behaviour of the two ASCs subpopulations under study, particularly regarding their potential to differentiate into the osteogenic lineage, and allowed the indentification of ASCs

Selective Cell Recruitment and Spatially Controlled Cell Attachment on Instructive Chitosan Surfaces Functionalized with Antibodies

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Adipose stem cell-derived osteoblasts sustain the functionality of endothelial progenitors from the mononuclear fraction of umbilical cord blood

as the best subpopulation for bone tissue‐engineering applications. Copyright

Posterior talar process as a suitable cell source for treatment of cartilage and osteochondral defects of the talus.

In recent years several studies have been supporting the existence of a close relationship in terms of function and progeny between Mesenchymal Stem Cells (MSCs) and Pericytes. This concept has opened new perspectives for the application of MSCs in Tissue Engineering (TE), with special interest for the pre-vascularization of cell dense constructs. In this work, cell sheet technology was used to create a scaffold-free construct composed of osteogenic, endothelial and perivascular-like (CD146+) cells for improved in vivo vessel formation, maturation and stability. The CD146 pericyte-associated phenotype was induced from human bone marrow mesenchymal stem cells (hBMSCs) by the supplementation of standard culture medium with TGF-β1. Co-cultured cell sheets were obtained by culturing perivascular-like (CD146+) cells and human umbilical vein endothelial cells (HUVECs) on an hBMSCs monolayer maintained in osteogenic medium for 7 days. The perivascular-like (CD146+) cells and the HUVECs migrated and organized over the collagen-rich osteogenic cell sheet, suggesting the existence of cross-talk involving the co-cultured cell types. Furthermore the presence of that particular ECM produced by the osteoblastic cells was shown to be the key regulator for the singular observed organization. The osteogenic and angiogenic character of the proposed constructs was assessed in vivo. Immunohistochemistry analysis of the explants revealed the integration of HUVECs with the host vasculature as well as the osteogenic potential of the created construct, by the expression of osteocalcin. Additionally, the analysis of the diameter of human CD146 positive blood vessels showed a higher mean vessel diameter for the co-cultured cell sheet condition, reinforcing the advantage of the proposed model regarding blood vessels maturation and stability and for the in vitro pre-vascularization of TE constructs.