Andre C. R. Martins

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.

In vivo short-term and long-term host reaction to starch-based scaffolds.

The implantation of biomaterials may elicit a host response to this foreign body, and the magnitude of that reaction depends on the host and on the implanted material. The aim of this study was to compare the inflammatory response induced by the implantation of starch-based (SPCL) scaffolds in two implantation rat models: subcutaneous (SC) and intramuscular (IM). Moreover, two methodologies, wet spinning (WS) and fibre-bonding (FB), were used to prepare the scaffolds. The short-term inflammatory/immune host reaction was assessed by SC and IM implantations in rats after 1 and 2 weeks, and the long-term host response was addressed after 8 and 12 weeks of SC implantation of both types of SPCL scaffolds in rats. After each time period, the scaffolds, surrounding tissue and nearby lymph nodes were explanted, and used for histological analysis and molecular biology evaluation. The results showed that SPCL-WS scaffolds seem to induce a slight lower inflammatory/immune reaction in both types of implantation models. Nonetheless, comparing the two models, the IM implantation resulted in a slightly higher inflammatory response than the SC implantation with early activation of the lymph nodes. The overall data suggests a good integration of the materials in the host, independently of the tissue location with a normal progress of the reaction for all the conditions.

Cell sheet technology-driven re-epithelialization and neovascularization of skin wounds

Skin regeneration remains a challenge, requiring a well-orchestrated interplay of cell-cell and cell-matrix signalling. Cell sheet (CS) engineering, which has the major advantage of allowing the retrieval of the intact cell layers along with their naturally organized extracellular matrix (ECM), has been poorly explored for the purpose of creating skin substitutes and skin regeneration. This work proposes the use of CS technology to engineer cellular constructs based on human keratinocytes (hKC), key players in wound re-epithelialization, dermal fibroblasts (hDFb), responsible for ECM remodelling, and dermal microvascular endothelial cells (hDMEC), part of the dermal vascular network and modulators of angiogenesis. Homotypic and heterotypic three-dimensional (3-D) CS-based constructs were developed simultaneously to target wound re-vascularization and re-epithelialization. After implantation of the constructs in murine full-thickness wounds, human cells were engrafted into the host wound bed and were present in the neotissue formed up to 14 days post-implantation. Different outcomes were obtained by varying the composition and organization of the 3-D constructs. Both hKC and hDMEC significantly contributed to re-epithelialization by promoting rapid wound closure and early epithelial coverage. Moreover, a significant increase in the density of vessels at day 7 and the incorporation of hDMEC in the neoformed vasculature confirmed its role over neotissue vacularization. As a whole, the obtained results confirmed that the proposed 3-D CS-based constructs provided the necessary cell machinery, when in a specific microenvironment, guiding both re-vascularization and re-epithelialization. Although dependent on the nature of the constructs, the results obtained sustain the hypothesis that different CS-based constructs lead to improved skin healing.