Cryopreservation of Cell Sheets of Adipose Stem Cells: Limitations and Successes

Cell Sheets of hASCs (hASCs-CS) have been previously proposed for wound healing applications(1, 2) and despite the concern for production time reduction, the possibility of having these hASCs-CS off-the-shelf is appealing. The goal of this work was to define a cryopreservation methodology allowing to preserve cells viability and the properties CS matrix. hASCs-CS obtained from three different donors were created in UP-cell thermoresponsive dishes(Nunc, Germany) as previously reported(1,2). Different cryopreservation conditions were considered: i)FBS plus DMSO(5% and10%); ii)0.4M of Trehalose plus DMSO (5% and 10%); iii)cryosolution PLL (Akron Biotech, USA); and iv)vitrification. The cryopreservation effect was first assessed for cellular viability by flow cytometry using 7-AAD, and after dissociating the hASCs-CS with collagenase and trypsin-EDTA 0.25%. The expression (RT-PCR) and deposition (western blot and immunocytochemistry) of collagen type I, laminin and fibronectin, and the organization (TEM) of the extracellular matrix was further assessed before and after hASCs-CS cryopreservation to determine a potential effect of the method over matrix composition and integrity. The obtained results confirmed that cell viability is affected by the cryopreservation methodology, as shown before for different CS(3). Interestingly, the matrix properties were not significantly altered and the typical cell sheetâ s easiness of manipulation for transplantation was not lost.

Development of advanced cell/tissue culture systems, based on enhanced polymeric scaffolds and sophisticated bioreactors, for tissue engineering applications

Programa Doutoral em Engenharia Biomédica

Perspetivas de casais e de diretores de biobancos face à criopreservação do sangue do cordão umbilical em Portugal

Dissertação de mestrado em Crime, Diferença e Desigualdade

Current approaches and future perspectives on strategies for the development of personalized tissue engineering therapies

Personalized tissue engineering and regenerative medicine (TERM) therapies propose patient-oriented effective solutions, considering individual needs. Cell-based therapies, for example, may benefit from cell sources that enable easier autologous set-ups or from recent developments on IPS cells technologies towards effective personalized therapeutics. Furthermore, the customization of scaffold materials to perfectly fit a patientâ s tissue defect through rapid prototyping technologies, also known as 3D printing, is now a reality. Nevertheless, the timing to expand cells or to obtain functional in vitrotissue substitutes prior to implantation prevents advancements towards routine use upon patient´s needs. Thus, personalized therapies also anticipate the importance of creating off-the-shelf solutions to enable immediately available tissue engineered products. This paper reviews the main recent developments and future challenges to enable personalized TERM approaches and to bring these technologies closer to clinical applications.