Padronização do processo de descelularização para scaffolds biológicos

Abstract

Currently, in the field of regenerative medicine several studies have been developed seeking mechanisms that can treat injuries, as well as replace or regenerate tissues that do not show good functioning. Tissue engineering is based on the use of matrix or scaffold that serves as a support template for cells of interest so that growth factors as well as signaling molecules are preserved. The use of biological scaffolds has been very promising because they are natural matrices, however, there is the possibility of immunogenic reaction and rejection, being necessary the application of techniques for elimination of cells. Thus, the objective of this study was to develop a decellularization protocol that allows the application of biological scaffold from porcine sclera in regenerative medicine. For this, five different protocols were tested with the appropriate modifications based on a model protocol, aiming to standardize an assay that respects the requirements determined by Crapo, Gilbert and Badylak (2011). After verification of the efficiency of the decellularization, it was observed that the Protocols A, B and C (benzonase 1 U/mL, 5 U/mL and 10 U/mL, respectively) were not efficient, presenting genomic DNA content higher than recommended. The Protocols D (DNase/RNase 80 U/mL) and E (DNase/RNase 8 U/mL, washing with water after detergent buffer and rotations of 100 rpm) presented the quantification of extracted DNA in accordance with the limit for efficient decellularization, with absence of residual genomic DNA and histology compatible with decellularized material. However, the E protocol was considered ideal due to the possible elimination of SDS residues, making the biomaterial less cytotoxic and biocompatible. This protocol was reproducible for application with larger amounts of samples, proving to be effective in the application on an industrial scale and can still be considered a fast and cost-effective protocol