Quantification of adhesion of mesenchymal stem cells spread on decellularized vein scaffold

ABSTRACT Purpose: To evaluate methods that improve adipose-derived stem cells (ASCs) population in decellularized biological venous scaffold for tissue engineering in blood vessels, a model in rabbits. Methods: The ASC was expanded until the third passage. Inferior vena cava (IVC) was submitted to the decellularization process using 1% sodium dodecyl sulfate (SDS) or 2% sodium deoxycholate (SD) to compose 12 study groups (G): pure SD or SDS, exposed or not to 1% TritonX-100 (TX-100) and exposed or not to poly-l'lysine and laminin (PL). Scaffolds were covered with 1 × 105 or 1 × 106 ASCs diluted in 10 μL Puramatrix™. The histological analysis was done by cell counting in hematoxylin and eosin (HE) and nuclei count in immunofluorescence (IF) with 4',6-Diamidine-2'-phenylindole dihydrochloride (DAPI). Results: The study of groups in HE and IF showed similar results. For both analyses,IVC-SD-1 × 106 ASC and IVC-SD-PL-1 × 106 ASC provided the best results. The IF technique showed better sensitivity than HE, with a weak agreement between them. Conclusions: Decellularizing agent and the number of ASC influence scaffolds cellularization response and the best protocols as those ones using SD with or without the addition of PL.

Ultrastructural analysis and residual DNA evaluation of rabbit vein scaffold

Abstract Purpose: To investigate the ultrastructural characteristics and analysis of residual DNA in scaffold models, produced with decellularized vena cava in an experimental model with rabbits. Methods: Three groups were created for ultrastructural and residual DNA analysis: group 1 - control, consisting of samples of vena cava in natura; group 2 - SD, consisting of vein fragments submitted to 2% sodium deoxycholate decellularization by shaking (160rpm - Shaker News Brunswick Scientific®) for 1 hour at controlled temperature shaker at 37°C; group 3 - SDS, consisting of vein fragments submitted to 1% sodium dodecyl sulfate decellularization under the same previous condition, for 2 hours. Results: The ultrastructural matrix of the blood vessel maintained its vintegrity after either decellularization models. The results of the two quantification methods demonstrated a significant decrease in the DNA content of the decellularized vena cava samples as compared to the control samples and, differed statistically from each other, p <0.05. Conclusion: The 2% DS protocol for vein decellularization, in this experimental model, was considered the best protocol because it presented less amount of residual DNA without causing substantial destruction of the extracellular matrix.