Abstract
ABSTRACTPurpose: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.
Highlights
The tissue engineering of blood vessels (TEBV) represents a promising perspective of vascular substitutes in revascularization surgeries, such as coronary or peripheral arterial diseases
Given the histological analysis stained with hematoxylin and eosin (HE), the cell counting showed that, by the generalized linear model (GLM), there was a significant difference between the groups (p < 0.001), and, through the Wald chi-square test for multiple comparisons, the groups with the best cell adhesion were: group 8 (IVC-sodium deoxycholate (SD)-1 × 106 adipose-derived stem cells (ASCs)) and group 12 (IVC-SD-P/L-1 × 106 ASC), without a statistic difference between them (p = 0.526), followed by group 4 (IVC-sodium dodecyl sulfate (SDS)-TX-1 × 106 ASC), which showed statistical lower results than group 8 (SD-1 × 106 ASC) (p = 0.028) and showed no statistical difference to group 12 (IVC-SD-P/L-1 × 106 ASC) (p = 0.117), followed by group 10 (IVC-SD-TX-1 × 106 ASC), which was less than the previous three (p < 0.001)
Given the histological analysis by immunofluorescence (IF), the cellular nuclei counting showed that, by the GLM, there was a significant difference between the groups (p < 0.001), and, through the Wald chisquare test for multiple comparisons, the groups with better cell adhesion were: group 8 Inferior vena cava (IVC)-SD-1 × 106 ASC (p < 0.001), followed by group 12 IVC-SD-P/L-1 × 106 ASC (p < 0.001), by group 4 IVC-SDS-TX-1 × 106 ASC, which showed a statistical difference for the group (p = 0.04), that was group 10 IVC-SD-TX-1 × 106 ASC
Summary
The tissue engineering of blood vessels (TEBV) represents a promising perspective of vascular substitutes in revascularization surgeries, such as coronary or peripheral arterial diseases. On the other hand, concerning the scaffolds, additional factors must be considered, such as the maintenance of a three-dimensional (3D) structure, tissue permeability, sufficient strength to resist blood pressure, elasticity, high durability, facilitation of attachment, migration, proliferation, and cell interaction with adjacent tissues. This material should maintain the antithrombotic activity and provide a microenvironment that imitates the natural architecture of in-vivo tissue for the seeded cells[3,4,5,6]. The discovery of techniques that promote tissue differentiation, with high exhibitions of plasticity, shows that the cultivation of ASCs can contribute a great deal to TEBV7-12
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