Oral bone lost is an important issue in maxillo-facial and dental surgery. In our previous study we have characterized human mesenchymal stem cells from Bichat’s fat pad, a convenient tissue for dentists, and proposed them for future therapies of periodontal defects and oral bone regeneration. Considering the need for preclinical study and the swine as an accepted animal model in tissue engineering applications, here we compared the swine Adipose-derived Stem Cells (ASCs) from Buccal fat pad and from subcutaneous adipose tissue both alone and in combination with synthetic scaffolds. 5.5x104±3.3x104 and 3.0x104±9.3x103 cells/ml were isolated from interscapular subcutaneous adipose tissue (ScI) and buccal fat pad (BFP) of 6 swine. All the ASCs, with MSC fibroblast-like morphology, proliferated costantly during culture with an average doubling time (DT) of 82.9±11.5 hours and 72.5±8.2 hours for ScI- and BFP-ASCs, respectively, and showed a high clonogenic ability (+10.1±1.4% for ScI- and +8.9±1.5% for BFP-ASCs). By FACS analysis both cell populations appeared similar in size and granularity, and expressed CD90, whereas the CD14, CD45 and CD271 were not detectable. Furthermore, after osteogenic induction, all the ASCs showed an increase of either collagen and calcified extracellular matrix (ECM) production of about 87% and 118% for ScI-ASCs, and of about 254% and 116% for BFP-ASCs, compared to undifferentiated cells. Alkaline phosphatase activity (+126% and +201% for ScI- and BFP-ASCs, respectively) and osteonectin expression (+336% for ScI- and +306% for BFP-ASCs) were also upregulated, confirming their capacity to osteo-differentiate. When induced, both cell types were also able to differentiate towards the adipogenic and chondrogenic lineages, as revealed by lipid vacuoles production and GAGs deposition. Additionally, the osteoinduction of cells in the presence of titanium disks and silicon carbide–plasma-enhanced chemical vapor deposition (SIC-PECVD) fragments, significantly increased the amount of calcified ECM compared to control cells; moreover, titanium is osteoinductive per se on ASCs (+91 and +284% for ScI- and BFP-ASCs). Finally, porcine autologous or heterologous sera, did not improve cell growth compared to standard condition. No significant difference between cells from the different harvesting sites was observed, concerning all the features described. In conclusions, swine buccal fat pad contains mesenchymal stem cells, and we suggest them for preclinical studies of periodontal and bone defects regeneration.
Study in vitro on porcine stem cells from buccal fat pad and subcutaneous adipose tissue for periodontal and oral bone / L.M.J. Ferreira, S. Niada, E. Arrigoni, A.T. Brini. ((Intervento presentato al convegno AICC/GISM tenutosi a Brescia nel 2013.
Study in vitro on porcine stem cells from buccal fat pad and subcutaneous adipose tissue for periodontal and oral bone
L.M.J. Ferreira;S. Niada;E. Arrigoni;A.T. Brini
2013
Abstract
Oral bone lost is an important issue in maxillo-facial and dental surgery. In our previous study we have characterized human mesenchymal stem cells from Bichat’s fat pad, a convenient tissue for dentists, and proposed them for future therapies of periodontal defects and oral bone regeneration. Considering the need for preclinical study and the swine as an accepted animal model in tissue engineering applications, here we compared the swine Adipose-derived Stem Cells (ASCs) from Buccal fat pad and from subcutaneous adipose tissue both alone and in combination with synthetic scaffolds. 5.5x104±3.3x104 and 3.0x104±9.3x103 cells/ml were isolated from interscapular subcutaneous adipose tissue (ScI) and buccal fat pad (BFP) of 6 swine. All the ASCs, with MSC fibroblast-like morphology, proliferated costantly during culture with an average doubling time (DT) of 82.9±11.5 hours and 72.5±8.2 hours for ScI- and BFP-ASCs, respectively, and showed a high clonogenic ability (+10.1±1.4% for ScI- and +8.9±1.5% for BFP-ASCs). By FACS analysis both cell populations appeared similar in size and granularity, and expressed CD90, whereas the CD14, CD45 and CD271 were not detectable. Furthermore, after osteogenic induction, all the ASCs showed an increase of either collagen and calcified extracellular matrix (ECM) production of about 87% and 118% for ScI-ASCs, and of about 254% and 116% for BFP-ASCs, compared to undifferentiated cells. Alkaline phosphatase activity (+126% and +201% for ScI- and BFP-ASCs, respectively) and osteonectin expression (+336% for ScI- and +306% for BFP-ASCs) were also upregulated, confirming their capacity to osteo-differentiate. When induced, both cell types were also able to differentiate towards the adipogenic and chondrogenic lineages, as revealed by lipid vacuoles production and GAGs deposition. Additionally, the osteoinduction of cells in the presence of titanium disks and silicon carbide–plasma-enhanced chemical vapor deposition (SIC-PECVD) fragments, significantly increased the amount of calcified ECM compared to control cells; moreover, titanium is osteoinductive per se on ASCs (+91 and +284% for ScI- and BFP-ASCs). Finally, porcine autologous or heterologous sera, did not improve cell growth compared to standard condition. No significant difference between cells from the different harvesting sites was observed, concerning all the features described. In conclusions, swine buccal fat pad contains mesenchymal stem cells, and we suggest them for preclinical studies of periodontal and bone defects regeneration.Pubblicazioni consigliate
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