3D culture of murine adipose-derived multipotent mesenchymal stromal cells in hydrogel based on carbomer 974P

Abstract

Actual issues during tissue regeneration are to ensure the survival of transplanted cells at the site of their application and further activity, especially in case of local pathological alterations such as inflammation and ischemia. For this purpose, the matrices that can not only fill the defects of tissues, but also be scaffolds for cells are developed.The aim of this study was to evaluate the effectiveness of 3D cultivation of murine adipose-derived multipotent mesenchymal stromal cells (MSMCs) in hydrogel based on carbomer 974P.Materials and methods. MSMCs were obtained from the adipose tissue of FVB-Cg-Tg(GFPU)5Nagy/J mice transgenic for GFP gene. The cells were phenotyped by flow cytometry and directly differentiated into osteogenic and adipogenic direction to confirm multipotent phenotype. MMSCs were cultured and directly differentiated into osteogenic direction in three-dimensional hydrogel scaffolds. For hydrogel preparation we used carbomer 974P with composition of glycerol, propylene glycol, triethylamine and agarose in original proportion.Results. The three-dimensional hydrogel based on carbomer 974P for the further engraftment with MMSCs was obtained. Modified protocols for the preparation of hydrogels based on carbomer and agarose and their rehydration by culture media for the 3D cultivation of adipose-derived MMSCs have been developed. The optimal concentration of MSMCs and the injection method for engraftment of hydrogels of the required form and size are selected. It was shown that adipose-derived MMSCs in 3D carbomer hydrogel preserve the potential of directed osteogenic differentiation.Conclusion. Three-dimensional hydrogel based on carbomer 974P is capable to support cells, provide the necessary cytoarchitectonics, maintain intercellular interactions, which can promote further long-term survival and specialization of graft.

Adipose-Derived Stem Cells Reduce Lipopolysaccharide-Induced Myelin Degradation and Neuroinflammatory Responses of Glial Cells in Mice

“…There was detected the high level of CD44, CD73, CD90 expression, while the percentage of cells with hematopoietic markers CD45 and CD117 was less than 2%. The expression of CD34 at the early passages ranged from 8–12%, which is typical for ASCs [ 14 ]. According to these results, as well as confirmed potential of directed differentiation into the osteogenic and adipogenic direction, the obtained ASC culture met the minimal criteria for defining multipotent MSCs proposed by the International Society for Cellular Therapy [ 15 ].…”

“…Passaging was carried out at reaching 80% confluence of a monolayer. On the passage two, multiparameter immunophenotyping using flow cytometry as well as directed differentiation into osteogenic and adipogenic directions were performed, as described earlier [ 14 ].…”