Modulation of mesenchymal stromal cells properties by the microenvironment in 3D culture

Abstract

The aim of the research was to compare the shape, viability, metabolic and proliferative activity of mesenchymal stromal cells (MSCs) during cultivation in hydrogels and macroporous scaffolds. Materials and methods. Human adipose tissue MSCs were isolated from lipoaspirates of healthy adult donors after obtaining informed consent. Hydrogels were obtained from platelet-poor human blood plasma and alginate polymer, cross-linked with calcium ions in microspheres. Macroporous scaffolds were prepared from plasma by the cryotropic gelation method. Morphology and viability of cells within carriers were assessed using vital dyes. Metabolic and proliferative activity of MSCs was studied by the Alamar Blue test on the 1st, 3rd and 7th day of 3D culturing. Results. Three-dimensional blood plasma scaffolds had a branched pore structure with a size sufficient for cell proliferation and migration. When plasma proteins were cross-linked with L-cysteine, almost all MSCs were viable, attached to the pore surface, spread and proliferated, filling carrier cavities. In plasma hydrogels, MSCs occupied spaces and acquired a fibroblast-like morphology, maintaining viability. In alginate microspheres, MSCs were uniform distributed throughout the gel volume, kept their spherical shape, but had high viability. The highest metabolic activity of MSCs was observed in macroporous scaffolds, the lowest one in alginate microspheres. During cultivation, the activity of cells in macroporous scaffolds and plasma hydrogels increased significantly, which indirectly indicated the proliferation processes. Conclusions. Properties of MSCs during 3D cultivation significantly depend on the microenvironment: in blood plasma carriers, cells acquire a fibroblast-like morphology and proliferate, while in alginate microspheres, they remain spherical and do not proliferate.