Novel Adhesive Silicone‐Based Cell Culture Systems for Differentiation of Human Adipose Tissue‐Derived Mesenchymal Stromal Cells

Abstract

We established a protocol for the differentiation of hepatocyte‐like cells derived from human adipose tissue mesenchymal stromal cells (hAT‐MSC) in classical two‐dimensional cell culture, yet the physiological functions can be optimized. Classical two‐dimensional cell culture systems exhibit a high modulus of elasticity, yet, without representing the stiffness of human organs. A novel adhesive silicone was utilized to improve the elasticity of the cell culture substratum and to simulate the stiffness of organ tissue.Aim of the study was to analyse the differentiation ability of hAT‐MSC into cells of meso‐ and endodermal lineages on collagen‐coated adhesive silicone layers with low elastic modulus (500 Pa) in comparison with classical 2D cell cultures on polystyrene (103 MPa). hAT‐MSC were differentiated by a two‐step protocol and the progress was verified 8 and 21 days after initiation by the detection of specific histochemical hallmarks and biochemical characteristics.hAT‐MSC attached to the collagen‐coated adhesive silicon and formed cell layers. The formation of spheroids was observed with ongoing culture only in undifferentiated and hepatogenic differentiation, to lower extent also during osteogenic differentiation. Specific stainings on undifferentiated cells on adhesive silicone versus classical cell culture revealed no evident differences in fat content by Oil‐Red‐O staining (adipogenic), alkaline phosphatase by BCIP/NBT staining (osteogenic), and glycogen by PAS staining (hepatogenic). Differentiated cells cultured on adhesive silicone showed more pronounced staining under all three differentiation directions as compared to classically cultured cells. Fat deposit quantification of adipogenic differentiated cells displayed a 2.9‐fold increase in cells cultured on adhesive silicon as compared to the 1.9‐fold increase in cells classically cultured. The urea production rate [μg/10.000 cells*24h] was significantly higher in the culture on adhesive silicone as compared to the classical culture after 21 days of hepatogenic differentiation: 51 ± 13 vs. 0.33 ± 0.04.The results show the appropriateness of adhesive collagen‐coated silicone for the various differentiation directions of hAT‐MSC. The silicone supports the potential of hAT‐MSC to growth as spheroids. Thus, the collagen‐coated adhesive silicone represents an important tool for the development of in vitro culture systems mimicking E‐moduli of tissues and improving physiological functions.Support or Funding InformationThe study was supported by the European Regional Development Fund and the Free State of Saxony (SAB) 100282466 to J.H. and P.S.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.