External-Force-Offset Effects of ECM Coating Layers on hMSCs Subjected to External Physical Force.

Abstract

Mesenchymal stem cells (MSCs) used for cell-delivery-based therapy also undergo considerable external stresses upon entering the recipient site in the body. Here, we sought to develop a cell-protective barrier on the MSC surface that protects against stress-induced damage from physical external stresses. The barrier was fabricated from gelatin and hyaluronic acid (HyA) using a layer-by-layer (LbL) technique. In addition to assessing the stability and biological properties of extracellular matrix (ECM)-coated human bone marrow-derived MSCs (hMSCs) produced using the LbL, we also evaluated the cell-protective effects of this coating against 2 external stresses: low-attachment conditions and mechanical force induced by injection. Cell biological and morphological surface changes accompanying cell surface coating were analyzed using fluorescence-activated cell sorting and scanning electron microscopy. Viability and cell cycle characteristics were not substantially different between bare hMSCs and ECM-coated hMSCs with different numbers of layers after 7 days in culture. Stemness was also maintained, as reflected in >97.3% expression of positive markers and <0.5% expression of negative markers in 6-layered ECM-coated hMSCs, termed ECM-hMSCs. ECM-hMSCs showed 62.1% decrease in cell damage and 50.6% increase in DNA content after 3 days under low-attachment conditions. In addition, ECM-hMSCs injected at 100 and 200 kPa showed 27.2% and 41.8% higher viability, with damaged cells decreased by 54.9% and 45.6%, respectively, compared to bare hMSCs. These results show that LbL coating of hMSCs with gelatin and HyA does not impair the function of hMSCs and can physically protect cells from low-attachment conditions and the mechanical force associated with injection.