Effects of substrate stiffness on the polarization states of murine bone marrow-derived neutrophils

The substrate stiffness plays an important role in mediating the cellular behavior. Neutrophils predominate the early inflammatory response and initiate the regeneration. The neutrophil activation can be regulated by physical cues. However, it is not known how neutrophils respond to substrate stiffness, which is of significant importance in determining the outcomes of engineered tissue mimics. Herein, a three-dimensional culture system made of hydrogel for bone marrow-derived neutrophils was developed to explore the effects of varying stiffness (1.5, 2.6, and 5.7 kPa) on neutrophil phenotype and polarization states. Overall design: Murine bone marrow-derived neutrophils were encapsuled and cultured in GelMA hydrogel with different stiffnesses (1.5, 2.6, and 5.7 kPa) for 2 days. Then the hydrogel was lysed and released the cells, which were collected. RNA was extracted and differential gene expression was examined by RNAsequencing. Three experimental groups were set up in this study accoding to the substrate stiffness: Low (1.5kpa), middle (2.6kpa), high (5.7kpa). The group Low was set as control. Three replicates of RNA samples per experimental group were analyzed.