Figure 2. Macrophage morphology is dependent upon the stiffness of the culture substrate. (A) Naïve bone-marrow derived macrophage culture on 8kPA gel, 32kPA gel, or tissue culture plastic (TCP). Macrophages cultured on gels of lower stiffnesses tended to exhibit more round morphologies with few to no filapodial extensions (black arrows, 8kPA, 32kPA gel groups). Conversely, macrophages cultured on gels of increased stiffness, such as on tissue culture plastic exhibited more spread morphologies often with several filapodial extensions (white arrows, TCP). (B) Cell area was found to be significantly increased for macrophages cultured on tissue culture plastic as compared to cells cultured on gels of 8kPA or 32kPA stiffness. At least 70 cells were counted per field of view for 2-3 independent replicates. (C) Cells cultured on tissue culture plastic demonstrated a greater percentage of cells exhibiting filipodia per field of view. At least 70 cells were counted per field of view for 2-3 independent replicates. (D) Naïve bone marrow-derived macrophages cultured on 32kPA gel coated with either young (8-16wk) or aged (20-24mo), decellularized cardiac tissue displayed comparable cell morphologies. TCP= tissue culture plastic. Scale bars = 100um. Data reported represents mean values with boxes representing 25%/75% of mean. Error bars represent standard deviation. Black triangles signify data point outliers. ANOVA with Tukey HSD post-hoc analysis. * p<0.05, ** p<0.001.