Effect of varying cell densities on the rheological properties of the bioink

The rheological characterization of any biopolymer solution is crucial for evaluating the overall printability or injectability of the hydrogel. However, the effect of cells in the cell-laden hydrogel's rheological profile is often ignored. As a result, there is a significant difference in the predicted and experimental outcome in the structural stability of the construct as well as on the cell viability, proliferation, and differentiation potential of the embedded cells. Our present study has addressed the effect of different cell densities (0.1 million cells/ml, 0.5 million cells/ml, 1 million cells/ml and 2 million cells/ml) of TVA-BMSCs on the flow property, modulus behaviour, gelation kinetics and printability of our proprietary silk fibroin-gelatin (5SF-6G) bioink. The cell-laden hydrogels demonstrated a characteristic shear thinning behaviour (low initial viscosity), low storage modulus and increased gelation time when compared to the acellular 5SF-6G hydrogel. The printability analysis also portrayed a square pore geometry with low spreading ratio in 1 million cells/ml encapsulated 5SF-6G hydrogel comparable to the acellular hydrogel. We postulated that incorporation of cells in the bioink interfered with the gelation mechanism of the mushroom tyrosinase in the 5SF-6G bioink by masking the active sites. Additionally, the mechanistic crosstalk between the cell-surface integrins with the cell-attachment motifs of the biomaterial alters the cellular biomechanics of the cell that in-turn profoundly impacts the rheological properties of the polymer blend. Therefore, cell density of 1 million cells/ml was considered the best fit for extrusion-based 3D bioprinting owing to its optimum rheological traits and printability index akin the acellular hydrogel.