Microfluidic gradients reveal enhanced neurite outgrowth but impaired guidance within 3D matrices with high integrin ligand densities

Abstract

The density of integrin-binding ligands in an extracellular matrix (ECM) is known to regulate cell migration speed by imposing a balance of traction forces between the leading and trailing edges of the cell, but the effect of cell-adhesive ligands on neurite chemoattraction is not well understood. A platform is presented here that combines gradient-generating microfluidic devices with 3D protein-engineered hydrogels to study the effect of RGD ligand density on neurite pathfinding from chick dorsal root ganglia-derived spheroids. Spheroids are encapsulated in elastin-like polypeptide (ELP) hydrogels presenting either 3.2 or 1.6 mM RGD ligands and exposed to a microfluidic gradient of nerve growth factor (NGF). While the higher ligand density matrix enhanced neurite initiation and persistence of neurite outgrowth, the lower ligand density matrix significantly improved neurite pathfinding and increased the frequency of growth cone turning up the NGF gradient. The apparent trade-off between neurite extension and neurite guidance is reminiscent of the well-known trade-off between adhesive forces at the leading and trailing edges of a migrating cell, implying that a similar matrix-mediated balance of forces regulates neurite elongation and growth cone turning. These results have implications in the design of engineered materials for in vitro models of neural tissue and in vivo nerve guidance channels.

Keywords: 3D migration; chemotaxis; elastin-like polypeptides; microfluidics; nerve growth factor; neurite pathfinding.