Micropatterning a Chemotactic Gradient in a Cell-Laden Hydrogel to Direct Cell Migration

Microfluidic systems enable the creation of highly controlled chemical gradients through modulated flow and diffusional profiles to spatially direct cell migration events. These are usually conducted in closed systems; wherein accurate flow control is possible. However, for in vitro 3D culture systems, cells must be cultured in millimeter-scale hydrogel slabs to recapitulate the cell microenvironment, which is difficult to support in a closed microfluidic system. Open microfluidic systems can ameliorate this problem. But flow control in an open system is much harder to achieve. A tailor-made hydrogel can be patterned at the millimeter scale and integrated with a fluidic system to study cellular responses to a chemotactic gradient. Combining such a system with live-cell imaging would allow for the spatiotemporal quantification of the chemical gradient and the cellular reaction. Here, we present the design, patterning, and integration techniques to create a cell-laden hydrogel in an open-top culture. Bounded by adjoining open microfluidic channels, the system generates a chemotactic gradient to direct U87 glioma cell migration. The hydrogel is fabricated through a positive-negative-positive process with a positive 3D-printed mold, negative polydimethylsiloxane (PDMS) mold, and photo-crosslinking of the hydrogel structure. The composite hydrogel is composed of gelatin methacrylate (GelMA) and hyaluronic acid (HA) to mimic the properties of brain tissue. Stromal-derived factor 1 (SDF-1), or CXCL12, will be utilized as the chemoattractant for gradient generation across a specific section of the hydrogel. Fluorescence and brightfield imaging will be employed to quantify cellular migration and signaling.