Gelatin-Hyaluronan Click-Crosslinked Cryogels Elucidate Human Macrophage Invasion Behavior

Hydrogel models of metastasis traditionally focus on the invasion of cancer cells; however, other cells in the tumor microenvironment that are associated with metastasis also have the ability to migrate. Macrophage phenotype plays a key role in the tumor microenvironment, yet understanding their migration within tunable 3D in vitro models has been limited. To gain a greater understanding of macrophage invasive behavior, stable and transparent oxime‐crosslinked cryogels comprised of click-crosslinked gelatin-oxyamine and hyaluronan-aldehyde (GELox-HAa) are synthesized. Fibronectin-derived, oxyamine-modified PHSRN-RGDSP peptides are incorporated to further mimic the tumor extracellular matrix without impacting cryogel mechanics. It is found that primary human macrophages migrate to a greater depth in cryogels with greater porosity and pore size. To better understand the mechanism of migration, cells are treated with either inhibitors of matrix metalloproteinases (MMPs) or rho-associated protein kinase (ROCK) and a predominantly MMP‐mediated mechanism of invasion is found. Macrophage polarization studies reveal that anti‐inflammatory, interleukin-4/13 (IL4/IL13)-treated macrophages migrate through cryogels to a greater extent than pro-inflammatory, interferon-gamma/lipopolysaccharide (IFNγ/LPS)-treated cells. Interestingly, polarized macrophages move through cryogels using a combination of amoeboid and mesenchymal migration. These findings of macrophage invasion in this cryogel platform set the stage for their further study in a biomimetic tumor microenvironment.