In mammary
tumors, intravital imaging techniques have uncovered an essential role for macrophages during
tumor cell invasion and
metastasis mediated by an
epidermal growth factor (
EGF) / colony stimulating factor-1 (CSF-1) paracrine loop. It was previously demonstrated that mammary
tumors in mice derived from rat
carcinoma cells (MTLn3) exhibited high velocity migration on extracellular matrix (ECM) fibers. These cells form paracrine loop-dependent linear assemblies of alternating host macrophages and
tumor cells known as "streams." Here, we confirm by intravital imaging that similar streams form in close association with ECM fibers in a highly metastatic patient-derived orthotopic mammary
tumor (TN1). To understand the in vivo cell motility behaviors observed in streams, an in vitro model of fibrillar
tumor ECM utilizing adhesive 1D micropatterned substrates was developed. MTLn3 cells on 1D
fibronectin or
type I collagen substrates migrated with higher velocity than on 2D substrates and displayed enhanced lamellipodial protrusion and increased motility upon local interaction and pairing with bone marrow-derived macrophages (BMMs). Inhibitors of
EGF or
CSF-1 signaling disrupted this interaction and reduced
tumor cell velocity and protrusion, validating the requirement for an intact paracrine loop. Both TN1 and MTLn3 cells in the presence of BMMs were capable of co-assembling into linear arrays of alternating
tumor cells and BMMs that resembled streams in vivo, suggesting the stream assembly is cell autonomous and can be reconstituted on 1D substrates. Our results validate the use of 1D micropatterned substrates as a simple and defined approach to study fibrillar ECM-dependent cell pairing, migration and relay chemotaxis as a complementary tool to intravital imaging.