Cells Assemble Invadopodia-Like Structures and Invade into Matrigel in a Matrix Metalloprotease Dependent Manner in the Circular Invasion Assay

Abstract

The ability of tumor cells to invade is one of the hallmarks of the metastatic phenotype. To elucidate the mechanisms by which tumor cells acquire an invasive phenotype, in vitro assays have been developed that mimic the process of cancer cell invasion through basement membrane or in the stroma. We have extended the characterization of the circular invasion assay and found that it provides a simple and amenable system to study cell invasion in matrix in an environment that closely mimics 3D invasion. Furthermore, it allows detailed microscopic analysis of both live and fixed cells during the invasion process. We find that cells invade in a protease dependent manner in this assay and that they assemble focal adhesions and invadopodia that resemble structures visualized in 3D embedded cells. We propose that this is a useful assay for routine and medium throughput analysis of invasion of cancer cells in vitro and the study of cells migrating in a 3D environment.

Podosomes in space

“…133 It is also an intriguing speculation that podosomes and invadopodia, which present as similar but distinct structures in 2D systems, 39 might show convergence in their morphology and properties in cells that are present in a 3D context. The more invadopodia-like morphology and localization of podosomes in 3D, as well as the relevance of 3D podosomes and invadopodia for 3D infiltration of cells, 16,56,134,135 should provide a sufficient basis to stimulate respective research.…”

Invadopodia and basement membrane invasion in vivo

“…[34][35][36][37] Reflecting the growing interest in these structures, more than half of the approximately 350 scholarly articles on invadopodia have been published within the last 3 y. Although recent advances have been made in analyzing invadopodia-like protrusions in 3D culture conditions, 19,38 most mechanistic studies on invadopodia have been performed with cancer cell lines in 2D cell culture environments. The advantages in imaging, biochemical approaches, and genetic and pharmacological perturbations in 2D conditions have led to remarkable advances in our understanding of invadopodia.…”

Three-dimensional cell migration does not follow a random walk

“…However, cell migration in vivo often forces cells to remodel, exert pulling forces on, and move through a 3D collagen I-rich matrix. Recent work has demonstrated that mechanisms of 3D migration are often different from their 2D counterparts (9)(10)(11)(12)(13)(14)(15). Migration on 2D dishes, which induces a basal-apical polarization of the cell, is driven by actomyosin contractility of stress fibers between large focal adhesions and the formation of a wide lamellipodium terminated by thin filopodial protrusions at the leading cellular edge (4,16).…”