Use of a reconstituted basement membrane to measure cell invasiveness and select for highly invasive tumor cells.

Abstract

Malignant cells must traverse basement membranes during their migration to sites distant from the primary tumor. Since basement membranes are thought to be a critical barrier to the passage of tumor cells, we have constructed a model basement membrane-stromal matrix consisting of laminin and type IV collagen reconstituted onto a disk of type I collagen for use in an in vitro assay of invasiveness. Metastatic tumor cells and leukocytes are able to cross this barrier, whereas nonmetastatic tumor cells, fibroblasts, and epidermal cells cannot penetrate it. Those tumor cells that penetrate the barriers were found, when isolated and subcultured, to be more invasive and to produce more metastases than the parental population. This assay system should be useful for studying the invasiveness of tumor cells and for isolating highly invasive variants.

Active Detachment Involves Inhibition of Cell-Matrix Contacts of Malignant Melanoma Cells by Secretion of Melanoma Inhibitory Activity

“…In this assay melanoma cells are placed into the upper chamber, which is separated from a second compartment by a filter coated with different ECM molecules. The number of cells that attach to and invade through the filters can be quantified reliably (Albini et al, 1987;Terranova et al, 1986). The results from both Boyden chambers and attachment assays parallel closely and reveal consistently that MIA inhibits interaction of melanoma cells with fibronectin, laminin, tenascin, and matrigel ( Fig.…”

“…Invasion assays were performed in Boyden chambers containing polycarbonate filters with 8-m pore size (Costar, Bodenheim, Germany) essentially as described previously (Albini et al, 1987;Terranova et al, 1986;Wach et al, 1996). Filters were coated with a commercially available reconstituted basement membrane (Matrigel, diluted 1:3 in H 2 O; Becton Dickinson #356235, Heidelberg, Germany), fibronectin (1 g/ cm 2 , 12.5 nmol/filter; Becton Dickinson #40008), laminin-1 (5 g/cm 2 , 5 nmol/filter; Becton Dickinson #40232), tenascin (1 g/cm 2 , 1 nmol/filter; Becton Dickinson #40239), collagen type I (1 g/cm 2 , 6.5 nmol/filter; Becton Dickinson #40243), type II (1 g/ cm 2 , 6.5 nmol/filter; Becton Dickinson #40234), and type IV (1 g/cm 2 , 5 nmol/filter; Becton Dickinson #40233), vitronectin (50 ng/cm 2 , 1 nmol/filter; Becton Dickinson #40238), or HSPG (2 g/cm 2 , 1 nmol/filter; Becton Dickinson #40250).…”

Effects of extracellular matrix analogues on primary human fibroblast behavior

“…A more complete suite of matricellular proteins and growth factors is provided by Matrigel ™ , an ECM preparation extracted from Engelbreth-Holm-Swarm (EHS) mouse sarcoma [14]. Matrigel ™ has been successfully utilized for a variety of applications, such as cell growth and differentiation, angiogenesis and invasion assays, and promotes a natural cell morphology and behavior [14][15][16][17][18]. However, limited availability, batch-to-batch variability, pathogen transmission, immunogenicity, technical challenges in handling, and the inability to experimentally vary composition and compliance suggested the need for a more versatile ECM equivalent.…”

Colon cancer cells with high invasive potential are susceptible to induction of apoptosis by a selective COX‐2 inhibitor

“…In vitro Matrigel invasion assay was performed according to a previously published procedure. 18) Briefly, PVP (polyvinyl polymer)-free polycarbonate transwells (Nucleopore; Corning Costar Corp., Acton, MA), pore size 12 µm, were coated with 1.05 mg/ml of Matrigel matrix for 1 h at 37°C. Human SW620, HT-29, HT-29/Inv1, HT-29/Inv2, and HT-29/Inv3 cells were placed respectively on the top of the transwell at a dose of 5×10 4 .…”