PHAGOCYTOSIS OF CROSS-LINKED GELATIN MATRIX BY HUMAN BREAST-CARCINOMACELLS CORRELATES WITH THEIR INVASIVE CAPACITY

During invasion and metastasis, cancer cells interact closely with the extracellular matrix molecules by attachment, degradation, and migrat ion, We demonstrated previously the local degradation of fluorescently labeled gelatin matrix by cancer cells at invasive membrane protrusio ns, called invadopodia. Using the newly developed quantitative fluores cence-activated cell sorting-phagocytosis assay and image analysis of localized degradation of fluorescently labeled matrix, we document her e that degradation and site-specific removal of cross-linked gelatin m atrix is correlated with the extent of phagocytosis in human breast ca ncer cells, A higher phagocytic capacity is generally associated with increasing invasiveness, documented in other invasion and motility ass ays as well, Gelatin phagocytosis is time and cell density dependent, and it is mediated by the actin cytoskeleton, Most of the intracellula r gelatin is routed to actively acidified vesicles, as demonstrated by the fluorescent colocalization of gelatin with acidic vesicles, indic ating the intracellular degradation of the phagocytosed matrix in lyso somes, We show here that normal intracellular routing is blocked after treatment with acidification inhibitors, In addition, the need for pa rtial proteolytic degradation of the matrix prior to phagocytosis is d emonstrated by the inhibition of gelatin phagocytosis with different s erine and metalloproteinase inhibitors and its stimulation by conditio ned medium containing the matrix metalloproteinases MMP-2 and MMP-9. O ur results demonstrate that phagocytosis of extracellular matrix is an inherent feature of breast turner cells that correlates with and may even directly contribute to their invasive capacity, This assay is use ful for screening and evaluating potential anti-invasive agents becaus e it is fast, reproducible, and versatile.