Chitra Biswas

Tumor cell stimulation of collagenase production by fibroblasts

Abstract Cocultures of rabbit fibroblasts with mouse tumor cells of epithelial origin showed increased collagenase production when compared to cultures of the individual cells. The stimulatory effect was observed when B-16 melanoma and A-10 adenocarcinoma cells, but not fibroblastic tumor cells or normal epithelial cells, were added to the fibroblasts. The effect was dependent on the cellular ratio between the normal fibroblasts and tumor cells and was mediated by a soluble factor produced by the tumor cells.

Collagenase stimulation in cocultures of human fibroblasts and human tumor cells

Cocultures of human fibroblasts with 3 different human tumor cell lines exhibited high levels of collagenase activity against type I collagen compared to individual cultures of each cell type. For LX-1 lung carcinoma cells maximum stimulation occurred at a ratio of tumor cells to fibroblasts of 0.5:1. These cocultures contained 15-50 times more collagenase activity than the sum of the individual cultures. Conditioned medium from the LX-1 cells stimulated the fibroblasts to an extent similar to that obtained in the cocultures. Conditioned medium from the fibroblasts did not stimulate the tumor cells. Thus human tumor cells secrete a factor(s) which stimulates collagenase production by human fibroblasts.

Partial sequencing and characterization of the tumor cell-derived collagenase stimulatory factor☆

A tumor cell-derived, collagenase stimulatory factor (TCSF), previously isolated and purified from LX-1 human lung carcinoma cells and judged by immunoblotting and SDS-PAGE to contain a single protein of approximately 58 kDa, has been further analyzed for its biological activity and composition. Three significant new findings have been made. First, the biological activity of TCSF preparations was shown definitively to reside in the 58-kDa protein. This was achieved in two ways: (a) a polyclonal antibody was raised against the 58-kDa protein, after excision from an SDS-PAGE gel, and shown to inhibit the stimulation of fibroblast collagenase production by TCSF preparations; (b) the 58-kDa protein was eluted from a transblot of purified TCSF and shown to stimulate fibroblast collagenase production. Second, partial sequencing of the 58-kDa protein revealed no significant homologies with other known collagenase stimulatory factors. Third, purified TCSF was found, on transblotting to Immobilon, to contain a doublet of 58 kDa (TCSF1) and 54 kDa (TCSF2) proteins; the former was present in higher concentration than the latter. N-terminal amino acid sequencing of the two intact proteins and of four corresponding pairs of tryptic peptides derived from the two proteins showed identity in each case, indicating that TCSF1 and TCSF2 are very similar in composition. However, TCSF1 but not TCSF2 stimulated fibroblast collagenase production, confirming that the 58-kDa protein is the major active component of TCSF preparations.

Modulation of the Extracellular Matrix by Tumor Cell-Fibroblast Interactions

Tumor cell invasion and metastasis involve numerous cellular phenomena, e.g., changes in cytoskeletal and cell-surface characteristics, interactions between tumor cells and cells of the vascular and immune systems, and penetration of extracellular matrices. In relation to the last of these phenomena, two major classes of extracellular matrices are traversed by malignant tumor cells during invasion of host tissues, viz., interstitial matrices and basement membranes. It is apparent that successful penetration of extracellular matrices by tumor cells would depend on at least two types of alterations in the structure of these matrices: (1) destruction of preexisting extracellular matrix barriers, and (2) reconstruction of a suitable extracellular environment for cell movement and proliferation. Both of these requirements are, in turn, certain to be multifaceted. However, important in the former would be production of degradative enzymes, such as type I collagenase (Gross et al., 1981; Biswas, 1982a; Woolley, 1982, 1984), type IV collagenase (Liotta et al., 1983; Liotta, 1986), heparanase (Kramer et al., 1982; Nakajima et al., 1983), cathepsin B (Poole et al., 1978;Graf et al., 1981;Sloane et al., 1986), and plasminogen activator (Ossowski and Reich, 1983). Important in the latter is likely to be production of those extracellular macromolecules characteristic of the pericellular milieu of migratory and proliferating cells in embryonic tissues, e.g., hyaluronate (Toole, 1981; Toole et al., 1984) and