Comparison of basement membrane matrix degradation by purified proteases and by XXXX cells.

@entity5 @entity3 @entity27238 @entity291 @entity214
We have examined the nature of biochemical degradation of an isolated basement membrane matrix ( @entity291 lens capsule) using different methodologies. The first strategy was quantitation of the release of surface-bound 125I and a second the documentation by @entity214 -PAGE of the appearance of putative cleavage products and the loss of high-molecular-weight components from the matrix. Basement membrane matrix bands resolved on @entity214 -PAGE were identified by their protease sensitivities as well as by Western immunoblots using monoclonal antibodies developed for this study. Radioiodinated components were found predominantly at positions on the gel equivalent to 160-200 kd and 400 kd proteins. Since these labeled moieties were sensitive to bacterial collagenase digestion and stained with anticollagen type IV antibodies, they were determined to represent various configurations of collagen type IV. Several other lower-molecular-weight bands also stained with the anticollagen IV antibodies. Monoclonal antibodies reactive with laminin exhibited a complex staining pattern on the gels, which included the expected 200 and 400 kd components. We confirmed that lens capsule basement membrane contained only a single heparan sulfate glycosaminoglycan species, and @entity5 cell-induced glycosaminoglycan degradation within the basement membrane matrix was detected using @entity27238 electrophoresis. Distinctive putative cleavage products were resolved on @entity214 -PAGE gels from matrices subjected to digestion by a variety of purified proteases as well as by @entity3 cells or their conditioned media. @entity5 cells of different histiotypes produced different characteristic cleavage patterns, suggestive of the existence of several pathways of matrix degradation. Overall, primary @entity5 cells exhibited a greater degradative activity towards the basement membrane matrix than did long-term tissue culture-passaged cells. The same @entity5 cell line could exhibit considerably different patterns of both protein and glycosaminoglycan degradation depending on recent culture history. The relevance of these biochemical studies to the pathogenesis of @entity5 is shown by: 1) the evaluation of degradative activities of B16 @entity5 cell populations exhibiting enhanced lung-colonizing phenotypes, and 2) the ability of a known antimetastatic moiety with antiprotease activity (Haementeria leech species salivary gland extract) to protect matrix components from degradation by @entity5 cell-conditioned medium.
