"In vitro" digestion of intact bovine lens capsules by four human lysosomal cysteine-proteinases.
We have examined the biochemical degradation of an isolated basement membrane matrix (bovine lens capsule) by human liver cathepsins B, H and L and the cathepsin B-like proteinase from malignant ascitic fluid. This study was carried out using two different methods: The first strategy was to follow the liberation of soluble proteins and peptides as a function of time at different pHs. Then the digestion products were characterized, as collagen IV, fibronectin and laminin fragments, using monospecific polyclonal antibodies and a quantitative dot-blot analysis. From these results, the ability of the four proteinases to digest "in vitro" intact bovine lens capsule in the physiological pH range is demonstrated. Cathepsin L is the most powerful against the three membrane components studied. As shown by electroelution and immunochemical quantitation, the digestion would be a consequence of proteinases binding to the capsule. With intact basement membrane as a substrate a "in vitro" molecular analysis of this digestion process was possible by these methods. On this basis, the "in vivo" secretion of cysteine proteinases during malignancy would be related to the local basement membrane dissolution associated with tumor invasion.
Laboratoire de Biochimie, Faculté de Médecine Broussais Hôtel Dieu, Paris, France.
N
N
Guinec
M
M
Pagano
V
V
Dalet-Fumeron
R
R
Engler
D000818:Animals; D001485:Basement Membrane / Q000378:metabolism; D002401:Cathepsin B / Q000032:analysis; D002417:Cattle; D003094:Collagen / Q000378:metabolism; D003546:Cysteine Endopeptidases / Q000378:metabolism*; D005353:Fibronectins / Q000032:analysis; D006801:Humans; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D015151:Immunoblotting; D007120:Immunochemistry; D007797:Laminin / Q000032:analysis; D007908:Lens, Crystalline / Q000378:metabolism*; D008247:Lysosomes / Q000201:enzymology*; D008969:Molecular Sequence Data; D013050:Spectrometry, Fluorescence; D013056:Spectrophotometry, Ultraviolet
D005353:Fibronectins; D007797:Laminin; D003094:Collagen; D003546:Cysteine Endopeptidases; D002401:Cathepsin B