Proteolytic degradation of the glomerular basement membrane and immunochemical characterization of split products

Glomerular basement membranes (GBM) were isolated and subjected to enzymatic degradation with the protease trypsin (Serva), chymotrypsin (Serva), papain (Sigma), pepsin (Serva) and collagenase (Worthington) as well as a lysosomal preparation from glass adherent rat blood and peritoneal exudate cells. Split products were characterized by immunoelectrophoresis and cellulose acetate electrophoresis. Urine was obtained from healthy rats and rats with Masugi's experimental glomerulonephritis, dialyzed and concentrated and applied on immunoelectrophoresis, using anti-GBM antibody from rabbit. Urinary GBM split products from healthy and nephrotic rats showed two precipitation lines like digestion products obtained after chymotrypsin degradation. This finding was supported by characterizing individual antigenic degradation products obtained after inhibition of GBM degradation by the lysosomal preparation with ethylenediaminetetraacetic acid and Trasylol alone and in combination, as well as with o-phenanthrolin. It is concluded that GBM-antigens excreted into urine indicate limited digestion of GBM by chymotrypsin-type protease.