Abstract
A collagenase and a neutral protease have been insolated and characterized from primary cultures obtained from rheumatoid subcutaneous nodules. Release of both active enzymes was maximal between the 3rd and 7th days of culture and was stimulated by the presence of small amounts of colchicine (0.1 mug/ml) added to the culture medium. Both the protease and the collagenase from nodule tissue were active at physiologic pH and were inhibited by chelating agents, sulfhydryl compounds, and 1:40 dilutions of human serum. Both enzymes appeared to have a molcular size equivalent to similar enzymes found in cultures of rheumatoid synovium. The nodule collagenase was purified by chromatography on molecular sieve columns followed by affinity chromatography. The pure enzyme cleaved collagen in solution at 24 degrees C at the locus common for mammalian collagenases to act: three quarters of the distance from the amino-terminus. Under the same conditions the purified enzyme cleaved gelatin (denatured collagen) at the same locus. It is likely therefore that the collagenase in rheumatoid connective tissues functions to produce the initial cleavage of collagen and that after the initial reaction products are denatured, proteases digest them into smaller polypeptides more rapidly than does the collagenase itself. Since rheumatoid nodules grow centrifugally at the expense of the palisading fibroblast layer it seems possible that the central necrotic areas are caused by release of collagenase and protease from the highly cellular palisading zone resulting in the destruction of the extracellular collagen matrix.