Biomedical subjects

C L Mainardi

Publications and source records attributed to C L Mainardi.

48 records · Page 3Linked to original sources

PubMed1980

Degradation of type IV (basement membrane) collagen by a proteinase isolated from human polymorphonuclear leukocyte granules.

A serine esterase with potent proteolytic activity against native bovine lens capsule type IV collagen was isolated and purified from extract of human polymorphonuclear leukocytes (PMN). The type IV collagenolytic activity co-purified with N-t-benzyloxycarbonyl-L-alanine nitroanilidase, and was inhibited by phenylmethanesulfonyl fluoride and N-acetyl-Ala-Ala-Ala-Ala chloromethyl ketone. In addition, the purified enzyme had elastolytic activity, reacted with a specific antibody to PMN elastase, and, therefore, appeared to be identical with this enzyme. A simple, reproducible assay for the detection of type IV collagenase activity using insoluble bovine anterior lens capsule collagen was defined. Sodium dodecyl sulfate polyacrylamide gel electrophoresis revealed that the enzyme released large molecular weight peptides (greater than 30,000) from the insoluble substrate. The enzyme was also active against native, pepsin-solubilized type IV collagen; five reaction products could be identified. These data suggest that PMN elastase may be involved in the degradation of basement membrane collagen in physiologic and pathologic states.

Animals↗

PubMed1980

Collagen-platelet interaction. Type V(A-B) collagen induced platelet aggregation.

One of the more recently discovered collagens, type V (or A-B) collagen, in its native fibillar form mediates human platelet aggregation and the release of serotonin. In agreement with a recent report, it has no detectable effect on human platelets in the soluble or amorphous form. The possibility that the observed results might be due to contaminating interstitial collagens was eliminated by taking advantage of unusual solubility properties of type V collagen. Type V collagen dissolved in 0.1M acetic acid formed native-type fibrils when dialyzed against PBS and amorphous fibrils when dislyzed against 0.05M Tris/0.13M NaCl, pH 7.4, at 4 degrees C. Interstitial collagens remained in solution under both of these conditions. In addition, type V collagen treated with sufficient, purified synovial collagenase to digest all contaminating interstitial collagen retained its platelet-aggregating properties. The purity of type V collagen was confirmed by SDS-PAGE of CNBr digests. These data indicate that the quaternary structure of type V collagen is important in its recognition by platelet membranes.

Amino Acids↗

PubMed1979

Covalent structure of collagen: amino acid sequence of alpha 2-CB5 of chick skin collagen containing the animal collagenase cleavage site.

The amino acid sequence of the 112 residues from the amino terminus of alpha 2-CB5 from chick skin collagen was determined by automated sequential degradation of intact alpha 2-CB5 and several chymotryptic and tryptic peptides. This segment of the peptide includes the site of the action of animal collagenases. As compared to the sequence around the alpha 1 cleavage site, the alpha 2 sequence is notable for the remarkable constancy of the residues to the amino side and the relative abundance of hydrophobic residues to the carboxyl side of the cleavage site, suggesting that these features are important in the recognition by the enzyme. The sequence of this region of the alpha 2 chain is consistent with the Gly-X-Y triplet structure and the preference of certain residues for either the X or Y position in distribution. However, three of the six residues of leucine were found in the Y position rather than the X position. Leucine residues were found only once in the Y position in the alpha 1 (I) chain. This preference does not appear to hold in the alpha 2 chain.

Amino Acid Sequence↗

PubMed1979

Inhibitor of human collagenase from cultures of human tendon.

A potent inhibitor of human collagenases, released from human tendon explants in culture, has been purified and partially characterized. The tendon inhibitor has an estimated molecular weight of 25,000. It is relatively heat-stable but undergoes loss of activity following exposure to trypsin. It inhibits trypsin-activated rheumatoid synovial collagenase as well as the enzyme obtained from polymorphonuclear leukocytes. No inhibition of collagenase from Clostridium histolyticum (clostridiopeptidase A, EC 3.4.24.3) was noted. This collagenase inhibitor may be a factor in the regulation of extracellular connective tissue catabolism.

Cells, Cultured↗

PubMed1979

Collagenase production by rheumatoid synovial cells: morphological and immunohistochemical studies of the dendritic cell.

The dendritic cells of dissociated, adherent rheumatoid synovial cell cultures are recognised by their distinctive morphological features--compact cytoplasm around the nucleus and long, branched cytoplasmic extensions. Such cells usually composed approximately 10% of the total adherent cell population but could vary from as few as 2% to as many as 40% with different synovial specimens. Histological studies have shown the cells to contain many mitochondria and large, spherical cytoplasmic inclusions which often distort the dendritic extensions. Although lysosomes were observed, no evidence for phagocytic activity was obtained. Immunolocalisation studies by means of a monospecific antibody to human collagenase have shown that the dendritic cell attached to a collagenous substratum produces and releases this enzyme in vitro. In contrast collagenase was detected in only a few of the fibroblast- and macrophage-like cells, and it was always intracellular. It is proposed that the dendritic cell may have an important role in the pathophysiology of the rheumatoid joint, particularly with regard to collagenase-mediated cartilage destruction.

Arthritis, Rheumatoid↗

PubMed1979

Rheumatoid arthritis: failure of daily heat therapy to affect its progression.

Seventeen volunteers with symmetrical rheumatoid arthritis (RA) applied heat to 1 hand twice daily for 2 years. Joint swelling, joint tenderness and grip strength were measured at intervals. The proliferative aspect of the disease was assessed by roentgenograms using a scoring system. There was no difference between experimental hand and control hand in any of the factors measured. The patients found the heat soothing and comforting. Thus, daily heat therapy did not accelerate the proliferative lesion in RA and may remain as an adjunct to therapy.

Adult↗

PubMed1978

Collagenase immunolocalization in cultures of rheumatoid synovial cells.

Cultures of rheumatoid synovial cells that have been enzymatically dissociated and are adherent to a culture vessel are morphologically heterogeneous. When these cells are cultured on a collagenous substrate for 2 to 6 days at 37 degrees C in serum-free medium, they produce collagenase. A monospecific antibody to human collagenase has localized the enzyme extracellularly around cytoplasmic extensions of dendritic cells and intracellularly within a few macrophage-like and fibroblast-like cells.

Arthritis, Rheumatoid↗

PubMed1978

Binding of latent rheumatoid synovial collagenase to collagen fibrils.

Collagenase released from rheumatoid synovial cells in culture is in a latent form. Subsequently, it may be activated by limited proteolysis. This study was designed to determine whether latent enzyme could bind to collagen fibrils and await activation. The data showed that latent collagenase bound to fibrils equally well at 24 degrees C and 37 degrees C, but that this represented little more than half the binding achieved by active enzyme at temperatures lower than that at which fibrils can be degraded. Binding was not inhibited by the presence of alpha2 macroglobulin, the principal proteinase inhibitor of plasma which cannot complex with inactive or latent collagenase but readily complexes with active species of enzyme. The data support the hypotheses that inactive forms of collagenase accumulate in tissues by binding to substrate, and that activation by proteases such as plasmin initiates collagen breakdown.

Arthritis, Rheumatoid↗

PubMed1978

Cellular control of collagen breakdown in rheumatoid arthritis.

Remodelling of connective tissue and its destruction in rheumatoid arthritis is related to collagenolysis. Study of collagenase released by rheumatoid synovial cels has indicated that the enzyme is released as latent form from adherent synovial cells in culture. As a latent enzyme it is protected from complexing with alpha2 macroglobulin, the principal proteinase inhibitor. Activation in vivo is very likely caused by proteases which destroy or complex with a portion of collagenase responsible for its latency. Recent data suggest that the latent collagenase is an enzyme-inhibitor complex and not a true zymogen.

Arthritis, Rheumatoid↗

PubMed1978

Activation in vitro of rheumatoid synovial collagenase from cell cultures.

Rheumatoid synovial cells dissociated from matrix and adherent to culture dishes released a latent form of collagenase into culture medium. Previous studies have shown that the latent enzyme does not complex with alpha2-macroglobulin and binds to fibrillar substrate. We now show that serum-free culture medium of the synovial cells contains an inhibitor of collagenase as well as latent enzyme; the two were separated on a column of acrylamide/agarose. Latent collagenase (estimated mol wt 45,000-49,000) was transformed by trypsin to active collagenase of approximately equal to mol wt 33,000. When mixed with inhibitor the active enzyme formed an inactive complex again with approximately equal to mol wt 45,000-49,000. The inhibitor(s) itself was found in one major peak of mol wt 33,000-35,000 and several minor peaks eluting with lower apparent molecular weight. Mersalyl, an organic mercurial compound, effectively activated latent collagenase producing an active enzyme with approximately equal to mol wt 33,000. Bacterial collagenase did not activate latent enzyme. We suggest that latent rheumatoid synovial collagenase, as it is harvested from synovial cells in culture, is an enzyme-inhibitor complex.

Arthritis, Rheumatoid↗

PubMed1977

Endogenous activation of latent collagenase by rheumatoid synovial cells. Evidence for a role of plasminogen activator.

To elucidate the mechanism of synovial damage in rheumatoid arthritis, we studied the activation of latent collagenases released from adherent rheumatoid synovial cells in culture. Latent enzyme was not complexed with alpha2 macroglobulin, the prinicpal proteinase inhibitor in serum, and could be activated by trypsin in the presence of alpha2 macroglobulin if sufficient proteinase was added to saturate inhibitor. Latent collagenase bound half as effectively to collagen fibrils as active enzyme. Plasmin was a threefold better activator of latent enzyme than trypsin and could be generated by addition of plasminogen to synovial-cell cultures. Production of both collagenase and plasminogen activator was inhibited by dexamethasone (10(-9) M). These studies emphasize in importance of control of activation in regulation collagenase activity, It is likely that rheumatoid synovium produces both latent collagenase and plasminogen activator; plasmin is activated from its zymogen, plasminogen, present in inflamed tissues, and in turn activates collagenase.

Arthritis, Rheumatoid↗

PubMedDate not supplied

Proliferative synovitis in hemophilia: biochemical and morphologic observations.

The synovium removed from the knee of a 10-year-old with hemophilia A was characterized morphologically and biochemically. The specimen showed villous hypertrophy with hyperplasia of synovial lining cells which contained abundant intracytoplasmic granules of hemosiderin. Monolayer cultures prepared from enzymatically dispersed tissue were characterized by pigment-laden fibroblast-like cells and round cells. Both explants of synovium and adherent cells secreted a large amount of latent collagenase and neutral proteinase into the culture medium. The secretion of these enzymes dropped sharply and intracellular pigment decreased with passage of these cultures. Lysozyme was secreted by the explants but was not detected in the monolayer culture medium. These data establish the degradative potential of the synovitis found in hemophilia and support the concept that recurrent hemarthrosis without inflammation is sufficient in and of itself to produce proliferative synovitis.

Cells, Cultured↗