cathepsin-g and Osteoarthritis

Lubricin (PRG4) is a mucin type protein that plays an important role in maintaining normal joint function by providing lubrication and chondroprotection. Improper lubricin modification and degradation has been observed in idiopathic osteoarthritis (OA), while the detailed mechanism still remains unknown. We hypothesized that the protease cathepsin G (CG) may participate in degrading lubricin in synovial fluid (SF). The presence of endogenous CG in SF was confirmed in 16 patients with knee OA. Recombinant human lubricin (rhPRG4) and native lubricin purified from the SF of patients were incubated with exogenous CG and lubricin degradation was monitored using western blot, staining by Coomassie or Periodic Acid-Schiff base in gels, and with proteomics. Full length lubricin (∼300 kDa), was efficiently digested with CG generating a 25-kDa protein fragment, originating from the densely glycosylated mucin domain (∼250 kDa). The 25-kDa fragment was present in the SF from OA patients, and the amount was increased after incubation with CG. A CG digest of rhPRG4 revealed 135 peptides and 72 glycopeptides, and confirmed that the protease could cleave in all domains of lubricin, including the mucin domain. Our results suggest that synovial CG may take part in the degradation of lubricin, which could affect the pathological decrease of the lubrication in degenerative joint disease.

Topics: Aged; Aged, 80 and over; Cathepsin G; Female; Glycopeptides; Glycoproteins; Glycosylation; Humans; Male; Middle Aged; Osteoarthritis; Proteome; Synovial Fluid

The link protein components of proteoglycan aggregates in adult human articular cartilage show heterogeneity due to proteolysis. Cleavages near the N-terminus of the intact link proteins, before residues 17, 19 and 24, generate three proteins of slightly diminished size (LP3). Cleavages within the N-terminal disulphide-bonded loop, before residues 66 and 73 of the intact link proteins, generate proteins that yield smaller degradation products upon reduction (LP fragments). In vitro, modified link protein components of a similar size to LP3 can be generated by a variety of proteinases, but of the physiologically relevant enzymes only stromelysin, cathepsin B and cathepsin G have the ability to yield modified link proteins with N-termini identical with those observed in situ. None of the proteolytic agents tested was able to produce LP fragments with N-termini identical with those observed in situ, and the majority of proteinases were not able to cleave within the disulphide-bonded loops. Cathepsin L and hydroxyl radicals can cleave within the N-terminal disulphide-bonded loop, and have the potential of initially opening the loop to allow further proteolytic processing by other agents to generate the native cleavage sites.

Topics: Adult; Amino Acid Sequence; Arthritis, Rheumatoid; Cartilage, Articular; Cathepsin B; Cathepsin G; Cathepsin L; Cathepsins; Chymotrypsin; Cysteine Endopeptidases; Endopeptidases; Extracellular Matrix Proteins; Humans; Infant, Newborn; Matrix Metalloproteinase 3; Metalloendopeptidases; Middle Aged; Molecular Sequence Data; Osteoarthritis; Peptide Fragments; Proteins; Proteoglycans; Serine Endopeptidases

Peroxidase-anti-peroxidase (PAP) staining and specific antibodies against cathepsin G and elastase from polymorphonuclear leukocytes (PMN) were applied to pannus-free and microscopically intact superficial articular cartilage. Restricted local deposits containing cathepsin G and elastase were found in three of ten patients with seropositive rheumatoid arthritis (RA), in one of three patients with seronegative RA and in one patient with juvenile chronic arthritis (JCA). Similarly, localized deposits of IgG and C3 were found in the patients with seropositive RA and JCA, but not in the patient with seronegative RA. Adjacent sections exhibited esterase activity in and around the PMN. In proteinase-positive areas from patients with seropositive RA the inhibitors alpha 1-proteinase inhibitor (alpha 1-PI) and alpha 2-macroglobulin (alpha 2-MG) were present in two of three and one of three patients, respectively. In JCA only alpha 1-proteinase inhibitor was present, and in seronegative RA no inhibitors were found. No staining of articular cartilage was observed in a patient with psoriatic arthritis. One of three cases with osteoarthritis exhibited patchy superficial staining for IgG only. In articular cartilage covered by pannus, in three patients with seropositive RA, in one with seronegative RA and in the patient with JCA a few regions with variably dense PMN infiltrates were observed. Cathepsin G, elastase and esterase activity were found in and around the PMN. In one of the three patients with seropositive RA the adjacent cartilage-pannus junction exhibited distinct staining for cathepsin G and elastase, but not for IgG/C3 and proteinase inhibitors.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Arthritis, Juvenile; Arthritis, Rheumatoid; Cartilage, Articular; Cathepsin G; Cathepsins; Complement C3; Histocytochemistry; Humans; Immunoglobulin G; Immunohistochemistry; Neutrophils; Osteoarthritis; Pancreatic Elastase; Serine Endopeptidases

Topics: Aged; alpha 1-Antitrypsin; alpha-Macroglobulins; Arthritis, Rheumatoid; Blood Proteins; Cathepsin G; Cathepsins; Humans; Leukocytes; Middle Aged; Osteoarthritis; Pancreatic Elastase; Protease Inhibitors; Serine Endopeptidases; Synovial Fluid