Abstract
Lubricin, encoded by the gene PRG4, is the principal lubricant in articulating joints. We immunized mice genetically deficient for lubricin (Prg4-/-) with purified human lubricin, and generated several mAbs. We determined each mAb’s binding epitope, sensitivity, and specificity using biologic samples and recombinant lubricin sub-domains, and we also developed a competition ELISA assay to measure lubricin in synovial fluid and blood. We found the mAbs all recognized epitopes containing O-linked oligosaccharides conjugated to the peptide motif KEPAPTTT. By western blot, the mAbs detected lubricin in 1 μl of synovial fluid from several animal species, including human. The mAbs were specific for lubricin since they did not cross-react with other synovial fluid constituents from patients with camptodactyly-arthropathy-coxa vara-pericarditis syndrome (CACP), who genetically lack this protein. The competition ELISA detected lubricin in blood samples from healthy individuals but not from patients with CACP, indicating blood can be used in a diagnostic test for patients suspected of having CACP. Lubricin epitopes in blood do not represent degradation fragments from synovial fluid. Therefore, although blood lubricin levels did not differentiate patients with inflammatory joint disease from healthy controls, epitope-specific anti-lubricin mAbs could be useful for monitoring disease activity in synovial fluid.
Highlights
Lubricin is a large (>250kDa), secreted, highly glycosylated mucinous protein that is abundant in the synovial fluid of mammalian diarthrodal joints [1]
Lubricin is secreted by synovial fibroblasts [2] and superficial zone chondrocytes [3] as a monomer, dimer, or tetramer [4,5,6], and is responsible for boundary lubrication [7,8,9] and the protection of articular cartilage from friction-induced damage [10]
The precocious joint failure that occurs in patients with camptodactyly-arthropathy-coxa vara-pericarditis syndrome (CACP), a genetic deficiency of lubricin, reveals the importance of this protein for protecting joints [12]
Summary
Lubricin is a large (>250kDa), secreted, highly glycosylated mucinous protein that is abundant in the synovial fluid of mammalian diarthrodal joints [1]. Lubricin is secreted by synovial fibroblasts [2] and superficial zone chondrocytes [3] as a monomer, dimer, or tetramer [4,5,6], and is responsible for boundary lubrication [7,8,9] and the protection of articular cartilage from friction-induced damage [10]. Genetic absence of lubricin causes the camptodactyly-arthropathy-coxa vara-pericarditis syndrome (CACP) [12,13]. Among the clinical consequences of CACP are non-inflammatory synovial cell hyperplasia, precocious joint failure, adhesions between tendons and tendon sheaths, and restrictive pericarditis [12]. Mice that have been genetically engineered to lack lubricin (Prg4-/-) developed camptodactyly, synovial hyperplasia, and cartilage deterioration, similar to humans with CACP [14,15,16]. Due to enhanced degradation following injury or inflammatory disease, may contribute to the deterioration of the involved joint [17]; in vitro studies have shown that reducing lubricin between loaded and sliding cartilage explants leads to increases in the coefficient of friction and in chondrocyte apoptosis [18]
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