Synovial fluid 5′‐nucleotidase activity. Relationship to other purine catabolic enzymes and to arthropathies associated with calcium crystal deposition

Abstract

We measured 5'-nucleotidase (5NT) activity in synovial fluid from 159 patients with various diagnoses. The activity of 5NT was compared with activities of nucleotide pyrophosphohydrolase, alkaline and neutral phosphatases, and adenosine deaminase, in the same samples. Higher levels of 5NT activity occurred in synovial fluid from osteoarthritic joints than from joints of patients with gout, pseudogout, or rheumatoid arthritis. The highest levels of 5NT activity were found in synovial fluid from patients with Milwaukee shoulder syndrome and from osteoarthritis patients in whom deposition of calcium-containing crystals was also present.The metabolic derangements that underlie osteoarthritis (OA) and arthropathies associated with calcium-containing crystals, including calcium pyrophosphate dihydrate (CPPD) crystal deposition disease, basic calcium phosphate (BCP) crystal deposition arthropathy (1,2), and Milwaukee shoulder syndrome (MSS) (3), are poorly understood. Previous observations indicate that purine catabolic enzymes may play a role in these processes (4-14) (Figure 1). We measured the activity of 5'-nucleotidase (5NT) in synovial fluids from 159 patients with wellcharacterized arthropathies and compared that activity with the activity of synovial fluid nucleoside triphosphate pyrophosphohydrolase (NTPPPH), alkaline phosphatase (AP), neutral phosphatase (NP), and adenosine deaminase (ADA), in a broad selection of the same fluid samples. In contrast with the results reported by others (9), we observed higher 5NT activity in fluids from OA joints than from joints of patients with rheumatoid arthritis (RA). The highest levels of 5NT activity were found in the subset of fluids from osteoarthritic joints that also contained calcium crystal (CPPD and/or BCP) deposits, and in fluids from the shoulders of patients with MSS. of noninflammatory synovial fluid (leukocyte count 5 1,OOOl mm3), and 4) the presence of synovial fluid BCP crystals (3,181. Synovial fluid collection. Synovial fluid was drawn from the joint into a plastic syringe, and a portion of the sample was submitted for routine synovianalysis. The remainder was transferred immediately into a plastic tube containing 200 units of heparin (beef lung; Upjohn, Kalamazoo, MI) and 5 mg of hyaluronidase (beef testes; Worthington, Freehold, NJ), incubated at room temperature for 30 minutes, and centrifuged at 28,OOOg for 20 minutes. The supernatant fluids were stored at -70°C. The pellets were resuspended in 100 mM Tris HC1, pH 8.0, in a volume equal to that of the removed supernatant, and stored at -70°C until assay for BCP content, which was performed within 10 days. PATIENTS AND METHODSBCP assay. BCP crystals were detected and quantified in the centrifuged synovial fluid pellets, using the a:4182-20.ct14C-EHPP binding method (Proctor & Gamble, Cincinnati, OH) (I). The assay is specific for hydroxyapatite and, as performed, was able to quantitate BCP crystal mass equivalent to 2 3 pg of hydroxyapatite standard per ml.PP, assay. Synovial fluid PP, concentration ...

5′-Nucleotidase: molecular structure and functional aspects

“…We find that starting from a contribution of about 300% the amount of soluble (concanavalin A-binding) 5'-nucleotidase increases with post mortem time in bovine brain (Vogel et al, 1992). The observation that the activity of soluble 5'-nucleotidase in synovial fluid varies with specific clinical symptoms (Wortmann et al, 1991) or its presence in human seminal plasma (Fini et al, 1991) and also snake venoms supports the notion that GPI-anchored 5'-nucleotidase may be released by a physiological mechanism. Whether this involves release from the surface or constitutive secretion remains to be elucidated.…”

Soluble low-Km 5′-nucleotidase from electric-ray (Torpedo marmorata) electric organ and bovine cerebral cortex is derived from the glycosyl-phosphatidylinositol-anchored ectoenzyme by phospholipase C cleavage

“…It is noteworthy in this connection that human seminal plasma (Fini et al, 1991), as well as synovial fluid (Wortmann et al, 1991) the protein, which is thought to function not only as an enzyme but to have also the potential to act in cell-cell and cell-matrix interactions . Although the plasma membrane is the most likely source of soluble low-Km-5'-nucleotidase, other sources need to be considered.…”

Purinergic signalling in the musculoskeletal system

“…Thus, the augmentation of adenosine levels was suggested as a treatment of this condition [379]. Compared to the joints from RA patients, higher levels of ATP and 5′-nucleotidase activity were also observed in the synovial fluid from osteoarthritic joints; especially from osteoarthritic patients in whom deposition of calcium-containing crystals was also present [380,381]. Later, ATP and UTP were claimed to activate calciummobilizing P2U receptors to act synergistically with IL-1 to stimulate PGE 2 release from human rheumatoid synovial cells [382].…”