Serum nucleotide pyrophosphohydrolase activity; elevated levels in osteoarthritis, calcium pyrophosphate crystal deposition disease, scleroderma, and fibromyalgia.

OBJECTIVE: Quantification of serum nucleotide pyrophosphohydrolase (NTPPHase) activity in healthy subjects and in patients with various rheumatic diseases or with quad/hemiplegia, hemodialysis, or renal transplant. METHODS: Colorimetric assay of enzyme activity in serum. RESULTS: Serum NTPPHase activity in 85 healthy subjects was independent of age or sex and was highly reproducible in each individual. The biologic and methodologic coefficients of variation were nearly identical. Elevated enzyme levels were found in sera from patients with osteoarthritis/spondylosis, calcium pyrophosphate dihydrate (CPPD) crystal deposition, scleroderma, fibromyalgia, or hemodialysis. Renal transplant patients receiving cyclosporine had the highest enzyme activity of any group, whereas transplant patients not taking this drug had normal levels. Histograms of values in all groups showed a normal distribution. CONCLUSION: Serum NTPPHase activity levels were significantly elevated in patients with degenerative arthritis whether or not CPPD crystals were present, in patients with either scleroderma or fibromyalgia, and in patients receiving hemodialysis therapy or taking cyclosporine.

Nucleotide pyrophosphohydrolase in human synovial fluid.

OBJECTIVE: To identify the molecular forms of ectonucleotide pyrophosphohydrolase (NTPPHase) in human synovial fluid (SF). METHODS: We examined synovial fluids from 32 patients with various joint diseases [10 calcium pyrophosphate dihydrate (CPPD) deposition disease; 7 osteoarthritis (OA); 6 rheumatoid arthritis (RA); 3 after total knee arthroplasty (TKA); 6 olecranon bursa] and 3 normal joint fluids. Joint fluids were analyzed after sequential centrifugation for NTPPHase activity and by Western blot using polyclonal antibodies against 127 kDa porcine articular cartilage vesicle-associated NTPPHase and against PC-1 and 58 kDa, 2 other ecto-NTPPHases. Lysate from human synoviocytes, porcine chondrocytes, and their conditioned media were examined using antibodies to these ecto-NTPPHases. Radiographs of joints from which fluid was obtained were graded for degenerative changes 0-4 using a standard method. RESULTS: NTPPHase activity was found in all pathological and normal SF tested and correlated with the degree of radiographic degeneration (r = 0.55, p < 0.05). NTPPHase specific activity in ultracentrifugation pellets was highest in CPPD deposition disease fluids (p < 0.05). 127 kDa enzyme was found in both sedimentable and soluble fractions from CPPD, OA, TKA, and normal fluids, and was extensively degraded in all inflammatory fluids. Intact 115 kDa PC-1 was found only in the 2 CPPD fluids with the highest NTPPHase activity. 58 kDa enzyme was found in most fluids, predominantly in the soluble fraction. 127 kDa protein was identified in human synoviocyte conditioned media but not in cell lysate, while PC-1 and 58 kDa proteins were found in the cell lysate but not in the conditioned media. CONCLUSION: There was no disease specific association with any one ecto-NTPPHase. Total enzyme activity correlated with the degree of degenerative change. The specific activity of pelletable 127 kDa enzyme was higher in fluids containing CPPD crystals. All 3 ecto-NTPPHases or their presumed degradation products were detectable in some pathologic and normal fluids. A 200 kDa reactive band often accompanied reactivity to the 127 kDa enzyme. PC-1 and 127 kDa proteins were extensively degraded in inflammatory SF, while 58 kDa protein was not. The relative contribution of each of these enzymes to inorganic pyrophosphate production by human joint tissues remains unclear.

Identification of a nucleotide pyrophosphohydrolase from articular tissues in human serum.

OBJECTIVE: To characterize the nucleotide pyrophosphohydrolase (NTPPHase) in human serum. METHODS: NTPPHase activity and kinetic analysis were performed using thymidine monophosphate paranitrophenyl ester (TMPNP) or 32Pgamma-labeled ATP as substrate. Sera were chromatographed (dye column), and peak fractions were analyzed kinetically and by immunoblot using antibodies to 127-kd articular cartilage vesicle (ACV) NTPPHase as well as to PC-1 and to 58 kd, two plasma membrane ecto-NTPPHases. Enzyme activity was measured before and after sample ultracentrifugation. RESULTS: NTPPHase activity was found in all sera tested (2 normal subjects, 9 arthritis patients). Specific activity was increased 9-32-fold after chromatography; 60-80% of total activity was recovered in a single peak containing an approximately 100-kd soluble peptide related to the 127-kd ACV enzyme. The apparent Km of this peptide (TMPNP) was virtually identical to that of the porcine ACV 127-kd enzyme. No immunoreactivity against PC-1 or 58-kd NTPPHase was found. CONCLUSION: Human serum NTPPHase is derived from 127-kd ACV-related enzyme.

Specificity of a porcine 127-kd nucleotide pyrophosphohydrolase for articular tissues.

OBJECTIVE: To determine the tissue specificity of a porcine 127-kd nucleotide pyrophosphohydrolase (NTPPHase) found in vesicles derived from hyaline articular cartilage (ACV). METHODS: Homogenates of porcine brain, lung, liver, kidney, urinary bladder, pancreas, spleen, skin, vena cava, marrow, bone (cells), tendon (Achilles), ligament (anterior cruciate), elastic cartilage, meniscus, and hyaline cartilage were analyzed for NTTPHase activity (thymidine monophosphate paranitrophenyl ester substrate) and by Western blot using polyclonal antibodies against 127-kd NTPPHase and against recombinant PC-1, another ecto-NTPPHase. RESULTS: All tissues contained NTPPHase activity; the highest specific activity was found in hyaline articular cartilage, the lowest in brain. ACV-associated 127-kd NTTPHase was expressed in cartilage, ligament, and tendon. PC-1 was also expressed in those tissues and in skin, kidney, bone cells, and (probably) in liver and muscle. CONCLUSION: The 127-kd NTPPHase appears to be highly specific for articular tissues.

An indirect fluorescent antibody technique for detection of anti-Dermatophilus congolensis antibodies in sheep.

An indirect fluorescent antibody (IFA) technique has been developed for detection of anti-Dermatophilus antibodies in sheep. Sera from 25 bacteriologically confirmed clinically affected sheep and from 10 negative non affected lambs were used. Whole cell antigen from brain heart infusion cultures of D. congolensis was used and all sera were tested in the same way for cross-reactivity against antigens obtained from cultures of Actinomyces viscosus, Micrococcus luteus, Nocardia asteroides, and Corynebacterium pseudotuberculosis. Sera from Dermatophilus-infected sheep gave positive results with D. congolensis antigen and negative results with the antigens from other bacteria. The whole cell antigens employed were simple to prepare and easy to recognise by microscopy. Cross-reactivity was further tested using the D. congolensis culture whole cell antigen and 3 sera from sheep with bacteriologically confirmed natural infections due to Corynebacterium pseudotuberculosis, Actinomyces pyogenes and Nocardia asteroides. None of these sera showed positive reactions. The authors recommend this technique for serological surveys and research on dermatophilosis.

Osteoarticular infection in intravenous drug abusers: influence of HIV infection and differences with non drug abusers.

OBJECTIVES: To determine (a) the influence of HIV in developing osteoarticular infections in intravenous drug abusers (IVDAs) and (b) the differences between the clinical features of osteoarticular infections in IVDAs and a control group of non-IVDAs. METHODS: A comparative study of the clinical features of osteoarticular infections in all HIV positive and HIV negative IVDAs admitted to the departments of rheumatology and internal medicine during a 10 year period was carried out. The joint infections of all IVDAs, irrespective of HIV status, were compared with those of a control group of non-IVDAs lacking risk factors for HIV infection. RESULTS: A total of 482 HIV positive and 85 HIV negative IVDAs was studied, in whom 25 (5%) and six (7%) osteoarticular infections were found respectively. There were no differences in age, sex, joints affected, and causative agents between these two groups. A comparison of the 31 (5.5%) osteoarticular infections in all IVDAs with 21 infections in 616 (3.4%) non-IVDAs showed significant differences in the mean age (27.5 v 54), the frequency of affection of the axial joints (hip, sacroiliac, and sternocostal joints) (64.5% v 16.6%), and in the incidence of Candida albicans (19% v 0%). CONCLUSIONS: (1) HIV may not predispose to osteoarticular infections in IVDAs. (2) The hip, sacroiliac, and sternocostal joints (axial joints) were most commonly affected in IVDAs. (3) In Spain, unlike other countries, Gram positive bacteria and C albicans seem to be predominant agents in osteoarticular infections in IVDAs, with a low incidence of Gram negative bacteria.

Osteoarticular infection associated with the human immunodeficiency virus.

An analysis is presented of 20 patients with the human immunodeficiency virus (HIV) infection who were treated for skeletal infection in our hospital. Seventeen cases of septic arthritis and three of osteomyelitis were found. All patients were intravenous drug abusers, 75% were male and all were younger than 40 years old. Eighty-eight percent of the septic arthritis cases were monarticular, the hip joint being the area most commonly affected (29%), and Staphylococcus aureus and Candida albicans were the most frequently isolated agents (60% and 20%, respectively). Gram negative bacteria were not found. No correlation between the number of CD4 lymphocytes, affected joints, isolated microorganism or functional capacity was found. Four of 17 patients with septic arthritis and 2 of 3 with osteomyelitis fulfilled the CDC criteria for the diagnosis of acquired immunodeficiency syndrome (AIDS). All cases except one responded to conventional therapy. We conclude that skeletal infection in HIV-infected patients is more closely associated with the risk-group than with HIV itself.

Rheumatic manifestations in 556 patients with human immunodeficiency virus infection.

We studied in retrospect the rheumatic manifestations of 556 patients with human immunodeficiency virus (HIV) infection. Eighty percent were men. Eighty-six percent were intravenous drug abusers (IVDAs), 9% homosexual, 3% partners of high-risk persons having the infection, 0.4% hemophiliacs, and 2% had no known risk factors. We found rheumatic disorders in 63 (11%) patients. The most frequent findings were myalgias and/or arthralgias (4.5%; one patient had an inflammatory myopathy), skeletal infections (3.6%), and arthralgias (1.6%). Reiter's syndrome and seronegative arthritis were present only in 0.5%, and HIV-associated arthritis and vasculitis in 0.4%, respectively. Skeletal infections were caused predominantly by Staphylococcus aureus (60%) and Candida albicans (20%). All these patients were IV drug abusers whose clinical features were similar to those previously described in skeletal infections of non-HIV-infected IVDAs. Comparing these data with other studies composed primarily of homosexual men where Reiter's syndrome is the predominant rheumatic disorder, we conclude that the type of rheumatic complaint is more related to the risk factors than to HIV itself.