Bone turnover in untreated polymyalgia rheumatica.

BACKGROUND: Polymyalgia rheumatica (PMR) is a common condition in the elderly. A previous study demonstrated that it is associated with an increase in bone resorption. This effect was ameliorated by steroids, implying that inflammation is the cause of increased bone resorption and that this can be reduced by steroids. This is in keeping with accumulating evidence that systemic inflammation is associated with bone resorption and bone loss. We studied bone formation and resorption markers in 53 patients with PMR prior to any therapeutic intervention. METHODS: Bone resorption was measured by estimating urinary free pyridinoline (fPYD) and deoxypyridinoline (fDPD). Bone formation was estimated by measuring serum concentrations of procollagen type 1 N-terminal propeptide (P1NP). Disease activity was assessed using inflammatory markers (erythrocyte sedimentation rate and C-reactive protein). Patients had a baseline dual-energy X-ray absorptiometer scan to assess bone mineral density. RESULTS: Bone resorption markers were significantly increased and bone formation markers significantly decreased in PMR patients prior to treatment, compared with a control population matched for gender and age. CONCLUSIONS: This implies that bone turnover is uncoupled in PMR. This may lead to a decrease in skeletal mass in the long term due to the disease process alone. However, no significant loss of bone mineral density was detected. It is possible that, due to the acute onset of PMR, increased bone resorption is not present long enough to result in a detectable decrease in bone mineral density. The effects of steroid treatment on bone metabolism and the subsequent long-term outcome need to be investigated.

Insoluble and soluble immune complexes activate neutrophils by distinct activation mechanisms: changes in functional responses induced by priming with cytokines.

BACKGROUND: Rheumatoid synovial fluid contains both soluble and insoluble immune complexes that can activate infiltrating immune cells such as neutrophils. OBJECTIVES: To determine if these different complexes activate neutrophils through similar or different receptor signalling pathways. In particular, to determine the circumstances which result in the secretion of tissue damaging reactive oxygen metabolites and granule enzymes. METHODS: Blood neutrophils were incubated with synthetic soluble and insoluble immune complexes and the ability to generate reactive oxidants tested by luminescence or spectrophotometric assays that distinguished between intracellular and extracellular production. Degranulation of myeloperoxidase and lactoferrin was determined by western blotting. The roles of FcgammaRII (CD32) and FcgammaRIIIb (CD16) were determined by incubation with Fab/F(ab')(2) fragments before activation. The effect of cytokine priming was determined by incubation with GM-CSF. RESULTS: Insoluble immune complexes activated unprimed neutrophils, but most of the oxidants produced were intracellular. This activation required FcgammaRIIIb, but not FcgammaRII function. Soluble complexes failed to activate unprimed neutrophils but generated a rapid and extensive secretion of reactive oxygen metabolites when the cells were primed with granulocyte-macrophage colony stimulating factor (GM-CSF). This activity required both FcgammaRII and FcgammaRIIIb function. Insoluble immune complexes activated the release of granule enzymes from primed or unprimed neutrophils, but the kinetics of release did not parallel those of secretion of reactive oxygen metabolites. Only primed neutrophils released enzymes in response to soluble complexes. CONCLUSIONS: Soluble and insoluble immune complexes activate neutrophils by separate receptor signalling pathways. Profound changes in neutrophil responsiveness to these complexes occur after cytokine priming.

Fcgamma receptors in autoimmune diseases.

Fcgamma-receptors (Fcgamma-R) recognise the Fc portion of IgG and thus form a link between humoral and cellular immunity. These receptors are expressed by a variety of immune cells, and they function in the binding of immune complexes or IgG-opsonised particles, such as microbial pathogens. The are three major types of Fcgamma-R, namely Fcgamma-RI (CD64), Fcgamma-RII (CD32) and Fcgamma-RIII (CD16), and these differ in their ability to bind IgG and complexes. There are many isoforms of these receptors and a number of recently identified polymorphisms in their structure. This review describes the structure and function of these Fcgamma-Rs, and highlights how gene deficiencies and polymorphisms may contribute to the pathology of human diseases.

Severe hypercalcaemia in B-cell lymphoma: combined effects of PTH-rP, IL-6 and TNF.

Hypercalcaemia as the only manifestation of B-cell lymphoma is seen very rarely. Its pathophysiology is heterogenous and not well understood. We report a 73-year-old man who presented with severe hypercalcaemia before any signs of malignancy became evident. He was diagnosed with a B-cell lymphoma on bone marrow trephine biopsy. The hypercalcaemia was associated with high plasma concentrations of parathyroid-hormone-related protein, interleukin-6 and tumour necrosis factor. Our patient had markedly increased osteoclast and osteoblast activity as a result of synergistic effects between these factors, with consequent severe hypercalcaemia. This is the first reported example of such combined effects of these factors in humans.

Neutrophils from the synovial fluid of patients with rheumatoid arthritis express the high affinity immunoglobulin G receptor, Fc gamma RI (CD64): role of immune complexes and cytokines in induction of receptor expression.

Neutrophils isolated from the synovial fluid of 16/24 patients with rheumatoid arthritis expressed Fc gamma RI (CD64), the high-affinity receptor for monomeric immunoglobulin G (IgG), on their cell surface. Receptor expression ranged from 17% to 168% of the level of expression obtained after incubation of control blood neutrophils with 100 U/ml interferon-gamma (IFN-gamma) for 24 hr in vitro. Similarly, mRNA for Fc gamma RI was detected in synovial fluid neutrophils from 12/15 patients and transcript levels ranged from 5% to 200% of the values obtained after treatment of blood neutrophils with IFN-gamma for 4 hr in vitro. No surface expression nor mRNA were detected in freshly isolated blood neutrophils from either patients or from healthy controls. Addition of cell-free synovial fluid to control blood neutrophils induced both mRNA and surface expression of Fc gamma RI to levels that were comparable to those achieved after addition of IFN-gamma. Neither soluble nor insoluble immune complexes appeared to be involved in induction of Fc gamma RI expression in spite of the ability of these complexes to induce protein biosynthesis. Synovial fluid-induced expression of Fc gamma RI was partially blocked by incubation with neutralizing IFN-gamma antibodies, whilst neutralizing interleukin (IL)-6 antibodies had little effect. Levels of IFN-gamma measured within these synovial fluids ranged from 0 to 2.7 U/ml, well within the range known to induce neutrophil Fc gamma RI expression. These data thus indicate that gene expression in synovial fluid neutrophils is selectively activated as the cells enter the diseased joint. Furthermore, these data indicate that induced expression of Fc gamma RI may alter the ability of infiltrating neutrophils to respond to IgG-containing immune complexes present in these joints.

Biochemical markers of bone turnover in seronegative spondylarthropathy: relationship to disease activity.

To investigate bone turnover in patients with seronegative spondylarthropathy, a bone formation marker, type 1 procollagen carboxy-terminal propeptide (P1CP), and resorption markers, the pyridinium cross-links of collagen [urinary free (f) PYR and DPYR], were measured. The median f-PYR, f-DPYR and P1CP (+/-interquartile range) were 15.8 (6.00) nmol/mmol creatinine, 3.8 (2.2) nmol/mmol creatinine and 101.5 (38) micrograms/1, respectively. There was a positive correlation between resorption markers and acute-phase reactants such as C-reactive protein (r = 0.42 for PYR, r = 0.42 for DPYR, P < 0.05), and a negative correlation observed between P1CP and the erythrocyte sedimentation rate (r = -0.64, P < 0.05). In the subgroup of patients with an elevated CRP concentration, the concentration of PYR and DPYR was significantly increased (f-PYR 25.7 vs 15.8 and f-DPYR 6.6 vs 3.8, P < 0.01 for f-PYR, P < 0.05 for f-DPYR). This study suggests than an elevation in acute-phase response in patients with seronegative spondylarthropathy is associated with increased concentration of bone resorption markers with a tendency for reduction in bone formation markers. This may represent uncoupling of bone formation and resorption, leading to bone loss in such patients.

Fat suppressed contrast enhanced MR imaging in the assessment of sacroiliitis.

OBJECTIVE: To investigate the role of Gd-DTPA enhancement in the diagnosis of sacroiliitis. PATIENTS AND METHODS: Fifteen patients with definite sacroiliitis, five subjects from an 'equivocal group', and five controls were imaged with Fast STIR, T1-weighted with fat suppression (T1FS) before and after Gd-DTPA. The 'equivocal group' included those who had abnormally high signal on Fast STIR images only suggestive of early sacroiliitis. The enhancement factor was calculated for T1FS images. Patients graded their symptoms at the time of MR imaging. The extent of abnormal enhancement was defined as none, local, or extensive. RESULTS: Fourteen patients with definite sacroiliitis and one patient from the 'equivocal group' demonstrated abnormal Gd-DTPA enhancement of their sacroiliac joints or adjacent subchondral marrow. Correlation between the patients' symptoms and maximal enhancement of the subchondral marrow was r = 0.75, P < 0.01, and the extent of abnormal enhancement was r = 0.68, P < 0.01. CONCLUSION: Fat suppressed Gd-DTPA or Fast STIR images can be used to assess disease activity in patients with sacroiliitis. Abnormal Gd-DTPA enhancement in the 'equivocal group' is important additional evidence suggesting the diagnosis of early sacroiliitis.

A comparison of MR and CT in suspected sacroiliitis.

OBJECTIVE: A prospective study to compare the MR and CT images of patients with suspected sacroiliitis and to establish the optimal MR sequences to demonstrate the changes of sacroiliitis was conducted. MATERIALS AND METHODS: Thirty-nine patients and nine controls were imaged in the axial plane, with SE T1-, T2-weighted fast spin echo (T2), T1 with fat suppression (T1WFS), and fast short tau inversion recovery (fast STIR) sequences on a 1.5 T system. The sacroiliac joints of all patients were imaged with CT. The images were evaluated by two independent radiologists. Following the blinded reading, direct comparison of T1 and T1WFS, T2, and fast STIR of the CT positive group was made to determine the optimal MR sequences. RESULTS: The sensitivity and specificity of MR images for the detection of cortical erosions and subchondral sclerosis when compared to CT images were 100 and 94.3%, respectively; interobserver variation was low (k = 0.80). T1WFS and fast STIR images were superior to T1 and T2 images, respectively, in demonstrating the changes of sacroiliitis. CONCLUSION: MRI (T1WFS and fast STIR) can replace CT in cases with a strong clinical suspicion of sacroiliitis and equivocal or normal plain radiographs.

Interleukin-1 expression by neutrophils in rheumatoid arthritis.

OBJECTIVE: To determine if neutrophils from blood and synovial fluid of patients with rheumatoid arthritis and other joint arthropathies express interleukin-1 beta mRNA. METHODS: RNA was isolated from neutrophils from patient and control blood, and synovial fluid of patients, probed in northern blots, and quantified by densitometry. It was also isolated and analysed from control blood neutrophils after incubation in vitro with granulocyte macrophage colony stimulating factor (GM-CSF). RESULTS: Neutrophils from the synovial fluid of patients with rheumatoid arthritis contained low levels of mRNA for interleukin-1 beta--between 0.1 and 2% of those observed during stimulation of control neutrophils with GM-CSF for one hour. Higher levels (4-40% of the maximal GM-CSF values) were observed in blood neutrophils from patients with rheumatoid arthritis. CONCLUSIONS: Neutrophils contribute to the cytokine network in rheumatoid arthritis. In some circumstances, activation of transcription may occur within the circulation of these patients.

Role of Fc gamma receptors in the activation of neutrophils by soluble and insoluble immunoglobulin aggregates isolated from the synovial fluid of patients with rheumatoid arthritis.

OBJECTIVES: Synovial fluid from patients with rheumatoid arthritis contains both soluble and insoluble immunoglobulin aggregates which activate reactive oxidant production in human neutrophils. The objectives were to determine the roles played by Fc gamma receptors in activation of neutrophils by these complexes. METHODS: Pronase treatment was used to remove Fc gamma RIII from the neutrophil surface and blocking monoclonal antibodies were used to prevent the binding of complexes to Fc gamma RII and Fc gamma RIII. RESULTS: When Fc gamma RIII was removed from the cell surface by pronase treatment, activation by the soluble aggregates did not occur [mean (SD) inhibition 89 (16)%, n = 6] whereas activation via the insoluble aggregates was less affected [34 (16)%, n = 6]. Blocking the binding to Fc gamma RIII with antibodies decreased activation in response to the soluble aggregates [mean (SD) inhibition 71 (22)%, n = 8] but again had a lower effect on activation by the insoluble aggregates [40 (17)%, n = 9]. When binding to Fc gamma RII was blocked, activation via the soluble aggregates was substantially inhibited [mean (SD) 93 (13)%, n = 8] whereas that via the insoluble aggregates was inhibited to a much lesser extent [28 (38)%, n = 9]. When Fc gamma RII and III were simultaneously blocked, activation by the insoluble aggregates was only inhibited by 45% [(19), n = 5]. CONCLUSION: These data thus indicate that activation of human neutrophils by soluble immunoglobulin aggregates from rheumatoid synovial fluid occurs via cooperative occupancy of both Fc gamma RII and III: perturbation of binding to either of these receptor classes will abrogate activation.

A western blot approach to detection of human plasma protein conjugates derived from D-penicillamine.

OBJECTIVES: To develop and apply an immunochemical approach to the study of drug-plasma protein conjugates derived from the anti-arthritic drug D-penicillamine (DP). METHODS: An antiserum with specificity for protein-conjugated DP was raised in a rabbit. Plasma samples from patients receiving DP or from incubations of isolated normal plasma with DP were analysed for DP-derived conjugates by Western blotting using the anti-drug antibody. RESULTS: A single DP-positive protein band was detected in plasma samples from 15/16 patients with rheumatoid arthritis receiving DP but in none of 20 patients of similar disease status who had not taken DP. The positive band appeared in patients' plasma during the course of treatment with DP. It was seen under nonreducing but not reducing conditions indicating that the drug is disulphide linked to the protein. The drug-modified protein migrated to a position intermediate between the trailing edge of albumin and the leading edge of transferrin (both non-reduced) suggesting a molecular weight of between 66 and 77 kDa. Incubations of normal human plasma, but not purified albumin or transferrin, with low concentrations of DP generated the same distinct band plus several less intense DP-positive bands. CONCLUSIONS: Drug-plasma protein conjugates derived from DP in vivo and in vitro can be detected immunochemically by the Western blot method. Theories of DP immunotoxicity have implicated antigenicity of the drug, but this is the first immunochemical demonstration of a potential DP-derived antigen in human plasma. The method we describe may have application to studies of the relationship between DP antigenicity and toxicity.