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Activation of neutrophil reactive-oxidant production by synovial fluid from patients with inflammatory joint disease. Soluble and insoluble immunoglobulin aggregates activate different pathways in primed and unprimed cells.

ABSTRACT: Cell-free synovial fluid from patients with rheumatoid arthritis stimulated the NADPH oxidase activity in human neutrophils, which reached a peak 15-20 min after addition. Insoluble immunoglobulin aggregates isolated from these fluids activated a similar pattern of oxidase activity. However, when synovial fluid was added to neutrophil suspensions which had been previously exposed to granulocyte-macrophage colony-stimulating factor, the stimulated oxidase activity was biphasic, in that an additional transient activity was observed which reached a peak within 5 min of addition. The additional neutrophil-stimulating activity could not be sedimented by centrifugation at 330,000 g-min, and only activated oxidase activity in neutrophils which had previously been primed. The neutrophil-stimulating activity in this soluble fraction was removed by Protein A affinity chromatography, and activity was recovered in eluates from this column. Thus activity in this soluble fraction from synovial fluid is attributed to the presence of soluble immunoglobulin aggregates. Whereas oxidase activity stimulated by the isoluble immunoglobulin aggregates was inhibited by staurosporine (and hence largely dependent on the activity of protein kinase C), the activity stimulated by the soluble immunoglobulin aggregates was staurosporine-insensitive. The soluble immunoglobulin aggregates were present at significantly higher levels in synovial fluids from patients with rheumatoid arthritis compared with those from other joint arthropathies. Thus rheumatoid synovial fluids possess heterogeneous immunoglobulin aggregates which activate neutrophils via distinct molecular pathways. As neutrophils within rheumatoid joints are primed, the soluble immunoglobulin aggregates are likely to be of importance in disease pathology.

SUBMITTER: Robinson J

PROVIDER: S-EPMC1132903 | biostudies-other | 1992 Sep

REPOSITORIES: biostudies-other

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Project description:Recent in vitro studies reported the exceptional ability of some bacterial species to form biofilm-like aggregates in human and animal synovial fluids (SF), but evidences from infected clinical samples are still lacking. In this study, we investigated whether this bacterial phenotype was present in infected SFs collected from joint infections and if it was maintained in in vitro settings. SFs sent for culture to the Laboratory of Microbiology of our institute were directly analyzed by means of confocal laser scanning microscopy (CLSM), and the infective agents were isolated for further in vitro tests. Moreover, sterile SF was collected from patients who did not receive previous antibiotic therapy to investigate the formation of bacterial aggregates, together with biofilm and matrix production on a titanium surface. Finally, antibiotic susceptibility studies were performed by using bovine SF. Four Staphylococcus aureus, one Staphylococcus lugdunensis, and one Prevotella bivia strain were identified in the infected SFs. The CLSM analysis showed that all staphylococci were present as a mixture of single cells and bacterial clumps surrounded by an exopolymeric substance, which comprised SF-derived fibrin, while all P. bivia cells appeared separated. Despite that, differences in the ability to aggregate between S. aureus and S. lugdunensis were observed in clinical SFs. These different phenotypes were further confirmed by in vitro growth, even though the application of such ex vivo approach lead all staphylococci to form exceptionally large microbial aggregates, which are several folds bigger than those observed in clinical samples. Planktonic aggregates challenged for antibiotic susceptibility revealed a sharp increase of recalcitrance to the treatments. Although this is still at a preliminary stage, the present work confirmed the ability of staphylococci to form free-floating biofilm-like aggregates in infected SF from patients with joint infections. Furthermore, the obtained results pointed out that future in vitro research on joint infections will benefit from the use of human- or animal-derived SF. Even though this approach should be carefully validated in further studies comprising a larger microbial population, these findings pose new challenges in the treatment of infected native and prosthetic joints and for the approach to new investigations.

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Activation of neutrophil reactive-oxidant production by synovial fluid from patients with inflammatory joint disease. Soluble and insoluble immunoglobulin aggregates activate different pathways in primed and unprimed cells.

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