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1.  Naturally occurring and experimentally induced mycoplasmal arthritis of cattle. 
Journal of Clinical Microbiology  1975;2(3):165-168.
Mycoplasma agalactiae subsp. bovis strain Iowa 1136 was isolated from synovial fluids of a clinical case of arthritis in cattle on pasture in Iowa. When given to calves and cows by intra-articular or intravenous injection, it caused severe and persistent joint infections with fever, lameness, and swelling of the affected joints, plus synovitis, tendonitis, and fibrinous-purulent synovial fluids of high protein content. Intramammary administration of the organism caused severe mastitis. Calves nursing the cows developed severe mycoplasmal arthritis.
PMCID: PMC274164  PMID: 1176623
2.  Pronounced production of polyclonal immunoglobulin G1 in the synovial fluid of goats with caprine arthritis-encephalitis virus infection. 
Infection and Immunity  1983;41(2):805-815.
Infection of goats with caprine arthritis-encephalitis virus, a lentivirus, resulted in arthritis characterized by the production of intrasynovial immunoglobulin G1 concentrations that were 2 to 5.3 times the serum concentrations in the inoculated carpi at 6 months postinoculation. The intrasynovial immunoglobulin was polyclonal, and its presence was accompanied by increased albumin leakage into the joints. Synovial fluid immunoglobulin levels fluctuated temporally but remained elevated compared with medium-inoculated controls for 38 months after infection. Elevated immunoglobulin G1 concentrations correlated with focal sublumenal plasmacytic infiltrates in the synovia of inoculated carpi at 5 months postinoculation. Inflammation in the uninoculated joints of infected goats was also accompanied by increased intrasynovial immunoglobulin G1 levels. Antibody to systemically administered antigens was a greater proportion of the immunoglobulin population in sera than in synovial fluids of infected goats, suggesting that antibody production to local antigens was responsible for increased intrasynovial immunoglobulin G1 levels.
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PMCID: PMC264711  PMID: 6307882
3.  Distribution of Mycoplasma pneumoniae and Mycoplasma salivarium in the Synovial Fluid of Arthritis Patients▿  
Journal of Clinical Microbiology  2006;45(3):953-957.
By use of a very sensitive nested PCR method targeting part of the strongly conserved mycoplasmal 16S RNA genes, Mycoplasma pneumoniae was found in the synovial fluid of 19/24 (79%) of rheumatoid arthritis patients, 6/6 (100%) of patients with nonrheumatoid inflammatory arthritis, and 8/10 (80%) of osteoarthritis patients attending the rheumatology clinic for drainage of joint effusions. It was not found in the synovial exudates of 13 people attending the orthopedic clinic with traumatic knee injuries or undergoing surgery for knee replacement. However, M. pneumoniae was detected in 2/4 synovial biopsy specimens from orthopedic patients with traumatic knee injuries. M. pneumoniae was associated with the increased synovial fluids found in arthritic flares but was not found in the synovial fluids of trauma patients. Mycoplasma salivarium occurred sporadically. Mycoplasma fermentans had previously been isolated from patients with inflammatory cellular infiltrates, such as rheumatoid arthritis, but it was not detected for osteoarthritic patients from either clinic. It is possible that these organisms may contribute to chronic inflammation within the joints.
doi:10.1128/JCM.01973-06
PMCID: PMC1829110  PMID: 17122006
4.  Synthesis of antibodies, including antiviral antibodies, in the knee joints of patients with arthritis. 
Annals of the Rheumatic Diseases  1985;44(11):734-737.
Serum and synovial fluids from 16 patients with seronegative arthritis and eight with rheumatoid arthritis were studied for immunoglobulin levels and for antibody levels to five viruses. When allowances were made for the distribution of immunoglobulins between serum and synovial fluid there was evidence that in several patients antibody to one or more viruses was synthesised locally in the joint. IgG and especially IgM were present in greatly increased amounts in arthritic joints compared with normal joints. On the basis of serum/synovial fluid ratios inflammation and local immunoglobulin synthesis are discussed as possible causes. These results are compared with antiviral antibody and immunoglobulin ratios observed in the serum and cerebrospinal fluid of patients with multiple sclerosis.
PMCID: PMC1001759  PMID: 4062387
5.  Acute Immunologic Arthritis in Rabbits 
Mediators of acute immunologic injury have been studied in vivo by producing arthritis in rabbit knee joints. A reversed passive Arthus lesion was produced by injecting antibody into the joint space and antigen intravenously. Injury was assessed by measuring leakage of serum proteins and circulating radiolabeled proteins into the joint space and by the accumulation of neutrophils in the joint fluid. Inflammatory exudate was recovered for study by a standardized irrigation technique.
Maximal vascular permeability developed 2 hr after injection as neutrophils accumulated about immune complexes in venule walls to produce structural injury. After 5 hr the number of neutrophils in the joint space rose rapidly, followed by a second rise in permeability at 8 hr. Neutrophil depletion abolished both peaks of permeability. It was then possible to reconstitute the synovial lesion in neutrophil-depleted rabbits by intra-articular injection of purified suspensions of neutrophils.
A requirement for complement was demonstrated in development of the lesion. Rabbits genetically deficient in C6 showed delay in vascular permeability, appearance of neutrophils, and histologic lesions. The delay was longer in normal rabbits depleted of C3. In C6-deficient rabbits depleted of C3, still further reduction in injury occurred.
Evidence was obtained as well for a chemotactic attraction of neutrophils in vivo. Antigen-antibody-complement complexes in the walls of blood vessels attracted neutrophils placed in the joint space of neutrophil-depleted rabbits. Omission of either antigen or antibody from this replacement reaction prevented the migration of neutrophils.
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PMCID: PMC332928  PMID: 4257028
6.  Cytokines profiling by multiplex analysis in experimental arthritis: which pathophysiological relevance for articular versus systemic mediators? 
Introduction
We have taken advantage of the large screening capacity of a multiplex immunoassay to better define the respective contribution of articular versus systemic cytokines in experimental arthritis.
Methods
We performed a follow up (from 7 hours to 14 days) multiplex analysis of 24 cytokines in synovial fluid and sera of rats developing Antigen-Induced Arthritis (AIA) and confronted their protein level changes with molecular, biochemical, histological and clinical events occurring in the course of the disease.
Results
The time-scheduled findings in arthritic joints correlated with time-dependent changes of cytokine amounts in joint effusions but not with their blood levels. From seven hours after sensitization, high levels of chemokines (MCP-1, MIP1α, GRO/KC, RANTES, eotaxin) were found in synovial fluid of arthritic knees whereas perivascular infiltration occurred in the synovium; local release of inflammatory cytokines (IFNγ, IL-1β, IL-6) preceded the spreading of inflammation and resulted in progressive degradation of cartilage and bone. Finally a local overexpression of several cytokines/adipocytokines poorly described in arthritis (IL-13, IL-18, leptin) was observed.
Conclusions
Distinct panels of cytokines were found in arthritic fluid during AIA, and the expected effect of mediators correlated well with changes occurring in joint tissues. Moreover, multiplex analysis could be helpful to identify new pathogenic mediators and to elucidate the mechanisms supporting the efficacy of putative targeted therapies.
doi:10.1186/ar3774
PMCID: PMC3446427  PMID: 22414623
7.  Local synovial synthesis of oligoclonal measles virus antibodies and of smooth muscle antibodies in a case of atypical rheumatoid arthritis. 
Annals of the Rheumatic Diseases  1977;36(4):302-310.
A patient with atypical rheumatoid arthritis (RA) and local synovial synthesis of oligoclonal IgG in an arthritic knee joint is described. Measles virus-specific antibodies isolated from the synovial fluid (SF) were carried by oligoclonal IgG proteins but constituted only a fraction of the total oligoclonal IgG of the SF. Smooth muscle antibodies were markedly increased in the SF compared with the serum and were associated with an electrophoretically restricted fraction of IgG. The results indicate that a local synovial synthesis of measles virus-specific antibodies and of smooth muscle antibodies occurred within the affected joint in our patient.
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PMCID: PMC1006690  PMID: 901028
8.  Mast cells contribute to autoimmune inflammatory arthritis via their tryptase•heparin complexes1 
Although mast cells (MCs) often are abundant in the synovial tissues of patients with rheumatoid arthritis (RA), MC’s contribution to joint inflammation and cartilage loss remains poorly understood. MC-restricted tryptase•heparin complexes have pro-inflammatory activity, and significant amounts of hTryptase-β are present in RA synovial fluid. Mouse MC protease-6 (mMCP-6) is the ortholog of hTryptase-β, and this serine protease is abundant in the synovium of arthritic mice. We now report that C57BL/6 (B6) mice lacking their tryptase•heparin complexes have attenuated arthritic responses, with mMCP-6 as the dominant tryptase responsible for augmenting neutrophil infiltration in the K/B×N mouse serum-transfer arthritis model. While inflammation in this experimental arthritis model was not dependent on protease activated receptor-2, it was dependent on the chemokine receptor CXCR2. In support of the latter data, exposure of synovial fibroblasts to hTryptase-β•heparin or mMCP-6•heparin complexes resulted in expression of the neutrophil chemotactic factors CXCL1/KC, CXCL5/LIX, and CXCL8/IL-8. Our proteomics, histochemistry, and immunohistochemistry data also revealed substantial loss of cartilage-derived aggrecan proteoglycans in the arthritic joints of wild-type B6 mice but not mMCP-6-null B6 mice. These observations demonstrate the functional contribution of MC-restricted tryptase•heparin complexes in the K/B×N mouse arthritis model and connect our mouse findings with RA pathophysiology.
PMCID: PMC2610352  PMID: 19109198
mast cell; rheumatoid arthritis; inflammation; chemokines; transgenic/knockout mice
9.  Serum transfer of collagen-induced arthritis in mice 
Immunization of DBA/1 mice with native chick type II collagen resulted in development of polyarthritis 4-5 wk later. Sera of these mice contained high levels of anticollagen antibodies, and immunoglobulin concentrates of their sera transferred arthritis to unimmunized recipients. Histopathologically, this passively transferred arthritis resembled the early disease of immunized donors. Immunofluorescence studies demonstrated the deposition of IgG and C3 on the articular surface but not in synovial tissue of arthritic joints. Transferred, isotopically labeled anticollagen antibodies rapidly localized to the limbs and to other cartilage-containing tissues. When transfer concentrate was administered to arthritis-resistant strains, they also developed arthritis. Indeed, immunoglobulin concentrates from rats with collagen-induced arthritis transferred arthritis to naive mice. The amount of concentrate required for transfer to B10.D2 resistant mice was reduced by immunizing them with collagen 4 wk before transfer. Although susceptibility to arthritis from immunization is H-2 linked, these studies clearly demonstrate that passive transfer of arthritis depends upon injection of specific antibody and not on other host factors.
PMCID: PMC2187334  PMID: 6886622
10.  Experimental Arthritis in Mice with Mycoplasma pulmonis 
Journal of Bacteriology  1969;100(1):5-10.
A Mycoplasma pulmonis strain, recovered from the arthritic joints of mice employed in the serial passage of a chemically induced tumor, was found to be arthritogenic for mice under experimental conditions. Some joint involvement occurred in all mice challenged intravenously with this strain, and M. pulmonis was recovered frequently from the enlarged joints. The arthritis was migratory, appearing first in the radiocarpal joints and later in the tibiotarsal joints. There was little evidence of a generalized mycoplasmal infection as a consequence of the experimental challenge. Histopathologically, the early stages of the infection in the joints was characterized by an inflammatory response in the synovium and periarticular tissues. Exudate in the joint space contained about equal numbers of polymorphonuclear and mononuclear cells. The polyarthritis resolved slowly, but some residual joint enlargement was noted for as long as 4 months. Two other M. pulmonis strains were also observed to be arthritogenic for mice. Rats were not susceptible to M. pulmonis challenge. Characteristics of the nonsuppurative M. pulmonis arthritis in mice were compared to M. arthritidis joint infections in rats.
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PMCID: PMC315349  PMID: 5344119
11.  Immunoelectrophoretic Analysis of Mycoplasma mycoides var. mycoides 
Infection and Immunity  1973;7(6):922-930.
Acrylamide gel electrophoresis was used to show the similarities and differences in the membrane proteins of two vaccine and two virulent strains of Mycoplasma mycoides var. mycoides. Immunoelectrophoretic (IEP) analysis was also used to partially characterize the associated antigens. Antibody spectra to the antigens of M. mycoides differ in rabbit, pig, and cattle sera. Rabbits produce better precipitating antibody against the anodic migrating protein mycoplasma antigens than cattle and pigs as seen in IEP. However, rabbit anti-M. mycoides serum did not show precipitating antibody against the heat-stable carbohydrate antigen. As judged by IEP, the major carbohydrate antigen extracted from the media, or boiled whole organism, is similar to that present in the sera-infected cattle and knee joints of calves. This carbohydrate antigen has a cathodic migration in IEP at pH 8.6. Periodate oxidation, classically used to destroy carbohydrate, also destroys most of the protein antigens. Heating the antigens to 56 C for 10 min destroys many of the noncarbohydrate antigens and 100 C eliminates all but the carbohydrate antigen. Extraction of M. mycoides with chloroform-methanol, phenol, ethanol, or ethanol-acetone reduced or eliminated most of the protein antigens. Some of the isolated antigenic fractions of M. mycoides were tested to determine their activity in the diagnostic complement fixation test for contagious bovine pleuropneumonia and their inhibitory effect in this test by using bovine anti-M. mycoides antisera having precipitating antibody and circulating antigen. The complement fixation antigen is not the galactan, cannot be extracted by chloroform-methanol, but is stable to boiling at 100 C and may be extracted by phenol and partially precipitated by ethanol-acetone.
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PMCID: PMC422784  PMID: 4577417
12.  A New Phenylpyrazoleanilide, Y-320, Inhibits Interleukin 17 Production and Ameliorates Collagen-Induced Arthritis in Mice and Cynomolgus Monkeys 
Pharmaceuticals  2013;7(1):1-17.
Interleukin (IL)-15 and IL-17 are thought to play an important role in the pathogenesis of rheumatoid arthritis (RA) because both pro-inflammatory cytokines are found in synovial fluid of RA patients. In this study, we examined the pharmacological profiles of Y-320, a new phenylpyrazoleanilide immunomodulator. Y-320 inhibited IL-17 production by CD4 T cells stimulated with IL-15 with IC50 values of 20 to 60 nM. Oral administration of Y-320 (0.3 to 3 mg/kg) significantly inhibited the development and progression of arthritis and joint destruction with reduction of IL-17 mRNA expression in arthritic joints of type II collagen-induced arthritis (CIA) in DBA/1J mice. Y-320 in combination with anti-murine tumor necrosis factor-α monoclonal antibody showed a synergistic effect on mouse CIA. Moreover, therapeutic treatment with Y-320 (0.3 and 1 mg/kg orally) ameliorated CIA in cynomolgus monkeys. Our results suggest that Y-320, an orally active inhibitor for IL-17 production, provides a useful therapy for RA.
doi:10.3390/ph7010001
PMCID: PMC3915191  PMID: 24366113
interleukin 15 (IL-15); interleukin 17 (IL-17); phenylpyrazoleanilide; Y-320; immunomodulator; collagen-induced arthritis (CIA); rheumatoid arthritis (RA)
13.  Epithelial neutrophil activating peptide-78: a novel chemotactic cytokine for neutrophils in arthritis. 
Journal of Clinical Investigation  1994;94(3):1012-1018.
We and others have shown that cells obtained from inflamed joints of rheumatoid arthritis (RA) patients produce interleukin-8, a potent chemotactic cytokine for neutrophils (PMNs). However, IL-8 accounted for only 40% of the chemotactic activity for PMNs found in these synovial fluids. Currently, we have examined the production of the novel PMN chemotactic cytokine, epithelial neutrophil activating peptide-78 (ENA-78), using peripheral blood, synovial fluid, and synovial tissue from 70 arthritic patients. RA ENA-78 levels were greater in RA synovial fluid (239 +/- 63 ng/ml) compared with synovial fluid from other forms of arthritis (130 +/- 118 ng/ml) or osteoarthritis (2.6 +/- 1.8 ng/ml) (P < 0.05). RA peripheral blood ENA-78 levels (70 +/- 26 ng/ml) were greater than normal peripheral blood levels (0.12 +/- 0.04 ng/ml) (P < 0.05). Anti-ENA-78 antibodies neutralized 42 +/- 9% (mean +/- SE) of the chemotactic activity for PMNs found in RA synovial fluids. Isolated RA synovial tissue fibroblasts in vitro constitutively produced significant levels of ENA-78, and this production was further augmented when stimulated with tumor necrosis factor-alpha (TNF-alpha). In addition RA and osteoarthritis synovial tissue fibroblasts as well as RA synovial tissue macrophages were found to constitutively produce ENA-78. RA synovial fluid mononuclear cells spontaneously produced ENA-78, which was augmented in the presence of lipopolysaccharide. Immunohistochemical localization of ENA-78 from the synovial tissue of patients with arthritis or normal subjects showed that the predominant cellular source of this chemokine was synovial lining cells, followed by macrophages, endothelial cells, and fibroblasts. Synovial tissue macrophages and fibroblasts were more ENA-78 immunopositive in RA than in normal synovial tissue (P < 0.05). These results, which are the first demonstration of ENA-78 in a human disease state, suggest that ENA-78 may play an important role in the recruitment of PMNs in the milieu of the inflamed joint of RA patients.
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PMCID: PMC295150  PMID: 8083342
14.  Pulmonary lesions induced by Pasteurella haemolytica in neutrophil sufficient and neutrophil deficient calves. 
The role of neutrophils in the development of peracute lung lesions of bovine pneumonic pasteurellosis was investigated. Eight calves were divided into two groups of four calves each. Group I was treated with intravenous phosphate-buffered saline and served as the neutrophil sufficient calves. Group II was treated with intravenous hydroxyurea which produced a state of neutropenia. When peripheral blood neutrophil numbers dropped below 300 cells/microL in group II, all calves were challenged with an intrabronchial bolus of Pasteurella haemolytica in the log phase of growth. An acute inflammatory process occurred in both groups of calves indicated by a rise in body temperature. While pulmonary lesions occurred in both groups by six hours postinoculation, they varied in pathological characteristics. Pulmonary lesions in the neutrophil sufficient calves consisted of fibrinopurulent alveolitis-bronchiolitis with associated alveolar septal necrosis, interlobular edema, and intravascular thrombi. The neutrophil deficient calves had extensive intra-alveolar edema, interlobular edema, intraalveolar hemorrhage, atelectasis, and focal areas of alveolar septal necrosis. These results show that P. haemolytica can induce severe pulmonary tissue damage through both neutrophil dependent and neutrophil independent mechanisms.
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PMCID: PMC1255428  PMID: 3370555
15.  Phagocytic Function of Polymorphonuclear Leukocytes in Rheumatic Diseases 
Journal of Clinical Investigation  1973;52(7):1632-1635.
Phagocytosis of yeast particles by peripheral blood and synovial fluid neutrophils was compared in the sera and synovial fluids from 16 osteoarthritis, 23 rheumatoid arthritis, and 12 miscellaneous arthritis patients. Phagocytosis by normal peripheral blood neutrophils was decreased equally and significantly in all synovial fluids. All synovial fluid neutrophils demonstrated decreased phagocytic capacity in all media. Rheumatoid arthritis synovial fluid neutrophils showed significantly less phagocytosis than miscellaneous arthritis synovial fluid neutrophils. Normal peripheral blood neutrophils which in vitro had previously ingested monosodium urate crystals or oil red O, subsequently exhibited a normal yeast phagocytic capacity. Normal peripheral blood neutrophils, which had ingested preformed immunoglobulin G-rheumatoid factor complexes exhibited significantly less yeast phagocytic capacity than control cells or cells preincubated with the individual complex components. There was a significant correlation between the log of the reciprocal of the rheumatoid factor titer in sera used to produce complexes and the phagocytic capacity exhibited by test neutrophils. Ingestion of immunoglobulin G-rheumatoid factor complexes may be important in the production of the cellular phagocytic defect which this study has demonstrated in rheumatoid arthritis synovial fluid neutrophils.
PMCID: PMC302436  PMID: 4578156
16.  The association between serological evidence of mycoplasma infection and respiratory disease in feedlot calves. 
Calves from five Ontario feedlots were bled on arrival and approximately 28 days later. Calves treated during this interval for undifferentiated respiratory disease were classified as cases and untreated calves were classified as controls. Serum was titrated blindly for antibodies to Mycoplasma bovis and Mycoplasma dispar. Indirect hemagglutination titers of 1:20 or more were assumed to reflect recent or current exposure, whereas 1:10 or less were not. The titers to M. bovis increased in all feedlots indicating active infection. The initial titers to M. dispar were higher than the titers against M. bovis, yet they increased in all feedlots except one, suggesting widespread infection with this organism. There was an increased risk (although not statistically significant) of being treated if the titer against M. bovis rose during the period. Calves with low M. dispar titers on arrival were at increased risk of being treated and titer increases were strongly associated with treatment (statistically significant). Thus, the serological results indicate high prevalence of M. bovis and M. dispar in the feedlot calves and that calves with increasing titers in particular to M. dispar are at increased risk of being treated for respiratory disease.
PMCID: PMC1255186  PMID: 3756671
17.  Rubella Virus and Chronic Joint Disease: Is There an Association? 
Journal of Clinical Microbiology  1998;36(12):3524-3526.
Synovial fluid samples and/or biopsies from 79 patients with various chronic inflammatory joint diseases or traumatic joint injury were tested for rubella virus (RV) in order to confirm or refute results from other studies that suggested RV as a cause of chronic inflammatory joint disease. Sixty-eight of the 72 patients tested had RV antibodies. RV RNA was detected by reverse transcription-PCR in the synovial fluid cells from two patients. RV was also isolated by cell culture from the synovial fluid of one of these two patients. This patient was a 42-year-old female with common variable immune deficiency and Mycoplasma hominis arthritis, while the other was a 68-year-old female with rheumatoid arthritis. While these results fail to confirm that RV is associated with chronic inflammatory joint disease, they suggest that RV may persist within a joint and be reactivated when cell-mediated immunity is suppressed.
PMCID: PMC105233  PMID: 9817866
18.  Tumour necrosis factor in synovial exudates. 
Annals of the Rheumatic Diseases  1988;47(9):768-772.
The actions of tumour necrosis factor (TNF) include resorption of bone and cartilage, suggesting a potential role in the pathogenesis of arthritis. TNF activity was looked for in synovial fluids from 137 patients with different rheumatic diseases. Unfractionated samples were tested in the L929 bioassay. Significant TNF activity that was neutralised by monoclonal antibody to TNF alpha occurred in 13 (30%) of 44 samples. Raised TNF levels were not associated with any particular disease type or routine laboratory markers of inflammation but were related to disease duration in osteoarthritis. The finding of biologically active TNF in symptomatic joints of arthritic patients supports the idea that it may contribute to the pathogenesis of joint damage in chronic rheumatic diseases.
PMCID: PMC1003595  PMID: 3263088
19.  New models of chronic synovitis in rabbits induced by mycoplasmas: microbiological, histopathological, and immunological observations on rabbits injected with Mycoplasma arthritidis and Mycoplasma pulmonis. 
Infection and Immunity  1977;16(1):382-396.
A dose-dependent chronic synovitis was induced in rabbit knees after the intra-articular injection of both Mycoplasma arthritidis and Mycoplasma pulmonis. The inflammation progressed from an initial acute phase at 1 week characterized by edema, infiltration of the synovium with monocytes and heterophils, and desquamation of lining cells, to a more chronic phase at 1 and 3 months, in which villus hyperplasia, lymph "nodules," mononuclear cell infiltration, fibroplasia, and collagen deposition were prominent. With one exception, mycoplasmas could no longer be cultivated from the joints 1 month postinoculation. Both mycoplasma species evoked a humoral antibody response that was more marked in synovial fluids than in peripheral blood. A cell-mediated immune reaction, as evidence by enhanced uptake by [3H]thymidine by sensitized blood, spleen, or node lymphocytes in the presence of homologous antigen, was detected only in rabbits injected with M. pulmonis. Lymphocytes taken from arthritic rabbits were no more cytotoxic toward synovial cells derived from normal or arthritic rabbits than were normal lymphocytes. The models of synovitis described in this study offer a convenient probe for determining the mechanisms of mycoplasma-induced inflammation, since they require only a single injection of the initiating agent and, in addition, utilize an animal host large enough for detailed investigation into the nature of mycoplasma/synovium interactions.
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PMCID: PMC421532  PMID: 873616
20.  Effect of repeated arthrocentesis and single joint lavage on cytologic evaluation of synovial fluid in 5 young calves 
The objective of this study was to evaluate the effect of repeated arthrocentesis and a single joint lavage on cytologic variables of synovial fluid. The left tarsi of 5 healthy Holstein calves were selected for the study. Samples of synovial fluid were collected daily for 4 d, then every 4 d until day 24. On day 2, joint lavage was performed with lactated Ringer’s solution in all the calves. Cytologic examinations, performed by the same clinical pathologist, included the determination of total protein concentration, total leukocyte count, and differential counts (of neutrophils, lymphocytes, and monocytes). The presence of lameness or swelling and other results of physical examination were recorded regularly during the study. No clinical signs of joint disease were observed during the study. Bacterial cultures of specimens collected on day 2 were negative for all the calves. All cytologic values but 1 peaked on day 2 and progressively returned to normal. In comparison with the results for day 1, the synovial fluid total leukocyte, neutrophil, and monocyte counts were significantly increased on days 2 and 3, and the total leukocyte and monocyte counts were also significantly increased on day 4. The monocyte and lymphocyte percentages were significantly decreased until day 4, whereas the neutrophil percentages were significantly increased until day 8. The total protein concentrations were significantly increased until day 3. There were no significant differences between values for specimens taken 4 d apart. This study demonstrated that, although arthrocentesis induces a moderate inflammatory response, the joints seem to rapidly adapt. A 4-d interval between arthrocenteses is suitable when studying cellular components of the synovial fluid. However, when arthrocentesis is repeated daily, a minimal interval of 8 d should be respected.
PMCID: PMC1829182  PMID: 17479776
21.  Effect of intravenous iron-dextran (Imferon) infusion on antigen induced monarticular arthritis in rabbits. 
Annals of the Rheumatic Diseases  1992;51(11):1237-1241.
The effect of intravenously infused iron-dextran (Imferon) on the progression of antigen induced monarticular arthritis in rabbits was studied. A rapid deposition of iron and apoferritin in the synovia of arthritis joints occurred after infusion of iron-dextran during either the acute or chronic phases of the disease. This coincided with the appearance of catalytic (bleomycin reactive) iron in the synovial fluid. There was no evidence, however, for an exacerbation of the antigen induced arthritis as a result of the iron-dextran, and synovial and bleomycin reactive iron concentrations decreased with time after administration, indicating a redistribution of the synovial iron load. Thus although intravenously infused iron-dextran appears to 'prime' the rabbit arthritic joint transiently with the potential for iron stimulated oxygen free radical damage, other factors may determine its occurrence.
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PMCID: PMC1012462  PMID: 1466600
22.  The frequency, distribution and effects of antibodies, to seven putative respiratory pathogens, on respiratory disease and weight gain in feedlot calves in Ontario. 
During 1983-85, 279 calves requiring treatment for bovine respiratory disease and 290 comparison (control) animals from 15 different groups of feedlot calves were bled on arrival and again at 28 days postarrival. Their sera were then analyzed for antibodies to seven putative respiratory pathogens. On arrival, the prevalences of indirect agglutination titers to Pasteurella haemolytica, P. haemolytica cytotoxin, Mycoplasma bovis and M. dispar were greater than 50%, the prevalence of titers to bovine virus diarrhea virus (BVDV) was approximately 40%, and the prevalences of titers to infectious bovine rhinotracheitis virus (IBRV), bovine respiratory syncytial virus (RSV) and parainfluenza virus type 3 (PIV3) were all below 25%. Seroconversion during the first month after arrival occurred in more than half the calves to P. haemolytica cytotoxin, PIV3 and RSV. Seroconversion of agglutination titers to P. haemolytica, Mycoplasma and BVDV occurred in about 40% of calves, and seroconversion to IBRV was infrequent (less than 5%). Initial titers were negatively correlated to subsequent titer changes within organism. Initial titers, and titer changes between organisms were essentially independent. Light calves had an increased risk of being selected for treatment for respiratory disease. Seroconversion to P. haemolytica cytotoxin, RSV and BVDV were predictive of respiratory disease cases, explaining approximately 69% of all respiratory disease cases in the feedlots. It was not possible to accurately predict weight gain or relapse from the serological data.
PMCID: PMC1255725  PMID: 2766158
23.  Concentrations of glycosaminoglycans in synovial fluids and their relation with immunological and inflammatory mediators in rheumatoid arthritis. 
Annals of the Rheumatic Diseases  1990;49(5):301-307.
The dimethylmethylene blue assay showed higher concentrations of glycosaminoglycans in many synovial fluids from patients with rheumatoid arthritis (RA) than in autologous sera or sera or synovial fluids from normal subjects. These results were taken to suggest that the glycosaminoglycans in RA synovial fluid were abnormally raised and derived from cartilage. To discover what stimulated such glycosaminoglycan release in RA joints relations were sought between synovial fluid concentrations of glycosaminoglycans and immunological and inflammatory mediators. It was shown that RA synovial fluid glycosaminoglycan concentrations correlated with synovial fluid C3d concentrations but not with synovial fluid rheumatoid factor concentrations, polymorphonuclear leucocyte numbers, myeloperoxidase concentrations, or the ability of the synovial fluids to release free radicals from normal polymorphonuclear leucocytes. A correlation was found between synovial fluid C3d and interleukin 1 concentrations as judged by both lymphocyte activating factor activity and immunoassay, but no significant correlation was detected between interleukin 1 and glycosaminoglycan concentrations. It is suggested that in the rheumatoid joint locally produced cytokines, in addition to interleukin 1, together stimulate glycosaminoglycan release from cartilage and render it vulnerable to attack by other processes.
PMCID: PMC1004073  PMID: 2344209
24.  Isolation and identification of mycoplasma agalactiae subsp. bovis from arthritic cattle in Iowa and Nebraska. 
Journal of Clinical Microbiology  1975;2(3):169-172.
Two strains of Mycoplasma were isolated from synovial fluids of arthritic feeder cattle and were identified as Mycoplasma agalactiae subsp. bovis by growth inhibition and fluorescent antibody tests. The strains (Iowa 1136 and Nebraska 2) could not be distinguished from known strains (Donetta and California 01) by immunoelectrophoresis or by agar gel precipitation.
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PMCID: PMC274165  PMID: 1176624
25.  Bonding the foe – NETting neutrophils immobilize the pro-inflammatory monosodium urate crystals 
In the presence of sodium, uric acid from purine metabolism precipitates as monosodium urate (MSU) needles and forms renal calculi or causes gouty arthritis in kidneys and joints, respectively. The latter is characterized by red, hot, and swollen arthritic joints. Here we report the in vitro effect of MSU crystals on blood granulocytes and analyze their contribution to granuloma formation and neutrophil extracellular traps (NETs) formation (NETosis) in synovial fluid of patients with gouty arthritis in vivo. We observed that MSU crystals induce NETosis in vitro in a reactive oxygen species (ROS)-dependent manner. Indeed, blocking ROS (e.g., the oxidative burst) by various anti-oxidants partially inhibited NETosis induced by MSU crystals. Analyses of synovial fluids and of tissue sections of patients suffering from gout revealed that NETs are also formed in vivo, especially during acute gouty flares and/or granuloma formation. Since prolonged exposure to NETs carries the risk for the development of chronic inflammation we also studied the opsonization of NETs, as a prerequisite for their clearance. The established dead cells’ opsonins C3b, galectin-9, and CRP decorated the residual dead cells’ corpses and opsonized these for disposal. Surprisingly, all three soluble pattern recognizing molecules spared the spread NET structures. We conclude that (i) MSU crystals are strong inducers of ROS-dependent NETosis and (ii) that the prolonged presence of NET-pathogen or NET-crystal aggregates observed in patients with systemic autoimmunity, especially in those with low serum DNase-1 activity, cannot be compensated by CRP, complement, and galectin-mediated phagocytic clearance.
doi:10.3389/fimmu.2012.00376
PMCID: PMC3517988  PMID: 23233855
NETosis; NETs; MSU; opsonins; inflammation; ROS; gout

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