Qualitative and quantitative comparison of allergen component-specific to birch and grass analyzed by ImmunoCAP assay and Euroline immunoblot test.

Summary: Background.In the diagnostic work up of allergy, determining allergen component-specific immunoglobulin E (IgE) is important for diagnosis, prognosis and choice of treatment. The purpose of this study was to evaluate the performance of the immunoblotting assay (Euroline) in detection of IgE antibodies against timothy grass and birch pollen allergen components compared to fluorescent enzyme assay (ImmunoCAP, Phadia 250). Methods. A total of 128 serum samples from patients allergic to timothy grass and birch pollen were analysed. The levels of IgE antibodies to timothy grass and birch pollen were measured using Euroline DPA-Dx pollen 1 and ImmunoCAP assay. The two methods were then compared on binary (positive vs negative), semi-quantitative (IgE classes) and quantitative (concentration) levels. The two methods were also compared to results from skin prick testing. Results. The Euroline method showed a positive percentage agreement of 93% and negative percentage agreement of 94% with an overall accuracy of 94% when compared to ImmunoCAP. Kappa analysis showed moderate strength of agreement between the methods in determining IgE classes for 7/11 components tested. All components showed a positive correlation when analysed using Spearman's rank correlation. Conclusions. Overall, we found that there is good correlation between the Euroline and ImmunoCAP methods in measuring IgE sensitization.

Pre-treatment allergen-specific IgE analysis and outcomes of allergen immunotherapy.

Summary: Background. Patients show varied results to allergen immunotherapy (AIT. The reason for this variability is unclear. Objective. To describe the relationship between AIT efficacy and demographic characteristics, as well as pre-treatment plasma levels of specific IgE-antibodies to grass and birch pollen. Methods. A retrospective study was performed based on medical records of 128 patients who received AIT. The patients completed a questionnaire and pre-AIT plasma levels of allergen-specific IgE to grass and birch pollen were measured using EUROLINE DPA-Dx pollen 1 method. Results. Seventy percent of patients classified their allergic symptoms as less severe after AIT. Twenty-seven percent had received AIT targeting only grass pollen, 19% targeting only birch pollen, and 55% targeting both grass and birch. A total of 35 different IgE profiles were found across our study population. On comparison of the demographic characteristics and concentration of allergen-specific IgE-antibodies, no statistically significant differences could be found.Conclusions. The majority of patients rated their allergic symptoms as less severe after AIT. No clear relationship could be demonstrated between pre-treatment allergen-specific IgE concentration, or demographic characteristics, and effect of AIT. There may be other factors underlying the different responses to AIT.

Chironomid midge sensitization in sewage workers: case study.

Non-biting chironomid midges (Diptera: Chironomidae) may cause sensitization and allergic reactions in humans and have recently been identified as a potential health problem in Swedish municipal sewage treatment plants. To investigate, on a pilot scale, the allergenic potential of chironomids in sewage workers, all workers (n = 8) at a sewage treatment plant and local controls (n = 16) completed a symptom questionnaire, underwent measurement of the fraction of nitric oxide in exhaled air, spirometry, and provided serum samples for the determination of atopy status and the prevalence of specific immunoglobulin E (IgE) antibodies against Chironomus thummi (Chi t) using a commercial fluorescence enzyme immunoassay (FEIA). Three sewage workers (38%) but no controls (0%) were FEIA positive for C. thummi-specific IgE antibodies (P < 0.05). No other health-related findings were significantly different between the groups. The study suggested that occupational exposure to Chironomids may cause sensitization with circulating IgE-antibodies in sewage workers.

Toll-like receptor cross-hyporesponsiveness is functional in interleukin-1-receptor-associated kinase-1 (IRAK-1)-deficient macrophages: differential role played by IRAK-1 in regulation of tumour necrosis factor and interleukin-10 production.

Signalling downstream Toll-like receptors (TLR) is regulated at several levels in order to activate the immune response and prevent excessive inflammation. Altered intracellular signalling may be one reason that repeated stimulation of various TLRs results in hyporesponsiveness and cross-tolerance. We report that TLR cross-tolerance is inducible in the absence of interleukin-1 receptor-associated kinase-1 (IRAK-1) in peritoneal macrophages. Similar to wild-type macrophages, IRAK-1-deficient macrophages respond with decreased tumour necrosis factor (TNF) production to a secondary TLR stimulation, but in opposite to IRAK-1(+/+), IRAK-1(-/-) macrophages display increased interleukin (IL)-10 production at TLR restimulation. IRAK-1-deficient peritoneal macrophages have a defective TNF and IL-10 production in response to lipoteichoic acid stimulation as well as a defective IL-10-but a normal TNF production in response to high concentration of lipopolysaccharide. Our results demonstrate that IRAK-1 is not necessary for induction of TLR cross-tolerance as judged by TNF production.

Serum concentration of interleukin-18 is up-regulated in patients with ANCA-associated vasculitis.

We investigated circulating interleukin-18 concentrations in patients with ANCA-associated vasculitis (ASV) and healthy control subjects, and included a group of hemodialysis patients, a patient group previously reported to show high IL-18 plasma levels. Anti-proteinase 3 (PR3) and anti-myeloperoxidase (MPO) serum levels were also measured. Interestingly we found significantly increased serum IL-18 concentrations in ASV patients as compared to healthy controls, 437 vs. 185 pg/ml (p < 0.0001). The increase of IL-18 production was similar irrespective of presence of autoantibodies to PR3 or MPO. As expected the hemodialysis patients also showed significantly increased circulating IL-18 concentrations as compared to control subjects.

Serum interleukin-1 receptor antagonist is an early indicator of colitis onset in Galphai2-deficient mice.

AIM: To study the serum concentration of IL-1beta, IL-1 receptor antagonist (IL-1Ra) and IL-18 in Galphai2-deficient mice at the age of 6 (healthy), 12 (pre-colitic) and 24 wk (colitic) and in healthy control mice. METHODS: At the time of killing, serum samples were collected and IL-1beta, IL-1Ra and IL-18 levels were measured using enzyme-linked immunosorbent assays. RESULTS: Serum concentration of IL-1Ra was significantly increased in pre-colitic (median: 524 ng/L; P=0.02) and colitic (450 ng/L; P=0.01), but not in healthy (196 ng/L) Galphai2-deficient mice as compared with controls (217 ng/L). Serum concentrations of IL-1beta did not differ between Galphai2-deficient mice and their controls, irrespective of age, IL-18 was significantly increased in colitic, but not in pre-colitic mice compared with controls (510 ng/L vs 190 ng/L; P=0.05). CONCLUSION: The increased serum concentrations of IL-18 and IL-1Ra in established diseases are suggested as markers of ongoing colitis. Interestingly, the significantly increased serum concentration of IL-1Ra in pre-colitic mice is found to be an early marker of disease progression.

Acellular Bordetella pertussis vaccine enhances mucosal interleukin-10 production, induces apoptosis of activated Th1 cells and attenuates colitis in Galphai2-deficient mice.

Mice deficient for the inhibitory G protein subunit alpha2 (Galphai2(-/-)) spontaneously develop a progressive inflammatory bowel disease resembling ulcerative colitis, and have a T helper 1 (Th1)-dominated immune response prior to onset of colitis, which is further augmented after the onset of disease. The present study was performed to investigate whether the Galphai2(-/-) mice were able to down-regulate the Th1-dominated inflammatory mucosal immune response and/or induce an anti-inflammatory Th2/T regulatory response and thereby diminish the severity of colitis following treatment with acellular Bordetella pertussis vaccine. The acellular vaccine against B. pertussis, the causative agent of whooping cough, has been demonstrated to induce a Th2-mediated response in both man and mice. We therefore treated Galphai2(-/-) mice intraperitoneally with a three-component acellular B. pertussis vaccine. The treated Galphai2(-/-) mice showed significantly increased interleukin (IL)-10 production in intestinal tissue, associated with significantly reduced colitis and decreased mortality, compared to untreated Galphai2(-/-) mice. The attenuation of colitis in Galphai2(-/-) mice was due, at least partly, to the B. pertussis surface antigen filamentous haemagglutinin (FHA), which almost completely inhibited proliferation of CD4(+) T cells and stimulated apoptosis of activated CD4(+) T helper 1 cells. In conclusion, the three-component acellular B. pertussis vaccine containing filamentous haemagglutinin increases the production of IL-10 in the intestinal mucosa, induces apoptosis of activated Th1 cells and attenuates colitis in Galphai2(-/-) mice.

Is interleukin-18 useful for monitoring rheumatoid arthritis?

OBJECTIVE: Interleukin-18 (IL-18) is a proinflammatory regulator of immune responses. Its similarities to IL-1beta and ability to induce tumour necrosis factor-alpha (TNF-alpha) make it potentially important in the pathogenesis of rheumatoid arthritis (RA). METHODS: The level of IL-18 was assessed in matched pairs of blood and synovial fluid samples from 90 RA patients (47 erosive, 43 non-erosive) by an enzyme-linked immunosorbent assay (ELISA), and the results compared to 40 healthy controls. RESULTS: In RA patients with erosive joint disease, the IL-18 level was higher than that in non-erosive RA [(median+/-QR) blood: 385+/-200 vs. 235+/-183 pg/mL, p = 0.02; synovial fluid: 392+/-392 vs. 224+/-324 pg/mL, p = 0.05]. IL-18 levels in blood of RA patients were similar and closely related to the local, intra-articular level (r = 0.96). The IL-18 level was not related to other markers of inflammation, to the duration of RA, or to the treatment modality. The IL-18 level in RA patients was similar to that of the controls (278+/-234 vs. 344+/-179 pg/mL, not significant). CONCLUSIONS: An increased IL-18 level is associated with erosive joint disease, but the measurement of IL-18 does not help to distinguish between RA patients and healthy controls.

Leptin consumption in the inflamed joints of patients with rheumatoid arthritis.

BACKGROUND: Leptin has been shown to participate in bone remodelling and leptin substitution reported to have a protective effect in experimental septic arthritis. OBJECTIVE: To assess leptin levels in inflamed joints and plasma of patients with RA. MATERIAL AND METHODS: Leptin concentrations were assessed in matched blood and synovial fluid samples from 76 patients with RA. Blood samples from 34 healthy subjects acted as additional controls. Results were analysed and correlated with duration and activity of RA, x ray changes, and treatment at time of sampling. RESULTS: In patients with RA, leptin levels were significantly higher in plasma than in synovial fluid samples obtained simultaneously and higher than in control samples. Plasma and synovial fluid leptin levels correlated strongly. Locally in the joint, leptin levels were related to WBC count. Such a relation was not seen in the bloodstream. Leptin levels were not related to sex, age, or disease duration. Difference between leptin levels in plasma and synovial fluid was greater in non-erosive arthritis (5.1 (SEM 1.2) v 3.7 (0.9) ng/ml, p=0.006), than in patients with erosive joint disease (6.2 (1.0) v 5.4 (0.8) ng/ml, NS). Methotrexate treatment was associated with relatively high plasma leptin levels, while treatment with other DMARDs was associated with lower leptin levels than in patients receiving no DMARD treatment (p=0.0005). CONCLUSIONS: Leptin production was significantly increased in patients with RA compared with healthy controls. Synovial fluid leptin levels were significantly lower than in matched plasma samples, suggesting an in situ consumption of this molecule.

Interleukin-10 ameliorates the outcome of Staphylococcus aureus arthritis by promoting bacterial clearance.

Staphyllococcus aureus-induced infections often result in high mortality and permanent joint destruction, despite treatment with antibiotics. IL-10 is typically regarded as an anti-inflammatory cytokine because it promotes a T helper cell type 2 response, and subsequently down-regulates cell mediated immune functions. To investigate the role of IL-10 in S. aureus-induced arthritis and sepsis, Balb/c mice, intact or defective with respect to IL-10 gene were intravenously inoculated with bacteria. IL-10-/- mice develop a more frequent and destructive arthritis compared to their congeneic controls. The mechanisms regulating such outcome may be due not only to the anti-inflammatory properties of IL-10 but also, directly or indirectly, to antibacterial features of this molecule. Indeed, inoculation of staphylococci to IL-10-/- mice resulted in higher bacterial load in blood and kidneys compared to congeneic controls. Altogether our data indicate that IL-10 is essential for efficient elimination of bacteria and thereby for protection against septic arthritis.

Leptin in septic arthritis: decreased levels during infection and amelioration of disease activity upon its administration.

Weight loss is typically found during severe infections, e.g. septic arthritis. The aim of our study was to evaluate the role of leptin, regulator of food intake and energy expenditure, for the development of Staphylococcus aureus-triggered arthritis. Leptin production was found to be decreased during murine S. aureus-induced arthritis. Treatment with recombinant leptin neither restored the basal leptin levels nor affected the weight loss during the disease, but it significantly decreased the severity of septic arthritis. Exogenous leptin did not affect the staphylococcal load as measured in blood, joints and kidneys. Preceding the effects on joint manifestations, serum levels of interleukin-6 decreased in leptin-treated mice. In conclusion, the treatment with recombinant leptin reduced both the severity of joint manifestations in S.aureus-induced arthritis and the inflammatory response, as measured by serum IL-6 levels, without affecting the survival of bacteria in vivo.

Model systems: modeling human staphylococcal arthritis and sepsis in the mouse.

The staphylococci have been recognized as serious pathogens for over a century and are the etiological agent of a variety of diseases ranging from mild cutaneous infections to often fatal forms of septic arthritis, endocarditis, toxic shock syndrome and sepsis. Despite intensive efforts to halt their spread, they remain the most common cause of community- and nosocomially acquired bacteremia. Murine models of Staphylocococus aureus-mediated arthritis and sepsis exist and are being used to gain a better understanding of the host-bacterium relationship as well to develop better methods of prevention and treatment.

Impact of transcription factors AP-1 and NF-kappaB on the outcome of experimental Staphylococcus aureus arthritis and sepsis.

Staphylococcus aureus infection is, despite adequate antibiotic treatment, a disease characterized by high mortality. The bacterium triggers an exaggerated immune response in the host, which on the one hand acts as an efficient defense, but on the other hand gives rise to tissue damage. In this study we have modulated the hosts response to S. aureus by inhibition of nuclear factor kappaB (NF-kappaB) and activator protein-1 (AP-1)-triggered release of pro-inflammatory cytokines and tissue-destructive proteins, respectively. Mice were administered with antisense oligonucleotides (ODN) to the p65 subunit of NF-kappaB and/or a double-stranded oligonucleotide (mCoAP-1) with homology to the murine AP-1 binding site of collagenase IV gene (metalloproteinase-9; MMP-9), solely or in combination with antibiotics. In mice systemically treated with antisense ODN to NF-kappaB p65 alone, the bacterial burden in the kidneys was significantly increased (P = 0.04) The same tendency was seen when mCoAP-1 was administered either alone or in combination with antibiotics. We also found significantly (P = 0.04) elevated levels of IL-6 in p65 antisense treated mice. Surprisingly, this p65 antisense therapy approach, which has turned out to be highly efficient in amelioration of aseptic arthritis and colitis, failed to change the clinical course of either septic arthritis or sepsis. We suggest that interaction with transcription factors leads to increased bacterial burden in vivo, abrogating the potential benefits of the anti-inflammatory properties exerted by these compounds.

Role of IL-12 in Staphylococcus aureus-triggered arthritis and sepsis.

The present study demonstrates that endogenous production of IL-12 is crucial for survival in Staphylococcus aureus-induced arthritis in mice. Staphylococcal load is enhanced in several organs, because of lack of IL-12. This might be due to decreased production of IFN-gamma in IL-12-deficient mice. Although IL-12-deficient mice were exposed to higher staphylococcal load, they demonstrated no increased severity of arthritis as compared with control animals.

Are B lymphocytes of importance in severe Staphylococcus aureus infections?

To investigate the role of B cells in experimental, superantigen-mediated Staphylococcus aureus arthritis and sepsis, we used gene-targeted B-cell-deficient mice. The mice were inoculated intravenously with a toxic shock syndrome toxin 1 (TSST-1)-producing S. aureus strain. The B-cell-deficient and thus agamma-globulinemic mice showed striking similarities to the wild-type control animals with respect to the development of arthritis, the mortality rate, and the rate of bacterial clearance. Surprisingly, we found that the levels of gamma interferon in serum were significantly lower (P < 0. 0001) in B-cell-deficient mice than in the controls, possibly due to impaired superantigen presentation and a diminished expression of costimulatory molecules. In contrast, the levels of interleukin-4 (IL-4), IL-6, and IL-10 in serum were equal in both groups. Our findings demonstrate that neither mature B cells nor their products significantly contribute to the course of S. aureus-induced septic arthritis.

Outcome of Staphylococcus aureus-triggered sepsis and arthritis in IL-4-deficient mice depends on the genetic background of the host.

Disruption of the IL-4 gene in two inbred mouse strains revealed a dual role of IL-4 in Staphylococcus aureus sepsis and arthritis depending on the host's genetic background. IL-4 was protective in 129SV mice, since 5 days after S. aureus inoculation IL-4(-/-) mice displayed 70% mortality as compared to survival of all 129SV wild-type counterparts. On the other hand, IL-4 was detrimental in C57BL/6 mice, since survival of IL-4(-/-) C57BL/6 mice was increased, as compared to wild-type controls, due to decreased staphylococcal growth. Altogether, our results show the dual role of IL-4 in S. aureus sepsis and arthritis, depending on the genetic background of the host.

TNF/lymphotoxin-alpha double-mutant mice resist septic arthritis but display increased mortality in response to Staphylococcus aureus.

To evaluate the importance of the proinflammatory cytokines TNF and lymphotoxin-alpha (LT alpha) in an experimental model of Staphylococcus aureus sepsis and arthritis, we used TNF/LT alpha-double-deficient mice raised on the C57BL/6 background. Mice were i.v. inoculated with a toxic shock syndrome toxin-1 (TSST-1)-producing S. aureus strain, LS-1. Intravenous inoculation of a high dose of bacteria (1 x 10(7)/mouse) resulted in 67% mortality in TNF/LT alpha-deficient mice, whereas none of the controls died (p = 0.009). Those results correlated to a significantly decreased phagocytosis in vitro and inefficient bacterial clearance in vivo in mice lacking capacity to produce TNF/LT alpha. Thus, at day 6 after inoculation, S. aureus could not be found in the bloodstream of controls, but bacteremia developed in all TNF/LT alpha-deficient mice examined (p = 0.02). Interestingly, upon infection with a lower dose of staphylococci (3 x 10(6)/mouse) the mortality was overall low, but the frequency of arthritis was clearly higher in the wild-type group as compared with the TNF/LT alpha-deficient mice (40% vs 13%). Histopathologic examination revealed a lower frequency of synovitis (38% vs 90%, p < 0.05) and erosivity (25% vs 60%, NS) in TNF/LT alpha-deficient mice as compared with wild-type counterparts. Our results show the importance of TNF/LT alpha in defense against systemic S. aureus infections and point out the detrimental role of these cytokines as mediators of inflammatory response in S. aureus arthritis.

Staphylococcus aureus-induced septic arthritis and septic death is decreased in IL-4-deficient mice: role of IL-4 as promoter for bacterial growth.

Lack of IL-4 has been shown to be protective in some experimental models of infectious diseases in mice such as cutaneous leishmaniasis. At the same time IL-4, together with other Th2 cytokines, including IL-10 and IL-13, is known as an anti-inflammatory cytokine with the potential to down-regulate proinflammatory cytokine production. To investigate the role of IL-4 in experimental Staphylococcus aureus-induced and T lymphocyte-mediated arthritis, IL-4-deficient C57BL/6 mice (IL-4(-/-)) and their congenic controls (IL-4(+/+)) were inoculated with a toxic shock syndrome toxin-1-producing S. aureus strain. In IL-4(+/+) mice, arthritis peaked 14 days after bacterial inoculation, whereas, at that time, IL-4(-/-) mice displayed significantly less frequent (p < 0.05) joint inflammation. Paralleling lower frequency of arthritis, IL-4-deficient mice showed a decreased bacterial burden in joints (p = 0.014) and kidneys (p = 0.029), as well as lower infection-triggered weight decrease and mortality. In vitro, IL-4 inhibited intracellular killing of S. aureus in infected macrophages, without affecting phagocytosis. This finding may explain the enhanced staphylococcal clearance observed in IL-4(-/-) mice in vivo. Our results suggest that IL-4 and IL-4-dependent Th2 responses promote septic arthritis and sepsis-related mortality by inhibition of bacterial clearance during S. aureus infection.