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The increasing age of populations throughout the world means that healthcare services are faced with new challenges, not least regarding the provision of food during hospital stay. There is a lack of knowledge of how hospital mealtimes are experienced by older patients, and so the aim of this article was to review current knowledge regarding mealtimes in hospitals from the perspectives of older patients. A literature search was performed using seven databases: PubMed, Web of Science, Scopus, Sociological Abstracts, SweMed+, ASSIA and CINAHL with no limits regarding publication date. The inclusion criteria were peer-reviewed articles in English or Swedish that used qualitative methods to examine older patients' (>65 years) mealtime experiences. The Five Aspect Meal Model (FAMM) served as a framework for understanding the complexity behind a mealtime experience. Qualitative content analysis was used as a guide when analysing the material. The search produced 415 studies, 14 of which were included in the review. The findings generated three main themes for understanding how older patients experience mealtimes while in hospital: (1) the food and the food service, (2) mealtime assistance and commensality during mealtimes and (3) the importance of retaining one's independence. The review also clearly indicated a shortage of studies that solely focus on older patients' experiences of their mealtime. More research is therefore needed to be fully able to understand the complex task of providing meals in hospitals., (© 2020 The Authors. Scandinavian Journal of Caring Sciences published by John Wiley & Sons Ltd on behalf of Nordic College of Caring Science.)

Background: Signalling through insulin-like growth factor 1 receptor (IGF-1R) is essential for cell survival, but may turn pathogenic in uncontrolled tissue growth in tumours. In rheumatoid arthritis (RA), the IGF-1R signalling is activated and supports expansion of the inflamed synovia., Aim: In the present study, we assess if disruption of IGF-1R signalling resolves arthritis., Material and Methods: Clinical associations of IGF-1R expression in leukocytes of the peripheral blood were studied in 84 RA patients. Consequences of the IGF-1R signalling inhibition for arthritis were studied in mBSA immunised Balb/c mice treated with NT157 compound promoting degradation of insulin receptor substrates., Results: In RA patients, high expression of IGF-1R in leukocytes was associated with systemic inflammation as verified by higher expression of NF-kB, serum levels of IL6 and erythrocyte sedimentation rate, and higher pain perception. Additionally, phosphorylated IGF-1R and STAT3 enriched T cells infiltrate in RA synovia. Treatment with NT157 inhibited the phosphorylation of IGF-1R and STAT3 in synovia, and alleviated arthritis and joint damage in mice. It also reduced expression of IGF-1R and despaired ERK and Akt signalling in spleen T cells. This limited IL-6 production, changed RoRgt/FoxP3 balance and IL17 levels., Conclusion: IGF-1R signalling contributes to T cell dependent inflammation in arthritis. Inhibition of IGF-1R on the level of insulin receptor substrates alleviates arthritis by restricting IL6-dependent formation of Th17 cells and may open for new treatment strategies in RA., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Osteoclasts are bone-resorbing cells that accumulate in the joints of patients with rheumatoid arthritis causing severe bone damage. Fms-like tyrosine kinase 3 ligand is enriched in the synovial fluid of patients with rheumatoid arthritis, and local exposure to Fms-like tyrosine kinase 3 ligand aggravates arthritis in mice. Because Fms-like tyrosine kinase 3 ligand has been suggested to facilitate osteoclast differentiation, we asked whether Fms-like tyrosine kinase 3 ligand affects bone remodeling in arthritis. The effect of Fms-like tyrosine kinase 3 signaling on osteoclast development was studied by immunohistochemistry in methylated bovine serum albumin-induced arthritis using mice that lack the gene for Flt3l (Flt3L(-/-)) and by an in vitro assay. Bone and joint changes were studied morphologically and by microcomputer tomography. We found that Flt3L(-/-) mice had increased accumulations of osteoclasts in the periarticular area of the arthritic joint. This triggered bone destruction and trabecular bone loss. The increased number of osteoclasts in Flt3L(-/-) mice may be a consequence of insufficient expression of interferon regulatory factor 8. Treatment of Flt3L(-/-) mice with Fms-like tyrosine kinase 3 ligand increased expression of interferon regulatory factor 8, reduced the number of osteoclasts in arthritic mice, and promoted trabecular bone formation. Finally, the reduced number of regulatory T cells in the bone marrow of Flt3L(-/-) mice could further contribute to the increased osteoclastogenesis by reducing the ratio of regulatory T cells to T helper 17 cells. This study shows that Fms-like tyrosine kinase 3 ligand may serve as a negative regulator of osteoclast development by promoting transcription of interferon regulatory factor 8 and sustaining a balance between protective regulatory T cells and pathogenic T helper 17 cells in the pathogenesis of arthritis., (© Society for Leukocyte Biology.)

Switched antibody classes are important for efficient immune responses. Aberrant antibody production to otherwise harmless antigens may result in autoimmunity. The protein kinase fms-like tyrosine kinase 3 receptor (Flt3) has an important role during early B-cell development, but the role of Flt3 in peripheral B cells has not been assessed before. Herein we describe a previously unappreciated role for Flt3 in IgG1 class-switch recombination (CSR) and production. We show that Flt3 is reexpressed on B-cell lymphoma 6(+) germinal center B cells in vivo and following LPS activation of peripheral B cells in vitro. Absence of Flt3 signaling in Flt3 ligand-deficient mice results in impaired IgG1 CSR and accumulation of IgM-secreting plasma cells. On activated B cells, Flt3 is coexpressed and functions in synergy with the common-gamma chain receptor family. B cells from Flt3 ligand-deficient mice have impaired IL-4R signaling, with reduced phosphorylation of signal transducer and activator of transcription (Stat) 6, and demonstrate a failure to initiate CSR to IgG1 with low expression of γ1 germ-line transcripts, resulting in impaired IgG1 production. Thus, functional synergy between Flt3 and IL-4R signaling is critical for Stat-mediated regulation of sterile γ1 germ-line transcripts and CSR to IgG1.

Survivin is a proto-oncogene that regulates cell division and apoptosis. It is a molecular marker of cancer. Recently, survivin has emerged as a feature of RA, associated with severe joint damage and poor treatment response. The present study examined if inhibition of survivin affects experimental arthritis, which was induced in mBSA-immunized mice by an injection of mBSA in the knee joint or developed spontaneously in collagen type II-immunized mice. The inhibition of survivin transcription by a lentivirus shRNA construct alleviated joint inflammation and reduced bone damage. The inhibition of survivin reduced the levels of metalloproteinases, β-catenin, and vimentin, limiting the invasive capacity of synovia, while no inhibition of osteoclastogenesis could be found. The inhibition of survivin led to a p53-independent reduction of T cell proliferation and favored the transcription and activity of Blimp-1, which limited IL-2 production and facilitated formation of regulatory Foxp3(+)CD4(+) and effector CD8(+) T cells. The study shows that the inhibition of survivin is sufficient to reduce joint inflammation and bone damage in preclinical models of arthritis. Antiarthritic effects of survivin inhibition are related to p53-independent control of lymphocyte proliferation., (© Society for Leukocyte Biology.)

Objectives: To evaluate the role of S100A4, a calcium-binding regulator of nonmuscle myosin assembly, for T-cell responses in rheumatoid arthritis., Methods: Arthritis was induced in the methylated bovine serum albumin (mBSA)-immunized mice lacking the entire S100A4 protein (S100A4KO) and in wild-type counterparts treated with short hairpin ribonucleic acid (shRNA)-lentiviral constructs targeting S100A4 (S100A4-shRNA). The severity of arthritis was evaluated morphologically. T-cell subsets were characterized by the expression of master transcription factors, and functionally by proliferation activity and cytokine production. The activity of the Scr-kinases Fyn and Lck was assessed by the autophosphorylation of C-terminal thyrosine and by the phosphorylation of the CD5 cytodomain. The interaction between S100A4 and the CD5 cytodomain was analysed by nuclear magnetic resonance spectrophotometry., Results: S100A4-deficient mice (S100A4KO and S100A4-shRNA) had significantly alleviated morphological signs of arthritis and joint damage. Leukocyte infiltrates in the arthritic joints of S100A4-deficient mice accumulated Foxp3(+) Treg cells, while the number of RORγt(+) and (pTyr705)STAT3(+) cells was reduced. S100A4-deficient mice had a limited formation of Th17-cells with low retinoic acid orphan receptor gamma t (RORγt) mRNA and IL17 production in T-cell cultures. S100A4-deficient mice had a low expression and activity of T-cell receptor (TCR) inhibitor CD5 and low (pTyr705)STAT3 (signal transducer and activator of transcription 3), which led to increased (pTyr352)ZAP-70 (theta-chain associated protein kinase of 70kDa), lymphocyte proliferation and production of IL2. In vitro experiments showed that S100A4 directly binds Lck and Fyn and reciprocally regulates their kinase activity towards the CD5 cytodomain. Spectrometry demonstrates an interaction between the CD5 cytodomain and EF2-binding sites of S100A4., Conclusion: The present study demonstrates that S100A4 plays an important part in the pathogenesis of arthritis. It controls CD5-dependent differentiation of Th17 cells by regulating the activity of the Src-family kinases Lck and Fyn., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Objective: S100A4 is a Ca-binding protein that regulates cell growth, survival, and motility. The abundant expression of S100A4 in rheumatiod arthritis contributes to the invasive growth of joint tissue and to bone damage. In the present study, we analysed the role of S100A4 in bone homeostasis., Methods: Peripheral quantitative computed tomography and histomorphometric analysis were performed in mice lacking the entire S100A4 protein (S100A4KO) and in wild-type (WT) counterparts treated with shRNA-lentiviral constructs targeting S100A4 (S100A4-shRNA). Control groups consisted of sex-matched WT counterparts and WT mice treated with a non-targeting RNA construct., Results: S100A4 deficiency was associated with higher trabecular and cortical bone mass, increased number and thickness of trabeculi combined with larger periosteal circumference and higher predicted bone strength. S100A4 inhibition by shRNA led to an increase in cortical bone in WT mice. S100A4-deficieny was associated with a reduced number of functional osteoclasts. S100A4KO and S100A4-shRNA-treated bone marrow progenitors gave rise to a large number of small TRAP+ cells with few nuclei and few pseudopodial processes. Poor osteoclastogenesis and the low resorptive capacity in S100A4Ko mice may be linked to low levels of surface integrins, impaired adhesion capacity, and poor multinucleation in S100A4-deficient osteoclasts, as well as a low content of proteolytic enzymes cathepsin K and MMP3 and MMP9 to break down the organic matrix., Conclusion: S100A4 emerges as a negative regulator of bone metabolism potentially responsible for the excessive bone turnover in conditions marked by high levels of S100A4 protein, such as inflammation and rheumatoid arthritis., (© 2013.)

S100A4 is a Ca-binding protein participating in regulation of cell growth, survival, and motility. Here we studied the role of S100A4 protein in sex hormone-regulated bone formation. Bone mineral density in the trabecular and cortical compartments was evaluated in female S100A4 knockout (KO), in matched wild-type (WT) counterparts, and in WT mice treated with lentiviral small hairpin RNA construct inhibiting the S100A4 gene transcription or with a nontargeting construct, by peripheral quantitative computed tomography. The effect of sex hormones on bone was measured 5 weeks after ovariectomy (OVX) and/or dehydroepiadrosterone treatment. S100A4KO had an excessive trabecular and cortical bone formation compared with the age- and sex-matched WT mice. S100A4KO had an increased periosteal circumference (P = .001), cortical thickness (P = .056), and cortical area (P = .003), which predicted 20% higher bone strength in S100A4KO (P = .013). WT mice treated with small hairpin RNA-S100A4 showed an increase of the cortical bone parameters in a fashion identical with S100A4KO mice, indicating the key role of S100A4 in the changed bone formation. S100A4KO mice had higher serum levels of osteocalcin and a higher number of osteocalcin-positive osteoblasts under the periosteum. OVX-S100A4 resulted in the loss of the cortical bone supported by high CTX-I levels, whereas no such changes were observed in OVX-WT mice. S100A4KO mice resisted the dehydroepiadrosterone -induced bone formation observed in the WT counterparts. Our study indicates that S100A4 is a regulator of bone formation, which inhibits bone excess in the estrogen-sufficient mice and prevents the cortical bone loss in the estrogen-deprived mice.

Fms-like tyrosine kinase 3 ligand (Flt3L) is known as the primary differentiation and survival factor for dendritic cells (DCs). Furthermore, Flt3L is involved in the homeostatic feedback loop between DCs and regulatory T cell (Treg). We have previously shown that Flt3L accumulates in the synovial fluid in rheumatoid arthritis (RA) and that local exposure to Flt3L aggravates arthritis in mice, suggesting a possible involvement in RA pathogenesis. In the present study we investigated the role of Flt3L on DC populations, Tregs as well as inflammatory responses in experimental antigen-induced arthritis. Arthritis was induced in mBSA-immunized mice by local knee injection of mBSA and Flt3L was provided by daily intraperitoneal injections. Flow cytometry analysis of spleen and lymph nodes revealed an increased formation of DCs and subsequently Tregs in mice treated with Flt3L. Flt3L-treatment was also associated with a reduced production of mBSA specific antibodies and reduced levels of the pro-inflammatory cytokines IL-6 and TNF-α. Morphological evaluation of mBSA injected joints revealed reduced joint destruction in Flt3L treated mice. The role of DCs in mBSA arthritis was further challenged in an adoptive transfer experiment. Transfer of DCs in combination with T-cells from mBSA immunized mice, predisposed naïve recipients for arthritis and production of mBSA specific antibodies. We provide experimental evidence that Flt3L has potent immunoregulatory properties. Flt3L facilitates formation of Treg cells and by this mechanism reduces severity of antigen-induced arthritis in mice. We suggest that high systemic levels of Flt3L have potential to modulate autoreactivity and autoimmunity.

Proteases of Staphylococcus aureus have long been considered to function as important virulence factors, although direct evidence of the role of particular enzymes remains incomplete and elusive. Here, we sought to provide a collective view of the prevalence of extracellular protease genes in genomes of commensal and pathogenic strains of S. aureus and their expression in the course of human and mouse infection. Data on V8 protease, staphopains A and B, aureolysin, and the recently described and poorly characterized group of six Spl proteases are provided. A phylogenetically diverse collection of 167 clinical isolates was analyzed, resulting in the comprehensive genetic survey of the prevalence of protease-encoding genes. No correlation between identified gene patterns with specific infections was established. Humoral response against the proteases of interest was examined in the sera derived from human patients and from a model mouse infection. The analysis suggests that at least some, if not all, tested proteases are expressed and secreted during the course of infection. Overall, the results presented in this study support the hypothesis that the secretory proteases as a group may contribute to the virulence of S. aureus., (© 2012 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.)

Staphylococcus aureus is a highly virulent bacterial pathogen capable of causing a variety of ailments throughout the human body. It is a major public health concern due to the continued emergence of highly pathogenic methicillin resistant strains (MRSA) both within hospitals and in the community. Virulence in S. aureus is mediated by an array of secreted and cell wall associated virulence factors, including toxins, hemolysins and proteases. In this work we identify a leucine aminopeptidase (LAP, pepZ) that strongly impacts the pathogenic abilities of S. aureus. Disruption of the pepZ gene in either Newman or USA300 resulted in a dramatic attenuation of virulence in both localized and systemic models of infection. LAP is required for survival inside human macrophages and gene expression analysis shows that pepZ expression is highest in the intracellular environment. We examine the cellular location of LAP and demonstrate that it is localized to the bacterial cytosol. These results identify for the first time an intracellular leucine aminopeptidase that influences disease causation in a Gram-positive bacterium., (Copyright © 2012 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.)

Background: Lysostaphin and the catalytic domain of LytM cleave pentaglycine crossbridges of Staphylococcus aureus peptidoglycan. The bacteriocin lysostaphin is secreted by Staphylococcus simulans biovar staphylolyticus and directed against the cell walls of competing S. aureus. LytM is produced by S. aureus as a latent autolysin and can be activated in vitro by the removal of an N-terminal domain and occluding region., Results: We compared the efficacies of the lysostaphin and LytM catalytic domains using a newly developed model of chronic S. aureus infected eczema. Lysostaphin was effective, like in other models. In contrast, LytM was not significantly better than control. The different treatment outcomes could be correlated with in vitro properties of the proteins, including proteolytic stability, affinity to cell wall components other than peptidoglycan, and sensitivity to the ionic milieu., Conclusions: Although lysostaphin and LytM cleave the same peptide bond in the peptidoglycan, the two enzymes have very different environmental requirements what is reflected in their contrasting performance in mouse eczema model.

Chronic granulomatous disease (CGD) is an inherited disorder characterized by recurrent life-threatening bacterial and fungal infections. CGD results from defective production of reactive oxygen species by phagocytes caused by mutations in genes encoding the NADPH oxidase 2 (NOX2) complex subunits. Mice with a spontaneous mutation in Ncf1, which encodes the NCF1 (p47(phox)) subunit of NOX2, have defective phagocyte NOX2 activity. These mice occasionally develop local spontaneous infections by Staphylococcus xylosus or by the common CGD pathogen Staphylococcus aureus. Ncf1 mutant mice were more susceptible to systemic challenge with these bacteria than were wild-type mice. Transgenic Ncf1 mutant mice harboring the wild-type Ncf1 gene under the human CD68 promoter (MN(+) mice) gained the expression of NCF1 and functional NOX2 activity specifically in monocytes/macrophages, although minimal NOX2 activity was also detected in some CD11b(+)Ly6G(+) cells defined as neutrophils. MN(+) mice did not develop spontaneous infection and were more resistant to administered staphylococcal infections compared with MN(-) mice. Most strikingly, MN(+) mice survived after being administered Burkholderia cepacia, an opportunistic pathogen in CGD patients, whereas MN(-) mice died. Thus, monocyte/macrophage expression of functional NCF1 protected against spontaneous and administered bacterial infections.

Background: Staphylococcus aureus is the most common pathogen causing septic arthritis in humans. The affected joints are often rapidly and permanently damaged despite antibiotic treatment, indicating that the elicited host immune response contributes substantially to joint destruction. Bacterial formylated peptides are important chemotactic molecules mediating neutrophil recruitment into infected tissues as an important first step of host defense against invading bacteria. The role of formylated peptides in S. aureus infections has been unknown., Methods: Mice were intravenously inoculated with wild-type S. aureus strain RN4220 or its isogenic mutant strain (Δfmt) lacking the ability to produce formylated peptides. The development of arthritis was followed clinically and histopathologically., Results: Mice inoculated with the formyl peptide-producing wild-type strain showed a significantly increased frequency and severity of arthritis and subsequent joint destruction as compared with Δfmt mutant strain-inoculated mice. The wild-type S. aureus strain also induced significantly more weight loss than the Δfmt mutant strain. The recruitment of neutrophils into infected kidneys and synovial tissue was significantly higher in mice inoculated with the wild-type strain., Conclusions: Our data show that formylated peptides function as important virulence factors in S. aureus arthritis, partly by mediating neutrophil recruitment, which contributes substantially to the joint damage.

The neuroendocrine impact on rheumatoid arthritis is not yet fully described although numerous neurotransmitters are shown to act as inflammatory modulators. One of these is the excitatory transmitter glutamate (Glu). In this study, the influence of the Glu receptor (GluR)-mediated effects on collagen-induced arthritis (CIA) was investigated. CIA was induced in DBA/1 mice by immunization with chicken collagen type II (CII). Mice were exposed to the following GluR antagonists: group 1, the N-methyl-D-aspartic acid (NMDA) receptor channel blocker memantine; group 2, the metabotropic GluR antagonist AIDA, and group 3, the excitatory amino acid receptor antagonist kynurenic acid (KA). Arthritis was evaluated clinically and histologically and compared to PBS-treated controls. The effects of treatment on T cell populations and the levels of anti-CII and anti-citrullinated peptide antibodies were evaluated. Memantine treatment significantly improved the course of CIA, reducing synovitis (p = 0.007) and the frequency of erosions (p = 0.007). Memantine treatment up-regulated the expression of Foxp3 in spleen CD4+ T cells followed by an increase in CD4+CD25+ regulatory T cells. The other GluR antagonists, AIDA and KA, had no effect on CIA. These results demonstrate that blockade of the NMDA receptor channel with memantine delays and attenuates the development of arthritis, probably by promoting the development of regulatory T lymphocytes., (Copyright © 2011 S. Karger AG, Basel.)

Objective: Human resistin has proinflammatory properties that activate NF-κB-dependent pathways, whereas its murine counterpart is associated with insulin resistance. The aim of this study was to examine potential cross-talk between resistin and insulin/insulin-like growth factor (IGF) signaling in rheumatoid arthritis (RA)., Methods: Levels of IGF-1, IGF binding protein 3, and resistin were measured in the blood and synovial fluid of 60 patients with RA and 39 healthy control subjects. Human RA synovium was implanted subcutaneously into SCID mice, and the mice were treated with resistin-targeting small interfering RNA. Primary synovial fibroblasts from patients with RA, as well as those from patients with osteoarthritis, and the human fibroblast cell line MRC-5 were stimulated with resistin. Changes in the IGF-1 receptor (IGF-1R) signaling pathway were evaluated using histologic analysis, immunohistochemistry, and reverse transcription-polymerase chain reaction., Results: Resistin and IGF-1R showed different expression profiles in RA synovia. Low levels of IGF-1 in RA synovial fluid were associated with systemic inflammation and inversely related to the levels of resistin. Stimulation of synovial fibroblasts with resistin induced phosphorylation of IGF-1R to a degree similar to that with insulin, and also induced phosphorylation of transcription factor Akt. This was followed by gene expression of GLUT1, IRS1, GSK3B, and the Akt inhibitors PTPN and PTEN. Abrogation of resistin expression in vivo reduced the expression of IGF-1R, the phosphorylation of Akt, and the expression of PTPN and PTEN messenger RNA in RA synovium implanted into SCID mice., Conclusion: Resistin utilizes the IGF-1R pathway in RA synovia. Abrogation of resistin synthesis in the RA synovium in vivo leads to reductions in the expression of IGF-1R and level of phosphorylation of Akt., (Copyright © 2011 by the American College of Rheumatology.)

Background: Efficient host defense mechanisms are crucial for survival in sepsis and septic arthritis. S100 proteins are reported to have proinflammatory and bactericidal properties. The aim of this study was to investigate the role of S100A4 in staphylococcal arthritis., Methods: S100A4 knockout mice (S100A4KO) and wild-type counterparts (WT) were intravenously and intra-articularly challenged with Staphylococcus aureus strain LS-1. Clinical and morphological signs of arthritis and sepsis, phagocytosis, bone mineral density (BMD), and bone metabolism were then monitored in S100A4 and WT mice., Results: S100A4KO mice had a lower bacterial load in the kidneys than WT mice (P < .05) but developed more severe clinical signs of arthritis (P < .001) and had higher levels of interleukin 6 and L-selectin (P = .002). S100A4KO mice had fewer morphological signs of synovitis and cartilage/bone destruction following intra-articular instillation of bacteria. S100A4KO mice were protected from loss of BMD and had lower levels of RANKL, MMP3, and MMP9 (P < .05). S100A4 was not bactericidal in vitro., Conclusions: In staphylococcal infection, S100A4 regulates bacterial clearance as well as systemic and local inflammatory responses.

Background: Despite advances in medical practices, in recent decades permanent reductions in joint function have not been achieved, and the high mortality rate of patients with staphylococcal septic arthritis has not substantially improved., Methods: We evaluated the effects of a combined tumor necrosis factor (TNF) inhibitor and antibiotic therapy on the course of Staphylococcus aureus arthritis and sepsis in mice., Results: Treatment with the combination of a TNF inhibitor and an antibiotic resulted in a quicker relief of clinical arthritis in mice with septic arthritis, compared with an antibiotic monotherapy. Both histopathologically verified synovitis and the extent of joint destruction were reduced by this combined treatment. Importantly, anti-TNF treatment significantly improved the survival rate of mice with S. aureus sepsis and staphylococcal enterotoxin shock syndrome; this effect might be the result of a partial restoration of the hemostatic balance between coagulation and fibrinolysis. Finally, we demonstrated that anti-TNF treatment downregulates high-mobility group protein B1 in staphylococcal enterotoxin shock syndrome., Conclusions: Thus, simultaneous systemic TNF inhibition and antibiotic therapy has beneficial effects on the outcome of S. aureus arthritis and sepsis in a mouse model, suggesting that the combination of a TNF inhibitor and antibiotics represents a novel therapeutic strategy for the treatment of staphylococcal infections.

Background: Ecs is an ATP-binding cassette (ABC) transporter present in aerobic and facultative anaerobic gram-positive Firmicutes. Inactivation of Bacillus subtilis Ecs causes pleiotropic changes in the bacterial phenotype including inhibition of intramembrane proteolysis. The molecule(s) transported by Ecs is (are) still unknown., Methodology/principal Findings: In this study we mutated the ecsAB operon in two Staphylococcus aureus strains, Newman and LS-1. Phenotypic and functional characterization of these Ecs deficient mutants revealed a defect in growth, increased autolysis and lysostaphin sensitivity, altered composition of cell wall proteins including the precursor form of staphylokinase and an altered bacterial surface texture. DNA microarray analysis indicated that the Ecs deficiency changed expression of the virulence factor regulator protein Rot accompanied by differential expression of membrane transport proteins, particularly ABC transporters and phosphate-specific transport systems, protein A, adhesins and capsular polysaccharide biosynthesis proteins. Virulence of the ecs mutants was studied in a mouse model of hematogenous S. aureus infection. Mice inoculated with the ecs mutant strains developed markedly milder infections than those inoculated with the wild-type strains and had consequently lower mortality, less weight loss, milder arthritis and decreased persistence of staphylococci in the kidneys. The ecs mutants had higher susceptibility to ribosomal antibiotics and plant alkaloids chelerythrine and sanguinarine., Conclusions/significance: Our results show that Ecs is essential for staphylococcal virulence and antimicrobial resistance probably since the transport function of Ecs is essential for the normal structure and function of the cell wall. Thus targeting Ecs may be a new approach in combating staphylococcal infection.

Haemostatic balance shifts towards pro-coagulation during infection. Plasminogen, a key molecule of fibrinolysis, may play an important role in the pathogenesis of staphylococcal infections. In the present study, we assessed the impact of inhibition of plasminogen activation by tranexamic acid on the course of staphylococcal sepsis and septic arthritis in mice. We found significantly down-regulated plasmin activity and increased D-dimer levels in the blood from the mice with staphylococcal sepsis. Treatment with tranexamic acid significantly increased the severity and mortality of staphylococcal infection. In addition, tranexamic acid reduced the survival rate in a murine model for staphylococcal enterotoxin A-induced death. The aggravation of diseases by tranexamic acid was due neither to the pro-inflammatory cytokine network, nor to impairment of bacterial clearance. Modulation of fibrinolysis, either by supplement of fibrinolytic molecules (tissue plasminogen activator or plasmin) or by fibrinogen depletion, did not reduce the mortality of staphylococcal sepsis. In conclusion, we report that treatment with tranexamic acid led to distinct aggravation of staphylococcal septic arthritis and sepsis in mice, suggesting the clinical importance of fibrinolytic balance in staphylococcal infection.

Proto-oncogene survivin has recently been identified as a prognostic marker distinguishing patients with destructive rheumatoid arthritis (RA). In the present material of 132 RA patients and 82 controls, the levels of survivin correlated to urokinase (uPA) (r= 0.46), a plasminogen activator over-expressed in inflamed joints and known to exhibit potent arthritogenic properties. Here we evaluate the functional relationship between these proteins using primary synovial fibroblasts and leucocytes of RA patients, human monocytic (THP-1) and fibroblast (MRC-5) cell lines. Using inhibitors of intracellular signalling, we show that uPA and survivin share common transduction pathways in synovial fibroblasts being dependent on the activity of tyrosine kinases, phosphatidylinositide 3 kinase and mitogen effector kinase. Moreover, uPA production is significantly reduced in fibroblasts if survivin synthesis has been silenced by siRNA. Importantly, silencing of survivin in fibroblasts prevented their invasive growth in knee joints of severe combined immune deficient mice. Interaction of uPA with receptor up-regulates survivin expression in leucocytes. In turn, survivin is required for the up-regulation of uPA receptor on the cell surface. These findings indicate that survivin is an essential mediator of arthritogenic properties of uPA regulating its synthesis in synovial fibroblasts and uPAR expression in leucocytes. Close correlation between survivin and uPA levels in patients with RA supports the importance of this connection for the pathogenesis of arthritis.

Introduction: Dichloroacetate (DCA) has been in clinical use for the treatment of lactacidosis and inherited mitochondrial disorders. It has potent anti-tumor effects both in vivo and in vitro, facilitating apoptosis and inhibiting proliferation. The pro-apoptotic and anti-proliferative properties of DCA prompted us to investigate the effects of this compound in arthritis., Methods: In the present study, we used DCA to treat murine collagen type II (CII)-induced arthritis (CIA), an experimental model of rheumatoid arthritis. DBA/1 mice were treated with DCA given in drinking water., Results: Mice treated with DCA displayed much slower onset of CIA and significantly lower severity (P < 0.0001) and much lower frequency (36% in DCA group vs. 86% in control group) of arthritis. Also, cartilage and joint destruction was significantly decreased following DCA treatment (P = 0.005). Moreover, DCA prevented arthritis-induced cortical bone mineral loss. This clinical picture was also reflected by lower levels of anti-CII antibodies in DCA-treated versus control mice, indicating that DCA affected the humoral response. In contrast, DCA had no effect on T cell- or granulocyte-mediated responses. The beneficial effect of DCA was present in female DBA/1 mice only. This was due in part to the effect of estrogen, since ovariectomized mice did not benefit from DCA treatment to the same extent as sham-operated controls (day 30, 38.7% of ovarectomized mice had arthritis vs. only 3.4% in sham-operated group)., Conclusion: Our results indicate that DCA delays the onset and alleviates the progression of CIA in an estrogen-dependent manner.

Introduction: Recent epidemiologic studies have implicated smoking as an environmental risk factor for the development of rheumatoid arthritis (RA). The aim of the present study is the evaluation of the role of cigarette smoke (CS) in the pathogenesis of collagen-induced arthritis in mice., Methods: DBA/1 mice exposed to CS for 16 weeks (n = 25) and mice exposed to nicotine in drinking water (n = 10) were immunized with collagen type II (CII). Severity of arthritis was evaluated clinically and morphologically and compared with control mice (n = 35). Intensity of inflammation was evaluated by serum IL-6 and TNF-alpha levels. Additionally, antibody response to CII (anti-CII) and citrullinated peptides (aCCP) was measured., Results: Clinical evaluation of arthritis showed a delayed onset of arthritis in CS-exposed mice compared with non-smoking controls (P < 0.05). Histologic index and weight changes were comparable between the groups; however, smoking mice presented less weight loss during the acute phase of the disease and gained weight significantly faster in the recovery phase (P < 0.05). Similar results were obtained in the mice exposed to nicotine. Nicotine also showed a direct anti-inflammatory effect diminishing IL-6 production by stimulated splenocytes in vitro (P < 0.001). Additionally, smoking mice had lower levels of aCCP and anti-CII antibodies compared with non-smoking (P < 0.05)., Conclusions: Neither smoking nor nicotine exposure aggravates development of CII-induced arthritis in mouse model. Moreover, CS exposure was associated with a lower level of anti-CII antibodies, providing a possible explanation for a delay of arthritis onset in this group.

Staphylococcus aureus is an important human commensal and opportunistic pathogen responsible for a wide range of infections. Long chain unsaturated free fatty acids represent a barrier to colonisation and infection by S. aureus and act as an antimicrobial component of the innate immune system where they are found on epithelial surfaces and in abscesses. Despite many contradictory reports, the precise anti-staphylococcal mode of action of free fatty acids remains undetermined. In this study, transcriptional (microarrays and qRT-PCR) and translational (proteomics) analyses were applied to ascertain the response of S. aureus to a range of free fatty acids. An increase in expression of the sigma(B) and CtsR stress response regulons was observed. This included increased expression of genes associated with staphyloxanthin synthesis, which has been linked to membrane stabilisation. Similarly, up-regulation of genes involved in capsule formation was recorded as were significant changes in the expression of genes associated with peptidoglycan synthesis and regulation. Overall, alterations were recorded predominantly in pathways involved in cellular energetics. In addition, sensitivity to linoleic acid of a range of defined (sigB, arcA, sasF, sarA, agr, crtM) and transposon-derived mutants (vraE, SAR2632) was determined. Taken together, these data indicate a common mode of action for long chain unsaturated fatty acids that involves disruption of the cell membrane, leading to interference with energy production within the bacterial cell. Contrary to data reported for other strains, the clinically important EMRSA-16 strain MRSA252 used in this study showed an increase in expression of the important virulence regulator RNAIII following all of the treatment conditions tested. An adaptive response by S. aureus of reducing cell surface hydrophobicity was also observed. Two fatty acid sensitive mutants created during this study were also shown to diplay altered pathogenesis as assessed by a murine arthritis model. Differences in the prevalence and clinical importance of S. aureus strains might partly be explained by their responses to antimicrobial fatty acids.

Septic arthritis and sepsis are common and feared complications of staphylococcal infections, and the increasing antibiotic resistance among staphylococci urge the extended research for virulence factors involved in these diseases. Staphylcoccus aureus produces a number of virulence factors controlled by several global regulatory genes including agr and sarA. MgrA is a recently identified global regulator, belonging to the SarA subfamily, which upregulates expression of several virulence factors including capsule and sortase. In addition, MgrA has been shown to regulate antibiotic resistance and decrease bacterial autolysis. In this study we have assessed the role of mgrA gene expression on induction and progression of septic arthritis and sepsis. Mice inoculated with the mgrA mutant displayed significantly less severe arthritis and showed a significantly better weight development, than wild-type inoculated mice. Importantly, all 10 mice inoculated with the mgrA mutant survived as compared to 70% mortality in the wild-type inoculated mice (p=0.003). In addition, the mgrA mutant showed significantly less bacterial persistence in kidneys as compared to the wild-type strain. We conclude that mgrA regulates virulence factors important for establishment and progression of septic arthritis and sepsis.

Dendritic cells (DC) express a functional NADPH oxidase and produce reactive oxygen species (ROS) upon interaction with microbes and T cells. Exposure to ROS leads to DC activation and maturation, as evidenced by phenotypic and functional changes. We have evaluated how endogenous ROS production affects the cytokine secretion pattern and T cell-activating capacity of bone marrow-derived murine DC. DC treated with ROS scavengers, as well as DC from mice that lack a functional NADPH oxidase (and thereby inherently deficient in ROS production) produced significantly increased levels of IL-1beta, IL-6, TNF-alpha and TGF-beta in response to microbial activation. DC deficient in ROS production induced high levels of IFN-gamma and IL-17 in responding T cells after Ag-specific or superantigen-induced activation. Finally, we show that ROS deficiency affected the induction of a T cell-dependent inflammatory condition, collagen-induced arthritis (CIA). C57BL/6 mice that lack a functional NADPH oxidase developed a severe and erosive CD4-dependent CIA, whereas the majority of the congenic wild-type animals remained healthy. These data suggest that ROS act as immunomodulators in DC-driven T cell activation and perhaps also in T cell-dependent immunopathology.

Introduction: TNFalpha and high mobility group box chromosomal protein 1 (HMGB1) are two potent proinflammatory cytokines implicated as important mediators of arthritis. Increased levels of these cytokines are found in the joints of rheumatoid arthritis patients, and the cytokines trigger arthritis when applied into the joints of naïve mice. HMGB1 is actively released from immune cells in response to TNFalpha; once released, HMGB1 in turn induces production of several proinflammatory cytokines--including IL-6 and TNFalpha--by macrophages. Whether HMGB1-induced arthritis is mediated via the TNFalpha pathway, however, is unknown. The purpose of the present study was to investigate whether the arthritis-inducing effect of HMGB1 is dependent on TNFalpha expression in vivo and to assess whether TNFalpha deficiency affects a proinflammatory cytokine response to HMGB1 in vitro., Methods: TNFalpha knockout mice and backcrossed control animals on a C57Bl6 background were injected intraarticularly with 5 microg HMGB1. Joints were dissected 3 days after intraarticular injection and were evaluated histologically by scoring the frequency and severity of arthritis. For in vitro studies, mouse spleen cultures from TNFalpha knockout mice and from control mice were incubated with different doses of HMGB1, and cell culture supernatants were collected at different time points for analysis of IL-6., Results: Intraarticular injection of HMGB1 into healthy mouse joints resulted in an overall frequency of 32% to 39% arthritic animals. No significant differences were found with respect to the severity and incidence of synovitis between mice deficient for TNFalpha (seven out of 18 mice with arthritis) in comparison with control TNFalpha+/+ animals (six out of 19). No significant differences were detected between spleen cells from TNFalpha+/+ mice versus TNFalpha-/- mice regarding IL-6 production upon stimulation with highly purified HMGB1 after 24 hours and 48 hours. Upon stimulation with a suboptimal dose of recombinant HMGB1, however, the splenocytes from TNFalpha+/+ animals released significantly more IL-6 than cells from the knockout mice (602 +/- 112 pg/ml and 304 +/- 50 pg/ml, respectively; P < 0.05)., Conclusion: Our data show that HMGB1-triggered joint inflammation is not mediated via the TNF pathway. Combined with our previous study, we suggest that HMGB1-triggered arthritis is probably mediated through IL-1 activation.

Staphylococcus aureus is the most common cause of joint infections. It also contributes to several other diseases such as pneumonia, osteomyelitis, endocarditis, and sepsis. Bearing in mind that S. aureus becomes rapidly resistant to new antibiotics, many studies survey the virulence factors, with the aim to find alternative prophylaxis/treatment regimens. One potential virulence factor is the bacterial ability to survive at different oxygen tensions. S. aureus expresses ribonucleotide reductases (RNRs), which help it to grow under both aerobic and anaerobic conditions, by reducing ribonucleotides to deoxyribonucleotides. In this study, we investigated the role of RNR class III, which is required for anaerobic growth, as a virulence determinant in the pathogenesis of staphylococcal arthritis. The wild-type S. aureus strain and its isogenic mutant nrdDG mutant were inoculated intravenously into mice. Mice inoculated with the wild-type strain displayed significantly more severe arthritis, with significantly more synovitis and destruction of the bone and cartilage versus mutant strain inoculated mice. Further, the persistence of bacteria in the kidneys was significantly more pronounced in the group inoculated with the wild-type strain. Together these results indicate that RNR class III is an important virulence factor for the establishment of septic arthritis.

Bacterial cell wall peptidoglycan is a dynamic structure requiring hydrolysis to allow cell wall growth and division. Staphylococcus aureus has many known and putative peptidoglycan hydrolases, including two likely lytic transglycosylases. These two proteins, IsaA and SceD, were both found to have autolytic activity. Regulatory studies showed that the isaA and sceD genes are partially mutually compensatory and that the production of SceD is upregulated in an isaA mutant. The expression of sceD is also greatly upregulated by the presence of NaCl. Several regulators of isaA and sceD expression were identified. Inactivation of sceD resulted in impaired cell separation, as shown by light microscopy, and "clumping" of bacterial cultures. An isaA sceD mutant is attenuated for virulence, while SceD is essential for nasal colonization in cotton rats, thus demonstrating the importance of cell wall dynamics in host-pathogen interactions.

The success of Staphylococcus aureus as a pathogen can largely be attributed to the plethora of genetic regulators encoded within its genome that temporally regulate its arsenal of virulence determinants throughout its virulence lifestyle. Arguably the most important of these is the two-component, quorum-sensing system agr. Over the last decade, the controversial presence of a second quorum-sensing system (the TRAP system) has been proposed, and it has been mooted to function as the master regulator of virulence in S. aureus by modulating agr. Mutants defective in TRAP are reported to be devoid of agr expression, lacking in hemolytic activity, essentially deficient in the secretion of virulence determinants, and avirulent in infection models. A number of research groups have questioned the validity of the TRAP findings in recent years; however, a thorough and independent analysis of its role in S. aureus physiology and pathogenesis has not been forthcoming. Therefore, we have undertaken such an analysis of the TRAP locus of S. aureus. We found that a traP mutant was equally hemolytic as the wild-type strain. Furthermore, transcriptional profiling found no alterations in the traP mutant in expression levels of agr or in expression levels of multiple agr-regulated genes (hla, sspA, and spa). Analysis of secreted and surface proteins of the traP mutant revealed no deviation in comparison to the parent. Finally, analysis conducted using a murine model of S. aureus septic arthritis revealed that, in contrast to an agr mutant, the traP mutant was just as virulent as the wild-type strain.

Oxidative-stress resistance in Staphylococcus aureus is linked to metal ion homeostasis via several interacting regulators. In particular, PerR controls the expression of a regulon of genes, many of which encode antioxidants. Two PerR regulon members, ahpC (alkylhydroperoxide reductase) and katA (catalase), show compensatory regulation, with independent and linked functions. An ahpC mutation leads to increased H2O2 resistance due to greater katA expression via relief of PerR repression. Moreover, AhpC provides residual catalase activity present in a katA mutant. Mutation of both katA and ahpC leads to a severe growth defect under aerobic conditions in defined media (attributable to lack of catalase activity). This results in the inability to scavenge exogenous or endogenously produced H2O2, resulting in accumulation of H2O2 in the medium. This leads to DNA damage, the likely cause of the growth defect. Surprisingly, the katA ahpC mutant is not attenuated in two independent models of infection, which implies reduced oxygen availability during infection. In contrast, both AhpC and KatA are required for environmental persistence (desiccation) and nasal colonization. Thus, oxidative-stress resistance is an important factor in the ability of S. aureus to persist in the hospital environment and so contribute to the spread of human disease.

Environmental factors are thought to play a major role in the development of rheumatoid arthritis. Because the use of ethanol is widespread, we assessed the role of ethanol intake on the propensity to develop chronic arthritis. Collagen type II-immunized mice were given water or water containing 10% (vol/vol) ethanol or its metabolite acetaldehyde. Their development of arthritis was assessed, as well as the impact of ethanol on leukocyte migration and activation of intracellular transcription factors. Mice exposed daily to this dose of ethanol did not display any liver toxicity, and the development of erosive arthritis was almost totally abrogated. In contrast, the antibody-mediated effector phase of collagen-induced arthritis was not influenced by ethanol exposure. Also, the major ethanol metabolite, acetaldehyde, prevented the development of arthritis. This antiinflammatory and antidestructive property of ethanol was mediated by (i) down-regulation of leukocyte migration and (ii) up-regulation of testosterone secretion, with the latter leading to decreased NF-kappaB activation. We conclude that low but persistent ethanol consumption delays the onset and halts the progression of collagen-induced arthritis by interaction with innate immune responsiveness.

Objective: Receptor for advanced glycation end products (RAGE) is a cell surface molecule that binds a variety of ligands, including high mobility group box chromosomal protein 1 (HMGB-1), a potent proinflammatory cytokine. RAGE-ligand interaction leads to an inflammatory response. A truncated form of the receptor, soluble RAGE (sRAGE), has been suggested to function as a decoy abrogating cellular activation, but its endogenous activity is not fully understood. We undertook this study to assess the properties of sRAGE in vivo and in vitro and to analyze the role of sRAGE in HMGB-1-induced arthritis., Methods: Mice were injected intraarticularly with HMGB-1 and treated systemically with sRAGE prior to histologic joint evaluation. All animals were subjected to peritoneal lavage to assess the local effect of sRAGE treatment. For in vitro studies, mouse splenocytes were incubated with sRAGE followed by assessment of NF-kappaB activation and cytokine production. The chemotactic properties of sRAGE were investigated using in vitro migration assay., Results: Soluble RAGE was determined to have proinflammatory properties since it gave rise to production of interleukin-6, tumor necrosis factor alpha, and macrophage inflammatory protein 2. This effect was triggered by interaction with leukocyte beta2 integrin Mac-1 and was mediated via NF-kappaB. Systemic treatment with sRAGE significantly down-regulated HMGB-1-triggered arthritis, but the observed effect was due to a deviation of the inflammatory response from the joint to the peritoneal cavity rather than a genuine antiinflammatory effect. Apart from its proinflammatory properties, sRAGE was proven to act as a chemotactic stimulus for neutrophils., Conclusion: We conclude that sRAGE interacts with Mac-1, thereby acting as an important proinflammatory and chemotactic molecule.

Background: Streptococcus agalactiae (group B streptococcus) is an important human pathogen that causes neonatal pneumonia, sepsis, septic arthritis, and meningitis, as well as severe infections in immunocompromised adult patients. The streptococci produce several molecules important for virulence., Methods: We used a murine model of sepsis and septic arthritis to assess the role of FbsA, a fibrinogen-binding adhesin of S. agalactiae as a virulence determinant. NMRI mice were inoculated intravenously with S. agalactiae strains isogenic for the expression of FbsA., Results: Inoculation with wild-type (wt) streptococci resulted in significantly higher mortality, more-pronounced weight decrease, and more-severe arthritis, compared with inoculation with the FbsA mutant isogenic strain. Neither active nor passive immunization with FbsA or FbsA-specific antibodies, respectively, resulted in any protection against subsequent infection with the S. agalactiae wt strain., Conclusion: Our results clearly indicate that the expression of FbsA by Streptococcus agalactiae is a significant virulence determinant in septic arthritis and septicemia. However, because blocking of the fibrinogen binding properties did not protect the host against the action of FbsA-expressing streptococci, we believe that the FbsA molecule has some other presently unknown biological in vivo properties.

Haemophilus ducreyi, the etiologic agent of the sexually transmitted disease chancroid, produces a cytolethal distending toxin (HdCDT) that inhibits cultured cell proliferation, leading to cell death. A rabbit model of dermal infection was used to investigate the roles of H. ducreyi bacteria and HdCDT in the development, clinical appearance, and persistence of infection. A non-toxin producing H. ducreyi strain, and for comparison purposes a non-capsulated Haemophilus influenzae strain, were inoculated intradermally, with and without co-administration of purified HdCDT. Co-administration of HdCDT resulted in significant aggravation of H. ducreyi-induced inflammatory lesions, and development of ulcers in rabbit skin. Less pronounced inflammatory lesions and lack of epithelial eruption were observed after inoculation with H. influenzae. Histopathological sections of the H. ducreyi-induced lesions, in both the presence and absence of HdCDT, showed dense infiltrates of the same type inflammatory cells, with the exception of a prominent endothelial cell proliferation noted in sections from lesions caused by H. ducreyi and toxin. Signs of chronic inflammation with involvement of T cells, macrophages, eosinophils, and granuloma formation were observed after H. ducreyi inoculation both with and without toxin. In conclusion, H. ducreyi causes a pronounced, chronic inflammation with involvement of T cells and macrophages, and in combination with HdCDT production of ulcers in the rabbit model. These pathogenic mechanisms may promote the development and persistence of chancroid ulcers.

Objectives: The aim of the study was to assess the role of extracellular cytochrome c as an inducer of joint inflammation and to examine its levels in sera and synovial fluids of rheumatoid arthritis (RA) patients., Methods: Mice were injected intra-articularly with different doses of cytochrome c and joints were evaluated histopathologically and immunohistochemically 3 and 10 days later. In addition, mouse spleen cells were stimulated with different concentrations of cytochrome c, followed by assessment of NF-kappaB activation and cytokine production. Sera and synovial fluid from RA patients and sera from healthy individuals were assessed with respect to cytochrome c levels by an enzyme-linked immunoassay technique., Results: Histopathological signs of arthritis were evident in 75% of animals following intra-articular injection of cytochrome c. Synovitis was characterized by influx of Mac-1+ cells. In vivo depletion of neutrophils and monocytes led to abrogation of arthritis. Stimulation of mouse spleen cells in vitro with cytochrome c resulted in activation of NF-kappaB and release of proinflammatory cytokines and chemokines. Cytochrome c levels in RA patients' sera were significantly lower than in healthy controls. Further, cytochrome c levels in synovial fluid were significantly lower than in corresponding blood samples., Conclusions: Our findings demonstrate that extracellular cytochrome c displays direct proinflammatory properties mediated by activation of NF-kappaB and causing neutrophil and monocyte triggered inflammation. We hypothesize that decreased levels of cytochrome c in RA patients reflect consumption of this molecule in the synovial tissue, decreasing apoptosis and shifting the balance towards inflammation.

The prototype Staphylococcus aureus strain 8325-4 produces high levels of hemolysins and proteases. Recently it has been shown that this property depends on a deficiency of sigma factor B (SigB) activity controlling the activation of regulatory genes such as agr and sarA. SigB deficiency is in turn due to a mutation in the rsbU gene, which is required for posttranslational activation of SigB. The rsbU defect of strain 8325-4 has recently been repaired, and we used this strain (SH1000), along with its isogenic sigB-negative mutant, to investigate the contributions of RsbU and SigB in a murine model of septic arthritis. Intravenous inoculation with the rsbU-repaired isogenic strain SH1000 resulted in significantly more severe arthritis, weight decrease, and mortality compared to those of the parental strain 8325-4 (rsbU-negative) or the isogenic sigB-negative mutant (MJH502). SH1000 also persisted more in kidneys and joints of infected mice. Our data strongly suggest that RsbU and SigB regulate important virulence factors, thereby contributing significantly to the outcome of staphylococcal infection.

We report that mitochondrial DNA (mtDNA) is inflammatogenic in vitro and in vivo as a result of the presence of unmethylated CpG sequences and its oxidative status. Purified human and murine mtDNAs induced arthritis when injected intra-articularly (i.a.) in mice. Importantly, oligodeoxynucleotide that contained a single oxidatively damaged base also induced arthritis when injected i.a. in mice. In contrast, neither human nor murine nuclear DNA induced inflammation. mtDNA-induced arthritis was neither B cell- nor T cell-dependent but was mediated by monocytes/macrophages. mtDNA-induced nuclear factor-kappaB stimulation resulted in the production of tumor necrosis factor alpha, a potent, arthritogenic factor. Finally, extracellular mtDNA was detected in the synovial fluids of rheumatoid arthritis patients but not of control subjects. We conclude that endogenous mtDNA displays inflammatogenic properties as a result of its content of unmethylated CpG motifs and oxidatively damaged adducts.

The exoproteases of Staphylococcus aureus have been proposed as virulence factors during S. aureus infections. To investigate this, we used the wild-type S. aureus strain 8325-4 and its mutants devoid of aureolysin, serine protease, and cysteine protease, respectively, in a well-established model of septic arthritis in mice. The inactivation of the exoprotease genes did not affect the frequency or the severity of joint disease. We conclude that in the model of haematogenously spread staphylococcal arthritis, the bacterial proteases studied do not act as virulence factors.

Objective and Design: The aim of this study was to investigate the impact of the isoflavone genistein on in vivo cell-mediated responses. In addition, we wanted to study the influence of genistein on collagen induced arthritis (CIA) in mice., Methods: Delayed type hypersensitivity reaction (DTH) to oxazolone and the inflammatory response to olive oil were measured in mice treated with genistein. In addition, the impact of genistein treatment on disease progression and outcome of collagen induced arthritis (CIA) was examined., Results: The DTH reaction to oxazolone and the granulocyte-mediated response were significantly suppressed in genistein-treated as compared to control mice. Also, genistein treatment led to decreased levels of oxazolone-specific antibodies. Histologically, mice exposed to genistein and immunized with collagen II displayed somewhat lower degree of inflammation and joint destruction. In addition, serum levels of autoantibodies to collagen II were significantly lower following genistein-treatment in immunized mice., Conclusion: We conclude that genistein exerts evident anti-inflammatory properties affecting granulocytes, monocytes, and lymphocytes.

Gram-positive pathogenic bacteria display proteins on their surface that play important roles during infection. In Staphylococcus aureus, these surface proteins are anchored to the cell wall by two sortase enzymes, SrtA and SrtB, that recognize specific surface protein sorting signals. The role of sortase enzymes in bacterial virulence was examined using a murine septic arthritis model. Intravenous inoculation with any of the Delta(srtA), Delta(srtB) or Delta(srtAB) mutants resulted in significantly increased survival and significantly lower weight loss compared with the parental strain. Mice inoculated with the Delta(srtA) mutant did not express severe arthritis, while arthritis in mice inoculated with the Delta(srtB) mutant was not different from that seen in mice that were infected with the wild-type parent strain. Furthermore, persistence of staphylococci in kidneys and joints following intravenous inoculation of mice was more pronounced for wild-type and Delta(srtB) mutant strains than for Delta(srtA) or Delta(srtAB) variants. Together these results indicate that sortase B (srtB) plays a contributing role during the pathogenesis of staphylococcal infections, whereas sortase A (srtA) is an essential virulence factor for the establishment of septic arthritis.

Objective: To examine the potential role of high mobility group box chromosomal protein 1 (HMGB-1) in the pathogenesis of arthritis., Methods: Mice were injected intraarticularly with 1 microg or 5 microg of HMGB-1. Joints were dissected on days 4, 7, and 28 after injection and were evaluated histopathologically and immunohistochemically. To investigate the importance of different white blood cell populations for the development of arthritis, in vivo cell depletion procedures were performed. In addition, spleen cells were cultured in the presence of HMGB-1, and nuclear factor kappaB (NF-kappaB) activation was detected by electrophoretic mobility shift assay., Results: Injection of recombinant HMGB-1 (rHMGB-1) into different mouse strains resulted in an overall frequency of arthritis in 80% of the animals. The inflammation was characterized by mild to moderate synovitis and lasted for at least 28 days. The majority of cells found in the inflamed synovium were Mac-1+ macrophages, whereas only a few CD4+ lymphocytes were detected. Pannus formation was observed in some cases 7 and 28 days after HMGB-1 injection. No significant differences were found with respect to incidence and severity of arthritis between mice depleted of monocytes, granulocytes, or lacking T/B lymphocytes. However, combined removal of monocytes and neutrophils resulted in a 43% lower incidence of arthritis. Mice rendered deficient in the interleukin-1 (IL-1) receptor did not develop inflammation upon challenge with HMGB-1. In vitro data corroborate this finding, showing that rHMGB-1 activated NF-kappaB, a major pathway leading to IL-1 production., Conclusion: Our results indicate that HMGB-1 is not a mere expression of inflammatory responses, but on its own, it triggers joint inflammation by activating macrophages and inducing production of IL-1 via NF-kappaB activation.

Persistence of Staphylococcus aureus during invasive infections has been associated with a small-colony variant (SCV) phenotype. SCVs are frequently auxotrophic for menadione or hemin, two compounds involved in the biosynthesis of the electron transport chain. SCVs have been shown to be more resistant to antibiotics such as aminoglycosides, grow slowly and persist intracellularly. The aim of this study was to assess the virulence of an hemB mutant, which has been shown to display the typical characteristics of clinical SCVs, in a murine model of septic arthritis. NMRI mice were inoculated intravenously with either the wild type strain Newman or with its mutant displaying the SCV phenotype. The clinical, bacteriological, and histopathological progression of the disease was studied. Mice inoculated with the hemB mutant displayed a higher frequency and a significantly higher severity of arthritis than mice inoculated with the wild type Newman strain. Despite that, the mutant inoculated mice displayed significantly lower bacterial burden in their kidneys and joints compared with mice exposed to the wild parental strain. Notably, the hemB mutant produced almost 20 times more protease in vitro than the parental strain. We conclude that the small colony variants of S. aureus are more virulent on a per organism basis than its isogenic parental strain in the model of septic arthritis. This can at least in part be explained by the ability of SCV to produce high amounts of destructive proteases.

Microbial superantigens represent a group of molecules that is able to cause massive activation of the host immune system. Human diseases originating from superantigen-secreting bacterial agents are characterized by shock, which continues to pose major health problems. Presently, the treatment of superantigen-mediated infections is limited to the administration of antibiotics and handling of the state of shock. However, the development of multiple antibiotic-resistant, superantigen-producing bacterial strains increases the threat of these infections, and prompts researchers to better understand and treat disease states in which exposure to superantigens is at least partly responsible for the outcome. In the past decade, significant understanding has been achieved regarding the molecular mechanisms of superantigen-host interactions. Based on this understanding, a variety of promising strategies directed against superantigens have been developed. In this review, we discuss some of these strategies, as well as the potential for therapeutic applications of superantigens for the benefit of the host.

Interactions between staphylococci and the joint tissues of the host lead typically to rapidly progressing and highly destructive processes. Staphylococci possess a vast arsenal of components and products that contribute to the pathogenesis of joint infection. Occasionally these compounds have overlapping activities and act either in concert or alone. Host responsiveness to staphylococcal infection displays an even more complex pattern. Most of the cells and molecules that participate in the innate immune system protect the host against bacteria. However, the staphylococci have developed systems that counteract endogenous protective mechanisms. Interestingly, certain cells and molecules of the acquired immune system potentiate the severity of infection by triggering exaggerated responses to the staphylococcal danger signals. This review deals with the intricate host-bacterium interactions that occur during experimental septic arthritis, and outlines potential preventive and treatment modalities., (Copyright 2002 Federation of European Microbiological Societies)

Background: Collagen-induced arthritis (CIA) is the most commonly used model of rheumatoid arthritis (RA). In both CIA and RA there is an increase in the cellular content of the synovium, this being dominated by macrophages., Objective: To assess the impact of etoposide, a topoisomerase II antagonist known to induce monocyte apoptosis, on the development of CIA., Methods: Mice were primed and booster immunised against collagen II (CII). One group of mice was treated with etoposide two days before CII immunisation and then on four consecutive days weekly until the end of the experiment. The second group of mice was injected with etoposide on four consecutive days a week starting 40 days after CII priming. The third group of mice were controls receiving phosphate buffered saline (PBS). The mice were examined for development of arthritis, numbers of circulating leucocytes, serum CII antibody, and cytokine concentrations., Results: None of the mice given etoposide before CII immunisation developed arthritis. Serum concentrations of anti-CII antibodies were undetectable in these mice, whereas they displayed significantly increased concentrations of interferon gamma and interleukin 6. In addition, the CII specific B cell responses in the draining lymph nodes were highly suppressed. Also, mice treated with etoposide at the onset of clinical arthritis showed reduced frequency of their disease by 50%., Conclusion: There was a striking disease alleviating impact of topoisomerase II antagonist on the course of CII-induced arthritis.