Rheumatoid arthritis (RA) is a chronic inflammatory disease char- acterized by the infiltration of T and B cells, macrophages, and neu- trophils into the synovial lining and fluid of the periarticular spaces. Chemokines act as ligands for highly related chemokine receptors that are expressed on distinct leukocyte cell populations, including the cell types recruited to the joint in RA (1). For example, the CC chemokine receptors (CCR) CCR5 (receptor for CCL5, CCL3, and CCL7) and CCR2 (receptor for CCL2, CCL12) are expressed on T cells and monocytes infiltrating the synovium of RA patients, respectively (2, 3). Supporting the role of chemokine receptors in RA pathogenesis, murine experimental arthritis can be partially ame- liorated by blocking CCR5 and CCR2 using selective receptor antag- onists (4–7). Thus, the prevailing paradigm is that the cellular influx to the inflamed joint in RA is choreographed through the temporal and spatially regulated expression of chemokines and their cognate receptors, prompting the hope of targeting specific members of the chemokine system for therapeutic purposes (1, 8). However, given the plethora of candidate chemokine–chemokine receptors medi-
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containing 5 mg/mL Mycobacterium tuberculosis. On day 7, a booster injection of 100 μg mBSA in incomplete Freund’s adjuvant (Difco) was given at the base of the tail. On day 21, arthritis was induced by intra-articular injection of 100 μ g mBSA in 10 μ L PBS into the left knee joint of mBSA-immunized mice, the right knee being injected with sterile PBS alone. The anti-IL-36R antibody M616 (150 μ g/mouse, i.p.), the isotype- matched control antibody 9B5 (150 μ g/mouse, i.p.) or PBS were injected 1 h before intra-articular mBSA injec- tion. Mice were sacrificed 8 or 14 days after induction of arthritis, the latter group receiving a second injection of antibodies or PBS on day 4. The development of arthri- tis was followed by measuring 99m Technetium (Tc) uptake in the knees on days 1, 3 and 7 after intra-articu- lar mBSA injection, as previously described . A sec- ond AIA experiment was performed on adult male IL- 36R -/- and wild-type (WT) C57BL/6 mice. 99m Tc uptake was measured in the knees on days 1, 3 and 7 after intra-articular mBSA injection and the mice were killed on day 8.
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trafficking indirectly by interfering with the function of JAM-C [6,28]. Alternatively, the anti-JAM-C antibody may act directly on resident synovial fibroblasts to disrupt retention of granulo- cytes within lesions. Synovial fibroblasts are indeed potent producers of cytokines, adhesion molecules, and chemokines that attract and retain large numbers of leukocytes in the inflamed synovium, thus sustaining chronic inflammation and preventing the resumption of normal tissue homeostasis . The mouse AIA model can be divided into two phases. Intra- articular antigen injection first results in an acute inflammatory reaction, characterized by joint swelling and leukocyte infiltra- tion, which later proceeds to a chronic destructive arthritis with
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The disease onset is frequent between the ages of 40 and 70, but can influence people of any age 2 . Inflammation is central to progression of RA. Many inflammatory cytokines have been recognized to play pivotal role in the pathogenesis of the disease 3 . TNF-α and IL-1β are main cytokines which are abundant in an inflamed synovium 4 . Dysregulated expression of TNF- in experimental arthritis has been reported to cause destructive arthritis 5 .
ERA-63 in vehicle, or vehicle alone. As a positive control for suppression of arthritis, animals were treated orally with 1.5 or 3 mg/kg prednisolone in vehicle. All experi- mental treatments were conducted in a blinded fashion. The clinical severity of arthritis (arthritis score) was graded (a scale of 0 to 2 for each paw). Mice were scored on alternative days, resulting in mean scores with a maxi- mum of 2 for each paw, and an overall maximum of 8 per animal. To assess the effects of treatment, the area under the curve (AUC) of mean arthritis score of each animal with baseline correction (subtracting baseline AUC of arthritis score on day 0) was used. At the end of the experiment (21 days of treatment) knee synovial biopsies, hind paws and serum samples were obtained. Hind paws were evaluated using X-ray analysis  to assess bone destruction. X-ray photographs were examined with a Faxitron X-ray MX-20 (0.02 mm resolution) and bone destruction was scored on a scale from 0 to 5 ranging from no damage to complete destruction . For histo- pathological analysis (infiltration and cartilage destruc- tion) knee joints were fixed in 4% formaldehyde, decalcified in 5% formic acid and processed and evalu- ated as described . Hematoxylin and eosin-stained sections (7 μM) were used to study joint inflammation. The severity of inflammation in joints was scored on a scale of 0 to 3 (0 = no cells, 1 = mild cellularity, 2 = mod- erate cellularity and 3 = maximal cellularity). To study proteoglycan depletion from the cartilage matrix, sec- tions were stained with safranin O. Depletion of proteo- glycan was scored on an arbitrary scale of 0 to 3 ranging from normal fully stained cartilage to destained cartilage. To analyze cytokine levels with Luminex (Bio-rad, Her- cules, CA, USA) technology, knee synovial biopsies were isolated as described , frozen in liquid nitrogen and stored at -70°C until use (see section cytokine and chemokine protein levels by Luminex).
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Osteoarthritis is a joint disorder associated with chronic inflammation. Targeting the in- flammation is often the initial step for effec- tive management of the condition. In the cur- rent study the anti-inflammatory potential of an Ayurveda polyherbal formulation used for topical application was evaluated. Both in vi- tro and in vivo experimental protocols were followed and significant effects were obtained. Considering that oral medications have their limitations, there is scope for exploring the potential of herbal topical applications.
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Treatment with anti-tumor necrosis factor-alpha monoclonal antibody suppresses TIARP expression To test the therapeutic efficacy of anti-TNFα mAb, we injected anti-TNFα mAb after clinical onset of arthritis at day 8. A single injection of 100 μg of anti-TNFα mAb at day 8 ameliorated the disease, as indicated by a rapid fall in the semiquantitative score of arthritis (Figure 3a) . To explore the relevance of the therapeutic effect of anti-TNFα mAb on TIARP expression, we evaluated TIARP expression after injection of anti-TNFα mAb in mice with GPI-induced arthritis. Treatment of mice with anti-TNFα mAb resulted in downregulation of TIARP expres- sion in spleen relative to control Ig injection, although no treat- ment-related change in TIARP expression was noted at day 14 (P = 0.03) (Figure 3b, top panel). However, in joints, expres- sion of TIARP mRNA was almost comparable between the treatment with anti-TNFα mAb and control Ig. These results suggest that TNF antagonism induces TIARP downregulation and results in the amelioration of arthritis.
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In this study, we have shown that a 1.4-Mbp locus lo- cated on mouse chromosome 5, comprising three pro- tein coding genes and a number of non-protein coding genetic elements (Fig. 1), controls the severity of clin- ical arthritis in CIA along with anti-CII antibody titers. Based on a literature survey and the link to other bone diseases in humans, we selected Tbx3 and Tbx5 , as po- tential candidate genes for CIA. We demonstrated vari- ations in the regulatory part of the Tbx3 gene, which may affect the observed difference in expression level of Tbx3 in splenic B lymphocytes, in addition to the in- creased in vitro B cell activation and proliferation upon stimulation through cell-bound IgM. Moreover, our functional data showed that TBX3 activity is inversely related to B cell activation both in vitro and in the CIA model. This suggests that TBX3 is a player in CIA pathogenesis by modulating B cell proliferative re- sponses, but that it does not affect the development of B lymphocytes.
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and humans . It has been difficult to determine the etiological agents of rheumatoid arthritis in humans due to its multifactorial nature, but animal models have proved to be helpful in these regards. Animal models have been useful to determine the role of mycoplasmas as ar- thritogenic agents but the outcome of these experimental models depends on several factors such as the inmmuno- logical state of the host, the virulence of the microorgan- ism inoculation site, size and concentration of the inoculum . Washburn et al. showed that rabbit is a good model to test the arthritogenic ability of M. pulmonis and M. arthritidis. In a previous study we used the rab- bit model to show that M. pneumoniae was able to induce arthritis . This arthritogenic ability is important be- cause M. pneumoniae may produce extra-respiratory com- plications in human beings . The pathogenic role of M. fermentans has not been completely defined. The aim of this study was to test and compare the arthritogenic ability of a clinical isolate of the respiratory tract with a type strain and to test if M. fermentans P-140 when inject- ed in the trachea is able to reach the joints and induce ar- thritis. M. fermentans P-140 injected in the joints induced a more severe arthritis than the one induced by M. fermen- tans PG-18 strain during the first days postinoculation showing a greater ability to migrate to the uninoculated left knee joint than PG-18 strain. The histological study of synovial tissue revealed few changes in the knee joints in- jected with PG-18. However, when rabbits were injected with the P-140 strain a marked local inflammatory re- sponse with fibroblast and cellular infiltration, constitut- ed by lymphocytes and plasma cells were shown. M. fermentans P-140 has been isolated from the human respi- ratory tract , it has had few passages in the laboratory Figure 1
hematopoietic progenitor cells [142–144]. Conversely, the neutralization of CSF-1R in adult mice leads to a reduction of mature monocytes in blood and bone marrow, without affecting precursors . With regard to experimental arthritis, it is interesting to note that the lack/blockade of CSF-1 as well as the blockade of CSF-1R is associated with less severe methylated bovine serum al- bumin (mBSA)-induced arthritis and CIA [146–148]. In RA patients, all available studies pointed to increased serum and SF levels of IL-34 with respect to normal and disease controls (OA, PsA, ankylosing spondylitis (AS)) [149–153]. Of interest, ser0075m IL-34 levels correlated with immunological markers of more severe disease in- cluding rheumatoid factor (RF), anticyclic citrullinated peptide antibody (anti-CCP) titers, erythrocyte sedimenta- tion rate (ESR), C-reactive protein (CRP), and with disease activity and smoking [149–152]. In this regard, serum IL- 34 levels have been also associated with radiographic pro- gression and appear to be good predictors of radiographic damage in RA patients [150, 152]. Interestingly, treatment with DMARDs or TNF-α inhibitors is able to reduce serum IL-34 levels [151, 154]. IL-34 levels are also higher in RA SF compared to OA SF and increased in RA pa- tients with higher disease activity [149, 153]. Of interest, SF IL-34 levels are directly correlated with those of SF RANK-L, further supporting the link between IL-34 and RANK-L mediated osteoclastogenesis . Finally, IL-34 is also consistently expressed in RA ST, mainly in the sub- lining and the intimal lining layer, with its expression be- ing associated to synovitis severity [148, 154, 155]. All these observations about IL-34 raise the question whether the blockade of its pro-inflammatory and bone remodeling effects are worth the loss also of the strongly suppressive IL-34 driven Treg cells. Therefore additional data are needed to clarify its therapeutic potential in RA.
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these sites, as binding capacities of cortical and trabecular bone are only slightly different. However, both decrease significantly if bone matrix is demineralized . There- fore, if bones with cancellous and noncancellous struc- tures are located near regions of interest, data analyses require a high degree of anatomical accuracy, which can be achieved with μCT. In quantification of experimental arthritis severity, we found that the effect of high baseline signaling in tibial and radial bones was of negligible rele- vance for 18 F-fluoride PET imaging, because signaling hotspots of arthritis pathogenesis were located in tarsal, metatarsal and carpal and metacarpal bones. Another source of a high and pathological 18 F-fluoride baseline PET signaling independent of autoimmune disease patho- genesis might be the preoccurrence of osteoarthritis resulting in mechanical bone erosion. Whereas this aspect is not relevant for imaging in experimental arthritis models, it should be considered in RA bone imaging, where age is a risk factor.
Other studies have used PSL as an inhibitor of inflamma- tion in carrageenan-induced arthritis model in mice (100 mg/kg, intraperitoneal) , for ischemia preven- tion in mice (5 mg/kg) , as an inhibitor of im- munogenicity of recombinant molecules [20–22], for myocardial infarction in mice (5 mg/kg) and even in magnetic resonance imaging (MRI) to evaluate the delivery routes of encapsulated drugs in mice, in addition to the use of pegylated PSLs (PEG-PSL) to maximize the anti-inflammatory effects . More- over, the consistent findings of Wu et al.  and Ma et al.  showed the intervention of PSL (5 mg/kg, intramuscular) in experimental arthritis in rats by inhi- biting inflammation and bone loss in rats with adjuvant- induced arthritis.
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Finally, we investigated the immunomodulatory effects of LAP in a second model of experimental arthritis. To this end, we employed the antigen-induced arthritis (AIA) model in C57BL/6 mice, which also requires a T-cell re- sponse for the generation of the acute articular inflamma- tion . Briefly, mice were treated orally with LAP (10 mg/kg) once a day over 9 days, beginning 12 days after the first immunization with the antigen mBSA. On day 21 after the first immunization, arthritis was induced by intra- articular injection of mBSA into the knees of immunized mice. We did not find differences in the serum levels of anti-mBSA total IgG between mBSA-immunized mice treated or not (vehicle) with LAP (Additional file 7: Figure S4). However, mice treated with LAP exhibited a remark- able reduction in leucocyte infiltration into the knee joint 6 h after mBSA challenge when compared to vehicle- treated mice (Fig. 4a). We then evaluated the recall re- sponses by cells from mBSA-immunized mice treated or not with LAP. The mBSA-specific production of IL-17 and IFN-γ by draining lymph node cells and splenocytes was significantly reduced in mice treated with LAP (Fig. 4b and c). IL-4 was not detected in the supernatant of stimulated cells (data not shown). Collectively, these findings show the marked immunomodulatory effects of LAP in two models of experimental arthritis.
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One factor that differentiates krill oil from fish oil is the presence of phospholipids in krill oil. More than 40% of the oil consists of phospholipids, and the majority of these phospholipids contain (n-3) fatty acids (data to be published). Unfortunately, little scientific attention has been paid to the absorption and distribution of dietary phospholipids, and almost exclusively, all studies on (n-3) fatty acids in human and experimental animal models have been performed using such fatty acids in either tria- cylglycerol form, free fatty acid form, or as fatty acid ethyl esters. There is some evidence, however, that the absorp- tion and the distribution of fatty acids from dietary phos- pholipids can be different than the corresponding fatty acids from triacylglycerol [39-41]. Fatty acids from dietary phospholipids have been reported to be taken up in the brain 2.1-fold more efficiently than fatty acids from triacylglycerol . It has been reported a difference in the pharmacokinetic properties between fatty acids origi- nating from phospholipids versus triacylglycerol , and the absorption, intestinal metabolism and distribution of (n-3) fatty acids from dietary phospholipids might be dif- ferent than (n-3) fatty acids from triacylglycerol. Indeed, in Zucker rats, EPA and DHA from krill oil were more efficiently incorporated into heart phospholipids, com- pared to EPA and DHA from fish oil . Moreover, a higher incorporation of DHA into brain was also reported after krill oil administration to Zucker rats . Whether this is relevant for the effects observed in the current study is not known, but one possibility is that (n- 3) fatty acids from krill oil and fish oil have different effects on inflammation. These mechanisms are only left to speculations, but can for example be related to a differ- ent uptake of (n-3) fatty acids in neutrophils and, conse- quently, an altered lipid composition and function of neutrophils . The presence of EPA, DHA and arachi- donic acid in neutrophil phospholipids after krill oil and fish oil administration should be investigated in future studies.
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Specific antigens recognized by the humoral response to B. burgdorferi differ in DBA/1 and C3H/HeJ mice. Having deter- mined differences in antibody isotype responses in DBA/1 and C3H/HeJ mice infected by B. burgdorferi, we sought to identify what specific differences in B. burgdorferi antigen recognition might account for the varying antibody production. Figure 5A shows Western blots of pooled DBA/1 and C3H/HeJ mouse sera against B. burgdorferi membrane proteins at 42 days postinfection. Individual arrows highlight obvious differences in reactivity be- tween the two strains. Because one-dimensional SDS-PAGE sep- aration did not allow specific identification of antigenic targets, we performed 2-dimensional electrophoresis using an immobilized pH gradient (IPG) for first-dimension separation by isoelectric point and traditional SDS-PAGE for second-dimension separa- tion by mass. Using Western blotting of 2-dimensional gels, Fig. 5B and C show the immunoreactivities of pooled DBA/1 and C3H/HeJ murine sera with B. burgdorferi membrane proteins sep- arated by pI and mass. Using data from our previous mapping of FIG 2 DBA/1 and C3H/HeJ mice develop similar histologic evidence of arthritis in tibiotarsal joints. DBA/1 and C3H/HeJ (n ⫽ 10 each) tibiotarsal joints were collected at 42 days after B. burgdorferi infection or sham infection and then decalcified and H&E stained. (A to D) Representative images at a magnification of ⫻ 10 are shown forB. burgdorferi-infected (A) or sham-infected (B) DBA/1 mice and for B. burgdorferi-infected (C) or sham-infected (D) C3H/HeJ mice. b, bone; s, synovium. Proliferative synovitis with leukocyte infiltrates is indicated by arrows. (E) Histologic scoring of arthritis severity was not different for the two strains. n.s., not significant (P ⫽ 0.66).
DOI: 10.4236/ojapps.2018.81001 2 Open Journal of Applied Sciences reactions. In an autoimmune reaction, antibodies and immune cells accidently target the bodies’ healthy tissues, which signaling the body to attack them. Re- searchers reported that there are more than 80 illnesses caused by autoimmunity . An autoimmune disease affects approximately 8% of the world population, 78% of whom are women . Studies have shown that autoimmunity causes pathogenesis of more than 100 diseases. Approximately 1-2% of the world’s population has rheumatoid arthritis and affects 45 out of every 3000 people who have autoimmune diseases. Although there are many diseases that cause chronic arthritis, rheumatoid arthritis (RA) is a chronic systemic disease, which damages the connective tissues irreversibly, gradually leading to a loss of ability to work and causing disability. This terminal illness requires permanent treatment which is highly expensive  .
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A second unexpected experimental observation provided further evidence that persistent TNF signalling in T cells could perturb TCR/CD3 expression at the cell surface. Immunoblotting analysis of unstimulated and TNF-stimu- lated T cells revealed that the expression of the novel transmembrane adaptor protein TRIM (T-cell-receptor- interacting molecule) was markedly downregulated by TNF treatment [59; Isomäki and Cope, unpublished data]. Closer examination revealed that TRIM expression was reduced by TNF before changes in TCRζ expression could be detected, and that reconstitution of both TRIM and TCRζ expression was required to fully restore TCR responsiveness in TNF-treated cells. The implications of these findings have only recently become apparent, through studies of TRIM expression in human peripheral blood and Jurkat T cells. In collaborative studies with Dr Burkhart Schraven, it was found that the half-life of TCR/CD3 complexes in stable Jurkat clones overexpress- ing TRIM is increased . This in turn leads to increased cell-surface expression of TCR and enhanced signalling responses as determined by intracellular calcium mobilisa- tion. We can conclude from these experiments that sus- tained TNF signals in T cells impair TCR/CD3 assembly not only through its effects on TCR ζ expression, but also by reducing the half-life of assembled complexes at the cell surface by downregulating the expression of TRIM (see Fig. 5). The kinetics of these changes, as well as the precise interactions between TCRζ and TRIM, are now being studied. Nevertheless, the findings provided a mole- cular basis for the profound hyporesponsiveness of T cells after TNF stimulation and predicted that downstream TCR signalling pathways might be significantly attenuated as a consequence of these structural constraints.
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We performed random-effects meta-analysis since we expected variability in prevalence estimates from different studies. We assessed heterogeneity through the use of both the Chi- square test and the I-square test statistic. We considered a p-value of less than 0.10 to be signifi- cant for the Chi-square test due to the low power of this test and an I-square of at least 50% to be significant heterogeneity . There were various case definitions from separate studies and we judged results to be clinically homogenous and we tested for statistical heterogeneity and only combined where heterogeneity was not statistically significant. We investigated sources of heterogeneity through subgroup analysis with respect to the country from which the study was done since prevalence of arthritis is known to have regional variation . The small number of studies in the meta-analyses did not allow us to perform sensitivity analyses with respect to risk of bias, diagnostic criteria or age range. Only studies with similar risk of bias assessment were pooled in a meta-analysis and studies with high risk of bias assessment were excluded from meta-analysis. Where there was significant heterogeneity, meta-analysis was not per- formed. We performed all meta-analyses using STATA version 13 and displayed results in the form of forest plots. We used the STATA command ‘metan’ to perform meta-analysis.
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arthritis’ (JRA), which includes JRA of pauciarticular, poly- articular and systemic onset . The European classification uses the term ‘juvenile chronic arthritis’ (JCA), which includes polyarticular arthritis, pauciarticular arthritis, sys- temic arthritis, juvenile ankylosing spondylitis, psoriatic arthritis and arthritis associated with inflammatory bowel disease. The more recent International League Against Rheumatism classification of ‘juvenile idiopathic arthritis’ (JIA) has seven subsets, including the spondyloarthro- pathies . The very fact that there are different classifica- tion criteria, and each classification scheme has several subtypes, suggests that juvenile arthritis is not a single entity with uniform clinical, laboratory and immunogenetic features. (While we have used the term JRA, the text reflects the different terminology used by the authors cited.) The revised American Rheumatism Association criteria for RA, proposed in 1987, state that patients fulfilling any four of seven criteria fulfill the requirement for RA . The terms ‘classic’ (or ‘definite’) RA and ‘probable’ RA were removed. Thus, RA in adults appears to be a single disease with dif- ferent manifestations, while patients with one subtype of JRA clearly differ from patients with the other subtypes.
antigen nonspecific ex vivo cytokine secretion or to cytokine mRNA levels [38,39]. In our recent studies in AA [30,36], we observed higher Th1 cytokine levels against Bhsp65 during the recovery phase compared with those at the onset of or the peak phase of the disease. The cytokine response to Rhsp65 was found to be similar to that against Bhsp65 . In general, the cytokine response of T cells against Bhsp65 and Rhsp65 as well as their defined antigenic determinants consisted predominantly of proinflammatory cytokines, IFNγ and TNF α . Furthermore, higher levels of proinflammatory cyto- kine secretion correlated with reduced severity of arthritis. As AA is a Th1-mediated disease, this observation pointed to the role of Th1 cytokines in disease regulation. This inference indeed was supported by the cytokine profiles of Mtb- immunized, AA-resistant WKY rats. These rats produced high levels of Th1 cytokines early following Mtb injection, and the overall shape of the cytokine profiles was almost the opposite of that of LEW rats when followed against time post Mtb injection. Furthermore, treatment of LEW rats with TNFα or with Rhsp65 peptide 465 to 479, which primes a Th1 response, resulted in downmodulation of the clinical course of AA in LEW rats, albeit employing different mechanisms [30,36]. The levels as well as the timing of production of Th1 cytokines in response to Bhsp65/Rhsp65 therefore have a significant influence on the outcome of Mtb challenge of LEW/WKY rats. Other investigators have reported differ- ences in the frequency/activity of T-helper type 2-cytokine secreting cells and CD8 + suppressor/regulatory T cells in