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ID PMID Title PublicationDate abstract
16418805 COX-2 inhibitors modulate IL-12 signaling through JAK-STAT pathway leading to Th1 response 2006 Jan Experimental allergic encephalomyelitis (EAE) is a Th1 cell-mediated autoimmune disease model of multiple sclerosis (MS). IL-12 plays a crucial role in the pathogenesis of EAE/MS and inhibition of IL-12 production or IL-12 signaling was effective in preventing EAE. Cyclooxygenase (COX-2) is a key enzyme promoting inflammation in rheumatoid arthritis and tumor induced angiogenesis. Recent studies have shown that COX-2 inhibitors prevent EAE, however, their mechanism of action is not fully understood. In this study, we show that in vivo treatment (i.p.) with 100 mug COX-2 selective inhibitors (LM01, LM08, LM11, and NS398), on every other day from day 0 to 30, significantly reduced the incidence and severity of EAE in SJL/J and C57BL/6 mice. Further analyses showed that the COX-2 inhibitors reduced neural antigen-induced IL-12 production, T cell proliferation and Th1 differentiation ex vivo and in vitro. The COX-2 inhibitors also decreased IL-12-induced T cell responses through blocking tyrosine phosphorylation of JAK2, TYK2, STAT3, and STAT4 proteins in T cells. These results demonstrate that COX-2 inhibitors ameliorate EAE in association with the modulation of IL-12 signaling through JAK-STAT pathway leading to Th1 differentiation and suggest their use in the treatment of MS and other Th1 cell-mediated autoimmune diseases.
16204471 Periplocoside E, an effective compound from Periploca sepium Bge, inhibited T cell activat 2006 Feb Periploca sepium Bge, a traditional Chinese herb medicine, is used for treating rheumatoid arthritis in China. Followed the bioactivity-guided isolation, the most potent immunosuppressive compound, periplocoside E (PSE), a pregnane glycoside, had been identified from P. sepium Bge. We investigated the immunosuppressive effects of PSE in vitro and in vivo. The results showed that PSE in a dose-dependent manner significantly inhibited the proliferation of splenocytes induced by concanavalin A and mixed lymphocyte culture reaction at no cytotoxic concentrations (<5 microM). Administration of PSE suppressed a delayed-type hypersensitivity reaction, and ovalbumin (OVA) induced antigen-specific immune responses in mice. In vivo treatment with PSE dose dependently suppressed OVA-induced proliferation and cytokine [interleukin (IL)-2 and interferon (IFN)-gamma] production from splenocytes in vitro. Purified T cells from OVA-immunized mice with PSE treatment showed its low ability for activation by OVA plus normal antigen presenting cell stimulation again in vitro. Further studies showed PSE dose dependently inhibited anti-CD3-induced primary T cell proliferation, activation for IL-2Ralpha (CD25) expression, and cytokine (IFN-gamma and IL-2) production also at the transcriptional level. PSE was highly specific and significantly inhibited the activation of extracellular signal-regulated kinase and Jun N-terminal kinase, whereas activation of p38 was not affected in T cells stimulated with anti-CD3. These results demonstrated that PSE is an immunosuppressive compound in P. sepium Bge, which directly inhibits T cell activation in vitro and in vivo. This study provided evidence to understand the therapeutic effects of P. sepium Bge and indicated that this herb is appropriate for treatment of T cell-mediated disorders, such as autoimmune diseases.
18843655 Single dose oral celecoxib for acute postoperative pain in adults. 2008 Oct 8 BACKGROUND: This is an update of a review published in The Cochrane Library, Issue 1, 2003. Celecoxib is a selective cyclo-oxygenase-2 (COX-2) inhibitor prescribed for the relief of chronic pain in osteoarthritis and rheumatoid arthritis. Celecoxib is believed to be associated with fewer upper gastrointestinal adverse effects than conventional non-steroidal anti-inflammatory drugs (NSAIDs). Its effectiveness in acute pain was demonstrated in the earlier review. Additional studies have now been published for the 400 mg dose, and this updated review provides more robust estimates of efficacy and harm. OBJECTIVES: To assess analgesic efficacy and adverse effects of a single oral dose of celecoxib for moderate to severe postoperative pain. SEARCH STRATEGY: Cochrane CENTRAL, MEDLINE, EMBASE, and the Oxford Pain Database. Most recent search: July 2008. SELECTION CRITERIA: Randomised controlled trials (RCTs) of adults prescribed any dose of oral celecoxib or placebo for acute postoperative pain were included. DATA COLLECTION AND ANALYSIS: Eight studies (1380 participants) met the inclusion criteria. Studies were assessed for quality and data extracted by two review authors. Summed pain relief (TOTPAR) or pain intensity difference (SPID) was converted into dichotomous information yielding the number of participants with at least 50% pain relief over four to six hours, and used to calculate the relative benefit (RB) and number-needed-to-treat-to-benefit (NNT) for one patient to achieve at least 50% pain relief with celecoxib who would not have done so with placebo. Information on use of rescue medication was used to calculate the proportion of participants requiring rescue medication and the weighted mean (WM) of the median time to use. MAIN RESULTS: The NNT for celecoxib 200 mg and 400 mg compared with placebo for at least 50% pain relief over four to six hours was 4.2 (CI 3.4 to 5.6) and 2.5 (2.2 to 2.9) respectively. The WM of the median time to use of rescue medication was 6.6 hours with celecoxib 200 mg, 8.4 with celecoxib 400 mg, and 2.3 hours with placebo. The WM proportion of participants requiring rescue medication over 24 hours was 74% with celecoxib 200 mg, 63% for celecoxib 400 mg, and 91% for placebo. The NNT to prevent one patient using rescue medication was 4.8 (3.5 to 7.7) and 3.5 (2.9 to 4.6) for celecoxib 200 mg and 400 mg respectively. One serious adverse event probably related to celecoxib was reported by the trialists. AUTHORS' CONCLUSIONS: Single dose oral celecoxib is an effective means of postoperative pain relief. The 400 mg dose has similar efficacy to ibuprofen 400 mg.
18382855 Testing human biologicals in animal host resistance models. 2008 Jan The purpose of immunotoxicity testing is to obtain data that is meaningful for safety assessment. Host resistance assays are the best measure of a toxicant's effect on the overall ability to mount an effective immune response and protect the host from infectious disease. An outline is presented for immunotoxicological evaluation using host resistance assays. The influenza virus host resistance model is useful to evaluate the overall health of the immune system and is one of the most thoroughly characterized host resistance models. Viral clearance requires all aspects of the immune system to work together and is the ultimate measure of the health of the immune system in this model. Mechanistic immune functions may be included while measuring viral clearance and include: cytokines, macrophage activity, natural killer (NK) cell activity, cytotoxic T-lymphocyte (CTL) activity, and influenza-specific IgM and IgG. Measurement of these immunological functions provides an evaluation of innate immunity (macrophage or NK activity), an evaluation of cell-mediated immunity (CMI) (CTL activity), and an evaluation of humoral-mediated immunity (HMI) (influenza-specific IgM or IgG). Measurement of influenza-specific IgM or IgG also provides a measurement of T-dependent antibody response (TDAR) since influenza is a T-dependent antigen. There are several targeted host resistance models that may be used to answer specific questions. Should a defect in neutrophil and/or macrophage function be suspected, Streptococcus pneumoniae, Pseudomonas aeruginosa, or Listeria monocytogenes host resistance models are useful. Anti-inflammatory pharmaceuticals or therapeutics for rheumatoid arthritis or Crohn's disease that target TNFalpha may also be evaluated for immunotoxicity using the S. pneumoniae intranasal host resistance assay. Marginal zone B (MZB) cells are required for production of antibody to T-independent antigens such as the polysaccharide capsule of the encapsulated bacteria that are so prominent in causing blood-borne infections and pneumonia. Intravenous infection with Streptococcus pneumoniae, an encapsulated bacterium, results in a blood-borne infection that requires MZB cells for clearance. The systemic S. pneumoniae host resistance assay evaluates whether a therapeutic test article exerts immunotoxicity on MZB cells and measures the T-independent antibody response (TIAR). Suppression of CMI or in some cases HMI may result in reactivation of latent virus that may result in a fatal disease such as progressive multifocal leukoencephalopathy (PML). The murine cytomegalovirus (MCMV) reactivation model may be used to evaluate a pharmaceutical agent to determine if suppression of CMI or HMI results in reactivation of latent virus. Candida albicans is another host resistance model to test potential immunotoxicity. Host resistance assays have been the ultimate measure of immunotoxicity testing for environmental chemicals and pharmaceutical small molecules. Human biologicals are now an important component of the drug development armamentarium for biotech and pharmaceutical companies. Many human biologicals are fusions of IgG, and/or target immune mediators, immunological receptors, adhesion molecules, and/or are indicated for diseases that have immune components. It is therefore necessary to thoroughly evaluate human biological therapeutics for immunotoxicity. Numerous biologicals that are pharmacologically active in rodents can be evaluated using well-characterized rodent host resistance assays. However, biologicals not active in rodents may use surrogate biologicals for testing in rodent host resistance assays, or may use host resistance assays in genetically engineered mice that mimic the effect of the human biological pharmacological agent.
18305142 PTPN22 Trp620 explains the association of chromosome 1p13 with type 1 diabetes and shows a 2008 Jun OBJECTIVE: The disease association of the common 1858C>T Arg620Trp (rs2476601) nonsynonymous single nucleotide polymorphism (SNP) of protein tyrosine phosphatase; nonreceptor type 22 (PTPN22) on chromosome 1p13 has been confirmed in type 1 diabetes and also in other autoimmune diseases, including rheumatoid arthritis and Graves' disease. Some studies have reported additional associated SNPs independent of rs2476601/Trp(620), suggesting that it may not be the sole causal variant in the region and that the relative risk of rs2476601/Trp(620) is greater in lower risk by HLA class II genotypes than in the highest risk class II risk category. RESEARCH DESIGN AND METHODS: We resequenced PTPN22 and used these and other data to provide >150 SNPs to evaluate the association of the PTPN22 gene and its flanking chromosome region with type 1 diabetes in a minimum of 2,000 case subjects and 2,400 control subjects. RESULTS: Due to linkage disequilibrium, we were unable to distinguish between rs2476601/Trp(620) (P = 2.11 x10(-87)) and rs6679677 (P = 3.21 x10(-87)), an intergenic SNP between the genes putative homeodomain transcription factor 1 and round spermatid basic protein 1. None of the previously reported disease-associated SNPs proved to be independent of rs2476601/Trp(620). We did not detect any interaction with age at diagnosis or sex. However, we found that rs2476601/Trp(620) has a higher relative risk in type 1 diabetic case subjects carrying lower risk HLA class II genotypes than in those carrying higher risk ones (P = 1.36 x 10(-4) in a test of interaction). CONCLUSIONS: In our datasets, there was no evidence for allelic heterogeneity at the PTPN22 locus in type 1 diabetes, indicating that the SNP rs2476601/Trp(620) remains the best candidate in this chromosome region in European populations. The heterogeneity of rs2476601/Trp(620) disease risk by HLA class II genotype is consistent with previous studies, and the joint effect of the two loci is still greater in the high-risk group.
17984291 Serum amyloid A mediates human neutrophil production of reactive oxygen species through a 2008 Feb Serum amyloid A (SAA) is one of the acute-phase reactants, a group of plasma proteins that increases immensely in concentration during microbial infections and inflammatory conditions, and a close relationship between SAA levels and disease activity in rheumatoid arthritis (RA) has been observed. RA is an inflammatory disease, where neutrophils play important roles, and SAA is thought to participate in the inflammatory reaction by being a neutrophil chemoattractant and inducer of proinflammatory cytokines. The biological effects of SAA are reportedly mediated mainly through formyl peptide receptor like-1 (FPRL1), a G protein-coupled receptor (GPCR) belonging to the formyl peptide receptor family. Here, we confirmed the affinity of SAA for FPRL1 by showing that stably transfected HL-60 cells expressing FPRL1 were activated by SAA and that the response was inhibited by the use of the FPRL1-specific antagonist WRWWWW (WRW4). We also show that SAA activates the neutrophil NADPH-oxidase and that a reserve pool of receptors is present in storage organelles mobilized by priming agents such as TNF-alpha and LPS from Gram-negative bacteria. The induced activity was inhibited by pertussis toxin, indicating the involvement of a GPCR. However, based on FPRL1-specific desensitization and use of FPRL1 antagonist WRW4, we found the SAA-mediated effects in neutrophils to be independent of FPRL1. Based on these findings, we conclude that SAA signaling in neutrophils is mediated through a GPCR, distinct from FPRL1. Future identification and characterization of the SAA receptor could lead to development of novel, therapeutic targets for treatment of RA.
17228672 [Intraoperative imaging to monitor prosthetic fixation for total hip arthroplasty]. 2006 Dec OBJECTIVE: To explore the intraoperative imaging to monitor the prosthetic fixation for the total hip arthroplasty (THA) and to increase the accuracy of the primary THA prosthetic fixation. METHODS: From April 2000 to August 2005, 69 patients (75 hips) underwent THA monitored by the imaging (the imaged THA group, Group I), and 72 patients (78 hips) underwent THA with the standard method (the standard THA group, Group S). There were 32 72 patients (78 hips) underwent THA with the standard method (the standard THA group s in Group S). There were 32 males and 37 females in Group S. The mean ages of the patients in Group I and Group S were 62. 3 years and 60. 5 years respectively, ranging 46-75 years in Group I and 43-75 years in Group S. Preoperative diagnoses were femoral neck fractal measure (Garden III, IV ) in 23 patients (23 hips) in Group and 2 5 patients (25 hips) in Group S, acetabular dysplasia Campbell I , II) in 9 patients (10 hips) in Group I and 11 patients (13 hips) in Group S osteoarthritis in 16 patients (17 hips) in Group I and 15 patients (15 hips) in Group S, femoral head osteonecrosis (Ficat III, IV ) 15 patients (16 hips) in Group I and 17 patients (17 hips) in Group S, and rheumatoid arthritis in 6 patients (9 hips) in Group I and 4 patients (8 hips) in Group S. There were 21 hips of cement prostheses in Group I and 22 hips in Group ferative 12 hips of cementless prostheses in Group I and 11 hips in Group S, 42 hips of cement and cementless prostheses in Group I and 45 hips in Group S. Group I used the standard THA and the intraoperative X-ray monitoring the prosthetic fixation in the numerical measure of abduction angle, anteversion angle, femoral neck length, and femoral offset distance. The items compared between Group I and Group S included incisional length, intraoperative bleeding, transfusion, operative time, frequency of X-ray imaging, infection, postoperative functional recovery, and prosthetic position of postoperative X-ray imaging. RESULTS The follow-up on 62 patients in Group I and 64 patients in Group S for 6-64 months averaged 42 months revealed that there were statistically significant differences in incisional length, intraoperative bleeding. transfusion, operative time, frequency of X-ray imaging, postoperative functional recovery, prosthetic position of postoperative X-ray imaging, and the Harris score between Group I and Group S one year after operation. The results of Group I were significatnly better than those of Group S. CONCLUSION: The intraoperative X-ray imaging can increase the accuracy of the THA prosthetic fixation and reduce the incidence of THA maloperation by The X-ray imaging can also be used in county hospitals if C-arm fluoroscopy can be provided.
16702601 Conjugated linoleic acid inhibits osteoclast differentiation of RAW264.7 cells by modulati 2006 Aug Bone destruction is a pathological hallmark of several chronic inflammatory diseases, including rheumatoid arthritis, periodontitis, and osteoporosis. Inflammation-induced bone loss of this sort results from increased numbers of bone-resorbing osteoclasts. Numerous studies have indicated that conjugated linoleic acid (CLA) positively influences calcium and bone metabolism. Gene-deletion studies have shown that receptor activator of nuclear factor-kappaB ligand (RANKL) is one of the critical mediators of osteoclastogenesis. In this report, we examine the ability of CLA to suppress RANKL signaling and osteoclastogenesis in RAW264.7 cells, a murine monocytic cell line. Treatment of these cells with RANKL activated nuclear factor-kappaB (NF-kappaB), and preexposure of the cells to CLA significantly suppressed RANKL-induced NF-kappaB activation, including phosphorylation of I-kappaBalpha, degradation of I-kappaBalpha, and nuclear translocation of p65. RANKL induced osteoclastogenesis in these monocytic cells, and CLA inhibited RANKL-induced tumor necrosis factor-alpha production and osteoclast differentiation, including osteoclast-specific genes such as tartrate-resistant acid phosphatase, cathepsin K, calcitonin receptor, and matrix metalloproteinase-9 expression and osteoclast-specific transcription factors such as c-Fos, nuclear factor of activated T-cells expression, and bone resorption pit formation. CLA also inhibited RANKL-induced activation of mitogen-activated protein kinase p38 but had little effect on c-Jun N-terminal kinase activation. Collectively, these data demonstrate for the first time that CLA inhibits osteoclastogenesis by modulating RANKL signaling. Thus, CLA may have important therapeutic implications for the treatment of bone diseases associated with enhanced bone resorption by excessive osteoclastogenesis.
16882186 Male New Zealand Black/KN mice: a novel model for autoimmune-induced permanent alopecia? 2006 Aug BACKGROUND: Irreversible, permanent and scarring alopecia is associated with several autoimmune diseases, including all autoimmune connective tissue disorders. The pathogenesis of autoimmune-induced permanent alopecia (APA) is still poorly understood, and instructive, simple mouse models for the study of APA are needed urgently. During the course of our studies in a well-established mouse model for chronic rheumatoid arthritis, the New Zealand Black/KN (NZB/KN) mouse, we noticed that ageing male NZB/KN mice developed spontaneous APA. OBJECTIVES: To study whether alopecia seen in ageing male NZB/KN mice displays key features of human APA and may, thus, be a useful new mouse model for clinically relevant APA research. METHODS: NZB/KN, the F1 hybrid of NZW/N Slc x NZB/KN (W/BKN F1), the F1 hybrid of NZB/KN x NZW/N Slc (BKN/W F1), and the F2 hybrid of W/BKN F1 x W/BKN F1 mice were employed in this study, in order to check which strain carries the highest risk of alopecia development. Besides routine histology, CD3, CD4 and CD8 expression as well as immunoglobulin (Ig) G and IgM deposition in hair follicles were investigated by immunohistology/immunofluorescence. Mast cell distribution/degranulation and Ki-67 (proliferation)/TUNEL (terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labelling) (apoptosis) positive cells were also analysed. RESULTS: Only F2 male NZB/KN mice were prone to develop alopecia, suggesting that Y chromosome-associated gene(s) are involved in the pathogenesis of APA, which incidence rises with increasing age. The lesional alopecia skin in 12-month-old male NZB/KN mice showed a sharp decline in hair follicle density, thus meeting a key criterion of permanent alopecia. Both macroscopically and histologically, the alopecia seen in these mice resembled in many respects different stages of clinical APA, such as alopecia associated with chronic discoid lupus erythematosus (DLE) in humans. Lesional APA hair follicles in mice displayed intrafollicular and perifollicular mononuclear cell infiltrates, as well as an increased number of activated (degranulated) perifollicular mast cells. In the fully developed lesion, many CD4+ cells were seen in perifollicular locations, including the epithelial stem cell region (bulge), and also contained a few CD8+ T cells. IgM deposits were found in the follicular basement membrane zone (BMZ). Both in the bulge and the hair matrix region of the affected anagen hair follicles, there were signs of massive keratinocyte apoptosis. CONCLUSIONS: Our currently available data suggest that male but not female NZB/KN mice may indeed represent a suitable mouse model for APA, with some similarities to the permanent alopecia seen in human DLE patients, although additional and confirmatory investigations are needed before this mouse strain can be accepted as a murine equivalent of APA in humans.
16755666 Implication of prostaglandin receptors in the accumulation of osteoprotegerin in human ost 2006 Jun OBJECTIVE: Prostaglandins are important mediators in bone metabolism and in pathologies such as rheumatoid arthritis and osteoarthritis. We investigated the roles of cyclooxygenases (COX) and prostaglandin receptors in the accumulation of osteoprotegerin (OPG) in the supernatants of human osteoblasts in culture. METHODS: Three different cellular models were used, the human osteosarcoma cell lines MG-63 and Saos-2, and primary cultures of human osteoblasts. OPG concentrations were determined by ELISA. RESULTS: RT-PCR analysis showed that, like primary human osteoblasts, MG-63 cells express DP, EP4, FP, IP, and TP receptors, whereas the Saos-2 cells lack IP. Concentration of OPG was highest in MG-63 cell supernatants (36 +/- 12.5 ng/ml), followed by human osteoblasts (12.77 +/- 2.2 ng/ml) and Saos-2 (3.6 +/- 0.76 ng/ml). COX inhibitors did not alter these values. Prostaglandin E2 and BW 245C (a synthetic DP receptor agonist) decreased OPG in the supernatants of human osteoblasts but not in immortalized cell lines. These effects were concentration-dependent and were inhibited by EP4 and DP receptor antagonists. Fluprostenol, an FP receptor agonist, increased the accumulation of OPG in MG-63 but not in primary human osteoblasts or Saos-2. CONCLUSION: Our results show that activation of EP4 or DP receptors decreased the accumulation of OPG in supernatants of osteoblasts in culture, and suggest that these receptors could be interesting pharmacological targets in bone diseases. They also demonstrate important differences between primary osteoblasts and immortalized cell lines, both in the distribution and in the effects mediated by prostaglandin receptors.
16644488 Reduction of lipopolysaccharide-induced interleukin-6 production by the kappa opioid U50,4 2006 Jun Several studies demonstrate that opioids modulate the immune response via opioid receptors expressed directly on the immune cells themselves. Recently, it has been suggested that the kappa opioid system has a modulatory role in various inflammatory diseases including rheumatoid arthritis. This modulation may occur via changes in cytokine secretion by monocyte-derived cells. To further study this opioid-immune relationship, we stimulated P388D1 cells, a mouse monocyte-like cell line, with lipopolysaccharide (LPS) in the presence or absence of the kappa opioid-selective ligand, U50,488. Pretreatment with U50,488 significantly reduced LPS-stimulated interleukin-6 (IL-6) production as measured by ELISA. This effect was mediated by the kappa opioid receptor, because nor-binaltorphimine (nor-BNI), a kappa-selective antagonist, blocked this inhibition. It is likely that this reduction of IL-6 protein by U50,488 treatment is attributed to decreases in IL-6 mRNA. RT-PCR experiments demonstrated that U50,488 treatment significantly reduced the LPS-mediated increase in IL-6 mRNA and that this effect was also blocked by nor-BNI. Understanding the mechanism behind the reduction of proinflammatory cytokine production by opioids may lead to the development of more effective therapeutics for inflammatory diseases.
16571771 IFN-beta-induced SOCS-1 negatively regulates CD40 gene expression in macrophages and micro 2006 May Costimulation between T cells and antigen-presenting cells is required for adaptive immune responses. CD40, a costimulatory molecule, is expressed in macrophages and microglia. The aberrant expression of CD40 is involved in human diseases including multiple sclerosis, rheumatoid arthritis, and Alzheimer's disease. CD40 expression is induced by a variety of stimuli, including IFN-gamma and lipopolysaccharide (LPS). In this study, we describe the molecular basis by which IFN-beta, a cytokine with immunomodulatory properties, regulates CD40 gene expression. IFN-beta induces CD40 expression in macrophages and microglia at the transcriptional level, and GAS elements in the CD40 promoter are required for IFN-beta-induced CD40 promoter activity. The critical role of signal transducers and activators of transcription-1alpha (STAT-1alpha) in this response was confirmed by utilizing primary microglia from STAT-1alpha deficient mice. IFN-beta induces suppressor of cytokine signaling-1 (SOCS-1) gene expression, which inhibits cytokine signaling by inhibiting activation of STAT proteins. The ectopic expression of SOCS-1 abrogates IFN-beta-mediated STAT-1alpha activation and inhibits IFN-beta-induced CD40 expression. IFN-beta-induced recruitment of STAT-1alpha and RNA Pol II and permissive histone modifications on the CD40 promoter are also inhibited by SOCS-1 overexpression. These novel results indicate that IFN-beta-induced SOCS-1 plays an important role in the negative regulation of IFN-beta-induced CD40 gene expression.
16525037 A selective small molecule IkappaB Kinase beta inhibitor blocks nuclear factor kappaB-medi 2006 Jun IkappaB kinase (IKK) beta is essential for inflammatory cytokine-induced activation of nuclear factor kappaB (NF-kappaB). NF-kappaB plays a pivotal role in the function of major cell types that contribute to the pathophysiological process of rheumatoid arthritis (RA). Here, we report the mechanism and the effect of the IKKbeta inhibitor N-(6-chloro-7-methoxy-9H-beta-carbolin-8-yl)-2-methylnicotinamide (ML120B), a beta-carboline derivative, on NF-kappaB signaling and gene activation in RA-relevant cell systems. ML120B is a potent, selective, reversible, and ATP-competitive inhibitor of IKKbeta with an IC50 of 60 nM when evaluated in an IkappaBalpha kinase complex assay. ML120B does not inhibit other IKK isoforms or a panel of other kinases. ML120B concentration-dependently inhibits tumor necrosis factor alpha (TNFalpha)-stimulated NF-kappaB signaling via inhibition of IkappaBalpha phosphorylation, degradation, and NF-kappaB translocation into the nucleus. For the first time, we have demonstrated that in human fibroblast-like synoviocytes, TNFalpha- or interleukin (IL)-1beta-induced monocyte chemoattractant protein-1 regulated on activation, normal T cell expressed and secreted and production is IKKbeta-dependent. In addition, for the first time, we have demonstrated that lipopolysaccharide- or peptidoglycan-induced cytokine production in human cord blood-derived mast cells is IKKbeta-dependent. In addition, in human chondrocytes, ML120B inhibited IL-1beta-induced matrix metalloproteinase production with an IC50 of approximately 1 microM. ML120B also blocked IL-1beta-induced prostaglandin E2 production. In summary, ML120B blocked numerous NF-kappaB-regulated cell responses that are involved in inflammation and destructive processes in the RA joint. Our findings support the evaluation of IKKbeta inhibitors as anti-inflammatory agents for the treatment of RA.
16418778 Human osteoblast precursors produce extracellular adenosine, which modulates their secreti 2006 Feb We showed that human osteoprogenitor cells produced adenosine and expressed ecto-5'-nucleotidase and all four adenosine receptor subtypes. Adenosine stimulated IL-6 but inhibited osteoprotegerin secretion, suggesting that adenosine is a newly described regulator of progenitor cell function. INTRODUCTION: Maintaining skeletal homeostasis relies on there being a balance between bone formation and resorption; an imbalance between these processes can lead to diseases such as osteoporosis and rheumatoid arthritis. Recent reports showed that locally produced ATP, acting through P2 receptors, has pronounced effects on bone formation. However, ATP can be enzymatically cleaved to adenosine that has little or no activity at P2 receptors but mediates its action through the P1 family of receptors. We studied whether adenosine may also have an important role in controlling bone cell differentiation and function. MATERIALS AND METHODS: Extracellular adenosine levels were analyzed by high-performance liquid chromatography in HCC1 and bone marrow stromal (BMS) cells. Ecto-5'-nucleotidase (CD73) expression and activity was determined by RT-PCR, immunocytochemistry, and the cleavage of etheno-AMP to ethenoadenosine. Adenosine receptor expression and activity were determined by RT-PCR and cAMP measurements. The effects of adenosine receptor agonists on IL-6, osteoprotegerin (OPG), and RANKL expression were determined by ELISA and QRT-PCR. RESULTS: HCC1 and BMS cells produce adenosine and express CD73 and all four adenosine receptor subtypes. The A2b receptor was shown to be functionally dominant in HCC1 cells, as determined by cAMP production and in its stimulation of IL-6 secretion. Adenosine receptor agonism also inhibited OPG secretion and OPG but not RANKL mRNA expression. CONCLUSIONS: Our findings show that HCC1 and primary BMS cells produce adenosine, express CD73 and all four adenosine receptor subtypes. In HCC1 cells, adenosine has a potent stimulatory action on IL-6 secretion but an inhibitory action on OPG expression. These data show for the first time that adenosine may be an important regulator of progenitor cell differentiation and hence an important local contributor to the regulation of bone formation and resorption.
19014637 IL-17 expression by breast-cancer-associated macrophages: IL-17 promotes invasiveness of b 2008 INTRODUCTION: IL-17 plays an important role in autoimmunity, promoting autoimmunity, inflammation and invasion in multiple sclerosis, rheumatoid arthritis and type I diabetes. The role of IL-17 in cancer is unclear, however, as there are few studies examining IL-17 protein expression in cancer. We therefore examined IL-17 protein expression in human breast cancer and modelled its potential biological significance in vitro. METHODS: Immunohistochemistry was used to determine IL-17 expression in breast cancers. Matrigel invasion assays were employed to examine the effect of IL-17 on cancer cell invasion by a panel of breast cancer cell lines. The role of matrix metalloproteinases (MMPs) was investigated with selective antagonists and immunoassays for MMP-2, MMP-3, MMP-9 and tissue inhibitor of MMP. RESULTS: IL-17-expressing cells with macrophage morphology were identified in the peritumoural area of a proportion of patients (8/19 patients). Macrophages were confirmed by CD68 staining on serial sections. With the exception of occasional lymphocytes, one patient with rare multinucleate giant cells and one patient with occasional expression of IL-17 in tumour cells, no other IL-17-positive cells were detected. Addition of IL-17 to cell lines in vitro stimulated marked invasion of Matrigel. In contrast, IL-17 did not promote the invasion of MCF7 or T47D cell lines. Invasion was initially thought to be dependent on MMPs, as evidenced by the broad-spectrum MMP inhibitor GM6001 and selective antagonists of MMP-2/MMP-9 and MMP-3. Measurement of MMP-2, MMP-3 and MMP-9, and tissue inhibitor of MMP 1 secretion, failed to reveal any changes in expression following IL-17 exposure. In contrast, TNF promoted secretion of MMPs but IL-17 did not augment TNF, indicating that IL-17 acts via an independent mechanism. CONCLUSIONS: The present study is the first to describe in situ expression of IL-17 protein in human breast tumours and to propose a direct association between IL-17 and breast cancer invasion. The precise effectors of IL-17-dependent invasion remain to be characterised but could include a range of proteases such as a disintegrin and metalloproteinase protein or astacins. Nevertheless, this work identifies a novel potential mechanism for breast cancer invasion and tumour progression, the prognostic implication of which is currently under investigation.
18683034 Decreased DNA methyltransferase 3A and 3B mRNA expression in peripheral blood mononuclear 2008 Sep OBJECTIVE: DNA methylation is known to play an important role in gene transcription and alterations of methylation contribute to the development of certain disorders such as cancer and immunodeficiency. Recent years have found an increasing interest in the role of epigenetic modifications in the etiology of human autoimmune diseases, such as systemic lupus erythromatosus (SLE) and rheumatoid arthritis (RA). DNA methyltransferases (DNMTs) are involved in the epigenetic control of DNA methylation processes. S-adenosylmethionine (SAM) and S-adenosylhomocysteine (SAH), as the substrate and product of essential cellular methyltransferase reactions, have important indicator action of cellular methylation status. The aim of this study is to explore if DNA methylation plays a role in the pathogenesis of idiopathic thrombocytopenic purpura (ITP). METHODS: DNMT1, DNMT3A, and DNMT3B mRNA expression in peripheral blood mononuclear cells (PBMCs) of adult ITP patients were analyzed by real-time quantitative polymerase chain reaction. Plasma SAM and SAH levels were assayed with reversed-phase high performance liquid chromatography (HPLC). RESULTS: DNMT3A and DNMT3B mRNA expressions were significantly lower in ITP patients than in healthy controls (p < 0.001), while DNMT1 mRNA expression was not significantly different between the two groups (p = 0.774). Plasma SAH concentration was significantly elevated in ITP patients than in healthy controls (p < 0.05), while the plasma SAM and SAM/SAH were not significantly different between the two groups (p = 0.133, p = 0.624 respectively). CONCLUSIONS: Our observations suggest that aberrant DNA methylation status reflected by increased plasma SAH concentration and decreased mRNA expression levels of DNMT3A and 3B are possibly involved in the pathogenesis of ITP although the precise mechanisms need further study.
18295198 A rat air pouch model for evaluating the efficacy and selectivity of 5-lipoxygenase inhibi 2008 Apr 14 The 5-lipoxygenase (5-LOX) pathway has been associated with a variety of inflammatory diseases including asthma, atherosclerosis, rheumatoid arthritis, pain, cancer and liver fibrosis. Several classes of 5-LOX inhibitors have been identified, but only one drug, zileuton, a redox inhibitor of 5-LOX, has been approved for clinical use. To better evaluate the efficacy of 5-LOX inhibitors for pharmacological intervention, a rat model was modified to test the in vivo efficacy of 5-LOX inhibitors. Inflammation was produced by adding carrageenan into a newly formed air pouch and prostaglandins produced. While macrophages and neutrophils are present in the inflamed pouch, little 5-LOX products are formed. Cellular 5-LOX activation was obtained by adding calcium ionophore (A23187) into the pouch thus providing a novel model to evaluate the efficacy and selectivity of 5-LOX inhibitors. Also, we described modifications to the in vitro 5-LOX enzyme and cell assays. These assays included a newly developed fluorescence-based enzyme assay, a 5-LOX redox assay, an ex vivo human whole blood assay and an IgE-stimulated rat mast cell assay, all designed for maximal production of leukotrienes. Zileuton and CJ-13,610, a competitive, non-redox inhibitor of 5-LOX, were evaluated for their pharmacological properties using these assays. Although both compounds achieved dose-dependent inhibition of 5-LOX enzyme activity, CJ-13,610 was 3-4 fold more potent than zileuton in all-assays. Evaluation of 5-LOX metabolites-by LC/MS/MS and ELISA confirmed that both compounds selectively inhibited all products downstream of 5-hydroperoxy eicosatetraenoic acid (5-HPETE), including 5-oxo-6,8,11,14-eicosatetraenoic acid (5-oxoETE), without inhibition of 12-lipoxygenase (12-LOX), 15-lipoxygenase (15-LOX), or cyclooxygenase (COX) products. In the rat air pouch model, oral dosing of CJ-13,610 and zileuton resulted in selective inhibition 5-LOX activity from pouch exudate and ex vivo rat whole blood with similar potency to in vitro assay. These data show that the rat air pouch model is a reliable and useful tool for evaluating in vivo efficacy of 5-LOX inhibitors and may aid in the development of the next generation of 5-LOX inhibitors, such as the non-redox inhibitors similar to CJ-13,610.
18073300 Interleukin (IL)-23 receptor is a major susceptibility gene for Graves' ophthalmopathy: th 2008 Mar CONTEXT: IL-23 and its receptor (IL-23R) guide T cells toward the T-helper 17 phenotype. IL-23R single nucleotide polymorphisms (SNPs) have been associated with several autoimmune diseases, including Crohn's disease and rheumatoid arthritis. OBJECTIVE: Our objective was to determine whether variants in the IL-23R gene are associated with Graves' disease (GD) and Graves' ophthalmopathy (GO). DESIGN AND PARTICIPANTS: A total of 216 North American Caucasian GD patients and 368 healthy controls were genotyped for four SNPs spanning the IL-23R gene. SNPs rs11209026 and rs7530511 were genotyped using the TaqMan allelic discrimination assays (Applied Biosystems, Foster City, CA), and SNPs rs2201841 and rs10889677 were genotyped using a fluorescent-based restriction fragment length polymorphism method. RESULTS: The A allele of rs2201841 was present in 78.8% of GD patients with GO and 64.7% of controls [P=1.1x10(-4); odds ratio (OR)=2.04]; the AA genotype was also significantly increased in GO patients compared with controls (62.5 and 41%, respectively; P=1.0x10(-4); OR=2.4). The C allele of rs10889677 was present in 78.6% of GO patients and 64.5% of controls (P=1.3x10(-4); OR=2.03), and the CC genotype was also significantly increased in GO patients vs. controls (62.1 and 41.0%, respectively; P=1.4x10(-4); OR=2.36). The TT genotype of rs7530511 was significantly associated with GD, but not specifically with GO; it was present in 2.5% of GD patients and 0.3% of controls (P=0.02; OR=9.4). The rs11209026 SNP, which is the most strongly associated with Crohn's disease, was not associated with GD or GO in our data set. CONCLUSIONS: Variants in the IL-23R gene are strongly associated with GO. These variants may predispose to GO by changing the expression and/or function of IL-23R, thereby promoting a proinflammatory signaling cascade.
17579055 Altered dynamics of Kv1.3 channel compartmentalization in the immunological synapse in sys 2007 Jul 1 Aberrant T cell responses during T cell activation and immunological synapse (IS) formation have been described in systemic lupus erythematosus (SLE). Kv1.3 potassium channels are expressed in T cells where they compartmentalize at the IS and play a key role in T cell activation by modulating Ca(2+) influx. Although Kv1.3 channels have such an important role in T cell function, their potential involvement in the etiology and progression of SLE remains unknown. This study compares the K channel phenotype and the dynamics of Kv1.3 compartmentalization in the IS of normal and SLE human T cells. IS formation was induced by 1-30 min exposure to either anti-CD3/CD28 Ab-coated beads or EBV-infected B cells. We found that although the level of Kv1.3 channel expression and their activity in SLE T cells is similar to normal resting T cells, the kinetics of Kv1.3 compartmentalization in the IS are markedly different. In healthy resting T cells, Kv1.3 channels are progressively recruited and maintained in the IS for at least 30 min from synapse formation. In contrast, SLE, but not rheumatoid arthritis, T cells show faster kinetics with maximum Kv1.3 recruitment at 1 min and movement out of the IS by 15 min after activation. These kinetics resemble preactivated healthy T cells, but the K channel phenotype of SLE T cells is identical to resting T cells, where Kv1.3 constitutes the dominant K conductance. The defective temporal and spatial Kv1.3 distribution that we observed may contribute to the abnormal functions of SLE T cells.
17204566 Effect of chondroitin sulphate in symptomatic knee osteoarthritis: a multicentre, randomis 2007 May OBJECTIVE: To evaluate the efficacy and tolerability of chondroitin sulphate (chondroitin sulphate) in knee osteoarthritis. PATIENTS AND METHODS: A 24-week, randomised placebo-controlled trial of chondroitin sulphate (1 g/day) in patients with symptomatic knee osteoarthritis as measured on a visual analogue scale. Pain on daily activities and Lequesne's Index were the primary efficacy criteria. Secondary outcomes included the rate of responders according to the outcome measures in rheumatoid arthritis clinical trials of the Osteoarthritis Research Society International (OMERACT-OARSI) criteria, quality of life, patient's/physician's global assessments and carry-over effect after treatment. Biochemical markers of bone (CTX-I), cartilage (CTX-II) and synovium (hyaluronic acid) metabolism were also measured. Safety was assessed by recording adverse events (AEs). Statistical analysis was performed on the inter-group differences in the intention-to-treat population. RESULTS: 307 patients were included in the study. 28 (9%) patients discontinued the study because of lack of efficacy or AEs. At the end of treatment, the decrease in pain was -26.2 (24.9) and -19.9 (23.5) mm and improved function was -2.4 (3.4) (-25%) and -1.7 (3.3) (-17%) in the chondroitin sulphate and placebo groups, respectively (p = 0.029 and 0.109). The OMERACT-OARSI responder rate was 68% in the chondroitin sulphate and 56% in the placebo group (p = 0.03). The investigator's assessments and short form 12 (SF-12) physical component reported improvement more frequently in the chondroitin sulphate than in the placebo group (p = 0.044 and 0.021, respectively). No significant difference was observed between treatment groups for changes in biomarkers over 24 weeks. However, there was a significant difference between non-responders and responders according to the OARSI criteria for 24-week changes of CTX-I (p = 0.018) and CTX-II (p = 0.014). Tolerance was considered to be satisfactory. CONCLUSION: This study failed to show an efficacy of chondroitin sulphate on the two primary criteria considered together, although chondroitin sulphate was slightly more effective than placebo on pain, OMERACT-OARSI response rate, investigator's assessment and quality of life.