Tumor necrosis factor-alpha triggers a cytokine cascade yielding postoperative cognitive decline.

Cognitive decline following surgery in older individuals is a major clinical problem of uncertain mechanism; a similar cognitive decline also follows severe infection, chemotherapy, or trauma and is currently without effective therapy. A variety of mechanisms have been proposed, and exploring the role of inflammation, we recently reported the role of IL-1ß in the hippocampus after surgery in mice with postoperative cognitive dysfunction. Here, we show that TNF-α is upstream of IL-1 and provokes its production in the brain. Peripheral blockade of TNF-α is able to limit the release of IL-1 and prevent neuroinflammation and cognitive decline in a mouse model of surgery-induced cognitive decline. TNF-α appears to synergize with MyD88, the IL-1/TLR superfamily common signaling pathway, to sustain postoperative cognitive decline. Taken together, our results suggest a unique therapeutic potential for preemptive treatment with anti-TNF antibody to prevent surgery-induced cognitive decline.

Nonsteroidal anti-inflammatory drugs increase TNF production in rheumatoid synovial membrane cultures and whole blood.

Nonsteroidal anti-inflammatory drugs (NSAIDs) inhibit cyclooxygenase activity and hence PG production. However, the ability of NSAIDs to ameliorate pain and tenderness does not prevent disease progression in rheumatoid arthritis, a disease whose pathogenesis is linked to the presence of proinflammatory cytokines, such as TNF-alpha. To understand this observation, we have examined the effect of NSAIDs on the production of clinically validated proinflammatory cytokines. We show that a variety of NSAIDs superinduce production of TNF from human peripheral blood monocytes and rheumatoid synovial membrane cultures. A randomized, double-blinded, crossover, placebo-controlled trial in healthy human volunteers also revealed that the NSAID drug celecoxib increased LPS-induced TNF production in whole blood. NSAID-mediated increases in TNF are reversed by either the addition of exogenous PGE(2) or by a PGE(2) EP2 receptor agonist, revealing that PGE(2) signaling via its EP2 receptor provides a valuable mechanism for controlling excess TNF production. Thus, by reducing the level of PGE(2), NSAIDs can increase TNF production and may exacerbate the proinflammatory environment both within the rheumatoid arthritis joint and the systemic environment.

Inhibitors of p38 suppress cytokine production in rheumatoid arthritis synovial membranes: does variable inhibition of interleukin-6 production limit effectiveness in vivo?

OBJECTIVE: The activity of p38 MAPK regulates lipopolysaccharide (LPS)-stimulated production of key proinflammatory cytokines such as tumor necrosis factor α (TNFα). Consequently, p38 MAPK inhibitors have attracted considerable interest as potential treatments of rheumatoid arthritis (RA), and studies in murine models of arthritis have yielded promising results. However, the performance of several compounds in human clinical trials has been disappointing. At present, the reason for this poor performance is unclear. The aim of this study was to examine the effects of p38 inhibitors on both diseased and normal human tissue and cells, in order to test whether this kinase still plays a critical role in cytokine production under conditions of chronic inflammation. METHODS: Proinflammatory and antiinflammatory cytokine production was monitored after treatment of primary human monocytes, macrophages, and RA synovial membrane cultures with p38 MAPK inhibitor compounds. The following 3 inhibitors were used in these studies: SB-203580 (inhibits the α and ß isoforms), BIRB-796 (inhibits the α, ß, γ, and δ isoforms), and a novel, structurally distinct p38 MAPK inhibitor, SB-731445 (inhibits the α and ß isoforms). RESULTS: SB-731445 and SB-203580 produced profound inhibition of spontaneous production of proinflammatory cytokines (TNFα and interleukin-1 [IL-1]) in both RA membrane cultures and LPS-stimulated primary human monocytes. However, this and other p38 MAPK inhibitors produced a significant increase in IL-6 production by LPS-stimulated primary human macrophages and a decrease in IL-10 production by all cell types examined. CONCLUSION: The potentially proinflammatory consequences of these activities (decreased IL-10 production and increased IL-6 production) may offer some explanation for the inability of p38 MAPK inhibitors to provide the therapeutic benefit that had been hoped for in RA.

SIGIRR/TIR-8 is an inhibitor of Toll-like receptor signaling in primary human cells and regulates inflammation in models of rheumatoid arthritis.

OBJECTIVE: Single-immunoglobulin interleukin-1 receptor-related (SIGIRR), which is also known as Toll/interleukin-1 receptor 8 (TIR-8), is a member of the TIR domain-containing family of receptors and was first characterized as an inhibitor of interleukin-1 receptor (IL-1R) and Toll-like receptor (TLR) signaling. In the Dextran sulfate sodium-induced colitis model, SIGIRR(-/-) mice were shown to have increased inflammation and to be more susceptible to endotoxin challenge. Increasing evidence implicates TLR and IL-1R signaling in the pathology of rheumatoid arthritis (RA). Therefore, the purpose of this study was to investigate the involvement of SIGIRR in regulating inflammation in disease-relevant models. METHODS: Primary human monocyte-derived macrophages and dendritic cells (DCs) were used to overexpress SIGIRR as well as to knock down endogenously expressed SIGIRR using small interfering RNAs. SIGIRR was also overexpressed in synovial cells derived from RA patients. To investigate the role of SIGIRR in vivo, zymosan-induced arthritis (ZIA) and collagen antibody-induced arthritis (CAIA) were induced in SIGIRR-knockout mice. RESULTS: SIGIRR overexpression inhibited TLR-induced cytokine production in macrophages and DCs, while SIGIRR knockdown resulted in increased cytokine production following TLR stimulation. Moreover, SIGIRR overexpression inhibited the spontaneous release of cytokines by human RA synovial cells. The role of SIGIRR as an inhibitor of inflammation was confirmed in vivo, since SIGIRR(-/-) mice developed a more severe disease in both the ZIA and CAIA models. CONCLUSION: Our study is the first to show the expression pattern and function of SIGIRR in primary human cells. Furthermore, this investigation defines the role of SIGIRR in disease-relevant cell types and demonstrates that SIGIRR is a potential therapeutic target for RA.

Oral activated charcoal prevents experimental cerebral malaria in mice and in a randomized controlled clinical trial in man did not interfere with the pharmacokinetics of parenteral artesunate.

BACKGROUND: Safe, cheap and effective adjunct therapies preventing the development of, or reducing the mortality from, severe malaria could have considerable and rapid public health impact. Oral activated charcoal (oAC) is a safe and well tolerated treatment for acute poisoning, more recently shown to have significant immunomodulatory effects in man. In preparation for possible efficacy trials in human malaria, we sought to determine whether oAC would i) reduce mortality due to experimental cerebral malaria (ECM) in mice, ii) modulate immune and inflammatory responses associated with ECM, and iii) affect the pharmacokinetics of parenteral artesunate in human volunteers. METHODS/PRINCIPAL FINDINGS: We found that oAC provided significant protection against P. berghei ANKA-induced ECM, increasing overall survival time compared to untreated mice (p<0.0001; hazard ratio 16.4; 95% CI 6.73 to 40.1). Protection from ECM by oAC was associated with reduced numbers of splenic TNF(+) CD4(+) T cells and multifunctional IFNgamma(+)TNF(+) CD4(+) and CD8(+) T cells. Furthermore, we identified a whole blood gene expression signature (68 genes) associated with protection from ECM. To evaluate whether oAC might affect current best available anti-malarial treatment, we conducted a randomized controlled open label trial in 52 human volunteers (ISRCTN NR. 64793756), administering artesunate (AS) in the presence or absence of oAC. We demonstrated that co-administration of oAC was safe and well-tolerated. In the 26 subjects further analyzed, we found no interference with the pharmacokinetics of parenteral AS or its pharmacologically active metabolite dihydroartemisinin. CONCLUSIONS/SIGNIFICANCE: oAC protects against ECM in mice, and does not interfere with the pharmacokinetics of parenteral artesunate. If future studies succeed in establishing the efficacy of oAC in human malaria, then the characteristics of being inexpensive, well-tolerated at high doses and requiring no sophisticated storage would make oAC a relevant candidate for adjunct therapy to reduce mortality from severe malaria, or for immediate treatment of suspected severe malaria in a rural setting. TRIAL REGISTRATION: Controlled-Trials.com ISRCTN64793756.

Toll-like receptor-2 mediates inflammation and matrix degradation in human atherosclerosis.

BACKGROUND: Inflammation and matrix degradation are the hallmarks of high-risk atherosclerosis that leads to myocardial infarction and stroke. Toll-like receptors (TLRs), key players in innate immunity, are upregulated in atherosclerotic lesions, but their functional role in human atherosclerosis is unknown. We explored the effects of blocking TLR-2, TLR-4, and myeloid differentiation primary response gene 88 (MyD88), a signaling adaptor shared by most TLRs and interleukin-1 receptor (IL-1R), in an in vitro model of human atherosclerosis. METHODS AND RESULTS: Carotid endarterectomies were obtained from patients with symptomatic carotid disease. Cells were isolated via enzymatic tissue dissociation and cultured in the presence or absence of TLR signaling blockers. A dominant-negative form of MyD88 (MyD88(DN)) decreased the production of monocyte chemotactic protein-1/CCL2 (P=0.000), IL-8/CXCL8 (P=0.006), IL-6 (P=0.002), matrix metalloproteinase-1 (MMP-1; P=0.002), and MMP-3 (P=0.000), as well as nuclear factor-kappaB activation (P<0.05) in atheroma cell cultures. IL-1R antagonist, TLR-4 blocking antibodies, or overexpression of a dominant-negative form of the TLR-4 signaling adaptor TRIF-related adaptor molecule reduced nuclear factor-kappaB activity but did not have a broad impact on the production of the mediators studied. In contrast, TLR-2 neutralizing antibodies inhibited nuclear factor-kappaB activation (P<0.05) and significantly reduced monocyte chemotactic protein-1/CCL2 (P=0.000), IL-8/CXCL8 (P=0.009), IL-6 (P=0.000), and MMP-1 (P=0.000), MMP-2 (P=0.004), MMP-3 (P=0.000), and MMP-9 (P=0.006) production. CONCLUSIONS: Our data indicate that TLR-2 signaling through MyD88 plays a predominant role in inflammation and matrix degradation in human atherosclerosis. TLR-2 blockade may represent a therapeutic strategy for atherosclerosis and its complications.

RelB/p50 regulates CCL19 production, but fails to promote human DC maturation.

DC, when fully matured are the APC best able to activate naïve T cells. Recently, we demonstrated using adenoviruses overexpressing IkappaBalpha and proteosome inhibitors that NF-kappaB is involved in DC activation, but the role of the individual subunits is still not clear. We investigated the role of the NF-kappaB subunits RelB and p50 in human DC activation using adenoviral vectors expressing RelB or p50. Nuclear RelB, in the form of RelB/p50, was active only in DC infected with both viruses, this induced the production of the soluble homeostatic chemokine CCL19, but not other homeostatic chemokines, particularly in LPS-matured DC. However, RelB/p50 did not affect the expression of costimulatory and antigen-presenting molecules, and increased the allogeneic mixed lymphocyte reaction only in LPS-matured DC. This enhanced mixed lymphocyte reaction is most likely due to enhanced CCL19 production, which sustains the interaction between mature DC and naïve T cells. In conclusion, we demonstrated that RelB/p50 was active only in DC expressing both RelB and p50, and induced CCL19 production, but not DC maturation.

Disordered macrophage cytokine secretion underlies impaired acute inflammation and bacterial clearance in Crohn's disease.

The cause of Crohn's disease (CD) remains poorly understood. Counterintuitively, these patients possess an impaired acute inflammatory response, which could result in delayed clearance of bacteria penetrating the lining of the bowel and predispose to granuloma formation and chronicity. We tested this hypothesis in human subjects by monitoring responses to killed Escherichia coli injected subcutaneously into the forearm. Accumulation of (111)In-labeled neutrophils at these sites and clearance of (32)P-labeled bacteria from them were markedly impaired in CD. Locally increased blood flow and bacterial clearance were dependent on the numbers of bacteria injected. Secretion of proinflammatory cytokines by CD macrophages was grossly impaired in response to E. coli or specific Toll-like receptor agonists. Despite normal levels and stability of cytokine messenger RNA, intracellular levels of tumor necrosis factor (TNF) were abnormally low in CD macrophages. Coupled with reduced secretion, these findings indicate accelerated intracellular breakdown. Differential transcription profiles identified disease-specific genes, notably including those encoding proteins involved in vesicle trafficking. Intracellular destruction of TNF was decreased by inhibitors of lysosomal function. Together, our findings suggest that in CD macrophages, an abnormal proportion of cytokines are routed to lysosomes and degraded rather than being released through the normal secretory pathway.

The role of transposable elements in the regulation of IFN-lambda1 gene expression.

IFNs lambda1, lambda2, and lambda3, or type III IFNs, are recently identified cytokines distantly related to type I IFNs. Despite an early evolutionary divergence, the 2 types of IFNs display similar antiviral activities, and both are produced primarily in dendritic cells. Although virus induction of the type I IFN-beta gene had served as a paradigm of gene regulation, relatively little is known about the regulation of IFN-lambda gene expression. Studies of virus induction of IFN-lambda1 identified an essential role of IFN regulatory factors (IRF) 3 and 7, which bind to a regulatory DNA sequence near the start site of transcription. Here, we report that the proximal promoter region of the IFN-lambda1 regulatory region is not sufficient for maximal gene induction in response to bacterial LPS, and we identify an essential cluster of homotypic NF-kappaB binding sites. Remarkably, these sites, which bind efficiently to NF-kappaB and function independently of the IRF3/7 binding sites, originate as transposable elements of the Alu and LTR families. We also show that depletion of the NF-kappaB RelA protein significantly reduces the level of the IFN-lambda1 gene expression. We conclude that IFN-lambda1 gene expression requires NF-kappaB, and we propose a model for IFN-lambda1 gene regulation, in which IRF and NF-kappaB activate gene expression independently via spatially separated promoter elements. These observations provide insights into the independent evolution of the IFN-lambda1 and IFN-beta promoters and directly implicate transposable elements in the regulation of the IFN-lambda1 gene by NF-kappaB.

Tyrosine kinases and inflammatory signalling.

The activity of tyrosine kinases is central to many cellular processes, and accumulating evidence suggests that their role in inflammation is no less profound. Three main tyrosine kinase families, the Src, Tec and Syk kinase families are intimately involved in TLR signalling, the critical first step in cellular recognition of invading pathogens and tissue damage. Their activity results in changes in gene expression in affected cells. Key amongst these genes are the cytokines, which orchestrate both the duration and extent of inflammation. Tyrosine kinases also play important roles in cytokine function, and are implicated in signalling through both pro- and anti-inflammatory cytokines such as TNF, IL-6 and IL-10. Thus, strategies to modulate tyrosine kinase activity have significant therapeutic potential in combating the chronic inflammatory state that is typical of many major health issues that face us today, including Rheumatoid Arthritis, Cardiovascular disease and cancer. Here we review current knowledge of the role of tyrosine kinases in inflammation with particular emphasis on their role in TLR signalling.

Bmx regulates LPS-induced IL-6 and VEGF production via mRNA stability in rheumatoid synovial fibroblasts.

Discordant cytokine production is characteristic of chronic inflammatory conditions like rheumatoid arthritis (RA), and anti-cytokine therapeutics are becoming routinely used to treat RA in the clinic. Fibroblasts from rheumatoid synovium have been shown to contribute to cytokine production in inflamed joints; likewise these cells also produce cytokines in response to inflammatory mediators signalling through Toll like receptors (TLRs). Tyrosine kinase activity is essential to LPS-induced cytokine production, and we have previously implicated a role for the Tec kinase, Bmx, in inflammatory cytokine production. Here we show that Bmx kinase activity in RASF is increased following LPS stimulation and that Bmx is involved in the regulation of LPS-induced IL-6 and VEGF production via mRNA stabilisation. This is an important insight into the regulation of VEGF, which is involved in a wide range of different pathologies, and may lead to more effective design of novel anti-inflammatory/angiogenic therapeutics for conditions such as RA.

Rac mediates TNF-induced cytokine production via modulation of NF-kappaB.

TNF is a key factor in a variety of inflammatory diseases. Here we report that TNF induced pro-inflammatory cytokine synthesis of IL-6 and IL-8 is mediated by the Rho GTPase Rac. TNF induces p42/p44, p54 and p38 MAPK kinase; these kinases have been implicated in control of cytokine synthesis. However, over-expression of a dominant negative form of Rac strongly inhibited TNF-induced p42/44 MAPK kinase activation, but had little effect upon JNK and no effect upon p38 MAPK activity. Another key signalling pathway controlling cytokine expression is NF-kappaB. When analyzing TNF-induced NF-kappaB activity via luciferase-reporter assays or via EMSA, we were able to show that the dominant negative version of Rac could completely abrogate TNF-induced NF-kappaB activity. In addition, we also observed that inhibition of the ERK pathway led to a reduction in TNF-induced NF-kappaB transcriptional activity; this was accompanied by an ablation of TNF-induced p65 phosphorylation at serine 276. This would suggest that TNF-induced activation of Rac, lies upstream of NF-kappaB activation, and that the inhibition of this pathway results in inhibition of cytokine production.

Bmx tyrosine kinase regulates TLR4-induced IL-6 production in human macrophages independently of p38 MAPK and NFkapp}B activity.

Chronic inflammation, as seen in conditions such as rheumatoid arthritis and Crohn disease, is in part driven by discordant production of inflammatory cytokines, such as tumor necrosis factor-alpha and interleukin-6 (IL-6). Tyrosine kinase activity is essential to lipopolysaccharide-induced cytokine production in monocytes, and previous studies by us and others have implicated a role for the Tec kinase Bruton's tyrosine kinase (Btk) in inflammatory cytokine production. Here we show that knockdown of Btk using RNA interference results in decreased tumor necrosis factor-alpha, but not IL-6 production. Further investigations into the signaling mechanisms regulating IL-6 production led to the discovery that the Tec kinase bone marrow tyrosine kinase gene in chromosome X (Bmx) regulates Toll-like receptor-induced IL-6 production. Our data further showed that Bmx-dependent super-induction of IL-6 does not involve nuclear factor-kappaB activity. More detailed investigations of pathways downstream of Bmx signaling revealed that Bmx targets the IL-6 3' untranslated region to increase mRNA stabilization via a novel, thus far undefined, p38 mitogen activated protein kinase-independent pathway. These data have important implications for the design of therapeutics targeted against specific cytokines and their regulators in inflammatory disease.

Chemical inhibition of Src family kinases affects major LPS-activated pathways in primary human macrophages.

Understanding the signalling mechanisms controlling inflammatory cytokine production is pivotal to the research of both acute and chronic immune disorders. Tyrosine phosphorylation is one of the earliest events to occur in response to an immune challenge yet the role of specific tyrosine kinases in inflammatory cytokine production has been difficult to ascribe due to conflicting literature. Here we show that the pyrazolo pyrimidine compound PP2, a selective inhibitor of Src family kinases (SFK), can inhibit LPS-induced TNF production as well as a number of other inflammatory cytokines. In addition, we show similar effects of PP2 on cytokine production when induced by other TLRs, (1, 2 and 5-8), indicating that SFK are important common regulators of TLR signalling. PP2 suppressed the activity of both TNF and IL-10 driven reporter genes, suggesting that this activity is mediated at the level of transcription. Interestingly, however, PP2 had no significant effect on the activation of NF-kappaB, or on p42/44 ERK, p46/54 JNK or p38 MAPK phosphorylation. In contrast, PP2 did inhibit AP-1 nuclear accumulation in response to LPS. Taken together, these findings show that the Src kinases are able to control inflammatory cytokine production at the transcriptional level independently of NF-kappaB, and highlight the role of the AP-1 family of transcription factors as downstream mediators of Src kinase action.

Could toll-like receptors provide a missing link in chronic inflammation in rheumatoid arthritis? Lessons from a study on human rheumatoid tissue.

In the last decade the development of a number of biological therapies has revolutionised the treatment of rheumatic diseases. The first and most widely used of these approaches, tumour necrosis factor (TNF) blockade (infliximab, entanercept, adalimumab), has now been administered to over a million patients. However, the success of these biological therapies has also highlighted their limitations. None of these treatments has shown a 100% patient response; normally responses are in the 50-70% range. As proteins, these drugs cannot be given orally and they are expensive to produce, a cost ultimately borne by the patient/health provider that can seriously limit the availability of these drugs. Lastly, these treatments, whether involving the systemic neutralisation of a cytokine (eg, TNF or IL6 receptor blockade (tocilizumab)), the ablation of a B cell population (anti-CD20, rituximab), or the potential disruption of important cellular interactions as with CTLA4-Ig (abatacept), can cause major perturbations of the immune system, the long-term effects of which are still unclear. At present, treatments such as TNF blockade can result in an increased infectious risk and the reactivation of tuberculosis can be a major issue in certain populations. As with all therapies, there is an increasing large refractory population over time. Therefore, despite the undoubted success of these therapies, there is room for improvement. Although it might be too much to expect any new treatment to affect a "cure" (all the current biological therapies require repeated administrations), there are definite gains to be made in terms of cost, oral bioavailability and a more selective interference with the immune-inflammatory response.

Adenoviral targeting of signal transduction pathways in synovial cell cultures.

Methods for high efficiency gene transfer into primary cells of various lineages and disease states are desirable, as they remove the uncertainties associated with using transformed cell lines. Adenoviruses have evolved to deliver their genes into cells with high efficiency and in recent years have been exploited as a gene transduction system. Prior to the discovery of adenoviruses, efficient expression of transgenes was only possible by cloning stably transfected cells; this was limited to cell lines and was not an option for primary cells. Here we describe a method of transgene expression, which enables previously untransfectable cells, such as primary myeloid cells or diseased synovium, to express protein at extremely high levels with nearly 100% of cells expressing the transgene. This allows us to examine the effect of target genes on signaling pathways in primary cells without the need for cell sorting or the simultaneous transfection of reporter genes. This is very important in studies of tissues such as rheumatoid synovium where sorting of cells will damage the biological value of the system.

Cutting edge: A transcriptional repressor and corepressor induced by the STAT3-regulated anti-inflammatory signaling pathway.

IL-10 regulates anti-inflammatory signaling via the activation of STAT3, which in turn controls the induction of a gene expression program whose products execute inhibitory effects on proinflammatory mediator production. In this study we show that IL-10 induces the expression of an ETS family transcriptional repressor, ETV3, and a helicase family corepressor, Strawberry notch homologue 2 (SBNO2), in mouse and human macrophages. IL-10-mediated induction of ETV3 and SBNO2 expression was dependent upon both STAT3 and a stimulus through the TLR pathway. We also observed that ETV3 expression was strongly induced by the STAT3 pathway regulated by IL-10 but not by STAT3 signaling activated by IL-6, which cannot activate the anti-inflammatory signaling pathway. ETV3 and SBNO2 repressed NF-kappaB- but not IFN regulatory factor 7 (IRF7)-activated transcriptional reporters. Collectively our data suggest that ETV3 and SBNO2 are components of the pathways that contribute to the downstream anti-inflammatory effects of IL-10.

IL-10 induces IL-10 in primary human monocyte-derived macrophages via the transcription factor Stat3.

IL-10 is an important immunosuppressive cytokine that can down-regulate expression of other cytokines and has been shown to down-regulate itself. We show, in this study, that treatment of human monocyte-derived macrophages with IL-10 induces IL-10 mRNA in a dose- and time-dependent manner with an optimum induction at 100 ng/ml and at 6 h, whereas IL-10-induced IL-10 protein can be detected at 18 h. In the same cells, IL-10 can partially suppress IL-10 mRNA induced by LPS, but only down to the level of IL-10-induced IL-10. An adenoviral luciferase reporter construct driven by the -195 IL-10 promoter, which contains a Stat motif, was readily induced by both IL-10 and LPS. Mutation of this Stat motif ablated IL-10 activation of this promoter, but not the LPS activation. Finally, we show that overexpression of a dominant-negative Stat3 protein will prevent IL-10 induction, but not LPS induction, of IL-10 mRNA. These data show that IL-10 induces IL-10 in monocyte-derived macrophages in an autocrine manner via activation of the transcription factor Stat3.

Inhibition of p38 mitogen-activated protein kinase is effective in the treatment of experimental crescentic glomerulonephritis and suppresses monocyte chemoattractant protein-1 but not IL-1beta or IL-6.

Activation of p38 mitogen-activated protein kinase (MAPK) is known to be important in cytokine production and cell survival in inflammation. This study examined the effect of inhibiting p38 MAPK after onset of renal injury in an experimental model of crescentic glomerulonephritis. Furthermore, this study investigated whether p38 MAPK inhibition would cause widespread suppression of the cytokine network in vivo or uncontrolled apoptosis. In the in vivo studies, daily treatment with a p38 MAPKalpha/beta inhibitor was started 1 h (early treatment study) or 4 d (late treatment study) after induction of nephrotoxic nephritis in Wistar Kyoto rats. The treated rats remained healthy with normal weight gain during the study. Both early and late treatment with p38 MAPK inhibitor reduced renal monocyte chemoattractant protein-1 (MCP-1) levels, the number of glomerular macrophages, the severity of tissue injury, and proteinuria compared with the vehicle group. Unexpected, treatment with p38 MAPK inhibitor did not suppress renal levels of IL-1beta or IL-6. In the in vitro study, the p38 MAPKalpha/beta inhibitor reduced production of MCP-1 and IL-6 by TNF-alpha-or IL-1beta-stimulated mesangial cells without any effect on cell viability or apoptosis. In conclusion, p38 MAPK inhibition is effective in reducing the severity of crescentic glomerulonephritis even when treatment is started after onset of disease. The therapeutic effect is associated with selective suppression of MCP-1, without widespread suppression of cytokine production or increased apoptosis. Therefore, p38 MAPK therapeutic blockade is a promising strategy in the treatment of antibody-mediated glomerulonephritis.

Expression of constitutively active STAT3 can replicate the cytokine-suppressive activity of interleukin-10 in human primary macrophages.

There is general agreement that signal transducer and activation of transcription 3 (STAT3) is required to mediate the anti-inflammatory activities of interleukin (IL)-10. However, STAT3 is activated by multiple factors that do not share the anti-inflammatory activity of IL-10. The question remains whether STAT3 is sufficient for the anti-inflammatory effects or whether there are other signals required, as had been suggested previously. We set out to map the human IL-10 receptor and to identify the key elements involved in transducing the cytokine-suppressive effects of IL-10. We were able to show an absolute requirement for both of the tyrosine residues found within the YXXQ-STAT3-docking site within the IL-10 receptor 1 and that no other signals appeared to be required. We used a constitutively active STAT3 to determine whether expression of this factor could suppress lipopolysaccharide-induced tumor necrosis factor and IL-6 production. Our data show that STAT3 activity can suppress both IL-6 and tumor necrosis factor production in lipopolysaccharide-stimulated macrophages. However, in synovial fibroblasts, STAT3 did not suppress IL-6 production, suggesting that the cellular environment plays an important role in dictating whether STAT3 drives a pro- or anti-inflammatory response.