The efficacy of indoximod upon stimulation with pro-inflammatory cytokines in triple-negative breast cancer cells.

BACKGROUND: Indoleamine 2,3-dioxygenase (IDO) inhibition has received much attention in cancer immunotherapy due to its role in immune escape in cancer cells. Additionally, changes in the pro-inflammatory cytokine levels can affect tumor growth and metastasis as well as the effectiveness of immunotherapy. The purpose of this study was for the first time to determine the effects of indoximod as an IDO inhibitor on triple-negative breast cancer (TNBC) and to assess the link between the efficacy of indoximod and IFN-γ or TNF-α stimulation. METHODS: The cytotoxic and apoptotic effects of indoximod alone or IFN-γ or TNF-α induction to mimic an inflammatory environment were evaluated by WST-1, Annexin V, cell cycle analysis, and acridine orange (AO)/ethidium bromide (EtBr) staining. Furthermore, the expression levels of IDO1 and PD-L1 expression were analyzed by RT-PCR. RESULTS: Our results demonstrated that indoximod significantly decreased the TNBC cell viability through apoptotic cell death (p < .05). The combination of indoximod and TNF-α was more effective than indoximod and IFN-γ stimulation or indoximod alone in TNBC cells. Additionally, PD-L1 expression level was significantly up-regulated after treatment with indoximod and TNF-α or IFN-γ combinations (p < .05). CONCLUSIONS: Indoximod exhibited a therapeutic potential in TNBC cells and pro-inflammatory cytokines could affect the effectiveness of indoximod. However, further studies are required to identify the role of the IDO-associated signaling pathways, the molecular mechanisms of indoximod induced apoptotic cell death, and the relationship between IDO inhibition by IDO inhibitors and pro-inflammatory cytokine levels.

Ribes nigrum Leaf Extract Preferentially Inhibits IFN-γ-Mediated Inflammation in HaCaT Keratinocytes.

Ribes nigrum L. (blackcurrant) leaf extracts, due to high levels of flavonols and anthocyanins, have been shown to exhibit beneficial effects in inflammatory diseases. However, whereas their traditional use has been investigated and validated in several models of inflammation and oxidative stress, the possible impact on skin disorders is still largely unknown. The purpose of this work was to elucidate the effects of R. nigrum leaf extract (RNLE) on keratinocyte-derived inflammatory mediators, elicited by a Th1 or Th2 cytokine milieu. HaCaT cells were challenged with TNF-α, either alone or in combination with the costimulatory cytokines IFN-γ or IL-4, and the release of proinflammatory cytokines and mediators (IL-8, IL-6, s-ICAM-1, and TSLP) was evaluated. The results showed that RNLE preferentially interferes with IFN-γ signaling, demonstrating only negligible activity on TNF-α or IL-4. This effect was attributed to flavonols, which might also account for the ability of RNLE to impair TNF-α/IL-4-induced TSLP release in a cAMP-independent manner. These results suggest that RNLE could have an antiallergic effect mediated in keratinocytes via mechanisms beyond histamine involvement. In conclusion, the discovery of RNLE preferential activity against IFN-γ-mediated inflammation suggests potential selectivity against Th1 type response and the possible use in Th1 inflammatory diseases.

Oncostatin M Is a Prognostic Biomarker and Inflammatory Mediator for Sepsis.

BACKGROUND: Oncostatin M (OSM) is a pleiotropic cytokine of the interleukin-6 family. The role of OSM in sepsis remains unknown. METHODS: Serum OSM level was determined and analyzed in septic patients on the day of intensive care unit (ICU) admission. Furthermore, the effects of OSM on polymicrobial sepsis induced by cecal ligation and puncture (CLP) were assessed. RESULTS: On the day of ICU admission, septic patients had significantly higher serum OSM levels when compared with ICU patient controls and healthy volunteers, which were related to the severity of sepsis, including parameters such as the sequential (sepsis-related) organ failure assessment score, procalcitonin level, and white blood cell number. A high serum OSM level on ICU admission was associated with 28-day mortality in septic patients. In CLP-induced polymicrobial sepsis, anti-OSM antibody decreased tissue inflammation and injury, and thus improved survival, while local and systemic bacterial dissemination was almost constant. Complementarily, supplementation with recombinant OSM protein in septic mice increased tissue injury, amplified inflammation, and worsened mortality after CLP, while it did not affect bacterial dissemination in septic mice. CONCLUSIONS: Sepsis results in an increased production of OSM, which might be a potential prognostic biomarker and therapeutic target for sepsis.

Anti-inflammatory or proinflammatory effect of an adenosine receptor agonist on the Th17 autoimmune response is inflammatory environment-dependent.

Adenosine is a key endogenous signaling molecule that regulates a wide range of physiological functions, including immune system function and inflammation. Studies have shown that adenosine receptor (AR) agonists can be either anti-inflammatory or proinflammatory in immune responses and in inflammation, and the clarification of the mechanisms causing these opposing effects should provide a better guide for therapeutic intervention. Whereas previous studies mostly examined the effects of AR agonists on Th1-type immune responses, in this study, we compared their effect on Th17 and Th1 autoimmune responses in experimental autoimmune uveitis, a mouse model of human uveitis induced by immunization with the human interphotoreceptor retinoid-binding protein peptides 1-20. We showed that injection of mice with a nonselective AR agonist, 5'-N-ethylcarboxamidoadenosine (NECA), at an early stage after immunization had an inhibitory effect on both Th1 and Th17 responses, whereas injection of the same amount of NECA at a late stage inhibited the Th1 response but had an enhancing effect on the Th17 response. We also showed that the effects of NECA on Th1 and Th17 responses were completely dissociated, that the enhancing effect of NECA on Th17 responses was modulated by γδ T cells, and that the response of γδ T cells to NECA was determined by their activation status. We conclude that the inflammatory environment has a strong impact on converting the effect of AR agonist on the Th17 autoimmune response from anti-inflammatory to proinflammatory. Our observation should help in the designing of better AR-targeted therapies.

Immune suppression via glucocorticoid-stimulated monocytes: a novel mechanism to cope with inflammation.

Glucocorticoids (GCs) are used as first-line therapies for generalized suppression of inflammation (e.g., allergies or autoimmune diseases), but their long-term use is limited by severe side effects. Our previous work revealed that GCs induced a stable anti-inflammatory phenotype in monocytes, the GC-stimulated monocytes (GCsMs) that we exploited for targeted GC-mediated therapeutic effects. We demonstrate that GCsMs interact with T cells in suppressing proliferation, as well as cytokine release of CD8(+) and, especially, CD4(+) T cells in vitro, and that they support generation of Foxp3(+) cells. Therefore, we tested their immunosuppressive potential in CD4(+) T cell-induced colitis in vivo. We found that injection of GCsMs into mice with severe colitis abolished the inflammation and resulted in significant clinical improvement within a few days. T cells recovered from GCsM-treated mice exhibited reduced secretion of proinflammatory cytokines IFN-γ and IL-17. Furthermore, clusters of Foxp3(+) CD4(+) T cells were detectable at local sites of inflammation in the colon. Thus, GCsMs are able to modify T cell responses in vitro and in vivo, as well as to downregulate and clinically cure severe T cell-mediated colitis.

Novel psoriasis therapies and patient outcomes, part 1: topical medications.

In recent years, advances in our understanding of inflammatory mediators and the underlying pathogenesis of psoriasis and psoriatic arthritis have shed light on potential therapeutic targets, which has led to the development of several new promising treatments. In this article, key clinical trials, mechanisms of action, patient outcomes, and relevant safety information for these novel topical medications will be evaluated. This article is the first in a 3-part series on treatments presently in the pipeline for the management of psoriasis and psoriatic arthritis including topical agents, biologic treatments, and systemic therapies in phase 2 and phase 3 clinical trials. With novel approaches to the disease process, these therapies may afford more targeted individualized treatment regimens and offer hope to patients with psoriasis and psoriatic arthritis who have reported a suboptimal therapeutic response to conventional therapies.

Intracranial injection of Gammagard, a human IVIg, modulates the inflammatory response of the brain and lowers Aß in APP/PS1 mice along a different time course than anti-Aß antibodies.

Gammagard IVIg is a therapeutic approach to treat Alzheimer's disease currently in phase 3 clinical trials. Despite the reported efficacy of the approach the mechanism of action is poorly understood. We have previously shown that intracranial injection of anti-Aß antibodies into the frontal cortex and hippocampus reveals important information regarding the time course of events once the agent is in the brain. In the current study we compared IVIg, mouse-pooled IgG, and the anti-Aß antibody 6E10 injected intracranially into the frontal cortex and hippocampus of 7-month-old APP/PS1 mice. We established a time course of events ranging from 1 to 21 d postinjection. IVIg and pooled mouse IgG both significantly reduced Aß deposition to the same degree as the 6E10 anti-Aß antibody; however, the clearance was much slower to occur, happening between the 3 and 7 d time points. In contrast, as we have previously shown, Aß reductions were apparent with the 6E10 anti-Aß group at the 1 d time point. Also, neuroinflammatory profiles were significantly altered by the antibody treatments. APP/PS1 transgenic mice at 7 months of age typically exhibit an M2a inflammatory phenotype. All antibody treatments stimulated an M2b response, yet anti-Aß antibody was a more rapid change. Because the neuroinflammatory switch occurs before the detectable reductions in amyloid deposition, we hypothesize that the IVIg and pooled mouse IgG act as immune modulators and this immune modulation is responsible for the reductions in amyloid pathology.

Metabolically induced liver inflammation leads to NASH and differs from LPS- or IL-1ß-induced chronic inflammation.

The nature of the chronic inflammatory component that drives the development of non-alcoholic steatohepatitis (NASH) is unclear and possible inflammatory triggers have not been investigated systematically. We examined the effect of non-metabolic triggers (lipopolysaccharide (LPS), interleukin-1ß (IL-1ß), administered by slow-release minipumps) and metabolic dietary triggers (carbohydrate, cholesterol) of inflammation on the progression of bland liver steatosis (BS) to NASH. Transgenic APOE3*Leiden.huCETP (APOE3L.CETP) mice fed a high-fat diet (HFD) developed BS after 10 weeks. Then, inflammatory triggers were superimposed or not (control) for six more weeks. Mouse livers were analyzed with particular emphasis on hallmarks of inflammation which were defined in human liver biopsies with and without NASH. Livers of HFD-treated control mice remained steatotic and did not progress to NASH. All four inflammatory triggers activated hepatic nuclear factor-κB (NF-κB) significantly and comparably (≥5-fold). However, HFD+LPS or HFD+IL-1ß did not induce a NASH-like phenotype and caused intrahepatic accumulation of almost exclusively mononuclear cells. By contrast, mice treated with metabolic triggers developed NASH, characterized by enhanced steatosis, hepatocellular hypertrophy, and formation of mixed-type inflammatory foci containing myeloperoxidase-positive granulocytes (neutrophils) as well as mononuclear cells, essentially as observed in human NASH. Specific for the metabolic inducers was an activation of the proinflammatory transcription factor activator protein-1 (AP-1), neutrophil infiltration, and induction of risk factors associated with human NASH, that is, dyslipidemia (by cholesterol) and insulin resistance (by carbohydrate). In conclusion, HFD feeding followed by NF-κB activation per se (LPS, IL-1ß) does not promote the transition from BS to NASH. HFD feeding followed by metabolically evoked inflammation induces additional inflammatory components (neutrophils, AP-1 pathway) and causes NASH.

D-series resolvin attenuates vascular smooth muscle cell activation and neointimal hyperplasia following vascular injury.

Recent evidence suggests that specialized lipid mediators derived from polyunsaturated fatty acids control resolution of inflammation, but little is known about resolution pathways in vascular injury. We sought to determine the actions of D-series resolvin (RvD) on vascular smooth muscle cell (VSMC) phenotype and vascular injury. Human VSMCs were treated with RvD1 and RvD2, and phenotype was assessed by proliferation, migration, monocyte adhesion, superoxide production, and gene expression assays. A rabbit model of arterial angioplasty with local delivery of RvD2 (10 nM vs. vehicle control) was employed to examine effects on vascular injury in vivo. Local generation of proresolving lipid mediators (LC-MS/MS) and expression of RvD receptors in the vessel wall were assessed. RvD1 and RvD2 produced dose-dependent inhibition of VSMC proliferation, migration, monocyte adhesion, superoxide production, and proinflammatory gene expression (IC50≈0.1-1 nM). In balloon-injured rabbit arteries, cell proliferation (51%) and leukocyte recruitment (41%) were reduced at 3 d, and neointimal hyperplasia was attenuated (29%) at 28 d by RvD2. We demonstrate endogenous biosynthesis of proresolving lipid mediators and expression of receptors for RvD1 in the artery wall. RvDs broadly reduce VSMC responses and modulate vascular injury, suggesting that local activation of resolution mechanisms expedites vascular homeostasis.

Pro-inflammatory effects of uric acid in the gastrointestinal tract.

Uric acid can be generated in the gastrointestinal (GI) tract from the breakdown of nucleotides ingested in the diet or from purines released from host cells as a result of pathogen-induced cell damage. Xanthine oxidase (XO) is the enzyme that converts hypoxanthine or xanthine into uric acid, a reaction that also generates hydrogen peroxide. It has been assumed that the product of XO responsible for the pro-inflammatory effects of this enzyme is hydrogen peroxide. Recent literature on uric acid, however, has indicated that uric acid itself may have biological effects. We tested whether uric acid itself has detectable pro-inflammatory effects using an in vivo model using ligated rabbit intestinal segments ("loops") as well as in vitro assays using cultured cells. Addition of exogenous uric acid increased the influx of heterophils into rabbit intestinal loops, as measured by myeloperoxidase activity. In addition, white blood cells adhered avidly to uric acid crystals, forming large aggregates of cells. Uric acid acts as a leukocyte chemoattractant in the GI tract. The role of uric acid in enteric infections and in non-infectious disorders of the GI tract deserves more attention.

Vasoactive intestinal peptide downregulates proinflammatory TLRs while upregulating anti-inflammatory TLRs in the infected cornea.

TLRs recognize microbial pathogens and trigger an immune response, but their regulation by neuropeptides, such as vasoactive intestinal peptide (VIP), during Pseudomonas aeruginosa corneal infection remains unexplored. Therefore, C57BL/6 (B6) mice were injected i.p. with VIP, and mRNA, protein, and immunostaining assays were performed. After VIP treatment, PCR array and real-time RT-PCR demonstrated that proinflammatory TLRs (conserved helix-loop-helix ubiquitous kinase, IRAK1, TLR1, TLR4, TLR6, TLR8, TLR9, and TNFR-associated factor 6) were downregulated, whereas anti-inflammatory TLRs (single Ig IL-1-related receptor [SIGIRR] and ST2) were upregulated. ELISA showed that VIP modestly downregulated phosphorylated inhibitor of NF-κB kinase subunit α but upregulated ST2 ~2-fold. SIGIRR was also upregulated, whereas TLR4 immunostaining was reduced in cornea; all confirmed the mRNA data. To determine whether VIP effects were cAMP dependent, mice were injected with small interfering RNA for type 7 adenylate cyclase (AC7), with or without VIP treatment. After silencing AC7, changes in mRNA levels of TLR1, TNFR-associated factor 6, and ST2 were seen and unchanged with addition of VIP, indicating that their regulation was cAMP dependent. In contrast, changes were seen in mRNA levels of conserved helix-loop-helix ubiquitous kinase, IRAK1, 2, TLR4, 9 and SIGIRR following AC7 silencing alone; these were modified by VIP addition, indicating their cAMP independence. In vitro studies assessed the effects of VIP on TLR regulation in macrophages and Langerhans cells. VIP downregulated mRNA expression of proinflammatory TLRs while upregulating anti-inflammatory TLRs in both cell types. Collectively, the data provide evidence that VIP downregulates proinflammatory TLRs and upregulates anti-inflammatory TLRs and that this regulation is both cAMP dependent and independent and involves immune cell types found in the infected cornea.

Recruitment of memory B cells to lymph nodes remote from the site of immunization requires an inflammatory stimulus.

Successful recall Ab responses require recruitment of quiescent memory B cells to secondary lymphoid organs. However, the cellular dynamics of memory cells responding to local antigenic challenge at lymphoid sites distal from the initial Ag encounter are not well understood. We show in this study that memory B cells generated following s.c. immunization in one footpad generate secondary responses to soluble Ag given i.p. but not to Ag given s.c. in the contralateral footpad unless LPS is coadministered. Memory B cells do not express CD62L, and CD62L(-ve) cells cannot enter lymph nodes unless LPS-mediated inflammation is induced there. Functional TLR4 is required on the B cells, as well as on non-B cells, in the lymph node to achieve full recruitment. Furthermore, splenectomized mice fail to respond to such inflammatory s.c. challenge in contralateral footpads, unlike lymphadenectomized mice lacking the original draining lymph nodes. Splenectomized mice also fail to respond to i.p. challenge with soluble Ag. Together, these data indicate that, unlike the central memory pool of T cells, which circulates through resting lymph nodes, the majority of long-lived memory B cells are spleen resident and require inflammatory signals for mounting recall responses at distal challenge sites.

Control of allergen-induced inflammation and hyperresponsiveness by the metalloproteinase ADAMTS-12.

A disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS) constitute a family of endopeptidases related to matrix metalloproteinases. These proteinases have been largely implicated in tissue remodeling associated with pathological processes. Among them, ADAMTS12 was identified as an asthma-associated gene in a human genome screening program. However, its functional implication in asthma is not yet documented. The present study aims at investigating potential ADAMTS-12 functions in experimental models of allergic airways disease. Two different in vivo protocols of allergen-induced airways disease were applied to the recently generated Adamts12-deficient mice and corresponding wild-type mice. In this study, we provide evidence for a protective effect of ADAMTS-12 against bronchial inflammation and hyperresponsiveness. In the absence of Adamts12, challenge with different allergens (OVA and house dust mite) led to exacerbated eosinophilic inflammation in the bronchoalveolar lavage fluid and in lung tissue, along with airway dysfunction assessed by increased airway responsiveness following methacholine exposure. Furthermore, mast cell counts and ST2 receptor and IL-33 levels were higher in the lungs of allergen-challenged Adamts12-deficient mice. The present study provides, to our knowledge, the first experimental evidence for a contribution of ADAMTS-12 as a key mediator in airways disease, interfering with immunological processes leading to inflammation and airway hyperresponsiveness.

ß-glucan curdlan induces IL-10-producing CD4+ T cells and inhibits allergic airway inflammation.

A number of studies have suggested a correlation between a decreased incidence in infectious diseases and an increased incidence of allergic diseases, including asthma. Although several pathogen-derived products have been shown to possess therapeutic potential for allergic diseases, it remains largely unknown whether ß-glucan, a cell wall component of a variety of fungi, yeasts, and bacteria, has a regulatory potential for allergic diseases. In this study, we examined the effect of curdlan, a linear ß-(1-3)-glucan, on the development of allergic airway inflammation. We found that i.p. injection of curdlan significantly inhibited Ag-induced eosinophil recruitment and Th2 cytokine production in the airways. The activation of CD4(+) T cells in the presence of curdlan induced IL-10-producing CD4(+) T cells with high levels of c-Maf expression. Curdlan-induced development of IL-10-producing CD4(+) T cells required the presence of APCs and ICOS/ICOS ligand interaction. Curdlan-induced development of IL-10-producing CD4(+) T cells also required intrinsic expression of STAT6. Furthermore, the transfer of Ag-specific CD4(+) T cells that were stimulated in the presence of curdlan inhibited Ag-induced eosinophil recruitment into the airways. Taken together, these results suggest that curdlan is capable of inducing IL-10-producing CD4(+) T cells and inhibiting the development of eosinohilic airway inflammation, underscoring the therapeutic potential of curdlan for allergic diseases.

B7-H3 participates in the development of experimental pneumococcal meningitis by augmentation of the inflammatory response via a TLR2-dependent mechanism.

In addition to a well-documented role in regulating T cell-mediated immune responses, B7-H3, a newly discovered member of the B7 superfamily, has been recently identified as a costimulator in the innate immunity-mediated inflammatory response. In this study, we further report that B7-H3 participates in the development of pneumococcal meningitis in a murine model. Exogenous administration of B7-H3 strongly amplified the inflammatory response, exacerbated blood-brain barrier disruption, and aggravated the clinical disease status in Streptococcus pneumoniae-infected C3H/HeN wild-type mice. Consistent with the in vivo findings, B7-H3 substantially augmented proinflammatory cytokine and chemokine production, upregulated NF-κB p65 and MAPK p38 phosphorylation, and enhanced the nuclear transactivation of NF-κB p65 at both TNF-α and IL-6 promoters in S. pneumoniae-stimulated primary murine microglia cells. These B7-H3-associated in vitro and in vivo effects appeared to be dependent on TLR2 signaling, as B7-H3 almost completely lost its amplifying actions in both TLR2-deficient microglial cells and TLR2-deficient mice. Furthermore, administration of the anti-B7-H3 mAb (MIH35) attenuated the inflammatory response and ameliorated blood-brain barrier disruption in S. pneumoniae-infected wild-type mice. Collectively, our results indicate that B7-H3 plays a contributory role in the development of S. pneumoniae infection-induced bacterial meningitis.

The T cell response to IL-10 alters cellular dynamics and paradoxically promotes central nervous system autoimmunity.

IL-10 is a critical anti-inflammatory cytokine, the deficiency of which leads to spontaneous autoimmunity. However, therapeutically administered or ectopically expressed IL-10 can either suppress or promote disease. Distinct lineage-specific activities may explain the contradictory effects of IL-10. To dissect the T cell-specific response to IL-10 during organ-specific autoimmunity, we generated mice with a selective deletion of IL-10Rα in T cells and analyzed its effects in an autoimmune model, experimental allergic encephalomyelitis (EAE). Surprisingly, the T cell response to IL-10 increased EAE severity. This did not result from altered T cell functional potential; T cell cytokine profile was preserved. IL-10 also diminished the proliferation of T cells in situ within the target organ, an effect that would be expected to restrain disease. However, IL-10 acted cell autonomously to sustain the autoreactive T cells essential for immunopathogenesis, promoting their accumulation and distorting the regulatory and effector T cell balance. Indeed, in chimeric mice and after adoptive transfer, wild type T cells showed a competitive advantage over cells deficient in IL-10Rα. Therefore, T cell specific actions of IL-10 can support autoimmune inflammation, and this appears to result from an overall increase in the long term fitness of pathologic T cells. Lineage-restricted, disease-promoting activities of IL-10 should be considered in the therapeutic manipulation of the IL-10 pathway.

Critical role for CCAAT/enhancer-binding protein ß in immune complex-induced acute lung injury.

C/EBPs, particularly C/EBPß and C/EBPδ, are known to participate in the regulation of many genes associated with inflammation. However, very little is known regarding the activation and functions of C/EBPß and C/EBPδ in acute lung inflammation and injury. In this study, we show that both C/EBPß and C/EBPδ activation are triggered in lungs and in alveolar macrophages following intrapulmonary deposition of IgG immune complexes. We further show that mice carrying a targeted deletion of the C/EBPß gene displayed significant attenuation of the permeability index (lung vascular leak of albumin), lung neutrophil accumulation (myeloperoxidase activity), total number of WBCs, and neutrophils in bronchoalveolar lavage fluids compared with wild-type mice. Moreover, the mutant mice expressed considerably less TNF-α, IL-6, and CXC/CC chemokine and soluble ICAM-1 proteins in bronchoalveolar lavage fluids, and corresponding mRNAs in the IgG immune complex-injured lung, compared with wild-type mice. These phenotypes were associated with a significant reduction in morphological lung injury. In contrast, C/EBPδ deficiency had no effect on IgG immune complex-induced lung injury. IgG immune complex-stimulated C/EBPß-deficient alveolar macrophages released significantly less TNF-α, IL-6, MIP-2, keratinocyte cell-derived chemokine, and MIP-1α compared with wild-type cells. Similar decreases in IgG immune complex-induced inflammatory mediator production were observed following small interfering RNA ablation of C/EBPß in a murine alveolar macrophage cell line. These findings implicate C/EBPß as a critical regulator of IgG immune complex-induced inflammatory responses and injury in the lung.

A2B adenosine receptor expression by myeloid cells is proinflammatory in murine allergic-airway inflammation.

Asthma is a chronic condition with high morbidity and healthcare costs, and cockroach allergens are an established cause of urban pediatric asthma. A better understanding of cell types involved in promoting lung inflammation could provide new targets for the treatment of chronic pulmonary disease. Because of its role in regulating myeloid cell-dependent inflammatory processes, we examined A(2B) R expression by myeloid cells in a cockroach allergen model of murine asthma-like pulmonary inflammation. Both systemic and myeloid tissue-specific A(2B) R deletion significantly decreased pulmonary inflammatory cell recruitment, airway mucin production, and proinflammatory cytokine secretion after final allergen challenge in sensitized mice. A(2B) R deficiency resulted in a dramatic reduction on Th2-type airways responses with decreased pulmonary eosinophilia without augmenting neutrophilia, and decreased lung IL-4, IL-5, and IL-13 production. Chemokine analysis demonstrated that eotaxin 1 and 2 secretion in response to repeated allergen challenge is myeloid cell A(2B) R dependent. In contrast, there were no differences in the levels of the CXC chemokines keratinocyte-derived chemokine and MIP-2 in the myeloid cell A(2B) R-deficient mice, strengthening A(2B) R involvement in the development of Th2-type airways inflammation. Proinflammatory TNF-α, IFN-γ, and IL-17 secretion were also reduced in systemic and myeloid tissue-specific A(2B) R deletion mouse lines. Our results demonstrate Th2-type predominance for A(2B) R expression by myeloid cells as a mechanism of development of asthma-like pulmonary inflammation.

Prevention of virus-induced type 1 diabetes with antibiotic therapy.

Microbes were hypothesized to play a key role in the progression of type 1 diabetes (T1D). We used the LEW1.WR1 rat model of Kilham rat virus (KRV)-induced T1D to test the hypothesis that the intestinal microbiota is involved in the mechanism leading to islet destruction. Treating LEW1.WR1 rats with KRV and a combination of trimethoprim and sulfamethoxazole (Sulfatrim) beginning on the day of infection protected the rats from insulitis and T1D. Pyrosequencing of bacterial 16S rRNA and quantitative RT-PCR indicated that KRV infection resulted in a transient increase in the abundance of Bifidobacterium spp. and Clostridium spp. in fecal samples from day 5- but not day 12-infected versus uninfected animals. Similar alterations in the gut microbiome were observed in the jejunum of infected animals on day 5. Treatment with Sulfatrim restored the level of intestinal Bifidobacterium spp. and Clostridium spp. We also observed that virus infection induced the expression of KRV transcripts and the rapid upregulation of innate immune responses in Peyer's patches and pancreatic lymph nodes. However, antibiotic therapy reduced the virus-induced inflammation as reflected by the presence of lower amounts of proinflammatory molecules in both the Peyer's patches and pancreatic lymph nodes. Finally, Sulfatrim treatment reduced the number of B cells in Peyer's patches and downmodulated adaptive immune responses to KRV, but did not interfere with antiviral Ab responses or viral clearance from the spleen, pancreatic lymph nodes, and serum. The data suggest that gut microbiota may be involved in promoting virus-induced T1D in the LEW1.WR1 rat model.

Retinoic acid attenuates rheumatoid inflammation in mice.

Retinoic acid is the active vitamin A derivative and is well-known to have diverse immunomodulatory actions. In this study, we investigated the impact of all-trans retinoic acid (ATRA), a biologic key metabolite of vitamin A, on the development of arthritis and the pathophysiologic mechanisms by which ATRA might have antiarthritic effects in animal model of rheumatoid arthritis (RA; collagen-induced arthritis [CIA] in DBA/1J mice). We showed that treatment with ATRA markedly suppressed the clinical and histologic signs of arthritis in the CIA mice. It reduced the expression of IL-17 in the arthritic joints. Interestingly, Foxp3(+) regulatory T cells were markedly increased and IL-17-producing CD4(+) T cells (Th17 cells) were decreased in the spleens of ATRA-treated mice. In vitro treatment with ATRA induced the expression of Foxp3 and repressed the IL-17 expression in the CD4(+) T cells in mice. ATRA suppressed the production of total IgG and IgG2a in splenocytes that were stimulated by LPS. It also reduced serum levels of total IgG and IgG2 anti-collagen Abs and germinal center formation in CIA mice. In addition, the ATRA-treated mice showed decreased osteoclast formation in arthritic joints. Moreover, ATRA downregulated the expression of receptor activator of NF-κB ligand, the leading player of osteoclastogenesis, in the CD4(+) T cells and fibroblast-like synoviocytes from patients with RA. Furthermore, ATRA prevented both human monocytes and mice bone marrow-derived monocytes/macrophage cells from differentiating into osteoclasts. These data suggest ATRA might be an effective treatment modality for RA patients.