Synthesis of DPIE [2-(1,2-Diphenyl-1H-indol-3-yl)ethanamine] Derivatives and Their Regulatory Effects on Pro-Inflammatory Cytokine Production in IL-1ß-Stimulated Primary Human Oral Cells.

Interleukin-1 beta (IL-1ß) has diverse physiological functions and plays important roles in health and disease. In this report, we focus on its function in the production of pro-inflammatory cytokines, including IL-6 and IL-8, which are implicated in several autoimmune diseases and host defense against infection. IL-1ß activity is markedly dependent on the binding affinity toward IL-1 receptors (IL-1Rs). Several studies have been conducted to identify suitable small molecules that can modulate the interactions between 1L-1ß and 1L-1R1. Based on our previous report, where DPIE [2-(1,2-Diphenyl-1H-indol-3-yl)ethanamine] exhibited such modulatory activity, three types of DPIE derivatives were synthesized by introducing various substituents at the 1, 2, and 3 positions of the indole group in DPIE. To predict a possible binding pose in complex with IL-1R1, a docking simulation was performed. The effect of the chemicals was determined in human gingival fibroblasts (GFs) following IL-1ß induction. The DPIE derivatives affected different aspects of cytokine production. Further, a group of the derivatives enabled synergistic pro-inflammatory cytokine production, while another group caused diminished cytokine production compared to DPIE stimulation. Some groups displayed no significant difference after stimulation. These findings indicate that the modification of the indole site could modulate IL-1ß:IL1R1 binding affinity to reduce or enhance pro-inflammatory cytokine production.

Immunomodulatory properties of characellide A on human peripheral blood mononuclear cells.

Marine sponges and their associated microbiota are multicellular animals known to produce metabolites with interesting pharmacological properties playing a pivotal role against a plethora of pathologic disorders such as inflammation, cancer and infections. Characellide A and B belong to a novel class of glycolipopeptides isolated from the deep sea marine sponge Characella pachastrelloides. In this study, we have evaluated the effects of characellide A and B on cytokine and chemokine release from human peripheral blood mononuclear cells (PBMC). Characellide A induces a concentration- and time-dependent CXCL8, IL-6 and TNF-α release from PBMC. This production is mediated by the induction of gene transcription. Moreover, cytokine/chemokine release induced by characellide A from PBMC is CD1d-dependent because a CD1d antagonist, 1,2-bis(diphenylphosphino)ethane [DPPE]-polyethylene glycolmonomethylether [PEG], specifically inhibits characellide A-induced activation of PBMC. In conclusion, characellide A is a novel modulator of adaptative/innate immune responses. Further studies are needed to understand its potential pharmacological application.

Cigarette smoke and electronic cigarettes differentially activate bronchial epithelial cells.

BACKGROUND: The use of electronic cigarettes (ECIGs) is increasing, but the impact of ECIG-vapor on cellular processes like inflammation or host defense are less understood. The aim of the present study was to compare the acute effects of traditional cigarettes (TCIGs) and ECIG-exposure on host defense, inflammation, and cellular activation of cell lines and primary differentiated human airway epithelial cells (pHBE). METHODS: We exposed pHBEs and several cell lines to TCIG-smoke or ECIG-vapor. Epithelial host defense and barrier integrity were determined. The transcriptome of airway epithelial cells was compared by gene expression array analysis. Gene interaction networks were constructed and differential gene expression over all groups analyzed. The expression of several candidate genes was validated by qRT-PCR. RESULTS: Bacterial killing, barrier integrity and the expression of antimicrobial peptides were not affected by ECIG-vapor compared to control samples. In contrast, TCIGs negatively affected host defense and reduced barrier integrity in a significant way. Furthermore ECIG-exposure significantly induced IL-8 secretion from Calu-3 cells but had no effect on NCI-H292 or primary cells. The gene expression based on array analysis distinguished TCIG-exposed cells from ECIG and room air-exposed samples. CONCLUSION: The transcriptome patterns of host defense and inflammatory genes are significantly distinct between ECIG-exposed and TCIG-treated cells. The overall effects of ECIGs on epithelial cells are less in comparison to TCIG, and ECIG-vapor does not affect host defense. Nevertheless, although acute exposure to ECIG-vapor induces inflammation, and the expression of S100 proteins, long term in vivo data is needed to evaluate the chronic effects of ECIG use.

Biased allosteric modulation of formyl peptide receptor 2 leads to distinct receptor conformational states for pro- and anti-inflammatory signaling.

BACKGROUND AND PURPOSE: Formyl peptide receptor 2 (FPR2) is a Class A G protein-coupled receptor (GPCR) that interacts with multiple ligands and transduces both proinflammatory and anti-inflammatory signals. These ligands include weak agonists and modulators that are produced during inflammation. The present study investigates how prolonged exposure to FPR2 modulators influence receptor signaling. EXPERIMENTAL APPROACH: Fluorescent biosensors of FPR2 were constructed based on single-molecule fluorescent resonance energy transfer (FRET) and used for measurement of ligand-induced receptor conformational changes. These changes were combined with FPR2-mediated signaling events and used as parameters for the conformational states of FPR2. Ternary complex models were developed to interpret ligand concentration-dependent changes in FPR2 conformational states. KEY RESULTS: Incubation with Ac2-26, an anti-inflammatory ligand of FPR2, decreased FRET intensity at picomolar concentrations. In comparison, WKYMVm (W-pep) and Aß42, both proinflammatory agonists of FPR2, increased FRET intensity. Preincubation with Ac2-26 at 10 pM diminished W-pep-induced Ca2+ flux but potentiated W-pep-stimulated ß-arrestin2 membrane translocation and p38 MAPK phosphorylation. The opposite effects were observed with 10 pM of Aß42. Neither Ac2-26 nor Aß42 competed for W-pep binding at the picomolar concentrations. CONCLUSIONS AND IMPLICATIONS: The results support the presence of two allosteric binding sites on FPR2, each for Ac2-26 and Aß42, with high and low affinities. Sequential binding of the two allosteric ligands at increasing concentrations induce different conformational changes in FPR2, providing a novel mechanism by which biased allosteric modulators alter receptor conformations and generate pro- and anti-inflammatory signals.

An impaired intrinsic microglial clock system induces neuroinflammatory alterations in the early stage of amyloid precursor protein knock-in mouse brain.

BACKGROUND: Disturbances in clock genes affect almost all patients with Alzheimer's disease (AD), as evidenced by their altered sleep/wake cycle, thermoregulation, and exacerbation of cognitive impairment. As microglia-mediated neuroinflammation proved to be a driver of AD rather than a result of the disease, in this study, we evaluated the relationship between clock gene disturbance and neuroinflammation in microglia and their contribution to the onset of AD. METHODS: In this study, the expression of clock genes and inflammatory-related genes was examined in MACS microglia isolated from 2-month-old amyloid precursor protein knock-in (APP-KI) and wild-type (WT) mice using cap analysis gene expression (CAGE) deep sequencing and RT-PCR. The effects of clock gene disturbance on neuroinflammation and relevant memory changes were examined in 2-month-old APP-KI and WT mice after injection with SR9009 (a synthetic agonist for REV-ERB). The microglia morphology was studied by staining, neuroinflammation was examined by Western blotting, and cognitive changes were examined by Y-maze and novel object recognition tests. RESULTS: CLOCK/BMAL1-driven transcriptional negative feedback loops were impaired in the microglia from 2-month-old APP-KI mice. Pro-inflammatory genes in microglia isolated from APP-KI mice were significantly higher than those isolated from WT mice at Zeitgeber time 14. The expression of pro-inflammatory genes was positively associated with NF-κB activation and negatively associated with the BMAL1 expression. SR9009 induced the activation of microglia, the increased expression of pro-inflammatory genes, and cognitive decline in 2-month-old APP-KI mice. CONCLUSION: Clock gene disturbance in microglia is involved in the early onset of AD through the induction of chronic neuroinflammation, which may be a new target for preventing or slowing AD.

Cannabidiol differentially regulates basal and LPS-induced inflammatory responses in macrophages, lung epithelial cells, and fibroblasts.

INTRODUCTION: Cannabidiol (CBD) containing products are available in a plethora of flavors in oral, sublingual, and inhalable forms. Immunotoxicological effects of CBD containing liquids were assessed by hypothesizing that CBD regulates oxidative stress and lipopolysaccharide (LPS) induced inflammatory responses in macrophages, epithelial cells, and fibroblasts. METHODS: Epithelial cells (BEAS-2B and NHBE), macrophages (U937), and lung fibroblast cells (HFL-1) were treated with varying CBD concentrations or exposed to CBD aerosols. Generated reactive oxygen species (ROS) and inflammatory mediators were measured. Furthermore, monocytes and epithelial cells were stimulated with LPS in combination with CBD or dexamethasone to understand the anti-inflammatory effects of CBD. RESULTS: CBD showed differential effects on IL-8 and MCP-1, and acellular and cellular ROS levels. CBD significantly attenuated LPS-induced NF-κB activity, IL-8, and MCP-1 release from macrophages. Cytokine array data depicted a differential cytokine response due to CBD. Inflammatory mediators, IL-8, serpin E1, CXCL1, IL-6, MIF, IFN-γ, MCP-1, RANTES, and TNF-α were induced, whereas MCP-1/CCL2, CCL5, eotaxin, and IL-2 were reduced. CBD and dexamethasone treatments reduced the IL-8 level induced by LPS when the cells were treated individually, but showed antagonistic effects when used in combination via MCPIP (monocytic chemotactic protein-induced protein). CONCLUSION: CBD differentially regulated basal pro-inflammatory response and attenuated both LPS-induced cytokine release and NF-κB activity in monocytes, similar to dexamethasone. Thus, CBD has a differential inflammatory response and acts as an anti-inflammatory agent in pro-inflammatory conditions but acts as an antagonist with steroids, overriding the anti-inflammatory potential of steroids when used in combination.

Shedding of membrane-associated LDL receptor-related protein-1 from microglia amplifies and sustains neuroinflammation.

In the CNS, microglia are activated in response to injury or infection and in neurodegenerative diseases. The endocytic and cell signaling receptor, LDL receptor-related protein-1 (LRP1), is reported to suppress innate immunity in macrophages and oppose microglial activation. The goal of this study was to identify novel mechanisms by which LRP1 may regulate microglial activation. Using primary cultures of microglia isolated from mouse brains, we demonstrated that LRP1 gene silencing increases expression of proinflammatory mediators; however, the observed response was modest. By contrast, the LRP1 ligand, receptor-associated protein (RAP), robustly activated microglia, and its activity was attenuated in LRP1-deficient cells. An important element of the mechanism by which RAP activated microglia was its ability to cause LRP1 shedding from the plasma membrane. This process eliminated cellular LRP1, which is anti-inflammatory, and generated a soluble product, shed LRP1 (sLRP1), which is potently proinflammatory. Purified sLRP1 induced expression of multiple proinflammatory cytokines and the mRNA encoding inducible nitric-oxide synthase in both LRP1-expressing and -deficient microglia. LPS also stimulated LRP1 shedding, as did the heat-shock protein and LRP1 ligand, calreticulin. Other LRP1 ligands, including α2-macroglobulin and tissue-type plasminogen activator, failed to cause LRP1 shedding. Treatment of microglia with a metalloproteinase inhibitor inhibited LRP1 shedding and significantly attenuated RAP-induced cytokine expression. RAP and sLRP1 both caused neuroinflammation in vivo when administered by stereotaxic injection into mouse spinal cords. Collectively, these results suggest that LRP1 shedding from microglia may amplify and sustain neuroinflammation in response to proinflammatory stimuli.

Preventive effect of apigenin against isoproterenol-induced apoptosis in cardiomyoblasts.

We investigated the effect of apigenin, a dietary flavonoid, on isoproterenol hydrochloride (ISO)-induced apoptotic signaling in cardiomyoblast H9C2 cells. The results showed that apigenin treatment (10 µM) prevented ISO (31.25 µM)-induced lipid peroxidative levels and antioxidants status in H9C2 cells. Furthermore, apigenin inhibited expression of inflammatory markers in ISO-treated cells. In addition, apigenin prevented ISO-induced DNA damage and apoptotic signaling through modulating the expression of Bax, caspase-3, -8 and -9, cytochrome c, and Fas proteins in H9C2 cells. It is concluded that apigenin prevents ISO-induced antioxidants depletion, oxidative DNA damage, inflammatory, and apoptotic signaling in H9C2 cells. Thus, the present results demonstrated that apigenin has a cardioprotective effect on cardiomyoblasts cells.

The Effect of Prim-O-Glucosylcimifugin on Tryptase-Induced Intestinal Barrier Dysfunction in Caco-2 Cells.

The intestinal barrier dysfunction is a critical pathological change in irritable bowel syndrome (IBS). The objective of this study was to evaluate the effect of Prim-O-glucosylcimifugin (POG) on intestinal barrier dysfunction and reveal possible molecular mechanisms. Human colon adenocarcinoma cell line (Caco-2) cell monolayers induced by tryptase (TRYP) were used to establish an intestinal barrier dysfunction model. Caco-2 cell monolayers from both functional and dysfunctional samples were treated with POG (30, 60 and 120 µg/mL) for 2, 8, 24, 36, 48 and 72 h. The Caco-2 cell monolayers were assessed by measurement of trans-epithelial electrical resistance (TEER) and percentage of fluorescein permeation (PFP). The expression of Protease Activated Receptor 2 (PAR-2) and myosin light chain kinase (MLCK) mRNA was analyzed by RT-PCR and the level of Zonula Occludens-1 (ZO-1) protein expression was determined by Western blot. In addition, the impact of POG on the distribution of the tight juction protein of Occludin was performed by immunofluorescence. Our results showed that POG elevated the TEER and decreased the PFP of the functional Caco-2 cell monolayers, as well as the dysfunctional Caco-2 cell monolayers. Furthermore, POG inhibited the expression of PAR-2 mRNA and MLCK mRNA and increased the level of ZO-1 protein expression in dysfunctional Caco-2 cells. The distribution of the Occludin proteins was ameliorated simultaneously. This study demonstrates that POG can enhance the intestinal barrier function of Caco-2 cell monolayers by inhibiting the expression of PAR-2 and MLCK and up-regulating the expression of ZO-1 protein, and ameliorated the distribution of Occludin protein.

Lnc-ing inflammation to disease.

Termed 'master gene regulators' long ncRNAs (lncRNAs) have emerged as the true vanguard of the 'noncoding revolution'. Functioning at a molecular level, in most if not all cellular processes, lncRNAs exert their effects systemically. Thus, it is not surprising that lncRNAs have emerged as important players in human pathophysiology. As our body's first line of defense upon infection or injury, inflammation has been implicated in the etiology of several human diseases. At the center of the acute inflammatory response, as well as several pathologies, is the pleiotropic transcription factor NF-κß. In this review, we attempt to capture a summary of lncRNAs directly involved in regulating innate immunity at various arms of the NF-κß pathway that have also been validated in human disease. We also highlight the fundamental concepts required as lncRNAs enter a new era of diagnostic and therapeutic significance.

Effects of oral exposure to naturally-occurring and synthetic deoxynivalenol congeners on proinflammatory cytokine and chemokine mRNA expression in the mouse.

The foodborne mycotoxin deoxynivalenol (DON) induces a ribotoxic stress response in mononuclear phagocytes that mediate aberrant multi-organ upregulation of TNF-α, interleukins and chemokines in experimental animals. While other DON congeners also exist as food contaminants or pharmacologically-active derivatives, it is not known how these compounds affect expression of these cytokine genes in vivo. To address this gap, we compared in mice the acute effects of oral DON exposure to that of seven relevant congeners on splenic expression of representative cytokine mRNAs after 2 and 6h. Congeners included the 8-ketotrichothecenes 3-acetyldeoxynivalenol (3-ADON), 15-acetyldeoxynivalenol (15-ADON), fusarenon X (FX), nivalenol (NIV), the plant metabolite DON-3-glucoside (D3G) and two synthetic DON derivatives with novel satiety-inducing properties (EN139528 and EN139544). DON markedly induced transient upregulation of TNF-α IL-1ß, IL-6, CXCL-2, CCL-2 and CCL-7 mRNA expressions. The two ADONs also evoked mRNA expression of these genes but to a relatively lesser extent. FX induced more persistent responses than the other DON congeners and, compared to DON, was: 1) more potent in inducing IL-1ß mRNA, 2) approximately equipotent in the induction of TNF-α and CCL-2 mRNAs, and 3) less potent at upregulating IL-6, CXCL-2, and CCL-2 mRNAs. EN139528's effects were similar to NIV, the least potent 8-ketotrichothecene, while D3G and EN139544 were largely incapable of eliciting cytokine or chemokine mRNA responses. Taken together, the results presented herein provide important new insights into the potential of naturally-occurring and synthetic DON congeners to elicit aberrant mRNA upregulation of cytokines associated with acute and chronic trichothecene toxicity.

Nicotine impairs cyclooxygenase-2-dependent kinin-receptor-mediated murine airway relaxations.

INTRODUCTION: Cigarette smoke induces local inflammation and airway hyperreactivity. In asthmatics, it worsens the symptoms and increases the risk for exacerbation. The present study investigates the effects of nicotine on airway relaxations in isolated murine tracheal segments. METHODS: Segments were cultured for 24h in the presence of vehicle, nicotine (10 µM) and/or dexamethasone (1 µM). Airway relaxations were assessed in myographs after pre-contraction with carbachol (1 µM). Kinin receptors, cyclooxygenase (COX) and inflammatory mediator expressions were assessed by real-time PCR and confocal-microscopy-based immunohistochemistry. RESULTS: The organ culture procedure markedly increased bradykinin- (selective B2 receptor agonist) and des-Arg9-bradykinin- (selective B1 receptor agonist) induced relaxations, and slightly increased relaxation induced by isoprenaline, but not that induced by PGE2. The kinin receptor mediated relaxations were epithelium-, COX-2- and EP2-receptor-dependent and accompanied by drastically enhanced mRNA levels of kinin receptors, as well as inflammatory mediators MCP-1 and iNOS. Increase in COX-2 and mPGES-1 was verified both at mRNA and protein levels. Nicotine selectively suppressed the organ-culture-enhanced relaxations induced by des-Arg9-bradykinin and bradykinin, at the same time reducing mPGES-1 mRNA and protein expressions. α7-nicotinic acetylcholine receptor inhibitors α-bungarotoxin and MG624 both blocked the nicotine effects on kinin B2 receptors, but not those on B1. Dexamethasone completely abolished kinin-induced relaxations. CONCLUSION: It is tempting to conclude that a local inflammatory process per se could have a bronchoprotective component by increasing COX-2 mediated airway relaxations and that nicotine could impede this safety mechanism. Dexamethasone further reduced airway inflammation together with relaxations. This might contribute to the steroid resistance seen in some patients with asthma.

Neuropeptide receptors as potential drug targets in the treatment of inflammatory conditions.

Cross-talk between the nervous, endocrine and immune systems exists via regulator molecules, such as neuropeptides, hormones and cytokines. A number of neuropeptides have been implicated in the genesis of inflammation, such as tachykinins and calcitonin gene-related peptide. Development of their receptor antagonists could be a promising approach to anti-inflammatory pharmacotherapy. Anti-inflammatory neuropeptides, such as vasoactive intestinal peptide, pituitary adenylate cyclase-activating polypeptide, α-melanocyte-stimulating hormone, urocortin, adrenomedullin, somatostatin, cortistatin, ghrelin, galanin and opioid peptides, are also released and act on their own receptors on the neurons as well as on different inflammatory and immune cells. The aim of the present review is to summarize the most prominent data of preclinical animal studies concerning the main pharmacological effects of ligands acting on the neuropeptide receptors. Promising therapeutic impacts of these compounds as potential candidates for the development of novel types of anti-inflammatory drugs are also discussed.

Pulmonary and hemostatic toxicity of multi-walled carbon nanotubes and zinc oxide nanoparticles after pulmonary exposure in Bmal1 knockout mice.

BACKGROUND: Pulmonary exposure to nanoparticles (NPs) may affect, in addition to pulmonary toxicity, the cardiovascular system such as procoagulant effects, vascular dysfunction and progression of atherosclerosis. However, only few studies have investigated hemostatic effects after pulmonary exposure. METHODS: We used Bmal1 (brain and muscle ARNT-like protein-1) knockout (Bmal1(-/-)) mice which have a disturbed circadian rhythm and procoagulant phenotype, to study the pulmonary and hemostatic toxicity of multi-walled carbon nanotubes (MWCNTs) and zinc oxide (ZnO) NPs after subacute pulmonary exposure. Bmal1(-/-) and wild-type (Bmal1(+/+)) mice were exposed via oropharyngeal aspiration, once a week, during 5 consecutive weeks, to a cumulative dose of 32 or 128 µg MWCNTs or 32 or 64 µg ZnO NPs. RESULTS: MWCNTs caused a pronounced inflammatory response in the lung with increased cell counts in the broncho-alveolar lavage and increased secretion of interleukin-1ß and cytokine-induced neutrophil chemo-attractant (KC), oxidative stress (increased ratio of oxidized versus reduced glutathione and decreased total glutathione) as well as anemic and procoagulant effects as evidenced by a decreased prothrombin time with increased fibrinogen concentrations and coagulation factor (F)VII. In contrast, the ZnO NPs seemed to suppress the inflammatory (decreased neutrophils in Bmal1(-/-) mice) and oxidative response (increased total glutathione in Bmal1(-/-) mice), but were also procoagulant with a significant increase of FVIII. The procoagulant effects, as well as the significant correlations between the pulmonary endpoints (inflammation and oxidative stress) and hemostasis parameters were more pronounced in Bmal1(-/-) mice than in Bmal1(+/+) mice. CONCLUSIONS: The Bmal1(-/-) mouse is a sensitive animal model to study the procoagulant effects of engineered NPs. The MWCNTs and ZnO NPs showed different pulmonary toxicity but both NPs induced procoagulant effects, suggesting different mechanisms of affecting hemostasis. However, the correlation analysis suggests a causal association between the observed pulmonary and procoagulant effects.

Extracellular ATP may contribute to tissue repair by rapidly stimulating purinergic receptor X7-dependent vascular endothelial growth factor release from primary human monocytes.

Extracellular ATP has been proposed to act as a danger signal to alert the immune system of cell damage. Release of high local concentrations of ATP activates the nucleotide receptor, purinergic receptor X7 (P2RX7), on monocytic cells, which promotes the processing/release of proinflammatory mediators. Although the proinflammatory actions of P2RX7 are well recognized, little is known regarding the potential function of P2RX7 in repair responses. Because the resolution of inflammation is characterized by monocytic cell-dependent production of proangiogenic factors, we evaluated the contribution of P2RX7 to this process. We observed that both short-term and long-term P2RX7 activation promotes the robust release of vascular endothelial growth factor from primary human monocytes. This vascular endothelial growth factor release is calcium dependent and associated with reactive oxygen species production. This previously unrecognized action of P2RX7 suggests that it may not only participate in inflammation and cell death, but that it is also likely to be important in the control of angiogenesis and wound repair.

Synergistic activation of NF-{kappa}B by bacterial chemoattractant and TNF{alpha} is mediated by p38 MAPK-dependent RelA acetylation.

In the host immune system, leukocytes are often exposed to multiple inflammation inducers. NF-κB is of considerable importance in leukocyte function because of its ability to activate the transcription of many proinflammatory immediate-early genes. Tremendous efforts have been made toward understanding how NF-κB is activated by various inducers. However, most research on NF-κB regulation has been focused on understanding how NF-κB is activated by a single inducer. This is unlike the situation in the human immune system where multiple inflammation inducers, including both exogenous and endogenous mediators, are present concurrently. We now present evidence that the formylated peptide f-Met-Leu-Phe (fMLP), a bacterial chemoattractant, synergizes with TNFα to induce NF-κB activation and the resultant inflammatory response in vitro and in vivo. The mechanism of synergistic activation of NF-κB by bacterial fMLP and TNFα may be involved in the induction of RelA acetylation, which is regulated by p38 MAPK. Thus, this study provides direct evidence for the synergistic induction of NF-κB-dependent inflammatory responses by both exogenous and endogenous inducers. The ability of fMLP to synergize with TNFα and activate NF-κB represents a novel and potentially important mechanism through which bacterial fMLP not only attracts leukocytes but also directly contributes to inflammation by synergizing with the endogenous mediator TNFα.

A TLR2 agonist in German cockroach frass activates MMP-9 release and is protective against allergic inflammation in mice.

The role of TLR2 in modulating experimentally induced asthma is not fully understood. We recently identified that German cockroach (GC) frass contains a TLR2 ligand allowing us to investigate the role of a TLR2 agonist in a complex real world allergen in mediating allergic airway inflammation. GC frass exposure significantly increased airway inflammation, airway hyperresponsiveness and serum IgE levels in wild-type mice; however the same exposure in TLR2-deficient mice resulted in greatly exaggerated serum IgE and eosinophilia but diminished airway neutrophilia, suggesting a protective role for TLR2. Since GC frass inhalation usually induces airway neutrophilia, we queried the effect of neutrophil depletion on airway responses. Inhibition of neutrophil recruitment into the airways of naive wild-type mice before intratracheal inhalation of GC frass resulted in significantly increased levels of serum IgE and eosinophilia. Neutrophils are a rich source of MMP-9, and we found that MMP-9 levels were significantly increased in the airways of mice following exposure to GC frass. Importantly the levels of MMP-9 were significantly decreased in neutrophil-depleted and TLR2-deficient mice after exposure to GC frass, suggesting that TLR2 regulated MMP-9 release from neutrophils. Functionally, MMP-9-deficient mice had more acute allergic inflammation than wild-type mice, suggesting that MMP-9 was protective against experimentally induced asthma. These data suggest that TLR2 activation of neutrophils leads to release of MMP-9 which decreases allergic responses to GC frass. This suggests a protective role for TLR2 activation and MMP-9 release in the context of experimentally induced asthma in mice.

The peroxisome proliferator-activated receptor gamma agonist rosiglitazone ameliorates murine lupus by induction of adiponectin.

Systemic lupus erythematosus (SLE) is an inflammatory autoimmune disease for which current therapy is suboptimal. SLE is characterized by autoantibody production, with renal disease and premature atherosclerosis being common and severe manifestations causing appreciable morbidity and mortality. Peroxisome proliferator-activated receptor gamma (PPARgamma) agonists are widely used in the treatment of diabetes mellitus for their insulin-sensitizing properties, but also have immunomodulatory effects. In this report, we show that the PPARgamma agonist rosiglitazone reduces autoantibody production, renal disease, and atherosclerosis in mouse models of SLE. The beneficial effect of rosiglitazone on SLE manifestations depends on the induction of adiponectin, because rosiglitazone has no effect on autoantibody production or renal disease in lupus mice that lack adiponectin. In addition, lupus mice that lack adiponectin develop more severe disease than adiponectin-sufficient lupus mice, indicating that endogenous adiponectin is involved in regulating disease activity. Furthermore, administration of exogenous adiponectin ameliorates disease. These experiments suggest that PPARgamma agonists may be useful agents for the treatment of SLE. They also demonstrate that induction of adiponectin is a major mechanism underlying the immunomodulatory effects of PPARgamma agonists.

The peroxisome proliferator-activated receptor gamma agonist pioglitazone improves cardiometabolic risk and renal inflammation in murine lupus.

Individuals with systemic lupus erythematosus (SLE) have a striking increase in the risk of premature atherosclerosis, a complication preceded by significant subclinical vascular damage. A proposed mechanism leading to accelerated vascular disease in SLE is an imbalance between vascular damage and repair, as patients with this disease display significant abnormalities in phenotype and function of endothelial progenitor cells. In addition, individuals with SLE have a higher incidence of insulin resistance which may further contribute to the increased cardiovascular risk. This study examined the role of the peroxisome proliferator activated receptor gamma agonist pioglitazone in improving endothelial function, endothelial progenitor cell numbers and functional capacity, metabolic parameters, and disease activity in the lupus-prone murine model New Zealand Black/New Zealand White (NZB x NZW)F(1). Ten-week-old prenephritic female NZB/NZW F(1) mice were exposed to 10 or 25 mg/kg/day of oral pioglitazone or vehicle for 15 or 24 wk. Mice exposed to pioglitazone exhibited pronounced enhancement in endothelial-dependent vasorelaxation of thoracic aortas and in endothelial progenitor cell function, as assessed by the capacity of bone marrow-derived endothelial progenitor cells to differentiate into mature endothelial cells. Pioglitazone-treated mice showed improvement in insulin resistance, adipokine, and lipid profile. Kidneys from pioglitazone-treated mice showed significant decreases in immune complex deposition, renal inflammation, T cell glomerular infiltration, and intrarenal synthesis of TNF-alpha, IL-1beta, and VCAM-1. These results indicate that peroxisome proliferator-activated receptor gamma agonists could serve as important tools in the prevention of premature cardiovascular disease and organ damage in SLE.

Peroxisome proliferator-activated receptor gamma agonist down-regulates IL-17 expression in a murine model of allergic airway inflammation.

Peroxisome proliferator-activated receptor gamma (PPARgamma) plays a critical role in the control of airway inflammation. Recently, IL-17 has been found to be implicated in many immune and inflammatory responses, including airway inflammation. However, no data are available concerning the effect of PPARgamma on IL-17 production in airway inflammatory diseases. In this study, we used a mouse model of asthma to evaluate the effect of two PPARgamma agonists, rosiglitazone or pioglitazone, on IL-17 expression in allergic airway disease. After OVA inhalation, mice developed the typical pathophysiological features of asthma, and the expression of IL-17 protein and mRNA in the lungs was increased. Administration of rosiglitazone or pioglitazone reduced the pathophysiological features of asthma and decreased the increased IL-17 protein and mRNA expression after OVA inhalation. In addition, the attenuating effect of PPARgamma agonist on allergic airway inflammation and bronchial hyperresponsiveness is abrogated by coadministration of rIL-17. This study also showed that the inhibition of IL-17 activity with anti-IL-17 Ab remarkably reduced the increased numbers of inflammatory cells of the airways, airway hyperresponsiveness, and the increased levels of IL-4, IL-5, and IL-13 in bronchoalveolar lavage fluid and OVA-specific IgE in serum. In addition, we found that administration of rosiglitazone or pioglitazone decreased the increased NF-kappaB activity and that a NF-kappaB inhibitor, BAY 11-7085, substantially reduced the increased IL-17 protein levels in the lung tissues after OVA inhalation. These findings suggest that the therapeutic effect of PPARgamma in asthma is partly mediated by regulation of IL-17 expression via NF-kappaB pathway.