Crystal-neutrophil interactions lead to interleukin-1 synthesis.

Normal human blood neutrophils were studied for their capacity to synthesize and release interleukin-1 (IL-1) species after phagocytosis of triclinic monosodium urate (MSU) and calcium pyrophosphate dihydrate crystals (CPPD). MSU crystals were more potent inducers of IL-1 generation than CPPD or unopsonized zymosan. Microcrystal-stimulated neutrophils characteristically secreted most of the newly synthesized IL-1. Colchicine partly inhibited the secretion of IL-1 by neutrophils during phagocytosis of solid particles. However, colchicine selectively inhibited IL-1 synthesis induced by microcrystals. These results suggest that neutrophil-derived IL-1 may contribute to the pathogenesis of crystal-induced arthritis.     

Resveratrol attenuates the MSU crystal-induced inflammatory response through the inhibition of TAK1 activity

TAK1 is closely associated with the NF-κB and MAPK signaling pathways. In the present study, we aimed to explore the relationship between TAK1 and gout as well as the effects of resveratrol on TAK1 activity and MSU crystal-induced inflammation. The expression levels of total TAK1 and phosphorylated TAK1 in gout patients were detected by western blotting. The influence of resveratrol on TAK1 activity and MSU crystal-induced inflammation was investigated in THP-1 cell and murine models of gout. The results showed that TAK1 and p-TAK1 were highly expressed in gouty arthritis patients. MSU crystals accelerated the expression of TAK1 and p-TAK1 in human PBMCs. The anti-inflammatory effects of resveratrol on MSU crystal-induced inflammation in vitro and in vivo included the alleviation of pro-inflammatory cytokines, inflammatory cell recruitment and foot swelling. Resveratrol limited the activation of TAK1 and its downstream signaling pathway, including the degradation of IκBα, the activation of NF-κB P65 and the phosphorylation of P38 and JNK. In conclusion, resveratrol may have a potential therapeutic effect on gouty arthritis by inhibiting TAK1 activity.     

Bee Venom Alleviated Edema and Pain in Monosodium Urate Crystals-Induced Gouty Arthritis in Rat by Inhibiting Inflammation

Bee venom (BV) acupuncture has anti-inflammatory and analgesic effects; therefore, it was used as a traditional Korean medicine for various musculoskeletal disorders, especially arthritis. In this study, we investigated the effect of BV on monosodium urate (MSU) crystal-induced acute gouty rats. An intra-articular injection of MSU crystal suspension (1.25 mg/site) was administered to the tibiotarsal joint of the hind paw of Sprague Dawley rats to induce MSU crystal-induced gouty arthritis. Colchicine (30 mg/kg) was orally administered 1 h before MSU crystal injection as a positive control, and BV (0.5 mg/kg) was injected into the tibiotarsal joint immediately after MSU crystal injection. The ankle thickness, mechanical allodynia, and expression of proinflammatory cytokines (TNF-α, IL-1β, IL6, COX2 and iNOS) and chemokines (MIP-1α, MIP-1β, MCP-1, GRO-α, MIP-2α) were then evaluated. BV reduced the expression of proinflammatory cytokines and chemokines, which are important mediators of MSU crystal-induced inflammatory responses. This anti-inflammatory effect was also confirmed histologically to attenuate synovitis and neutrophil infiltration. We demonstrated that BV markedly ameliorated ankle edema and mechanical allodynia in gouty rats. These results suggest that BV acupuncture is a potential clinical therapy for acute gouty management.     

Effect of pranoprofen on sodium urate crystal-induced inflammation

Effect of a non-steroidal anti-inflammatory drug, pranoprofen (PPF), on sodium urate crystal-induced inflammation was investigated in comparison with standard drugs for treating acute gout in experimental animals. PPF inhibited sodium urate crystal-induced paw edema in both rats (1-10 mg/kg, p.o.) and mice (5-25 mg/kg, p.o.) in a dose-dependent manner. On rat sodium urate crystal-induced paw edema, PPF was found to be almost equally active as indomethacin (IM) and colchicine. In addition, PPF (2.5-10 mg/kg, p.o.) inhibited the accumulation of exudate and decreased the leucocyte numbers and the amount of prostaglandin E2 (PGE2)-like substance in sodium urate crystal-induced pleurisy in rats dose-dependently, with a potency slightly greater than that of IM. The specific anti-gout agent colchicine (5 mg/kg, p.o.) also suppressed the accumulation of exudate and decreased the leucocyte numbers, without affecting the amount of PGE2-like substance. Moreover, in mouse peritonitis, PPF (1-10 mg/kg, p.o.) suppressed the sodium urate crystal-induced increase in vascular permeability in a dose-dependent manner. Furthermore, in experimental models of articular gout, PPF inhibited the pain response (abnormal gait) of sodium urate crystal-induced arthritis in both rats (0.25 and 1 mg/kg, p.o.) and dogs (3 mg/kg, p.o.), with a potency greater than that of IM and phenylbutazone, respectively. These results indicate that as an anti-gout agent, PPF is at least as effective as other standard drugs, so that it should have good clinical potential.     

Sites of action of IL-1 in the development of fever and cytokine responses to tissue inflammation in the rat

1. The objective of the present study was to determine the sites of action of the cytokine, interleukin-1 (IL-1), in the febrile response to local inflammation in the rat, by comparing the importance of IL-1 in the local tissues, the circulation and the brain. This was achieved by injecting lipopolysaccharide (LPS, 100 μg kg⁻¹) into a subcutaneous air pouch and testing the effects of blocking IL-1 action with the human recombinant interleukin-1 receptor antagonist (IL-1ra) injected either into the air pouch, intraperitoneally (1 mg kg⁻¹, 0+1 h, i.p.), or intracerebroventricularly (200 μg/rat, 0+1 h, i.c.v.).
2. To investigate the effect of IL-1ra on fever and the induction of local and circulating cytokines (IL-1 and IL-6), separate experiments were performed in which groups of animals were killed 1.5, 3 or 5 h after LPS injection. Plasma and pouch fluid samples were collected for bioassay of IL-1 and IL-6.
3. Injection of LPS into the air pouch significantly increased (1.5°C) body temperature, local (air pouch) concentrations of bioactive IL-1 and IL-6, and circulating bioactive IL-6, compared to saline-treated controls.
4. Injection of IL-1ra into the pouch significantly attenuated LPS fever (P<0.001). This decrease in body temperature was associated with significant inhibition of local IL-1 bioactivity 1.5 (96%), 3 (84%) and 5 h (72%), and in bioactive IL-6 in pouch lavage fluid 1.5 (45%) and 5 h (35%), after LPS injection. The concentration of bioactive IL-6 in the plasma was significantly reduced (39%) at 3 h, when temperature was approaching the maximal value.
5. Both systemic (i.p.) and central (i.c.v.) administration of IL-1ra significantly attentuated LPS fever (P<0.05). However, it had no effect on either local concentrations of bioactive IL-1 or IL-6, or circulating IL-6, at any of the sample points.
6. These data suggest that IL-1 is released locally, at the site of tissue inflammation and that it is an important mediator of the febrile response to local inflammation. The results also indicate that IL-1 produced locally may contribute to the production of IL-6 which is released into the circulation, and that IL-1 has important actions in the generation of fever at other sites, including the brain.

     

Inhibition of Inducible Nitric Oxide Synthase Attenuates Monosodium Urate-induced Inflammation in Mice

The present study elucidated the effect of the selective inducible nitric oxide synthase (iNOS) inhibitor N(6)-(1-iminoethyl)-L-lysine (L-NIL) on monosodium urate (MSU) crystal-induced inflammation and edema in mice feet. L-NIL (5 or 10 mg/kg/day) was administered intraperitoneally 4 h before injection of MSU (4 mg) into the soles of mice hindlimb feet. Twenty-four hours after MSU injection, foot thickness was increased by 160% and L-NIL pretreatment reduced food pad swelling in a dose dependent manner. Pretreatment of 10 mg/kg/day L-NIL significantly suppressed the foot pad swelling by MSU. Plasma level of nitric oxide (NO) metabolites and gene expression and protein level of iNOS in feet were increased by MSU, which was suppressed by L-NIL pretreatment. Similar pattern of change was observed in nitrotyrosine level. MSU increased the gene expression of tumor necrosis factor (TNF)-α and interleukin (IL)-1β and L-NIL pretreatment suppressed MSU-induced cytokines expression. The mRNA levels of superoxide dismutase and glutathione peroxidase1 were increased by MSU and L-NIL pretreatment normalized the gene expression. Phosphorylation of extracellular signal-regulated kinase 1/2 and p38 was increased by MSU, which was suppressed by L-NIL pretreatment. The mRNA levels of iNOS, TNF-α, and IL-1β were increased by MSU in human dermal fibroblasts, C2C12 myoblasts, and human fetal osteoblasts in vitro, which was attenuated by L-NIL in a dose dependent manner. This study shows that L-NIL inhibits MSU-induced inflammation and edema in mice feet suggesting that iNOS might be involved in MSU-induced inflammation.     

Quercetin inhibits gout arthritis in mice: induction of an opioid-dependent regulation of inflammasome

We investigated the anti-inflammatory and analgesic effects of quercetin in monosodium urate crystals (MSU)-induced gout arthritis, and the sensitivity of quercetin effects to naloxone, an opioid receptor antagonist. Mice were treated with quercetin, and mechanical hyperalgesia was assessed at 1–24 h after MSU injection. In vivo, leukocyte recruitment, cytokine levels, oxidative stress, NFκB activation, and gp91^phox and inflammasome components (NLRP3, ASC, Pro-caspase-1, and Pro-IL-1β) mRNA expression by qPCR were determined in the knee joints at 24 h after MSU injection. Inflammasome activation was determined, in vitro, in lipopolysaccharide-primed macrophages challenged with MSU. Quercetin inhibited MSU-induced mechanical hyperalgesia, leukocyte recruitment, TNFα and IL-1β production, superoxide anion production, inflammasome activation, decrease of antioxidants levels, NFκB activation, and inflammasome components mRNA expression. Naloxone pre-treatment prevented all the inhibitory effects of quercetin over MSU-induced gout arthritis. These results demonstrate that quercetin exerts analgesic and anti-inflammatory effect in the MSU-induced arthritis in a naloxone-sensitive manner.     

MicroRNA and long noncoding RNA involvement in gout and prospects for treatment

MicroRNAs (miRNAs) and long noncoding RNAs (lncRNAs) are both types of noncoding RNA. They have been demonstrated to be involved in the regulation of various human inflammatory diseases and can be used as biomarkers for disease diagnosis and prognosis, and even be developed into new drugs. Gout is an arthritic disease caused by the deposition of monosodium urate crystal (MSU) in the joints, which can lead to acute inflammation and damage adjacent tissue. Recent studies have shown that miRNAs and lncRNAs mediate the progress of gout. Based on the pathogenesis of gout, including hyperuricemia, MSU deposition, acute gouty arthritis and gouty bone erosion, this paper reviewed the role of miRNAs and lncRNAs in the processes and the possible therapeutic targets of miRNAs and lncRNAs in gout.     

Anti-inflammatory Activity of Methylene Chloride Fraction From Glehnia littoralis Extract via Suppression of NF-κB and Mitogen-Activated Protein Kinase Activity

Glehnia littoralis (Umbelliferae) has been used traditionally in Korean, Japanese, and Chinese medicine for the treatment of immune-related diseases; however, its anti-inflammatory activity and underlying mechanism remain to be defined. We investigated the anti-inflammatory effect and inhibitory mechanism on inflammation by the methylene chloride fraction from Glehnia littoralis extract (MCF-GLE), which was more effective than Glehnia littoralis extract (GLE). MCF-GLE inhibited 12-O-Tetradecanoyl-phorbol-13-acetate (TPA)–induced inflammation in an inflammatory edema mouse model. Also, MCF-GLE strongly inhibited the releases of nitric oxide (NO), prostaglandin E₂ (PGE₂), tumor necrosis factor-α (TNF-α), and interleukin-1β (IL-1β) and significantly suppressed the mRNA and protein expression of inducible nitric oxide synthase and cyclooxygenase-2 in lipopolysaccharide-stimulated RAW 264.7 macrophage cells in a dose-dependent manner. Furthermore, MCF-GLE suppressed NF-κB activation and IκB-α degradation. MCF-GLE also attenuated the activation of ERK and JNK in a dose-dependent manner. These results indicate that MCF-GLE has an inhibitory effect on the in vivo and in vitro inflammatory reaction and is a possible therapeutic agent. Our results suggest that the anti-inflammatory properties of MCF-GLE may result from the inhibition of pro-inflammatory mediators, such as NO, PGE₂, TNF-α, and IL-1β via suppression of NF-κB– and mitogen-activated protein kinases-dependent pathways.     

Dendritic cells treated with a prostaglandin I2 analog,iloprost, promote antigen-specific regulatory T cell differentiation in mice

Iloprost, a stable prostaglandin I₂ (PGI₂) analog, can inhibit allergic inflammation in an ovalbumin (OVA)-induced asthma model via inhibition of airway dendritic cell (DC) function. However, the underlying mechanism of PGI₂ signaling-mediated immunosuppression remains unclear. This study explored whether iloprost-treated DCs can suppress inflammation by promoting antigen-specific regulatory T cell (Treg) differentiation through PGI₂-G-protein-coupled receptor (IP). We established an allergic lung inflammation model using a hydrogel biomaterial delivery system and observed that iloprost significantly suppressed OVA-induced Th2 lung inflammation and increased the frequency of OVA-specific Tregs in vivo. We further observed that iloprost-treated DCs displayed tolerogenic characteristics, including low inflammatory cytokine (IL-12, TNF-α, IL-6, IL-23) expression levels, high anti-inflammatory cytokine (IL-10) production, and a semimature phenotype. In addition, iloprost-treated DCs increased OVA-specific CD4⁺Foxp3⁺ T cell differentiation from naïve T cells in an IP-dependent pathway in vitro and in vivo. Blocking experiments showed that iloprost-treated DCs promoted Treg differentiation, at least in part, through programmed death ligand 1 (PD-L1), whereas iloprost-induced PD-L1 expression in DCs was through the IP receptor. Furthermore, iloprost treatment suppressed DC-mediated airway inflammation and increased the frequency of OVA-specific Tregs through PD-L1 in vivo. Taken together, these results show that PGI₂-IP signaling mediated by iloprost in DCs may lead to immune tolerance, suggesting that the PGI₂ analog has the potential to be applied therapeutically for tolerogenic DC immunotherapy in autoimmune diseases or allergic asthma.     

Hydrogen sulfide inhibits cigarette smoke-induced inflammation and injury in alveolar epithelial cells by suppressing PHD2/HIF-1α/MAPK signaling pathway

Chronic obstructive pulmonary fibrosis (COPD) is a chronic and fatal lung disease with few treatment options. Sodium hydrosulfide (NaHS), a donor of hydrogen sulfide (H₂S), was found to alleviate cigarette smoke (CS)-induced emphysema in mice, however, the underlying mechanisms have not yet been clarified. In this study, we investigated its effects on COPD in a CS-induced mouse model in vivo and in cigarette smoke extract (CSE)-stimulated alveolar epithelial A549 cells in vitro. The results showed that NaHS not only relieved emphysema, but also improved pulmonary function in CS-exposed mice. NaHS significantly increased the expressions of tight junction proteins (i.e., ZO-1, Occludin and claudin-1), and reduced apoptosis and secretion of pro-inflammatory cytokines (i.e., TNF-α, IL-6 and IL-1β) in CS-exposed mouse lungs and CSE-incubated A549 cells, indicating H₂S inhibits CS-induced inflammation, injury and apoptosis in alveolar epithelial cells. NaHS also upregulated prolyl hydroxylase (PHD)2, and suppressed hypoxia-inducible factor (HIF)-1α expression in vivo and in vitro, suggesting H₂S inhibits CS-induced activation of PHD2/HIF-1α axis. Moreover, NaHS inhibited CS-induced phosphorylation of ERK, JNK and p38 MAPK in vivo and in vitro, and treatment with their inhibitors reversed CSE-induced ZO-1 expression and inflammation in A549 cells. These results suggest that NaHS may prevent emphysema via the suppression of PHD2/HIF-1α/MAPK signaling pathway, and subsequently inhibition of inflammation, epithelial cell injury and apoptosis, and may be a novel strategy for the treatment of COPD.     

Preclinical development of CAT-354, an IL-13 neutralizing antibody, for the treatment of severe uncontrolled asthma

BACKGROUND AND PURPOSE

IL-13 is a pleiotropic Th2 cytokine considered likely to play a pivotal role in asthma. Here we describe the preclinical in vitro and in vivo characterization of CAT-354, an IL-13-neutralizing IgG4 monoclonal antibody (mAb), currently in clinical development.

EXPERIMENTAL APPROACH

In vitro the potency, specificity and species selectivity of CAT-354 was assayed in TF-1 cells, human umbilical vein endothelial cells and HDLM-2 cells. The ability of CAT-354 to modulate disease-relevant mechanisms was tested in human cells measuring bronchial smooth muscle calcium flux induced by histamine, eotaxin generation by normal lung fibroblasts, CD23 upregulation in peripheral blood mononuclear cells and IgE production by B cells. In vivo CAT-354 was tested on human IL-13-induced air pouch inflammation in mice, ovalbumin-sensitization and challenge in IL-13 humanized mice and antigen challenge in cynomolgus monkeys.

KEY RESULTS

CAT-354 has a 165 pM affinity for human IL-13 and functionally neutralized human, human variant associated with asthma and atopy (R130Q) and cynomolgus monkey, but not mouse, IL-13. CAT-354 did not neutralize human IL-4. In vitro CAT-354 functionally inhibited IL-13-induced eotaxin production, an analogue of smooth muscle airways hyperresponsiveness, CD23 upregulation and IgE production. In vivo in humanized mouse and cynomolgus monkey antigen challenge models CAT-354 inhibited airways hyperresponsiveness and bronchoalveolar lavage eosinophilia.

CONCLUSIONS AND IMPLICATIONS

CAT-354 is a potent and selective IL-13-neutralizing IgG4 mAb. The preclinical data presented here support the trialling of this mAb in patients with moderate to severe uncontrolled asthma.     

6-Shogaol inhibits monosodium urate crystal-induced inflammation – An in vivo and in vitro study

Gout is a rheumatic disease that is manifestated by an intense inflammation secondary to monosodium urate crystal deposition in joints. In the present study, we assessed the effect of 6-shogaol (isolated active principle from ginger) on monosodium urate crystal-induced inflammation in mice; an experimental model for gouty arthritis and compared it with that of the non-steroidal anti-inflammatory drug, indomethacin. Paw volume and levels/activities of lysosomal enzymes, lipid peroxidation, anti-oxidant status and inflammatory mediator TNF-α were determined in control and monosodium urate crystal-induced mice. The levels of β-glucuronidase and lactate dehydrogenase were also measured in monosodium urate crystal-incubated polymorphonuclear leucocytes (PMNL) in vitro. The levels of lysosomal enzymes, lipid peroxidation, and inflammatory mediator tumour necrosis factor-α and paw volume were increased significantly and the activities of anti-oxidant status were in turn decreased in monosodium urate crystal-induced mice, whereas these changes were reverted to near normal levels upon 6-shogaol administration. In vitro, 6-shogaol reduced the level of β-glucuronidase and lactate dehydrogenase in monosodium urate crystal-incubated polymorphonuclear leucocytes in concentration dependent manner when compared to control cells. The present results clearly indicated that 6-shogaol exerted a strong anti-inflammatory effect and can be regarded as useful tool for the treatment of acute gouty arthritis.     

Overview of hyperuricaemia and gout

In most mammals purine degradation ultimately leads to the formation of allantoin. Humans lack the enzyme uricase, which catalyzes the conversion of uric acid to allantoin. The resulting higher level of uric acid has been hypothesized to play a role as an antioxidant. Hyperuricaemia is usually an asymptomatic condition which is hypothesized to play a role in cardiovascular disease and hypertension. Some hyperuricaemic individuals develop gout, an inflammatory arthritis caused by the deposition of monosodium urate crystals in joints. Over time, acute intermittent gouty arthritis can develop into a chronic condition with deposits of monosodium urate (MSU) crystals in joints and as tophi. The mechanisms by which MSU crystals lead to an acute inflammatory arthritis are under investigation and current knowledge is reviewed here. Treatment of gout includes management of acute flares with anti-inflammatory medications such as non-steroidal anti-inflammatory drugs or corticosteroids and long term management with urate-lowering therapy when indicated. Future directions in the treatment of gout, in part guided by a better understanding of pathophysiology, are discussed.     

Functional nano-vector boost anti-atherosclerosis efficacy of berberine in Apoe(−/−) mice

Atherosclerosis (AS) is the leading cause of heart attacks, stroke, and peripheral vascular disease. Berberine (BBR), a botanical medicine, has diversified anti-atherosclerotic effects but with poor absorption. The aim of this study was to develop an effective BBR-entrapped nano-system for treating AS in high-fat diet (HFD)-fed Apoe^(−/−) mice, and also explore the possible underlying mechanisms involved. Three d-α-tocopherol polyethylene glycol (PEG) succinate (TPGS) analogues with different PEG chain lengths were synthesized to formulate BBR-entrapped micelles. HFD-fed Apoe^(−/−) mice were administered with optimized formula (BBR, 100 mg/kg/day) orally for 5 months. The artery plaque onset and related metabolic disorders were evaluated, and the underlying mechanisms were studied. Our data showed that, BT₁₅₀₀M increased BBR deposition in liver and adipose by 107.6% and 172.3%, respectively. In the Apoe^(−/−) mice, BT₁₅₀₀M ameliorated HFD-induced hyperlipidemia and lipid accumulation in liver and adipose. BT₁₅₀₀M also suppressed HFD-induced chronic inflammation as evidenced by the reduced liver and adipose levels of interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β); and decreased plasma level of TNF-α, IL-6, IL-1β, interferon-γ (IFN-γ), monocyte chemotactic protein (MCP), and macrophage inflammatory factor (MIP). The mechanism study showed that BT₁₅₀₀M changed Ampk and Nf-κb gene expression, and interrupted a crosstalk process between adipocytes and macrophages. Further investigation proved that BT₁₅₀₀M decreased endothelial lesion and subsequent macrophage activation, cytokines release, as well as cholesteryl ester gathering in the aortic arch, resulting in ameliorated artery plaque build-up. Our results provide a practical strategy for treating AS using a BBR-entrapped nano-system.     

An in vivo and in vitro potential of Indian ayurvedic herbal formulation Triphala on experimental gouty arthritis in mice

In the present study, we have investigated the efficacy of Indian ayurvedic herbal formulation Triphala on monosodium urate crystal-induced inflammation in mice; an experimental model for gouty arthritis and compared it with that of the non-steroidal anti-inflammatory drug, Indomethacin. The anti-arthritic effect of Triphala was evaluated by measuring changes in the paw volume, lysosomal enzyme activities, lipid peroxidation, anti-oxidant status and inflammatory mediator TNF-alpha in control and monosodium urate crystal-induced mice. The levels of beta-glucuronidase and lactate dehydrogenase were also measured in monosodium urate crystal-incubated polymorphonuclear leucocytes (PMNL). Triphala treatment (1 gm/kg/b.w. orally) significantly inhibited the paw volume and the levels of lysosomal enzymes, lipid peroxidation and inflammatory mediator tumour necrosis factor-alpha; however the anti-oxidant status was found to be increased in plasma, liver and spleen of monosodium urate crystal-induced mice when compared to control mice. In addition, beta-glucuronidase and lactate dehydrogenase level were reduced in Triphala (100 microg/ml) treated monosodium urate crystal-incubated polymorphonuclear leucocytes. In conclusion, the results obtained clearly indicated that Triphala exerted a strong anti-inflammatory effect against gouty arthritis.     

Formosanin C attenuates lipopolysaccharide-induced inflammation through nuclear factor-κB inhibition in macrophages

Extended inflammation and cytokine production pathogenically contribute to a number of inflammatory disorders. Formosanin C (FC) is the major diosgenin saponin found in herb Paris formosana Hayata (Liliaceae), which has been shown to exert anti-cancer and immunomodulatory functions. In this study, we aimed to investigate anti-inflammatory activity of FC and the underlying molecular mechanism. RAW264.7 macrophages were stimulated with lipopolysaccharide (LPS) or pre-treated with FC prior to being stimulated with LPS. Thereafter, the macrophages were subjected to analysis of the expression levels of pro-inflammatory mediators, including nitric oxide (NO), prostaglandin E₂ (PGE), tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), and IL-6, as well as two relevant enzymes, inducible nitric oxide synthase (iNOS), and cyclooxygenase-2 (COX-2). The analysis revealed that FC administration blunted LPS-induced production of NO and PGE in a dose-dependent manner, while the expression of iNOS and COX-2 at both mRNA and protein levels was inhibited in LPS-stimulated macrophages pre-treated with FC. Moreover, LPS stimulation upregulated mRNA expression and medium release of TNF-α, IL-1β, and IL-6, whereas this effect was blocked upon FC pre-administration. Mechanistic studies showed that inhibitory effects of FC on LPS-induced inflammation were associated with a downregulation of IκB kinase, IκB, and p65/NF-κB pathway. Taken together, these data suggest that FC possesses an inflammation-suppressing activity, thus being a potential agent for the treatment of inflammation-associated disorders.     

Inflammatory parameters in Caco-2 cells: Effect of stimuli nature,concentration, combination and cell differentiation

Enterocytes regulate gut maintenance and defence by secreting and responding to inflammatory mediators and by modulating the intestinal epithelial permeability. In order to develop an in vitro model of the acute phase of intestinal inflammation, Caco-2 cells were exposed to the inflammatory mediators IL-1β, TNF-α, IFN-γ and LPS, and the importance of several experimental parameters, i.e. cell differentiation, stimulus nature, concentration and combination on the inflammatory response was assessed by measuring the production of IL-6, IL-8, PGE-2 and NO and by evaluating the monolayer permeability. A maximal increase in IL-8, IL-6 and PGE-2 production and monolayer permeability was observed when using the cytokines simultaneously at their highest level, but this relied mainly on IL-1β. The effects of TNF-α on IL-8 and IL-6 or NO production were stronger upon combination with IL-1β or IFN-γ, respectively, whereas cells were unaffected by the presence of LPS. Although NO production, induced by IFN-γ-containing combinations, was observed only in differentiated cells, general inflammatory response was higher in proliferating cells. The use of a mixture of IL-1β, TNF-α and IFN-γ thus accurately mimics intestinal inflammatory processes, but cell differentiation and stimuli combination are important parameters to take into account for in vitro studies on intestinal inflammation.     

Interleukin 17 sustains rather than induces inflammation

Interleukin (IL)17 is a novel cytokine that has been suggested to play a key role in sustaining chronic inflammation in autoimmune diseases. Since its discovery, much attention has been given to mediators and factors responsible for the development of IL-17-producing cells while very few studies have investigated the inflammatory properties of this cytokine. Here we aimed to characterize the inflammatory properties of IL-17 and to establish if this cytokine per se can initiate an inflammatory reaction or if its main role is to contribute to the exacerbation of ongoing inflammation. To this aim we used two different mouse models of inflammation: the paw oedema and the airpouch. Interestingly, injection of IL-17 in the hind paw did not cause oedema while administration in the pre-inflamed tissues of 6-day-old air pouches induced a long lasting inflammatory reaction that was sustained between 4 and 24 h post-injection. Phenotypic analysis of cellular infiltrates demonstrated selective PMN recruitment in exudates and pouch lining tissues. This event was accompanied by induction of a distinct set of pro-inflammatory cytokines including IL-1β, IL-6, TNF-α and chemokines KC and MCP-1. Co-administration of a neutralizing anti-KC antibody with IL-17 significantly reduced its inflammatory response suggesting a key role for this chemokine in mediating the inflammatory effects of IL-17. In conclusion these results demonstrate that IL-17 does not initiate an inflammatory reaction while, if injected in pre-inflamed tissues, is able to further amplify biochemical and cellular events characteristic of the early stages of the inflammatory reaction.