Polydatin Prevents Calcium Pyrophosphate Crystal-Induced Arthritis in Mice.

BACKGROUND: Polydatin is a stilbenoid with important antioxidant, anti-inflammatory, and immunomodulating properties. The aim of this study was to assess the anti-inflammatory preventive effect of polydatin in the mouse model of acute arthritis induced by calcium pyrophosphate (CPP) crystals. METHODS: Acute arthritis was induced by the injection of a suspension of sterile CPP crystals into the ankle joint of Balb/c mice. Animals were randomized to receive polydatin or colchicine (the control drug) according to a prophylactic and a therapeutic protocol. The primary outcome was the variation of ankle swelling obtained after crystal injection and treatment, while histological parameters such as leukocyte infiltration, IL-1ß and CXCL1 levels and tissue expression were considered as secondary outcomes. RESULTS: Prophylactic treatment with PD significantly diminished ankle swelling after 48 h from crystal injection. Secondary outcomes such as leukocyte infiltration, necrosis, edema, and synovitis were also decreased. PD caused a reduction in circulating levels of IL-1ß and CXCL1, as well as their tissue expression. By contrast, the therapeutic administration of PD did not have any beneficial effect. CONCLUSIONS: PD can effectively prevent acute inflammatory response to crystals in the mouse model of CPP crystal-induced arthritis. These results suggest that this bioactive compound might be used in the prevention of crystal-induced acute attacks in humans.

Monovalent antibody-conjugated lipid-polymer nanohybrids for active targeting to desmoglein 3 of keratinocytes to attenuate psoriasiform inflammation.

To improve the treatment of psoriasiform inflammation, we developed actively targeted nanocarriers loaded with the phosphodiesterase 4 inhibitor AN2728. Methods: Phospholipid-poly(lactic-co-glycolic acid) nanohybrids were prepared and conjugated with monovalent anti-desmoglein 3 antibody to bind keratinocytes. Results: The actively targeted nanohybrids were 229 nm in mean size with a nearly neutral surface charge. Flow cytometry and confocal microscopy showed a 9-fold increase in keratinocyte uptake of targeted nanohybrids relative to non-targeted nanoparticles. The nanoparticles localized mainly in lysosomes after internalization. AN2728-loaded antibody-conjugated nanocarriers inhibited cytokine/chemokine overexpression in activated keratinocytes without affecting cell viability. The targeted nanohybrids also suppressed neutrophil migration by reducing CXCL1 and CXCL2 release from keratinocytes. Following subcutaneous administration in mice, the nanohybrids distributed to the epidermis and hair follicles. In a psoriasis-like skin mouse model, the actively targeted nanoparticles were superior to free drug and non-targeted nanoparticles in mitigating skin inflammation. Intervention with the targeted nanosystem reduced the epidermal thickness of the psoriasiform lesion from 191 to 42 µm, decreased the Psoriasis Area Severity Index by 74%, restored barrier function, and returned chemokine levels to baseline. Conclusions: Our developed nanosystem was safe and demonstrated efficient targeting properties for the treatment of cutaneous inflammation.

RIP3 promotes colitis-associated colorectal cancer by controlling tumor cell proliferation and CXCL1-induced immune suppression.

Rationale: Necroptosis is a programmed form of non-apoptotic cell death that requires receptor-interacting protein 3 (RIP3). RIP3 has been shown to be relevant in multiple tumor types and has differential impact on tumor progression. We investigated whether RIP3 is involved in the progression of colitis-associated cancer (CAC) in mice. Methods: Tissues from colorectal cancer patients were examined for RIP3 expression. CAC was induced using azoxymethane (AOM) injection followed by dextran sodium sulfate (DSS) treatment in RIP3-deficient or wild-type mice. Colon tissues were collected and analyzed by Western blotting and gene expression profile analyses. Immune cell infiltration and CXCL1 expression were examined by flow cytometry and Real-time PCR, respectively. Results: RIP3 expression was upregulated in mouse CAC and human colon cancer. RIP3-deficient mice showed significantly attenuated colitis-associated tumorigenesis. Bone marrow transplantation experiments suggested that RIP3's function in hematopoietic cells primarily contributes to the phenotype. RIP3 supported epithelial proliferation and tumor growth via JNK signaling but had no effect on apoptosis. RIP3 deletion increased T cell accumulation and reduced infiltration by immunosuppressive subsets of myeloid cells during acute colitis and CAC. The immune-suppressive tumor microenvironment was dependent on RIP3-induced expression of the chemokine attractant CXCL1, and administration of recombinant CXCL1 during CAC restored tumorigenesis in Rip3-/- mice. Conclusion: Our results reveal an unexpected function of RIP3 in enhancing the proliferation of premalignant intestinal epithelial cells (IECs) and promoting myeloid cell-induced adaptive immune suppression. These two distinct mechanisms of RIP3-induced JNK and CXCL1 signalling contribute to CAC progression.

Anti-inflammatory effects of propranolol in the temporomandibular joint of female rats and its contribution to antinociceptive action.

BACKGROUND: ß-Blockers reduce temporomandibular joint (TMJ) pain. We asked whether they also reduce TMJ inflammation and, if so, whether this anti-inflammatory effect contributes to its analgesic action. METHODS: We measured many parameters of the inflammatory response after co-administration of the ß-blocker propranolol with the inflammatory agent carrageenan in the TMJ of female rats. We also hypothesized that the activation of ß-adrenoceptors in the TMJ induces nociception mediated, at least in part, by the inflammatory response. To test this hypothesis, we examined the nociceptive response induced by the activation of the ß-adrenoceptors in the TMJ in female rats pretreated with thalidomide and fucoidan. RESULTS: We found that the co-administration of propranolol with carrageenan in the TMJ of female rats significantly reduced several parameters of the inflammatory response induced by carrageenan such as plasma extravasation, neutrophil migration and the release of the pro-inflammatory cytokines TNF-α, IL-1ß and CINC-1. Furthermore, the injection of the ß-adrenergic receptor agonist isoproterenol in the TMJ induced nociception that was significantly reduced by thalidomide, fucoidan and by the co-administration of propranolol but not of the α-adrenergic receptor antagonist phentolamine. CONCLUSIONS: Propranolol has anti-inflammatory effects that contribute to its antinociceptive action in the TMJ of females. SIGNIFICANCE: ß-Blockers have an anti-inflammatory effect on temporomandibular joint (TMJ) that contributes to its analgesic effect. The results of this work suggest that ß-blockers can be used to treat the painful conditions of TMJ, especially when they are associated with an inflammatory process.

Proposed mechanisms for oligonucleotide IMT504 induced diabetes reversion in a mouse model of immunodependent diabetes.

Type 1 diabetes (T1D) originates from autoimmune ß-cell destruction. IMT504 is an immunomodulatory oligonucleotide that increases mesenchymal stem cell cloning capacity and reverts toxic diabetes in rats. Here, we evaluated long-term (20 doses) and short-term (2-6 doses) effects of IMT504 (20 mg·kg(-1)·day(-1) sc) in an immunodependent diabetes model: multiple low-dose streptozotocin-injected BALB/c mice (40 mg·kg(-1)·day(-1) ip for 5 consecutive days). We determined blood glucose, glucose tolerance, serum insulin, islet morphology, islet infiltration, serum cytokines, progenitor cell markers, immunomodulatory proteins, proliferation, apoptosis, and islet gene expression. IMT504 reduced glycemia, induced ß-cell recovery, and impaired islet infiltration. IMT504 induced early blood glucose decrease and infiltration inhibition, increased ß-cell proliferation and decreased apoptosis, increased islet indoleamine 2,3-dioxygenase (IDO) expression, and increased serum tumor necrosis factor and interleukin-6 (IL-6). IMT504 affected islet gene expression; preproinsulin-2, proglucagon, somatostatin, nestin, regenerating gene-1, and C-X-C motif ligand-1 cytokine (Cxcl1) increased in islets from diabetic mice and were decreased by IMT504. IMT504 downregulated platelet endothelial cell adhesion molecule-1 (Pecam1) in islets from control and diabetic mice, whereas it increased regenerating gene-2 (Reg2) in islets of diabetic mice. The IMT504-induced increase in IL-6 and islet IDO expression and decreased islet Pecam1 and Cxcl1 mRNA expression could participate in keeping leukocyte infiltration at bay, whereas upregulation of Reg2 may mediate ß-cell regeneration. We conclude that IMT504 effectively reversed immunodependent diabetes in mice. Corroboration of these effects in a model of autoimmune diabetes more similar to human T1D could provide promising results for the treatment of this disease.

Effects of Tocilizumab on Experimental Severe Acute Pancreatitis and Associated Acute Lung Injury.

OBJECTIVE: To examine the therapeutic effects of tocilizumab, an antibody against interleukin-6 receptor, on experimental severe acute pancreatitis and associated acute lung injury. The optimal dose of tocilizumab and the activation of interleukin-6 inflammatory signaling were also investigated. DESIGN: Randomized experiment. SETTING: Research laboratory at a university hospital. SUBJECT: Experimental severe acute pancreatitis in rats. INTERVENTIONS: Severe acute pancreatitis was induced by retrograde injection of sodium taurocholate (50 mg/kg) into the biliopancreatic duct. In dose-study, rats were administered with different doses of tocilizumab (1, 2, 4, 8, and 16 mg/kg) through the tail vein after severe acute pancreatitis induction. In safety-study, rats without severe acute pancreatitis induction were treated with high doses of tocilizumab (8, 16, 32, and 64 mg/kg). Serum and tissue samples of rats in time-study were collected for biomolecular and histologic evaluations at different time points (2, 6, 12, 18, and 24 hr). MEASUREMENTS AND MAIN RESULTS: 1) Under the administration of tocilizumab, histopathological scores of pancreas and lung were decreased, and severity parameters related to severe acute pancreatitis and associated lung injury, including serum amylase, C-reactive protein, lung surfactant protein level, and myeloperoxidase activity, were all significant alleviated in rat models. 2) Dose-study demonstrated that 2 mg/kg tocilizumab was the optimal treatment dose. 3) Basing on multi-organ pathologic evaluation, physiological and biochemical data, no adverse effect and toxicity of tocilizumab were observed in safety-study. 4) Pancreatic nuclear factor-κB and signal transducer and activator of transcription 3 were deactivated, and the serum chemokine (C-X-C motif) ligand 1 was down-regulated after tocilizumab administration. CONCLUSIONS: Our study demonstrated tocilizumab, as a marketed drug commonly used for immune-mediated diseases, was safe and effective for the treatment of experimental severe acute pancreatitis and associated acute lung injury. Our findings provide experimental evidences for potential clinical application of tocilizumab in severe acute pancreatitis and associated complications.

IL-1ß siRNA adenovirus benefits liver regeneration by improving mesenchymal stem cells survival after acute liver failure.

UNLABELLED:   BACKGROUND: Uncontrolled hapatic inflammatory response is regarded as the primary pathological mechanism of acute liver failure and impairs the regeneration of hepatocytes and stem cell grafts. Interleukin-1 plays a key role for activating immune and inflammatory response. Recently, siRNA has made quite a few progresses in treating inflammatory response. AIM: To assess the effect of IL-1? siRNA adenovirus on MSC and the therapeutic effect of MSC combined with IL-1? siRNA adenovirus in ALF. MATERIAL AND METHODS: We implanted MSC or/and IL-1? siRNA adenovirus via the tail vein, using CCl4-induced ALF in a mice model. Mice were sacrificed at different time points. Blood samples and liver tissues were collected. Hepatic injury, liver regeneration, cytokines (CXCL1, IL-1?, IL-10, IL-6, VEGF and HGF), animal survival and vital MSC were assessed after cell transplantation. RESULTS: MSC combined with IL-1? siRNA reduced the inflammatory levels and prevented liver failure. These animals administrated with MSC and IL-1? siRNA also exhibited improved liver regeneration and increased survival rates. Immunohistochemistry and fluorescence microscopy revealed the number of vital MSC in ALF + MSC + IL-1? siRNA group were significantly more than that in ALF + MSC group. CONCLUSION: IL-1? siRNA adenovirus could enhance MSC ability of tissue regeneration through increasing its survival rate. Accordingly, combination of IL-1? siRNA adenovirus and MSC had a synergistic effect on acute liver failure.

Autoantibodies to citrullinated proteins induce joint pain independent of inflammation via a chemokine-dependent mechanism.

OBJECTIVE: An interesting and so far unexplained feature of chronic pain in autoimmune disease is the frequent disconnect between pain and inflammation. This is illustrated well in rheumatoid arthritis (RA) where pain in joints (arthralgia) may precede joint inflammation and persist even after successful anti-inflammatory treatment. In the present study, we have addressed the possibility that autoantibodies against citrullinated proteins (ACPA), present in RA, may be directly responsible for the induction of pain, independent of inflammation. METHODS: Antibodies purified from human patients with RA, healthy donors and murinised monoclonal ACPA were injected into mice. Pain-like behaviour was monitored for up to 28 days, and tissues were analysed for signs of pathology. Mouse osteoclasts were cultured and stimulated with antibodies, and supernatants analysed for release of factors. Mice were treated with CXCR1/2 (interleukin (IL) 8 receptor) antagonist reparixin. RESULTS: Mice injected with either human or murinised ACPA developed long-lasting pronounced pain-like behaviour in the absence of inflammation, while non-ACPA IgG from patients with RA or control monoclonal IgG were without pronociceptive effect. This effect was coupled to ACPA-mediated activation of osteoclasts and release of the nociceptive chemokine CXCL1 (analogue to human IL-8). ACPA-induced pain-like behaviour was reversed with reparixin. CONCLUSIONS: The data suggest that CXCL1/IL-8, released from osteoclasts in an autoantibody-dependent manner, produces pain by activating sensory neurons. The identification of this new pain pathway may open new avenues for pain treatment in RA and also in other painful diseases associated with autoantibody production and/or osteoclast activation.

High-density lipoproteins inhibit urate crystal-induced inflammation in mice.

OBJECTIVES: To investigate the effects and mechanisms of action of high-density lipoproteins (HDL) in monosodium urate (MSU) crystal-induced inflammation -that is, gouty inflammation, in vivo. METHODS: Air pouches raised on the backs of mice were injected with MSU crystals or tumour necrosis factor (TNF) in the presence or absence of HDL and/or interleukin (IL)-1 receptor antagonist (IL-1Ra) for 3 h. Leucocyte count and neutrophil percentage in pouch fluids were measured using a haemocytometer and May-Grünwald-Giemsa staining. The cytokine production and expression in the pouch were measured by ELISA and quantitative RT-PCR. RESULTS: MSU crystals induced leucocyte infiltration, mostly neutrophils, and the release of IL-1ß, IL-6, chemokine (C-X-C motif) ligand 1 (CXCL1), chemokine (C-C motif) ligand 2 (CCL2) and IL-1Ra in pouch fluids. TNF remained under the detection limit. MSU crystals triggered IL-1ß, IL-6 and CXCL1 expression in both pouch exudates and membranes, whereas CCL2 and TNF mRNA were not modulated. The co-injection of MSU crystals and HDL inhibited leucocyte influx by 59% and neutrophil infiltration by 83% and, in turn, both protein and mRNA levels of all assessed proinflammatory cytokines were reduced, but not those of IL-1Ra. Similar results were obtained when mice were injected with MSU crystals pretreated with HDL or TNF instead of crystals. When HDL and IL-1Ra were added together they displayed additional inhibition, suggesting different mechanisms of action. CONCLUSIONS: This study demonstrated that HDL may represent an important factor in the modulation of gouty inflammation by acting on both tissue and infiltrating cells -that is, synovial tissue and synovial fluid cells. HDL display anti-inflammatory activity, in part, by interacting with crystals but also by directly acting on cells.

Dietary fish oil reduces the acute inflammatory response and enhances resolution of antigen-induced peritonitis.

Dietary n-3 polyunsaturated fatty acids (PUFA) influence the inductive phase of inflammation but less is known about their effects on the resolution phase. This study examined the effects of dietary fish oil on induction and resolution of antigen-induced inflammation in mice. Mice were fed a control diet with or without 2.8% fish oil, immunized twice with methylated BSA (mBSA) and inflammation induced by intraperitoneal injection of mBSA. Prior to and at different time points after mBSA administration, peritoneal cells were analyzed and expression of surface molecules determined by flow cytometry. Concentration of chemokines, cytokines and soluble cytokine receptors was determined by ELISA. Mice fed the fish oil diet had fewer peritoneal neutrophils, shorter resolution interval and lower levels of pro-inflammatory cytokines and chemokines than mice fed the control diet. In mice fed the fish oil diet there was an early peak in peritoneal levels of the immunosuppressive molecules sIL-6R and TGF-ß, that was not seen in mice fed the control diet. In the resolution phase, peritoneal macrophages from mice fed the fish oil diet expressed more of the atypical chemokine receptor D6 and peritoneal TGF-ß levels were higher than that in mice fed the control diet. Furthermore, in the late-resolution phase there were more peritoneal eosinophils and macrophages in mice fed the fish oil diet than in mice fed the control diet. These results demonstrate a suppressive effect of n-3 PUFA on the inductive phase of inflammation and indicate an enhancing effect of n-3 PUFA on resolution of inflammation.

Effects of tobacco on cytokine expression from human endothelial cells.

OBJECTIVE: To investigate the effects of nicotine and cigarette smoke condensate (CSC) exposure on cytokine expression from human endothelial cells in order to identify one possible mechanism that smoking plays in the pathogenesis of both periodontal disease (PDD) and cardiovascular disease (CVD). METHODS: Human endothelial cells (HUVECs) were exposed to different concentrations of nicotine and CSC to examine the effects that they have on cell proliferation and cytotoxicity. Non-toxic levels were then used to examine cytokine expression using cytokine protein arrays. RESULTS: Exposure to nicotine caused significant down-regulation in the expression of IL-10 (P = 0.046), growth-regulated oncogene (GRO)α (P = 0.036), MCP-1 (P = 0.046), and GMCSF (P = 0.004) compared with the control untreated HUVECs. Exposure to CSC caused significant down-regulation in the expression of GRO (P = 0.04), GROα (P = 0.01), IL-6 (P = 0.03), and MCP-1 (P = 0.04) compared with the control untreated HUVECs. CONCLUSIONS: Exposure of HUVECs to nicotine or CSC affects the levels of cytokine expression including reduction in anti-inflammatory and chemoattractant cytokines. This may subsequently affect the host defensive mechanisms of the tissues. The action of toxic chemicals in tobacco smoke on endothelial cells is a potential pathogenic mechanism that may in part explain the association between tobacco, PDD, and CVD.

Redox-active protein thioredoxin-1 administration ameliorates influenza A virus (H1N1)-induced acute lung injury in mice.

OBJECTIVES: Influenza virus infections can cause severe acute lung injury leading to significant morbidity and mortality. Thioredoxin-1 is a redox-active defensive protein induced in response to stress conditions. Animal experiments have revealed that thioredoxin-1 has protective effects against various severe disorders. This study was undertaken to evaluate the protective effects of recombinant human thioredoxin-1 administration on influenza A virus (H1N1)-induced acute lung injury in mice. DESIGN: Prospective animal trial. SETTING: Research laboratory. SUBJECTS: Nine-week-old male C57BL/6 mice inoculated with H1N1. INTERVENTION: The mice were divided into a vehicle-treated group and recombinant human thioredoxin-1-treated group. For survival rate analysis, the vehicle or recombinant human thioredoxin-1 was administered intraperitoneally every second day from day -1 to day 13. For lung lavage and pathological analyses, vehicle or recombinant human thioredoxin-1 was administered intraperitoneally on days -1, 1, and 3. MEASUREMENTS AND MAIN RESULTS: Lung lavage and pathological analyses were performed at 24, 72, and 120 hrs after inoculation. The recombinant human thioredoxin-1 treatment significantly improved the survival rate of H1N1-inoculated mice, although the treatment did not affect virus propagation in the lung. The treatment significantly attenuated the histological changes and neutrophil infiltration in the lung of H1N1-inoculated mice. The treatment significantly attenuated the production of tumor necrosis factor-α and chemokine (C-X-C motif) ligand 1 in the lung and oxidative stress enhancement, which were observed in H1N1-inoculated mice. H1N1 induced expressions of tumor necrosis factor-α and chemokine (C-X-C motif) ligand 1 in murine lung epithelial cells MLE-12, which were inhibited by the addition of recombinant human thioredoxin-1. The recombinant human thioredoxin-1 treatment started 30 mins after H1N1 inoculation also significantly improved the survival of the mice. CONCLUSIONS: Exogenous administration of recombinant human thioredoxin-1 significantly improved the survival rate and attenuated lung histological changes in the murine model of influenza pneumonia. The protective mechanism of thioredoxin-1 might be explained by its potent antioxidative and anti-inflammatory actions. Consequently, recombinant human thioredoxin-1 might be a possible pharmacological strategy for severe influenza virus infection in humans.

N-acetyl-S-farnesyl-l-cysteine suppresses chemokine production by human dermal microvascular endothelial cells.

Isoprenylcysteine (IPC) molecules modulate G-protein-coupled receptor signalling. The archetype of this class is N-acetyl-S-farnesyl-l-cysteine (AFC). Topical application of AFC locally inhibits skin inflammation and elicitation of contact hypersensitivity in vivo. However, the mechanism of these anti-inflammatory effects is not well understood. Dermal microvascular endothelial cells (ECs) are involved in inflammation, in part, by secreting cytokines that recruit inflammatory cells. We have previously shown that the sympathetic nerve cotransmitter adenosine-5'-triphosphate (ATP) and adenosine-5'-O-(3-thio) triphosphate (ATPγS), an ATP analogue that is resistant to hydrolysis, increase secretion of the chemokines CXCL8 (interleukin-8), CCL2 (monocyte chemotactic protein-1) and CXCL1 (growth-regulated oncogene α) by dermal microvascular ECs. Production of these chemokines can also be induced by the exposure to the proinflammatory cytokine TNFα. We have now demonstrated that AFC dose-dependently inhibits ATP-, ATPγS- and TNFα-induced production of CXCL1, CXCL8 and CCL2 by a human dermal microvascular EC line (HMEC-1) in vitro under conditions that do not affect cell viability. Inhibition of ATPγS- or TNFα-stimulated release of these chemokines was associated with reduced mRNA levels. N-acetyl-S-geranyl-l-cysteine, an IPC analogue that is inactive in inhibiting G-protein-coupled signalling, had greatly reduced ability to suppress stimulated chemokine production. AFC may exert its anti-inflammatory effects through the inhibition of chemokine production by stimulated ECs.

Attenuation of indomethacin-induced gastric mucosal injury by prophylactic administration of sake yeast-derived thioredoxin.

BACKGROUND: Indomethacin is one of the group of nonsteroidal anti-inflammatory drugs, which often cause gastric mucosal injury as a side effect. Infiltration and activation of inflammatory cells, production of proinflammatory cytokines and chemokines, generation of reactive oxygen species, and activation of apoptotic signaling are involved in the pathogenesis of indomethacin-induced gastric injury. We examined whether sake yeast-derived thioredoxin (a small redox-active protein with anti-oxidative activity and various redox-regulating functions) reduced indomethacin-induced gastric injury. METHODS: Gastric injury was produced by the intraperitoneal administration of indomethacin (40 mg/kg body weight) to C57BL/6 mice. Prior to the administration of indomethacin, the mice were offered food pellets containing non-genetically modified sake yeast-derived thioredoxin (thioredoxin 200 µg/g) for 3 days. Histological examinations, assessment of myeloperoxidase activity, and analysis of the gene expressions of proinflammatory cytokines and a chemokine (interleukin [IL]-1ß, IL-6, and CXCL1) were statistically evaluated. Indomethacin cytotoxicity was determined by lactate dehydrogenase release from murine gastric epithelial GSM06 cells induced by 24-h treatment with 200 and 400 µM indomethacin after 1-h preincubation with 100 µg/ml sake yeast-derived thioredoxin. RESULTS: Macroscopic (edema, hemorrhage, and ulcers) and histological (necrosis, submucosal edema, neutrophil infiltration) findings induced by indomethacin were significantly reduced by pretreatment with food pellets containing thioredoxin. Gastric myeloperoxidase activity and the gene expressions of proinflammatory cytokines (IL-1ß and IL-6) were also significantly reduced by this pretreatment compared with findings in the mice not pretreated with thioredoxin-containing food pellets. The administration of sake yeast-derived thioredoxin significantly reduced indomethacin-induced cytotoxicity in GSM06 cells. CONCLUSIONS: We conclude that oral administration of sake yeast-derived thioredoxin reduces indomethacin-induced gastric injury. Sake yeast-derived thioredoxin may have therapeutic potential against indomethacin-induced gastric injury.

Dietary supplementation of krill oil attenuates inflammation and oxidative stress in experimental ulcerative colitis in rats.

OBJECTIVE: To evaluate the effects of krill oil (KO) on inflammation and redox status in dextran sulfate sodium (DSS)-induced colitis in rats. MATERIALS AND METHODS: Thirty male Wistar rats were divided into three groups: Control, DSS, and DSS + KO 5% in a 4-week diet study. Colitis was induced by 5% DSS in the drinking water the last week of the experiment. Weight and disease activity index (DAI), colon length, histological combined score (HCS), colon levels of selected cytokines and prostaglandins, markers of protein oxidative damage, fatty acid profile, and expression of selected genes were measured. RESULTS: Rats in the DSS group increased their DAI and HCS compared with healthy controls. The colon length was significantly preserved after KO diet. Tumor necrosis factor (TNF)-α and interleukin (IL)-1ß were elevated in the DSS group compared with controls. Cytokines and HCS were nonsignificantly lower in the KO versus the DSS group. Prostaglandin (PG)E(3) increased significantly in the KO versus the other groups. Peroxisome proliferator-activated receptor (PPAR)-γ expression was nonsignificantly increased while PPAR-γ coactivator 1α (Pparg1α) expression increased significantly after KO. The levels of protein oxidation markers decreased significantly. CONCLUSIONS: KO showed protective potential against DSS colitis based on the preservation of colon length, reduction of oxidative markers and the consistent beneficial changes of HCS, cytokine, and (PG)E(3) levels, as well as PPAR-γ and Pparg1α expression compared with DSS alone. These findings indicate an anti-inflammatory and a protein antioxidant effect of KO.

IKK-2 inhibitor TPCA-1 represses nasal epithelial inflammation in vitro.

BACKGROUND: Nasal polyposis (NP) is considered a subgroup within chronic rhinosinusitis. NP can be further subdivided into aspirin sensitive- and aspirin tolerant types (ASNP/ ATNP). Although the true etiology of NP has not been identified so far, it is agreed that NP represents an inflammatory disease of the nasal mucosa. Alterations of cellular kinase activities including that of IKK-2 might play a role in this inflammatory process. METHODS: Paraffin sections of ASNP, ATNP and controls were immunostained with Phospho-IkB-α antibody that detects the direct IKK-2 product (IkB-α. Intensity of epithelial staining was analysed semi-quantitatively by two independent observers. In cultured nasal polyp epithelial cells (NPECs) epithelial derived cytokines IL-8 and GRO α were induced by TNF-α or Staphylococcal supernatants and subsequently repressed by IKK-2 inhibitor TPCA-1. RESULTS: Significant Phospho-IkB-α staining was observed in the nasal epithelium of ASNP compared to ATNP and controls suggesting strong IKK-2 activation in patients with ASNP in vivo. In vitro, pro-inflammatory cytokines IL-8 and GRO-α in NPECs were significantly repressed by TPCA-1. CONCLUSION: IKK-2 activity is increased in the subgroup of ASNP. IL-8 and GRO-α responses were repressed by IKK-2 inhibitor TPCA-1 in vitro. IKK-2 inhibitors might represent a potential target for anti-inflammatory intervention in ASNP.

Butyrate and type 1 diabetes mellitus: can we fix the intestinal leak?

OBJECTIVES: An intestinal permeability defect precedes type 1 diabetes mellitus and may be a permissive factor in its pathogenesis. Butyrate strengthens the intestinal tight junctions. We hypothesized that enteral administration of sodium butyrate (NaB) in preweaned rats would result in differences in the development of diabetes associated with decreased inflammation and pancreatic ß-cell destruction. MATERIALS AND METHODS: Using biobreeding diabetes-prone rat pups, oral NaB or saline was administered twice per day via micropipette from postnatal days 10 to 23. Rat pups were randomly assigned to 1 of 4 groups for the first experiment (control group, n = 7) and 3 different doses of butyrate groups (n = 8 for each group) and 2 groups for the second and third experiments (control n = 23; NaB at 400 mg · kg(-1) · day(-1), n = 20). Animals were studied into adulthood (up to day 140) for development of diabetes. RESULTS: The results showed that the survival rates were 28% versus 20% (butyrate vs control). No significant differences in survival were seen; however, there was a trend of delaying of onset of diabetes in the butyrate group. There were no differences of pancreatic histology score of islet inflammation between the 2 groups. Cytokine-induced neutrophil chemoattractant-1 was lower in the butyrate group at a dose of 400 mg · kg(-1) · day(-1) in the distal small intestine (P = 0.008) and in the liver (P = 0.01). There were no significant differences in the tracer flux measurements across the distal ileum and colon between the 2 animal groups. CONCLUSIONS: Oral NaB given during the preweaning period did not significantly decrease the subsequent development of death from diabetes in biobreeding diabetes-prone rats.

Effect of formoterol and budesonide on chemokine release, chemokine receptor expression and chemotaxis in human neutrophils.

Severe persistent asthma and chronic obstructive pulmonary disease (COPD) are associated with neutrophil influx into the airways. It is not clear whether neutrophil chemotaxis is influenced by beta(2)-agonists and glucocorticoids, drugs commonly used in treatment of asthma and COPD. The effect of a long-acting beta(2)-agonist (formoterol), and a glucocorticosteroid (budesonide) on chemokine/cytokine release (CXCL8, CXCL1, IL-6), regulation of chemokine receptors (CXCR1, CXCR2), and migration were assessed in neutrophils from 10 non-allergic, healthy donors. Formoterol enhanced and budesonide inhibited IL-6, CXCL8 and CXCL1 release from LPS-stimulated neutrophils. Formoterol up-regulated both CXCR1 and CXCR2 expression, whereas budesonide up-regulated the expression of CXCR2 only. Despite the effects on chemokine release and drug-induced up-regulation of CXCR1 and CXCR2, no influence on neutrophil chemotaxis could be demonstrated. We conclude that a beta(2)-agonist and a glucocorticoid, commonly used in the treatment of obstructive lung diseases, influence chemokine release and receptor sensitivity but the functional consequences of these findings remain unclear.

Mast cells are involved in the pathogenesis of indomethacin-induced rat enteritis.

BACKGROUND: Nonsteroidal antiinflammatory drug (NSAID)-induced enteropathy is clinically very important, but the pathological mechanisms remain unclear. Mast cells have been reported to play an important role in the pathogenesis of indomethacin-induced small intestinal injury. In this study, we investigated the role of mast cells in indomethacin-induced small intestinal injury using mast cell deficiency (Ws/Ws) rat. METHODS: Ws/Ws rats and control (W+/W+) rats were given indomethacin (15 mg/kg) subcutaneously, and the intestinal mucosal damage was estimated after 24 h. RESULTS: The area (mm2) of macroscopic visible lesions, the concentrations of thiobarbituric acid-reactive substances (TBARS) as an index of lipid peroxidation, myeloperoxidase (MPO) activity as an index of neutrophil accumulation, and the content of cytokine-induced neutrophil chemoattractant-1 (CINC-1) were significantly increased in indomethacin-treated groups compared with the sham groups. The development of intestinal lesions in response to indomethacin was prevented in Ws/Ws rats compared with W+/W+ rats, together with significant suppression of the increased levels of TBARS, MPO activities, and CINC-1 levels. CONCLUSIONS: These results indicate that mast cells are involved in the pathogenesis of the intestinal mucosal damage induced by indomethacin.

Geranylgeranylacetone prevents acute liver damage after massive hepatectomy in rats through suppression of a CXC chemokine GRO1 and induction of heat shock proteins.

BACKGROUND AND METHODS: Acute liver failure after massive hepatectomy remains a challenging problem. In this study, using a microarray designed to monitor the side effects of drugs, we examined changes in gene expression in the remnant liver during the 24 h after hepatectomy and the effects of a nontoxic heat shock protein (HSP) 70 inducer, geranylgeranylacetone (GGA), after 90% hepatectomy in rats. RESULTS: A single oral administration of 100 mg/kg GGA significantly suppressed the release of aminotransferases and improved survival compared with vehicle administration. The hepatectomy upregulated 74 genes and downregulated 95. Interestingly, ten cytokine genes were upregulated, while no cytokine-related gene was downregulated. Among the ten cytokine genes, a potent chemoattractant for neutrophils, GRO1, was most rapidly and markedly upregulated after 90% hepatectomy. GGA effectively suppressed the up-regulation of GRO1 messenger ribonucleic acid, and this was validated by Northern hybridization. Microarray and immunoblot analyses showed that, in addition to HSP70 and HSP27, GGA preferentially induced an endoplasmic reticulum chaperone, BIP. CONCLUSION: Considering hemodynamic and metabolic overloading as a primary cause of acute lever failure, the ER stress response enhanced by GGA may also play an important role in the prevention of overload-induced liver damage.