Ex-vivo mucolytic and anti-inflammatory activity of BromAc in tracheal aspirates from COVID-19.

COVID-19 is a lethal disease caused by the pandemic SARS-CoV-2, which continues to be a public health threat. COVID-19 is principally a respiratory disease and is often associated with sputum retention and cytokine storm, for which there are limited therapeutic options. In this regard, we evaluated the use of BromAc®, a combination of Bromelain and Acetylcysteine (NAC). Both drugs present mucolytic effect and have been studied to treat COVID-19. Therefore, we sought to examine the mucolytic and anti-inflammatory effect of BromAc® in tracheal aspirate samples from critically ill COVID-19 patients requiring mechanical ventilation. METHOD: Tracheal aspirate samples from COVID-19 patients were collected following next of kin consent and mucolysis, rheometry and cytokine analysis using Luminex kit was performed. RESULTS: BromAc® displayed a robust mucolytic effect in a dose dependent manner on COVID-19 sputum ex vivo. BromAc® showed anti-inflammatory activity, reducing the action of cytokine storm, chemokines including MIP-1alpha, CXCL8, MIP-1b, MCP-1 and IP-10, and regulatory cytokines IL-5, IL-10, IL-13 IL-1Ra and total reduction for IL-9 compared to NAC alone and control. BromAc® acted on IL-6, demonstrating a reduction in G-CSF and VEGF-D at concentrations of 125 and 250 µg. CONCLUSION: These results indicate robust mucolytic and anti-inflammatory effect of BromAc® ex vivo in tracheal aspirates from critically ill COVID-19 patients, indicating its potential to be further assessed as pharmacological treatment for COVID-19.

Effect of a single high dose of vitamin D3 on cytokines, chemokines, and growth factor in patients with moderate to severe COVID-19.

BACKGROUND: The modulating effect of vitamin D on cytokine concentrations in severe coronavirus disease 2019 (COVID-19) remains unknown. OBJECTIVES: We aimed to investigate the effect of a single high dose of vitamin D3 on cytokines, chemokines, and growth factor in hospitalized patients with moderate to severe COVID-19. METHODS: This is a post hoc, ancillary, and exploratory analysis from a multicenter, double-blind, placebo-controlled, randomized clinical trial. Patients with moderate to severe COVID-19 were recruited from 2 hospitals in São Paulo, Brazil. Of 240 randomly assigned patients, 200 were assessed in this study and randomly assigned to receive a single oral dose of 200,000 IU vitamin D3 (n = 101) or placebo (n = 99). The primary outcome was hospital length of stay, which has been published in our previous study. The prespecified secondary outcomes were serum concentrations of IL-1ß, IL-6, IL-10, TNF-α, and 25-hydroxyvitamin D. The post hoc exploratory secondary outcomes were IL-4, IL-12p70, IL-17A, IFN-γ, granulocyte-macrophage colony-stimulating factor (GM-CSF), IL-8, IFN-inducible protein-10 (IP-10), macrophage inflammatory protein-1ß (MIP-1ß), monocyte chemoattractant protein-1 (MCP-1), vascular endothelial growth factor (VEGF), and leukocyte count. Generalized estimating equations for repeated measures, with Bonferroni's adjustment, were used for testing all outcomes. RESULTS: The study included 200 patients with a mean ± SD age of 55.5 ± 14.3 y and BMI of 32.2 ± 7.1 kg/m2, of which 109 (54.5%) were male. GM-CSF concentrations showed a significant group-by-time interaction effect (P = 0.04), although the between-group difference at postintervention after Bonferroni's adjustment was not significant. No significant effects were observed for the other outcomes. CONCLUSIONS: The findings do not support the use of a single dose of 200,000 IU vitamin D3, compared with placebo, for the improvement of cytokines, chemokines, and growth factor in hospitalized patients with moderate to severe COVID-19.This trial was registered at clinicaltrials.gov as NCT04449718.

Modulation of the chemokine/chemokine receptor axis as a novel approach for glioma therapy.

Chemokines are a large subfamily of cytokines known for their ability to facilitate cell migration, most notably leukocytes, throughout the body. Chemokines are necessary for a functioning immune system in both health and disease and have received considerable attention for their roles in orchestrating temporal-spatial regulation of immune cell populations in cancer. Gliomas comprise a group of common central nervous system (CNS) primary tumors that are extremely challenging to treat. Immunotherapy approaches for highly malignant brain tumors offer an exciting new avenue for therapeutic intervention but so far, have seen limited successful clinical outcomes. Herein we focus on important chemokine/chemokine receptor systems in the regulation of pro- and anti-tumor mechanisms, highlighting potential therapeutic advantages of modulating these systems in malignant gliomas and other cancers.

Comparative effects of atorvastatin 80 mg and rosuvastatin 40 mg on the levels of serum endocan, chemerin, and galectin-3 in patients with acute myocardial infarction.

OBJECTIVE: Endocan, chemerin, and galectin-3 are discrete biomarkers associated with cardiovascular diseases and acting through different pathophysiological pathways. The aim of this study is to investigate and compare the effects of high doses of atorvastatin and rosuvastatin on serum endocan, chemerin, and galectin-3 levels in patients with acute myocardial infarction (AMI). METHODS: Sixty-three patients with AMI were randomized to receive atorvastatin (80 mg/day) or rosuvastatin (40 mg/day) after percutaneous revascularization. Serum levels of endocan, chemerin, and galectin-3 were evaluated at baseline and after 4-week therapy. RESULTS: Endocan levels were not decreased statistically significantly with atorvastatin 80 mg, but rosuvastatin 40 mg markedly decreased the levels of endocan according to baseline [from 110.27 (86.03-143.69) pg/mL to 99.22 (78.30-122.87) pg/mL with atorvastatin 80 mg and from 110.73 (77.28-165.22) pg/mL to 93.40 (70.48-115.13) pg/mL with rosuvastatin 40 mg, p=0.242 for atorvastatin 80 mg and p=0.014 for rosuvastatin 40 mg]. Chemerin levels significantly decreased in both groups according to baseline [from 264.90 (196.00-525.95) ng/mL to 135.00 (105.95-225.65) ng/mL with atorvastatin 80 mg and from 309.95 (168.87-701.27) ng/mL to 121.25 (86.60-212.65) ng/mL with rosuvastatin 40 mg, p<0.001, respectively, for both groups]. Galectin-3 levels did not change markedly with atorvastatin 80 mg, but they decreased with rosuvastatin 40 mg [from 17.00 (13.10-22.25) ng/mL to 19.30 (15.25-23.45) ng/mL with atorvastatin 80 mg, p=0.721, and from 18.25 (12.82-23.82) ng/mL to 16.60 (10.60-20.15) ng/mL with rosuvastatin 40 mg, p=0.074]. There were no significant between-group differences in terms of absolute and percentage changes of endocan, chemerin, and galectin-3 at 4 weeks. CONCLUSION: We reported that both statins similarly decreased the endocan levels, whereas rosuvastatin seems to have more prominent effects on the reduction of the chemerin and galectin-3 levels in patients with AMI.

The effect of carvacrol on inflammatory mediators and respiratory symptoms in veterans exposed to sulfur mustard, a randomized, placebo-controlled trial.

BACKGROUND: The aim of this study was to evaluate the effect of carvacrol on serum level of inflammatory mediators and respiratory symptoms in the veterans exposed to sulfur mustard (SM). METHODS: Twenty-one patients who were exposed to SM more than two decades' ago were divided to placebo and carvacrol (1.2 mg/kg/day) treated groups. Serum levels of Tumor Necrosis Factor-α (TNF-α), Monocyte chemotactic protein-1 (MCP-1), Vascular endothelial growth factor (VEGF), Epidermal growth factor (EGF), forced expiratory volume-one second (FEV1) and respiratory symptoms including; Chest wheeze (CW), night wheeze (NW), night cough (NC) and cough and wheeze during exercise (ECW) were assessed at the baseline (step 0), one and two months after starting treatment (step I and II, respectively). FINDINGS: FEV1 value was significantly increased in carvacrol treated group in step II compared to step 0 (p < 0.001) and also increased in step II compared to step I (p < 0.05). The respiratory symptoms including; CW and NW was significant decreased in carvacrol treated group in step I and II compared to step 0 (p < 0.01 to p < 0.001), NC and ECW were significantly decreased only in step II compared to step 0 (p < 0.01, for both cases). The serum levels of TNF-α, EGF and VEGF were decrease in carvacrol treated group in step I and II compared to step 0 (p < 0.05 to p < 0.001). The serum level of MCP-1 was decrease in carvacrol treated group only in the step II compared to step 0 (p < 0.05). INTERPRETATION: Two months' treatment with carvacrol reduced inflammatory cytokine and chemokine while increased anti-inflammatory cytokines and improved respiratory symptom and FEV1 value in SM exposed patients. CLINICAL TRIALS REGISTRY NUMBER: This trial was registered under IRCT2014031617020N1 at http://www.irct.ir/.

Ibudilast Inhibits Chemokine Expression in Rheumatoid Arthritis Synovial Fibroblasts and Exhibits Immunomodulatory Activity in Experimental Arthritis.

OBJECTIVE: Ibudilast is a well-tolerated, orally available phosphodiesterase 4 (PDE4) inhibitor used to treat asthma and stroke. Since PDE4 inhibition suppresses inflammatory mediator production and cell proliferation in leukocytes, ibudilast may be a valuable therapy for the treatment of inflammatory autoimmune diseases such as rheumatoid arthritis (RA). This study was undertaken to assess the therapeutic potential of ibudilast by measuring its capacity to modulate inflammation in human leukocytes and RA synovial fibroblasts (RASFs) and in experimental arthritis. METHODS: Using standard curve quantitative polymerase chain reaction, the effect of ibudilast on gene expression in activated human leukocytes and RASFs was measured. Ibudilast was used to treat DBA/1 mice with collagen-induced arthritis, and an adoptive transfer model was used to assess its tolerogenic capacity. RESULTS: Ibudilast inhibited the expression of TNF, IL12A, and IL12B and the secretion of tumor necrosis factor (TNF) and interleukin-12 (IL-12)/23p40 from leukocytes, and reduced the expression of CCL5 and CCL3 in activated RASFs. Treatment of experimental arthritis with ibudilast resulted in a reduction in IL-17-producing cells and inhibition of disease progression. When combined with a TNF inhibitor, ibudilast caused marked suppression of active disease. Exposure of leukocytes from type II collagen-immunized DBA/1 mice to ibudilast in vitro attenuated their ability to adoptively transfer arthritis to DBA/1J-PrkdcSCID mice, providing evidence of an immunomodulatory effect. CONCLUSION: Our findings indicate that ibudilast reduces the expression and/or secretion of inflammatory mediators from activated human leukocytes and RASFs, inhibits Th17 cell responses in vivo, and improves established arthritis. Given the established safety profile of ibudilast in humans, its clinical evaluation in RA, either alone or in combination with a TNF inhibitor, should be considered.

Role of Gut-Derived Endotoxin on Type I Collagen Production in the Rat Pancreas After Chronic Alcohol Exposure.

BACKGROUND: Pancreatic fibrosis is a key pathological feature of alcoholic chronic pancreatitis (ACP). Bacterial endotoxin lipopolysaccharide (LPS) is considered as an important cofactor in the fibrogenesis of ACP. However, there are limitations in the use of exogenous LPS for evaluating the role of endotoxin in ACP pathogenesis. In this study, we determined the relationship between the concentration of LPS in the portal vein and pancreatic type I collagen (Col1) content in chronic alcohol-fed rats. METHODS: Male Sprague Dawley rats were divided into 2 groups and fed with Lieber-DeCarli isocaloric control (CON) liquid diet or ethanol (EtOH) (15 g/kg/d) liquid diet. Eleven CON or EtOH rats were euthanized at the end of week 8, 9, or 10. The plasma LPS from portal vein was determined. Pancreatic inflammatory injury and fibrosis were assessed. Pancreatic stellate cells (PSCs) and macrophages were identified; pancreatic type I collagen alpha 1 (Col1A1) and Toll-like receptor (TLR4) mRNA and protein were examined; pancreatic chemokines and transforming growth factor-beta1 (TGF-ß1) were determined. RESULTS: Pancreatic inflammatory scores were increased in 10-week EtOH rats compared with CON rats, but there was no significant difference in collagen deposition between 2 groups. The levels of portal vein LPS and pancreatic TLR4 and Col1A1 mRNA and protein were increased in a time-dependent fashion in EtOH rats, with the highest levels occurring at 10 weeks. Additionally, by 8 weeks, pancreatic TLR4 and Col1A1 mRNA in EtOH rats were statistically increased as compared to CON rats, whereas portal vein LPS remained unchanged. The number of PSCs and macrophages and expression of chemokines (MCP-1, MIP-1α, and RANTES), TGF-ß1, or Col1A1 were significantly increased, each of which was positively correlated with the level of portal vein LPS in 10-week EtOH rats. CONCLUSIONS: These results suggest that LPS is associated with alcohol-induced fibrosis in pancreatitis and targeting of bacterial endotoxin may be a promising therapeutic strategy for ACP.

Prognostic significance of baseline metabolic tumor volume in relapsed and refractory Hodgkin lymphoma.

Identification of prognostic factors for patients with relapsed/refractory Hodgkin lymphoma (HL) is essential for optimizing therapy with risk-adapted approaches. In our phase 2 study of positron emission tomography (PET)-adapted salvage therapy with brentuximab vedotin (BV) and augmented ifosfamide, carboplatin, and etoposide (augICE), we assessed clinical factors, quantitative PET assessments, and cytokine and chemokine values. Transplant-eligible patients with relapsed/refractory HL received 2 (cohort 1) or 3 (cohort 2) cycles of weekly BV; PET-negative patients (Deauville score ≤2) proceeded to autologous stem cell transplantation (ASCT) whereas PET-positive patients received augICE before ASCT. Serum cytokine and chemokine levels were measured at baseline and after BV. Metabolic tumor volume (MTV) and total lesion glycolysis were measured at baseline, after BV, and after augICE. Sixty-five patients enrolled (45, cohort 1; 20, cohort 2); 49 (75%) achieved complete response and 64 proceeded to ASCT. Three-year overall survival and event-free survival (EFS) were 95% and 82%, respectively. Factors predictive for EFS by multivariable analysis were baseline MTV (bMTV) (P < .001) and refractory disease (P = .003). Low bMTV (<109.5 cm3) and relapsed disease identified a favorable group (3-year EFS, 100%). For patients who received a transplant, bMTV and pre-ASCT PET were independently prognostic; 3-year EFS for pre-ASCT PET-positive patients with low bMTV was 86%. In this phase 2 study of PET-adapted therapy with BV and augICE for relapsed/refractory HL, bMTV and refractory disease were independent prognostic factors for EFS. Furthermore, bMTV improved the predictive power of pre-ASCT PET. Future studies should optimize efficacy and tolerability of salvage therapy by stratifying patients according to risk factors such as bMTV.

Geldanamycin Reduces Acute Respiratory Distress Syndrome and Promotes the Survival of Mice Infected with the Highly Virulent H5N1 Influenza Virus.

Infections with lethal influenza viruses lead to acute lung injury (ALI) or acute respiratory distress syndrome (ARDS), which may be related to the activation of the host's immune system. Here, in our study, male C57BL/6 mice were infected with 10 LD50 of the H5N1 influenza virus and treated with geldanamycin or oseltamivir 2 h after infection. Lung injury was assessed by histopathology on days 4 and 7. The viral load was quantified by measuring the NP gene expression level on days 2, 4, and 7. Levels of cytokines and chemokines in bronchoalveolar lavage fluids and inflammatory cells were analyzed at different time points. Geldanamycin administration prolonged survival in mice and dramatically reduced lung injury and pulmonary inflammatory compared with other mice. Viral loads in geldanamycin-treated mice also significantly reduced compared with non-treated mice, but not to the extent as the oseltamivir-treated mice. Furthermore, the geldanamycin treatment markedly reduced the production of major proinflammatory cytokines and chemokines and attenuated the infiltration and activation of immune cells, but it did not alter the generation of virus-neutralizing antibodies. In conclusion, geldanamycin plays an important role in attenuating virus infection-induced ALI/ARDS by reducing the host's inflammatory responses and may provide an important reference for clinical treatments.

Decrease in Dengue virus-2 infection and reduction of cytokine/chemokine production by Uncaria guianensis in human hepatocyte cell line Huh-7

ABSTRACT BACKGROUND Dengue fever may present hemorrhages and cavitary effusions as result of exacerbated immune responses. We investigated hydro-alcoholic extracts from leaves (UGL) and bark (UGB) of the medicinal species Uncaria guinanensis with respect to antiviral effects in Dengue virus (DENV) infection and in immunological parameters associated with in vivo physiopathological features. METHODS Chemical profiles from UGB or UGL were compared in thin layer chromatography and 1H nuclear magnetic resonance using flavonoid compounds and a pentacyclic oxindole alkaloid-enriched fraction as references. DENV-2-infected hepatocytes (Huh-7) were treated with extracts. Cell viability, DENV antigens and immunological factors were detected by enzyme-linked immunosorbent assay (ELISA) or flow cytometry. FINDINGS The UGL mainly differed from UGB by selectively containing the flavonoid kaempferitrin. UGB and UGL improved hepatocyte viability. Both extracts reduced intracellular viral antigen and inhibited the secretion of viral non-structural protein (NS1), which is indicative of viral replication. Reduction in secretion of macrophage migration inhibitory factor was achieved by UGB, of interleukin-6 by UGL, and of interleukin-8 by both UGB and UGL. MAIN CONCLUSIONS The U. guianensis extracts presented, antiviral and immunomodulatory effects for DENV and possibly a hepatocyte-protective activity. Further studies may be performed to consider these products as potential candidates for the development of an herbal product for the future treatment of dengue.

B cells influence sex specificity of arthritis via myeloid suppressors and chemokines in humanized mice.

Rheumatoid arthritis (RA) occurs two times more often in women than men. B cell depletion has been shown to be efficacious in treating RA. Our previous studies suggested that antigen presentation via B cells results in a sex-specific immune response in DR4 and DR4/DQ8 mice. Here we evaluated the mechanism of efficacy of the B cell depletion in treating arthritis-susceptible DQ8 mice. The data show that arthritic DQ8 mice treated with anti-CD20 antibody in therapeutic protocols show milder disease severity in females as compared to males, which is associated with decreased antibodies to citrullinated proteins and reduced levels of IL-23 and CCL5. Treatment led to significantly increased numbers of T regulatory and monocyte-derived suppressor F4/80+Gr1hi cells in females as compared to male DQ8 mice. Our observations suggest that therapeutic strategies that target B cells may benefit females while functions of DCs might be relatively more important for men than women.

The therapeutic potential of targeting chemokine signalling in the treatment of chronic pain.

Chronic pain is a distressing condition, which is experienced even when the painful stimulus, whether surgery or disease related, has subsided. Current treatments for chronic pain show limited efficacy and come with a host of undesirable side-effects, and thus there is a need for new, more effective therapies to be developed. The mechanisms underlying chronic pain are not fully understood at present, although pre-clinical models have facilitated the progress of this understanding considerably in the last decade. The mechanisms underlying chronic pain were initially thought to be neurocentric. However, we now appreciate that non-neuronal cells play a significant role in nociceptive signalling through their communication with neurons. One of the major signalling pathways, which mediates neuron/non-neuronal communication, is chemokine signalling. In this review, we discuss selected chemokines that have been reported to play a pivotal role in the mechanisms underlying chronic pain in a variety of pre-clinical models. Approaches that target each of the chemokines discussed in this review come with their advantages and disadvantages; however, the inhibition of chemokine actions is emerging as an innovative therapeutic strategy, which is now reaching the clinic, with the chemokine Fractalkine and its CX3 CR1 receptor leading the way. This article is part of the special article series "Pain".

Dietary gossypol suppressed postprandial TOR signaling and elevated ER stress pathways in turbot (Scophthalmus maximus L.).

Gossypol is known to be a polyphenolic compound toxic to animals. However, its molecular targets are far from fully characterized. To evaluate the physiological and molecular effects of gossypol, we chose turbot (Scophthalmus maximus L.), a carnivorous fish, as our model species. Juvenile turbots (7.83 ± 0.02 g) were fed diets containing gradient levels of gossypol at 0 (G0), 600 (G1), and 1,200 (G2) mg/kg diets for 11 wk. After the feeding trial, fish growth, body protein, and fat contents were significantly reduced in the G2 group compared with those of the G0 group (P < 0.05). Gossypol had little impact on digestive enzyme activities and intestine morphology. However, gossypol caused liver fibrosis and stimulated chemokine and proinflammatory cytokine secretions. More importantly, gossypol suppressed target of rapamycin (TOR) signaling and induced endoplasmic reticulum (ER) stress pathway in both the feeding experiment and cell cultures. Our results demonstrated that gossypol inhibited TOR signaling and elevated ER stress pathways both in vivo and in vitro, thus providing new mechanism of action of gossypol in nutritional physiology.

Deregulated neddylation in liver fibrosis.

Hepatic fibrosis is a global health problem currently without effective therapeutic approaches. Even though the ubiquitin-like posttranslational modification of neddylation, that conjugates Nedd8 (neural precursor cell expressed developmentally downregulated) to specific targets, is aberrant in many pathologies, its relevance in liver fibrosis (LF) remained unexplored. Our results show deregulated neddylation in clinical fibrosis and both in mouse bileductligation- and CCl4 -induced fibrosis. Importantly, neddylation inhibition, by using the pharmacological inhibitor, MLN4924, reduced liver injury, apoptosis, inflammation, and fibrosis by targeting different hepatic cell types. On one hand, increased neddylation was associated with augmented caspase 3 activity in bile-acid-induced apoptosis in mouse hepatocytes whereas neddylation inhibition ameliorated apoptosis through reduction of expression of the Cxcl1 and Ccl2 chemokines. On the other hand, chemokine receptors and cytokines, usually induced in activated macrophages, were reduced after neddylation inhibition in mouse Kupffer cells. Under these circumstances, decreased hepatocyte cell death and inflammation after neddylation inhibition could partly account for reduction of hepatic stellate cell (HSC) activation. We provide evidence that augmented neddylation characterizes activated HSCs, suggesting that neddylation inhibition could be important for resolving LF by directly targeting these fibrogenic cells. Indeed, neddylation inhibition in activated HSCs induces apoptosis in a process partly mediated by accumulation of c-Jun, whose cullin-mediated degradation is impaired under these circumstances. CONCLUSION: Neddylation inhibition reduces fibrosis, suggesting neddylation as a potential and attractive therapeutic target in liver fibrosis. (Hepatology 2017;65:694-709).

The effect of an IκB-kinase-ß(IKKß) inhibitor on tobacco smoke-induced pulmonary inflammation.

PURPOSE OF THE STUDY: Inactivation of NF-κB with IKKß knockout mice reduces tobacco smoke-induced pulmonary inflammation. In this study, we investigated whether the IKKß inhibitor PS-1145 could attenuate the pulmonary inflammation induced by tobacco smoke. MATERIALS AND METHODS: We divided 30 mice into three groups: a control group, a smoking group, and a PS-1145 group. Mice from the smoking and PS-1145 groups were exposed for 2 weeks to tobacco smoke. PS-1145 was injected intraperitoneally before every tobacco smoke exposure. After 2 weeks, bronchoalveolar lavage (BAL) was performed for cell counting and measuring of inflammatory chemokines. We analyzed the correlation between NF-κB and NF-κB-regulated chemokines in BAL fluid and measured the neutrophils and macrophages by immunostaining in lung tissues. RESULTS: The PS-1145 group showed a significant reduction in the number of total cells, neutrophils, and macrophages, as well as the KC and MCP-1 level, in the BAL fluid compared to the smoking group. There was no significant difference in the level of MIP-1α. The level of NF-κB in BAL fluid was significantly positively correlated with KC and MCP-1 levels, but not with MIP-1α level. The PS-1145 group also showed a significant fewer neutrophils and macrophages in the lung tissue. CONCLUSIONS: We conclude that the IKKß inhibitor PS-1145 suppressed the NF-κB signaling pathway and reduced the recruitment of inflammatory cells and chemokines in pulmonary inflammation induced by tobacco smoke. IKKß inhibition offers a potential therapeutic target for tobacco smoke-induced pulmonary inflammation.

Successes and failures of chemokine-pathway targeting in rheumatoid arthritis.

Chemokines and chemokine receptors are involved in leukocyte recruitment and angiogenesis underlying the pathogenesis of rheumatoid arthritis (RA) and other inflammatory rheumatic diseases. Numerous chemokines, along with both conventional and atypical cell-surface chemokine receptors, are found in inflamed synovia. Preclinical studies carried out in animal models of arthritis involving agents targeting chemokines and chemokine receptors have yielded promising results. However, most human trials of treatment of RA with antibodies and synthetic compounds targeting chemokine signalling have failed to show clinical improvements. Chemokines can have overlapping actions, and their activities can be altered by chemical modification or proteolytic degradation. Effective targeting of chemokine pathways must take acount of these properties, and can also require high levels of receptor occupancy by therapeutic agents to prevent signalling. CCR1 is a promising target for chemokine-receptor blockade.

Hepatic stellate cells relay inflammation signaling from sinusoids to parenchyma in mouse models of immune-mediated hepatitis.

UNLABELLED: Hepatic stellate cells (HSCs) constitute the liver sinusoid with Kupffer cells and liver sinusoidal endothelial cells. While the sinusoid functions as the gateway to liver inflammation, whether HSCs contribute to liver inflammation and, if so, how they exert such functions remain elusive. Here, we found that mouse as well as human HSCs expressed DP1 receptor for prostaglandin D2 selectively in the liver. Pharmacological stimulation of DP1 by BW245C, a DP1-selective agonist, suppressed the activation of cultured HSCs by tumor necrosis factor-α at least in part through down-regulation of nuclear factor kappa-light-chain-enhancer of activated B cells signaling and inhibition of c-Jun N-terminal kinase phosphorylation. DP1 deficiency or BW245C administration in mice significantly enhanced or suppressed concanavalin A (ConA)-induced hepatitis, respectively. ConA injection induced tumor necrosis factor-α and interferon-γ expression in the sinusoid, which was suppressed by administration of BW245C. Coculture of spleen cells and liver nonparenchymal cells showed that ConA first activated spleen cells and that this activation led to activation of nonparenchymal cells to secondarily produce tumor necrosis factor-α and interferon-γ. Microarray analysis revealed ConA-induced expression of endothelin-1, tissue factor, and chemokines in the liver and inducible nitric oxide synthase in hepatocytes, resulting in flow stagnation, leukocyte adherence and migration to the parenchyma, and hepatocyte death. DP1 stimulation inhibits all these events in the liver. Therefore, HSCs mediate amplification of ConA-induced liver inflammation in the sinusoid, causing direct and indirect hepatocyte injury, and DP1 stimulation inhibits this HSC activation. CONCLUSIONS: HSCs integrate cytokine-mediated inflammatory responses in the sinusoids and relay them to the liver parenchyma, and these HSC actions are inhibited by DP1 stimulation.

Effects of new over-the-counter periodontal ointment-containing applicator with single-tuft brush on cytokine levels in gingival crevicular fluid during supportive periodontal therapy phase: a randomized double-blind clinical trial.

BACKGROUND AND OBJECTIVE: The biochemical effects of an over-the-counter (OTC) medication were studied, which consists of a single-tuft brush containing cetylpyridinium chloride as a bactericidal agent, dipotassium glycyrrhizate as an anti-inflammatory drug and allantoin as a promoter of cell proliferation and wound healing, for delivery to hardly brushed sites. MATERIAL AND METHODS: This randomized controlled double-blind study was performed in 61 subjects with chronic periodontitis in supportive periodontal therapy phase (test group: n = 27; placebo group: n = 28; dropout: n = 6). The OTC medication was self-applied twice a day for 12 wk to two molars with probing pocket depths of 4-6 mm. Biochemical indicators were evaluated at baseline and 12 wk using the suspension array system for eight cytokines and chemokines (interleukin [IL]-1ß, IL-1ra, IL-4, IL-6, IL-8, IL-10, monocyte chemoattractant protein-1 and tumor necrosis factor [TNF]-α) in gingival crevicular fluid. RESULTS: The levels of IL-1ß, IL-6, IL-8 and TNF-α remained significantly lower in the test group compared to the placebo group. In the placebo group, when the probing pocket depth at baseline was 4 mm, IL-1ß increased, particularly in the second molar tooth, and the greatest increase was seen when PPD at baseline was 5-6 mm. In the test group, IL-1ß decreased markedly in cases with furcation involvement and low bleeding on probing at baseline. In both groups, IL-1ß, IL-6 and TNF-α were closely correlated with each other. CONCLUSION: This OTC medication is biochemically effective for steady chronic periodontitis in the supportive periodontal therapy phase.

Effect of fosinopril on chemerin and VEGF expression in diabetic nephropathy rats.

As the most common and severe complication of diabetes, diabetic nephropathy (DN) has been known to be related with angiotensin converting enzyme inhibitor (ACEI), which can reduce proteinuria and protect renal function. This study analyzed the effect of ACEI analog-fosinopril-on the expression of chemerin and vascular epithelial growth factor (VEGF), in an attempt to reveal the mechanism of ACEI analog on renal protection. A total of 45 SD rats were induced by sreptozotocin for diabetes and were given fosinopril via intragastric cannulation for 12 weeks. After sacrifice, serum and renal chemerin and VEGF contents were quantified by enzyme linked immunosorbent assay (ELISA) and Western blot method, in addition to biochemical laboratory examinations. In diabetic model rats, blood glucose, creatinine, urea nitrogen, 24-hour urinary protein, chemerin and VEGF protein contents were all significantly elevated when compared to those in control group (P<0.05). After fosinopril treatment, blood creatinine, urea nitrogen, 24-hour urinary protein, Chemerin and VEGF protein concentrations were significantly depressed (P<0.05 compared to model group). Positive relationships existed between renal chemerin, VEGF and urea protein levels. Fosinopril may protect renal tissues in diabetes by suppressing chemerin and VEGF protein expression.

Mesenchymal Stem Cells Exhibit Regulated Exocytosis in Response to Chemerin and IGF.

Mesenchymal stem cells (MSCs) play important roles in tissue repair and cancer progression. Our recent work suggests that some mesenchymal cells, notably myofibroblasts exhibit regulated exocytosis resembling that seen in neuroendocrine cells. We now report that MSCs also exhibit regulated exocytosis. Both a G-protein coupled receptor agonist, chemerin, and a receptor tyrosine kinase stimulant, IGF-II, evoked rapid increases in secretion of a marker protein, TGFßig-h3. The calcium ionophore, ionomycin, also rapidly increased secretion of TGFßig-h3 while inhibitors of translation (cycloheximide) or secretory protein transport (brefeldin A) had no effect, indicating secretion from preformed secretory vesicles. Inhibitors of the chemerin and IGF receptors specifically reduced the secretory response. Confocal microscopy of MSCs loaded with Fluo-4 revealed chemerin and IGF-II triggered intracellular Ca2+ oscillations requiring extracellular calcium. Immunocytochemistry showed co-localisation of TGFßig-h3 and MMP-2 to secretory vesicles, and transmission electron-microscopy showed dense-core secretory vesicles in proximity to the Golgi apparatus. Proteomic studies on the MSC secretome identified 64 proteins including TGFßig-h3 and MMP-2 that exhibited increased secretion in response to IGF-II treatment for 30min and western blot of selected proteins confirmed these data. Gene ontology analysis of proteins exhibiting regulated secretion indicated functions primarily associated with cell adhesion and in bioassays chemerin increased adhesion of MSCs and adhesion, proliferation and migration of myofibroblasts. Thus, MSCs exhibit regulated exocytosis that is compatible with an early role in tissue remodelling.