CCL2 Concentrations Research Articles

ARRDC4 regulates enterovirus 71-induced innate immune response by promoting K63 polyubiquitination of MDA5 through TRIM65

Enterovirus 71 (EV71) is the main causative agent of hand, foot and mouth disease (HFMD), which induces significantly elevated levels of cytokines and chemokines, leading to local or system inflammation and severe complications, whereas the underlying regulatory mechanisms and the inflammatory pathogenesis remain elusive. ARRDC4 is one member of arrestins family, having important roles in glucose metabolism and G-protein-coupled receptors (GPCRs) related physiological and pathological processes, however, the function of ARRDC4 in innate immune system is largely unknown. Here we identified that ARRDC4 expression was increased after EV71 infection in THP-1-derived macrophages and verified in EV71-infected HFMD patients and the healthy candidates. The expression level of ARRDC4 was positively correlated with the serum concentration of IL-6, TNF-α and CCL3 in clinical specimens. ARRDC4 interacted with MDA5 via the arrestin-like N domain, and further recruited TRIM65 to enhance the K63 ubiquitination of MDA5, resulting in activation of the downstream innate signaling pathway and transcription of proinflammatory cytokines during EV71 infection. Our data highlight new function of ARRDC4 in innate immunity, contributing to the better understanding about regulation of MDA5 activation after EV71 infection, and also suggest ARRDC4 may serve as a potential target for intervention of EV71-induced inflammatory response.

Peripheral viral challenge triggers hippocampal production of inflammatory proteins.

Peripheral viral infections increase seizure propensity and intensity in susceptible individuals. We have modeled this comorbidity by demonstrating that intraperitoneal (ip) injection of the conventional viral mimetic, polyinosinic-polycytidylic acid (PIC), renders the brain hypersusceptible to seizures induced by kainic acid (KA). At the molecular level, the hippocampus, which is the ictal site of KA-induced seizures, exhibits upregulated expression of messages encoding several inflammatory genes. Here, we profiled temporal expression of these genes at the protein level. Briefly, eight-week old female C57BL/6 mice were ip injected with 12mg/kg of PIC and inflammatory proteins were quantified in the hippocampus and blood by ELISA. We found a robust but transient increase in blood concentration of IL-6, CXCL10, CCL2, CXCL9, CCL7 and CCL12 six hours after PIC challenge. CXCL1, IL1β, TNFα and CXCL2 featured a moderate increase. However, only four chemokines were increased in the hippocampus. CXCL10 showed the highest increase 6-12h after PIC challenge, and its level dwindled to the baseline by 48h. CXCL1, CXCl9 and CXCL2 were also transiently elevated but their maximal values were by an order of magnitude lower than the values for CXCL10. These results indicate that CXCL10 is the primary inflammatory protein generated in the hippocampus in response to PIC challenge, and that this chemokine may drive the development of seizure hypersusceptibility. In addition, the hippocampus featured a protracted increase in the levels of anaphylatoxins C3a and C5a, indicating the activation of the complement cascades.

Quantification of Chemokine CCL5 in Patients with Diabetes Mellitus Type 2 and/or Chronic Periodontitis: Preliminary Study

The purpose of the present study was to quantify the presence of chemokine CCL5 (RANTES) in gingival crevicular fluid (LCG) in patients with chronic periodontitis (PC) and / or type 2 diabetes mellitus (DM2). A comparative cross-sectional study was conducted in 40 patients. LCG was taken from periodontal pockets and gingival grooves from 4 patient groups (10 per study group); patients who received periodontal, antibiotic and anti-inflammatory treatment 6 months prior to the study or who had systemic disease other than DM2 were excluded. Concentrations of CCL5 were determined by LUMINEX® assays. Descriptive statistics, one-way ANOVA, Student's T, and Pearson's correlation were performed. The quantification of CCL5 was higher in the patients who presented both diseases, followed by the group with only PC, healthy and the group with only DM2. No significant difference was found between groups and there was no correlation between quantifications and glycemic indicators. Although the differences were not significant, the group of patients with both diseases had the highest CCL5 quantification. The expression of CCL5 in LGC should be considered as a potential inducer of periodontal destruction, its determination could be useful for monitoring the health/disease of periodontal tissues.

Structural and Electronic Noninnocence of α-Diimine Ligands on Niobium for Reductive C–Cl Bond Activation and Catalytic Radical Addition Reactions

A d0 niobium(V) complex, NbCl3(α-diimine) (1a), supported by a dianionic redox-active N,N'-bis(2,6-diisopropylphenyl)-1,4-diaza-2,3-dimethyl-1,3-butadiene (α-diimine) ligand (ene-diamido ligand) served as a catalyst for radical addition reactions of CCl4 to α-olefins and cyclic alkenes, selectively affording 1:1 radical addition products in a regioselective manner. During the catalytic reaction, the α-diimine ligand smoothly released and stored an electron to control the oxidation state of the niobium center by changing between an η4-(σ2,π) coordination mode with a folded MN2C2 metallacycle and a κ2-(N,N') coordination mode with a planar MN2C2 metallacycle. Kinetic studies of the catalytic reaction elucidated the reaction order in the catalytic cycle: the radical addition reaction rate obeyed first-order kinetics that were dependent on the concentrations of the catalyst, styrene, and CCl4, while a saturation effect was observed at a high CCl4 concentration. In the presence of excess amounts of styrene, styrene coordinated in an η2-olefinic manner to the niobium center to decrease the reaction rate. No observation of oligomers or polymers of styrene and high stereoselectivity for the radical addition reaction of CCl4 to cyclopentene suggested that the C-C bond formation proceeded inside the coordination sphere of niobium, which was in good accordance with the negative entropy value of the radical addition reaction. Furthermore, reaction of 1a with (bromomethyl)cyclopropane confirmed that both the C-Br bond activation and formation proceeded on the α-diimine-coordinated niobium center during transformation of the cyclopropylmethyl radical to a homoallyl radical. With regard to the reaction mechanism, we detected and isolated NbCl4(α-diimine) (6a) as a transient one-electron oxidized species of 1a during reductive cleavage of the C-X bonds; in addition, the monoanionic α-diimine ligand of 6a adopted a monoanionic canonical form with selective one-electron oxidation of the dianionic ene-diamido form of the ligand in 1a.

Contributions of CD8 T cells to the pathogenesis of mouse adenovirus type 1 respiratory infection

CD8 T cells are key components of the immune response to viruses, but their roles in the pathogenesis of adenovirus respiratory infection have not been characterized. We used mouse adenovirus type 1 (MAV-1) to define CD8 T cell contributions to the pathogenesis of adenovirus respiratory infection. CD8 T cell deficiency in β2m-/- mice had no effect on peak viral replication in lungs, but clearance of virus was delayed in β2m-/- mice. Virus-induced weight loss and increases in bronchoalveolar lavage fluid total protein, IFN-γ, TNF-α, IL-10, CCL2, and CCL5 concentrations were less in β2m-/- mice than in controls. CD8 T cell depletion had similar effects on virus clearance, weight loss, and inflammation. Deficiency of IFN-γ or perforin had no effect on viral replication or inflammation, but perforin-deficient mice were partially protected from weight loss. CD8 T cells promote MAV-1-induced pulmonary inflammation via a mechanism that is independent of direct antiviral effects.

Chemokine binding protein 'M3' limits atherosclerosis in apolipoprotein E-/- mice.

Chemokines are important in macrophage recruitment and the progression of atherosclerosis. The ‘M3’ chemokine binding protein inactivates key chemokines involved in atherosclerosis (e.g. CCL2, CCL5 and CX3CL1). We aimed to determine the effect of M3 on plaque development and composition. In vitro chemotaxis studies confirmed that M3 protein inhibited the activity of chemokines CCL2, CCL5 and CX3CL1 as primary human monocyte migration as well as CCR2-, CCR5- and CX3CR1-directed migration was attenuated by M3. In vivo, adenoviruses encoding M3 (AdM3) or green fluorescence protein (AdGFP; control) were infused systemically into apolipoprotein (apo)-E-/- mice. Two models of atherosclerosis development were used in which the rate of plaque progression was varied by diet including: (1) a ‘rapid promotion’ model (6-week high-fat-fed) and (2) a ‘slow progression’ model (12-week chow-fed). Plasma chemokine activity was suppressed in AdM3-infused mice as indicated by significantly less monocyte migration towards AdM3 mouse plasma ex vivo (29.56%, p = 0.014). In the ‘slow progression’ model AdM3 mice had reduced lesion area (45.3%, p = 0.035) and increased aortic smooth muscle cell α-actin expression (60.3%, p = 0.014). The reduction in lesion size could not be explained by changes in circulating inflammatory monocytes as they were higher in the AdM3 group. In the ‘rapid promotion’ model AdM3 mice had no changes in plaque size but reduced plaque macrophage content (46.8%, p = 0.006) and suppressed lipid deposition in thoracic aortas (66.9%, p<0.05). There was also a reduction in phosphorylated p65, the active subunit of NF-κb, in the aortas of AdM3 mice (37.3%, p<0.0001). M3 inhibited liver CCL2 concentrations in both models with no change in CCL5 or systemic chemokine levels. These findings show M3 causes varying effects on atherosclerosis progression and plaque composition depending on the rate of lesion progression. Overall, our studies support a promising role for chemokine inhibition with M3 for the treatment of atherosclerosis.

Abstract P6-01-11: Distinct recruitment of tumor-associated immune cells correlates with increased pro-malignant chemokines in tumors expressing epithelial atypical chemokine receptor 1 (ACKR1/DARC), indicating a unique tumor microenvironment

Abstract The breast tumor microenvironment is a complex assortment of cancer and host immune cells that interact to produce a myriad of clinical outcomes for breast cancer (BrCa) patients.Chemokine receptors, such asthe Atypical Chemokine Receptor 1 (ACKR1), play an important role in maintaining the homeostasis of pro-inflammatory chemokines, as well as influencing the migration of host immune cells. ACKR1 has the ability to modify the breast tumor microenvironment in multiple ways, including sequestering chemokine activity and affecting leukocyte migration. The purpose of this study is to characterize the variation in immune cell signaling between ACKR1 positive and negative tumors, and determine any correlation between ACKR1 and CCL2 (MCP-1) and CXCL8 (IL-8) in various BrCa cell types. Primary breast tumor samples were stained using immunohistochemistry for ACKR1, in addition to various immune cells including, T-cells, B-cells, dendritic cells, and macrophages. We determined levels of circulating chemokines using a Luminex assay kit on whole blood lysate. We also tested the localization of ACKR1-associated chemokines and the abundance of ACKR1 using immunofluorescence techniques. Pilot data collected from primary breast tumor tissue suggested that differential expression of ACKR1 from various tumor subtypes leads to the recruitment of a specific subset of host immune cells to the tumor microenvironment. In our cohort, ACKR1 positive tumors tended to recruit B cells and dendritic cells to the site of the tumor, whereas ACKR1 negative cells did not. We also detected a positive correlation between ACKR1 levels in tumor tissue with CCL2 and CXCL8 levels in the circulating blood of our study group. Finally, co-localization of ACKR1 with CCL2 and CXCL8 was observed in cultured mammalian breast cancer cells. Overall, our pilot data suggests that there is differential recruitment of immune cells within the ACKR1 positive and negative BrCa tumor microenvironments, and that circulating CCL2 and CXCL8 concentrations are positively correlated with ACKR1 levels in BrCa cells. Citation Format: Jenkins BD, Martini RN, Hire R, Monteil MA, Davis MB. Distinct recruitment of tumor-associated immune cells correlates with increased pro-malignant chemokines in tumors expressing epithelial atypical chemokine receptor 1 (ACKR1/DARC), indicating a unique tumor microenvironment [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr P6-01-11.

Global DNA hypomethylation has no impact on lung function or serum inflammatory and fibrosis cytokines in asbestos-exposed population.

To examine the effect of asbestos exposure on global DNA methylation and determine whether lung function and inflammatory and fibrosis biomarkers are correlated with the methylation state. A total of 26 healthy subjects without asbestos exposure (Group 1), 47 healthy subjects with exposure (Group 2), and 52 subjects with benign asbestos-related disorders (ARDs) (Group 3) participated in this cross-sectional study. Blood global 5-methylcytosine (5mC) and serum TNF-α, collagen IV, CCL5 and CC16 concentrations were analyzed using enzyme-linked immunosorbent assay-like assays. Spirometric maneuvers were performed to assess lung function. Decreased 5mC levels were observed in Groups 2 and 3 compared to Group 1, irrespective of lung function (p < 0.01). There was no significant change in 5mC between Groups 2 and 3. Overall, 5mC was negatively correlated with CCL5 and collagen IV (p < 0.05), but no significant inverse relationship was found between 5mC and CCL5 or collagen IV in each group. Additionally, both 5mC and CC16 were inversely associated with FEV1/FVC% (p = 0.001, adjusted R 2 = 0.145) for non-smokers, and consistently significant inverse relationships were found between CC16 and FEV1/FVC%, independent of asbestos exposure. Asbestos exposure causes global DNA hypomethylation. DNA hypomethylation has no influence on serum biomarkers and lung function in asbestos-exposed population with or without pleural and pulmonary parenchymal abnormalities.

Plasma Chemokines in Patients with Alcohol Use Disorders: Association of CCL11 (Eotaxin-1) with Psychiatric Comorbidity.

Recent studies have linked changes in peripheral chemokine concentrations to the presence of both addictive behaviors and psychiatric disorders. The present study further explore this link by analyzing the potential association of psychiatry comorbidity with alterations in the concentrations of circulating plasma chemokine in patients of both sexes diagnosed with alcohol use disorders (AUD). To this end, 85 abstinent subjects with AUD from an outpatient setting and 55 healthy subjects were evaluated for substance and mental disorders. Plasma samples were obtained to quantify chemokine concentrations [C–C motif (CC), C–X–C motif (CXC), and C–X3–C motif (CX3C) chemokines]. Abstinent AUD patients displayed a high prevalence of comorbid mental disorders (72%) and other substance use disorders (45%). Plasma concentrations of chemokines CXCL12/stromal cell-derived factor-1 (p < 0.001) and CX3CL1/fractalkine (p < 0.05) were lower in AUD patients compared to controls, whereas CCL11/eotaxin-1 concentrations were strongly decreased in female AUD patients (p < 0.001). In the alcohol group, CXCL8 concentrations were increased in patients with liver and pancreas diseases and there was a significant correlation to aspartate transaminase (r = +0.456, p < 0.001) and gamma-glutamyltransferase (r = +0.647, p < 0.001). Focusing on comorbid psychiatric disorders, we distinguish between patients with additional mental disorders (N = 61) and other substance use disorders (N = 38). Only CCL11 concentrations were found to be altered in AUD patients diagnosed with mental disorders (p < 0.01) with a strong main effect of sex. Thus, patients with mood disorders (N = 42) and/or anxiety (N = 16) had lower CCL11 concentrations than non-comorbid patients being more evident in women. The alcohol-induced alterations in circulating chemokines were also explored in preclinical models of alcohol use with male Wistar rats. Rats exposed to repeated ethanol (3 g/kg, gavage) had lower CXCL12 (p < 0.01) concentrations and higher CCL11 concentrations (p < 0.001) relative to vehicle-treated rats. Additionally, the increased CCL11 concentrations in rats exposed to ethanol were enhanced by the prior exposure to restraint stress (p < 0.01). Concordantly, acute ethanol exposure induced changes in CXCL12, CX3CL1, and CCL11 in the same direction to repeated exposure. These results clearly indicate a contribution of specific chemokines to the phenotype of AUD and a strong effect of sex, revealing a link of CCL11 to alcohol and anxiety/stress.

Plasma concentrations of CCL3 and CCL4 in the cardiac and digestive clinical forms of chronic Chagas disease

The aim of this study was to investigate the plasma levels of the CCL3 and CCL4 chemokines in patients with the cardiac and digestive clinical forms of chronic Chagas disease and in cardiac patients with and without left ventricular systolic dysfunction (LVSD). Plasma samples from 75 patients were evaluated by enzyme-linked immunosorbent assay (ELISA) to confirm infection by T. cruzi. Plasma levels of the CCL3 and CCL4 chemokines were measured using Milliplex® MAP assay (Millipore). There were no significant differences in the levels of CCL3 and CCL4 between patients with the digestive and cardiac clinical forms of Chagas disease. Moreover, no significant differences were found between patients without LVSD and those with LVSD. Higher CCL3 and CCL4 plasma levels were found in patients with LVSD compared to those with the digestive form of the disease. The CCL3 and CCL4 chemokines might not be involved in differential susceptibility to the digestive and cardiac clinical forms of chronic Chagas disease, and it seems they do not influence the development of LVSD.

Establishment and verification of detecting multiple biomarkers for ovarian cancer by suspension array technology

Objective: Establish and validation of combined detecting of CCL18, CXCL1, C1D, TM4SF1, FXR1, TIZ suspension array technology. Methods: (1)CCL18, CXCL1 monoclonal antibody and C1D, TM4SF1, FXR1, TIZ protein were coupled with polyethylene microspheres. Biotinylated CCL18, CXCL1 polyclonal antibody and sheep anti-human IgG polyclonal antibody were prepared simultaneously. The best packaged concentrations of CCL18, CXCL1 monoclonal antibody and C1D, TM4SF1, FXR1, TIZ antigens were optimized. The best packaged concentrations of CCL18, CXCL1 polyclonal antibodys and C1D, TM4SF1, FXR1, TIZ sheep anti-human IgG polyclonal antibody were optimized to establish a stable detected suspension array.(2)Sixty patients confirmed by pathological examination with ovarian cancer(ovarian cancer group)which treated in Affiliated Tumor Hospital of Guangxi Medical University, 30 patients with ovarian benign tumor(benign group)and 30 cases of healthy women(control group)were chosen between September 2003 and December 2003. Suspension array technology and ELISA method were used to detect expression of CCL18, CXCL1 antigen and C1D, TM4SF1, FXR1 and TIZ IgG autoantibody contented in 3 groups of serum, then to compare the diagnostic efficiency and diagnostic accuracy of two methods(coefficient of variation between batch and batch). Results: (1)This research successfully established stable detecting system of CCL18, CXCL1, C1D, TM4SF1, FXR1 and TIZ IgG autoantibody. The best concentration of CCL18, CXCL1 monoclonal antibody and C1D, TM4SF1, FXR1, TIZ antigen package were 8, 8, 12, 8, 4 and 8 μg/ml; the best detection of CCL18, CXCL1 biotin polyclonal antibody and C1D, TM4SF1, FXR1, TIZ sheep anti-huamne IgG polyclonal antibody were respectively 4, 2, 2, 4, 4 and 2 μg/ml.(2)Suspension array technology and ELISA method were used to detect CCL18, CXCL1 antigen and C1D, TM4SF1, FXR1, TIZ IgG autoantibody of three groups in serum were similar(P>0.05).(3)The comparison of two methods in the diagnosis of efficiency: the diagnostic accuracy of two methods were 99.2%(119/120)and 94.2%(113/120), the difference was statistically significant(P=0.031). The sensitivity of the diagnosis of ovarian cancer of two methods were 100.0%(60/60)and 93.3%(56/60), specific degrees were 100.0%(59/59)and 93.4%(57/61), positive predictive value was 100.0%(60/60)and 93.3%(56/60), negative predictive value was 98.3%(59/60)and 95.0%(57/60), the difference was statistically significant(P<0.05).(4)The detected results of CCL18, CXCL1 antigen and C1D, TM4SF1, FXR1, TIZ IgG autoantibody shown that the diagnostic accuracy of suspension array technology was superior to those of ELISA method(all P<0.05). Conclusion: The study has established the stable detection of suspension array technology, and the diagnostic efficiency and diagnostic accuracy was much better than that by ELISA.

Cerebrospinal Fluid Chemokine (C-C Motif) Ligand 2 Is an Early-Response Biomarker for Blast-Overpressure-Wave-Induced Neurotrauma in Rats.

Chemokines and their receptors are of great interest within the milieu of immune responses elicited in the central nervous system in response to trauma. Chemokine (C-C motif)) ligand 2 (CCL2), which is also known as monocyte chemotactic protein-1, has been implicated in the pathogenesis of traumatic brain injury (TBI), brain ischemia, Alzheimer's disease, and other neurodegenerative diseases. In this study, we investigated the time course of CCL2 accumulation in cerebrospinal fluid (CSF) after exposures to single and repeated blast overpressures of varied intensities along with the neuropathological changes and motor deficits resulting from these blast conditions. Significantly increased concentrations of CCL2 in CSF were evident by 1 h of blast exposure and persisted over 24 h with peak levels measured at 6 h post-injury. The increased levels of CCL2 in CSF corresponded with both the number and intensities of blast overpressure and were also commensurate with the extent of neuromotor impairment and neuropathological abnormalities resulting from these exposures. CCL2 levels in CSF and plasma were tightly correlated with levels of CCL2 messenger RNA in cerebellum, the brain region most consistently neuropathologically disrupted by blast. In view of the roles of CCL2 that have been implicated in multiple neurodegenerative disorders, it is likely that the sustained high levels of CCL2 and the increased expression of its main receptor, CCR2, in the brain after blast may similarly contribute to neurodegenerative processes after blast exposure. In addition, the markedly elevated concentration of CCL2 in CSF might be a candidate early-response biomarker for diagnosis and prognosis of blast-induced TBI.

Source proximity and meteorological effects on residential outdoor VOCs in urban areas: Results from the Houston and Los Angeles RIOPA studies

Concentrations of volatile organic compounds (VOCs) measured outside homes in Houston, TX and Los Angeles, CA were characterized by the effects of source proximity and meteorological factors. Benzene, toluene, ethylbenzene, m,p-xylene, o-xylene (BTEX), methyl tert butyl ether (MTBE), tetrachloroethylene (perchloroethylene, PCE), and carbon tetrachloride (CCl4) were examined. Multiple stepwise regression analysis converged the best-fit models with predictors from meteorological conditions and the proximity to specific point, area, and mobile sources on the residential outdoor VOC concentrations. Negative associations of wind speed with concentrations demonstrated the effect of dilution by high wind speed. Atmospheric stability increase was associated with concentration increase. Petrochemical source proximity was a significant predictor for BTEX and MTBE concentrations in Houston. Ethylbenzene and xylene source proximity was a significant predictor in Los Angeles. Close proximity to area sources such as scrap metal recycling or dry cleaning facilities increased the MTBE, PCE, and CCl4 concentrations in Houston and Los Angeles. Models for ethylbenzene, m,p-xylene, and MTBE in Houston, and benzene in Los Angeles explained that for the median values of the meteorological factors, homes closest to influential highways would have concentrations that were 1.7–2.2 fold higher than those furthest from these mobile emission sources. If the median distance to sources were used in the models, the VOC concentrations varied 1.7 to 6.6 fold as the meteorological conditions varied over the observed range. These results highlight that each urban area is unique and localized sources need to be carefully evaluated to understand potential contributions to VOC air concentrations near residences, which influence baseline indoor air concentrations and personal exposures. Results of this study could assist in the appropriate design of monitoring networks for community-level sampling. They may also improve the accuracy of exposure models linking emission sources with estimated pollutant concentrations at the residential level.

Inhibited early immunologic response is associated with hypertrophic scarring.

This study aimed to examine changes in the inflammatory response in early hypertrophic compared to normal wound healing. The immune system is thought to be involved in hypertrophic scar formation. However, the exact mechanism and time of onset of the derailment remain unknown. In a prospective observational study, skin biopsies were taken directly postwounding and 3hours later from patients who had elective cardiothoracic surgery. The skin biopsies were analysed for mRNA, proteins and cells involved in the early inflammatory phase of wound healing. The endpoint was scar outcome (hypertrophic (HTS) or normal (NTS)) at one year after surgery. There were significant differences between the NTS and HTS groups regarding the fold changes of mRNA expression of P-selectin during surgery. Postoperative skin concentrations of inflammatory proteins IL-6, IL-8 and CCL2 were significantly lower in the HTS compared to the NTS group. Also, a trend of higher pre-operative M2 macrophage numbers was observed in the HTS group. Neutrophil numbers increased equally during surgery in both groups. The increase of P-selectin mRNA in hypertrophic wound healing could affect leucocyte migration. The decreased concentrations of inflammatory proteins in hypertrophic wound healing indicate a reduced inflammatory response, which has consequences for the treatment of hypertrophic scarring during the early inflammatory phase. In a conclusion, alterations of wound healing associated with hypertrophic scarring are visible as early as 3hours postwounding and include a reduced rather than increased inflammatory protein response.

Association of semen cytokines with reactive oxygen species and histone transition abnormalities.

PurposeThe aim of this study is to investigate the relationships among reactive oxygen species (ROS) elevation, histone transition, and seminal cytokine concentrations.MethodsTotal levels of ROS in semen samples from 6560 men were measured. From this sample, 118 cases with high ROS and 106 controls were recruited. Basic semen parameters and histone-to-protamine ratios were analyzed, 400 semen cytokine and receptor alterations were assayed by protein chip, and finally 18 cytokines were validated in each sample using a Bio-Plex Cytokine assay.ResultsThe results showed that the seminal ROS concentration was associated with abnormalities in the sperm histone transition. Compared with controls, 93 cytokines had significant alterations in the high ROS cases, with 14 of them further verified in individual samples. The concentrations of CXCL5, CXCL16, CXCL8, IL-1b, IL-10, CSF3, CCL3, and TNF-α were significantly correlated with the histone transition ratio. In addition, IL-16 showed significantly different concentrations in controls, normal semen with high ROS levels, and abnormal semen with high ROS levels.ConclusionsSemen ROS are associated with abnormalities in sperm histone transition. CXCL5, CXCL8, IL-16, CCL8, CCL22, CCL20, CXCL16, IL-1B, IL-6, IL-7, IL-10, CSF3, CCL3, CCL4, and TNF-α all have elevated concentrations in semen with high ROS levels. These data might help to explain the mechanisms behind the increase in the levels of ROS and seminal cytokines and their relationship with defective spermatogenesis.

Baseline cytokine profiling identifies novel risk factors for invasive fungal disease among haematology patients undergoing intensive chemotherapy or haematopoietic stem cell transplantation

Invasive fungal disease (IFD) is a disease of immunocompromised hosts. Cytokines are important mediators of innate and adaptive immune system. The aim of this study was to identify cytokine profiles that correlate with increased risk of IFD. We prospectively enrolled 172 adult haematology patients undergoing intensive chemotherapy, immunosuppressive therapy, and haematopoietic stem cell transplantation. Pro-inflammatory cytokine profiling using 30-plex Luminex assay was performed at baseline and during treatment. Nine single nucleotide polymorphisms (TLR1, TLR2, TLR3, TLR4.1, TLR4.2, TLR6, CLEC7A, CARD9, and INFG) were investigated among transplant recipients and donors. The incidence of IFD in this cohort was 16.9% (29/172). Median baseline serum concentrations of IL-15, IL-2R, CCL2, and MIP-1α were significantly higher whilst IL-4 was lower in patients with proven/probable IFD compared to those with no evidence of IFD. Baseline high IL-2R and CCL2 were associated with increased risk of IFD in the multivariate analysis (adjusted hazard ratio 2.3 [95% CI 1.1-5.1; P=0.037], and hazard ratio 2.7 [95% CI 1.2-6.1; P=0.016], respectively). However, these differences were not significant in follow up measurements. Similarly, no significant independent prognostic value was associated with baseline cytokine profile. High baseline IL-2R and CCL2 concentrations were independent indicators of the risk of developing IFD and could be used to identify patients for enhanced prophylaxis and early antifungal therapy.

Plasma levels of chemokine ligand 20 and chemokine receptor 6 in patients with sepsis: A case control study.

Chemokine ligand 20 (CCL20) is a chemokine released by mainly liver and blood leucocytes. Particularly under pro-inflammatory circumstances it triggers chemotaxis of lymphocytes and dendritic cells via activating receptor chemokine receptor 6 (CCR6) that is specific to it. In experimental sepsis models, the chemokine-receptor pair has been identified as a potential pathophysiological axis affecting mortality. Measurement of CCL20 and CCR6 plasma levels in septic patients compared with postsurgical, nonseptic patients. Case control study. Surgical ICUs of the Department of Anaesthesiology, General Hospital of Vienna, Vienna, Austria. Plasma levels were measured in 46 patients with sepsis, severe sepsis or septic shock according to current American College of Chest Physicians/Society of Critical Care Medicine criteria at the day of sepsis onset. Plasma levels in 36 postsurgical controls without sepsis admitted to the ICU were investigated. Plasma concentrations were determined by using commercially available ELISA kits. Data are given as median and interquartile range (IQR). CCL20 and CCR6 plasma levels. CCL20 plasma levels were significantly increased in the sepsis group: 220.9 pg ml (IQR, 72.8 to 540.1) compared with the ICU controls: 37.0 pg ml (IQR 6.5 to 83.6) (P < 0.0001). Significantly elevated CCR6 levels were found in the sepsis group: 2.47 ng ml (IQR 0.92 to 5.54) compared with the controls: 0.59 ng ml (IQR 0.17 to 1.48) (P < 0.0001). Both CCL20 and CCR6 correlated with the maximum sequential organ failure assessment score (CCL20: P < 0.0001, CCR6: P < 0.0001). Length of ICU admission depended significantly on the logarithm of CCR6 (P = 0.008) and sequential organ failure assessment maximum (P < 0.0001). There were early increased plasma concentrations of CCL20 and CCR6 in patients with sepsis. CCL20 and CCR6 correlate with severity of illness in ICU patients. Levels of CCR6 predicted the length of patients' admission.

Effect of aspirin on models of retinal pigment epithelium pathology.

To characterize the effect of aspirin (ASA) in mouse models of choroidal neovascularization (CNV) and retinal degeneration. In vivo: Male C57BL/6 mice were given ASA in food or regular rodent diet. CNV was induced by argon laser photocoagulation. Subretinal injections of polyethylene glycol 400 (PEG-400) were administered to induce retinal degeneration. CNV size, laser spot area and mean intensity of VEGF in the laser injured zones were measured. In the PEG injected eyes the thickness of retinal pigment epithelium (RPE) and choroid was measured. In vitro: Human ARPE-19 cells were treated with 0.5 or 2.0 mM/L of ASA for 72 h. ELISA was used to measure the concentration of VEGF and CCL-2 in the supernatants. Additionally, damaged RPE monolayer was treated with ASA (0.5 or 2.0 mM/L) and vehicle separately. Size of damaged area was measured. ELISA was used to measure secretion of VEGF-A and CCL-2 by damaged cells after 24 h. No statistically significant effect of ASA on CNV size, laser spot size or VEGF expression was noted in CNV model. In the PEG model, ASA did not have any effect on RPE and choroid thickness; however, a significant increase in RPE atrophy was observed (P = 0.02 + 38%). In addition, ASA had a significant effect on the ability of the RPE cells to regenerate and become confluent after mechanical damage. ASA at doses consumed clinically for various medical causes may not worsen CNV in human subjects. However, ASA may increase RPE atrophy when consumed over long periods of time.

Pro-inflammatory chemokines CCL2, chemerin, IP-10 and RANTES in human serum during an oral lipid tolerance test

BackgroundThere is a strong coincidence of obesity and a chronic state of modest inflammation. Secretion of pro-inflammatory cytokines from adipocytes and immune cells represents a key mechanism in this process and is affected by fatty acids. Material and methodsA study cohort of 100 overnight fasted healthy volunteers underwent an oral lipid tolerance test (OLTT) by ingestion of 160ml of a protein- and sugar-free lipid emulsion of defined composition. Venal blood was drawn at 0h (fasting) and at 2, 4, and 6h after lipid ingestion. Subjects were characterized by anthropometric and standard laboratory parameters. Serum concentrations of CCL2, IP-10, chemerin, and RANTES were measured by enzyme-linked immunosorbent assay (ELISA). Murine 3T3-L1 adipocytes were stimulated with free fatty acids (FA) and with sex steroids and concentrations of CCL2 and chemerin in cell culture supernatants were measured by ELISA. ResultsA significant reduction of circulating CCL2, IP-10, and chemerin concentrations was observed as a consequence of triglyceride ingestion whereas RANTES levels were increased. CCL2 serum concentrations were positively correlated with resistin and visfatin levels and with LDL/HDL ratio and negatively with adiponectin. There were significant differences in chemerin and RANTES serum concentrations in female and male subjects. CCL2 secretion from 3T3-L1 adipocytes was inhibited by treatment with linoleic (LA) and oleic acid (OA) whereas chemerin secretion was induced. Chemerin release from 3T3-L1 adipocytes was inhibited by testosterone. ConclusionsOral lipid loading is linked to reduced circulating pro-inflammatory chemokines CCL2, IP-10, and chemerin and to increased RANTES levels, suggesting that dietary lipids affect immune function.

Analgesic effects evoked by a CCR2 antagonist or an anti-CCL2 antibody in inflamed mice.

Chemokine CCL2, also known as monocyte chemoattractant protein-1 (MCP-1), is a molecule that in addition to its well-established role in chemotaxis can also act as nociceptor sensitizer. The upregulation of this chemokine in inflamed tissues could suggest its involvement in inflammatory hypernociception. Thus, we have measured CCL2 levels in mice with acute or chronic inflammation due to the intraplantar (i.pl.) injection of carrageenan or complete Freund's adjuvant (CFA), respectively, and we have studied whether inflammatory hyperalgesia or allodynia could be attenuated by blocking CCR2 receptors or neutralizing CCL2 with an anti-CCL2 antibody. A remarkable increase in CCL2 concentration was detected by ELISA in paw homogenates coming from carrageenan- or CFA-inflamed mice, being its expression mainly localized in macrophages, as shown by immunohistochemical assays. The s.c. (0.3-3 mg/kg) or i.pl. (0.3-3 μg) administration of the CCR2 antagonist, RS 504393, dose dependently inhibited thermal hyperalgesia measured in acutely or chronically inflamed mice, whereas s.c. administration of this drug did not reduce inflammatory mechanical allodynia. Furthermore, the inhibition of inflammatory hyperalgesia after the administration of an anti-CCL2 antibody (0.1-1 μg; i.pl.) suggests that CCL2 could be the endogenous chemokine responsible for CCR2-mediated hyperalgesic effects. Besides, the acute administration of the highest antihyperalgesic dose of RS 504393 assayed did not reduce paw tumefaction or modify the presence of inflammatory cells. These results indicate that the blockade of the CCL2/CCR2 system can counteract inflammatory hyperalgesia, being this antinociceptive effect unrelated to a decrease in the inflammatory reaction.