Interleukin-10 containing normal human serum inhibits granzyme B release but not perforin release from alloreactive and EBV-specific T cell clones

Interleukin-10 (IL-10), also known as cytokine synthesis inhibitory factor, has pleiotropic effects in immunoregulation and inflammation. It is capable of inhibiting synthesis of pro-inflammatory cytokines like interferon γ (IFNγ), IL-2, IL-3, tumor necrosis factor α(TNFα) and granulocyte macrophage colony stimulating factor (GM-CSF) made by cells such as macrophages and T helper Type 1 cells. We observed that normal human serum, derived from a healthy individual but containing large amounts of IL-10 (arbitrarily designated as "IL-10 serum"), inhibited cytotoxic activity and interfered with granzyme B release from alloreactive cytotoxic T cell (CTL) clones _in vitro_, but did not affect perforin release. The addition of normal human serum containing high levels of anti-IL-10 IgG (arbitrarily designated as "anti-IL-10 IgG serum") neutralized the inhibitory effects of IL-10 serum. Moreover, we have identified that cytotoxic activity and granzyme B release from an Epstein-Barr virus (EBV)-specific CTL clone was similarly inhibited in the presence of IL-10 serum, while perforin release was unaffected. Anti-IL-10 IgG serum also appeared to neutralize the inhibitory effect of IL-10 serum on an EBV-specific CTL clone

Effects of Creosote-Contaminated Sediment Extracts on Mouse Macrophage Physiology and Function

The southern branch of the Elizabeth River, at Portsmouth Virginia is one of the most polluted systems in North America. This harbor estuary system is also home to the Atlantic Wood Superfund site, which is heavily contaminated with creosote from the Atlantic Wood (AW) preservative company that ceased production in the later 1990s. Creosote is a mixture of polyaromatic hydrocarbons (PAHs), metals, and numerous aliphatic hydrocarbons, and well known to be carcinogenic. The toxicity of sediments and pore waters from the AW site has been studied extensively using Fundulus heteroclitus, also known as the mummichog, or Atlantic killifish. Most adult killifish from AW have hepatic lesions, ranging from focal hyperplasia to highly malignant tumors. As embryos and juveniles, these fish are resistant to developmental toxicity of PAHs relative to a reference population on King\u27s Creek (KC) located near Mobjack Bay on the York River. Moreover, the resistance of AW fish to developmental toxicity is associated with altered aryl-hydrocarbon receptor (AhR) functions and signaling, leading to recalcitrance in CYP1A induction by PAHs. To determine the potential of AW sediment pore water to modulate immune functions in mammals, this study used the mouse macrophage cell line RAW264.7 as a model for pro-inflammatory functions. Compared to KC sediment pore water, AW pore water induces more iNOS and COX-2 protein expression, nitric oxide and IL-6 secretion, in RAW264.7 cells. This enhanced pro-inflammatory property of AW extracts is due, in large part, to high levels of endotoxin, as polymyxin-B ameliorates this property. Moreover, AW sediment extracts have quantitatively higher endotoxin as measures by the limulus lysate assay

Relationship between antipyretic effects and cytokine levels in uncomplicated falciparum malaria during different treatment regimes

We have previously shown that both chloroquine and paracetamol (acetaminophen) have antipyretic activity during treatment of acute uncomplicated Plasmodium falciparum malaria in children 1-4 years old. Here, we studied if this effect was accompanied by changes in plasma cytokine levels. The 104 children were treated with either chloroquine or sulfadoxine/pyrimethamine (SP) alone, SP + chloroquine or SP + paracetamol for 4 days. Cytokine levels were determined days 0, 2 and 3, body temperature every sixth hour until 72 h and parasitemia once daily for 4 days. At admission, body temperature correlated with levels of IL-10, IFN-γ and IL-6, and parasitemia correlated with IL-10 and IL-6. Except for TNF-α and IL-1β, where no significant effect was found, all cytokine levels (IL-10, IFN-γ, IL-6, IL-12, IL-13, IL-18 and IL-4) decreased up to day 2 (p \u3c 0.05). IL-6 levels continued to fall from days 2 to 3 (p \u3c 0.05), whereas increased levels were found for several cytokines (IL-12, IL-13, IL-18 and IL-1β) (p \u3c 0.05). The antipyretic effects of chloroquine and paracetamol could not be related to any specific changes in the evaluated cytokine production or in Th1/Th2 or inflammatory/anti-inflammatory cytokine ratios. Alternative mechanisms for antipyretic effects and associations between fever and cytokine levels during uncomplicated P. falciparum malaria are therefore discussed. © 2006 Elsevier B.V. All rights reserved

Differences in leukocyte profile, gene expression, and metabolite status of dairy cows with or without sole ulcers

peer-reviewedSole ulcers are one of the most severe pathologies causing lameness in dairy cows and are associated with abnormal behavior and impaired production performance. However, little is known about how or whether lameness caused by sole ulcers affects the cow systemically. This study compared hematology profile, leukocyte gene expression, and physiological responses [metabolite, cortisol, the endogenous steroid hormone dehydroepiandrosterone (DHEA), and haptoglobin concentrations] of cows with sole ulcers and healthy cows. Twelve clinically lame cows (lame) were identified as having at least one sole ulcer and no other disorder, and matched with a cow that had good locomotion and no disorders (sound), using days in milk, liveweight, body condition score, and diet. Blood samples were taken from all 24 cows within 24 h of sole ulcer diagnosis. Leukocyte counts were obtained using an automated cell counter, cortisol and DHEA concentration by ELISA, and plasma haptoglobin, urea, total protein, creatine kinase, and glucose were analyzed on an Olympus analyzer. Expression of 16 genes associated with lameness or stress were estimated using reverse transcription-PCR. Data were analyzed using the MIXED procedure in SAS software (version 9.3; SAS Institute Inc., Cary, NC). Lame cows had a higher neutrophil percentage, a numerically lower lymphocyte percentage, and tended to have a higher neutrophil:lymphocyte ratio than sound cows. Serum cortisol and DHEA concentrations were higher in lame than in sound cows. Lame cows also tended to have higher haptoglobin and glucose levels than sound, as well as higher protein yet lower urea levels. Sound cows tended to have higher relative expression of the gene coding for colony-stimulating factor 2 than lame, but in all other cases where differences were detected in cytokine gene expression (IL-1α, IL-1β, CXCL8, and IL-10), relative gene expression in sound cows tended to be, or was, lower than in lame. Relative expression of MMP-13, GR-α, Fas, haptoglobin, and CD62L were, or tended to be, higher in lame than sound cows. A high neutrophil:lymphocyte ratio in combination with higher cortisol levels in cows with ulcers is indicative of physiological stress. Moreover, increased DHEA and a higher cortisol:DHEA ratio, as well as a tendency for higher haptoglobin levels and increased haptoglobin mRNA expression, are indicative of systemic inflammation. Increased cytokine mRNA expression indicates activation of the immune system compared with healthy cows. Increased expression of MMP-13 mRNA has been found in cows with impaired locomotion and thus could be implicated in development of claw horn disorders.This study was funded by a Marie Curie Intra-European Fellowship (FP7-People 2009-IEF; grant agreement number: 252611) to Keelin O'Driscoll

CD69 Signaling in Eosinophils Induces IL-10 Production and Apoptosis via the Erk1/2 and JNK Pathways, Respectively

INTRODUCTION: Eosinophils contribute to the pathogenesis of allergic diseases, including asthma, allergic rhinitis, and atopic dermatitis. We previously reported that human tissue eosinophils have high CD69 expression compared to blood eosinophils, and its expression is correlated with disease severity and the number of infiltrated eosinophils. However, biological CD69 signaling activity in eosinophils remains unclear. METHODS: CD69 expression on lung tissue eosinophils obtained from mice with ovalbumin-induced asthma was measured using flow cytometry. CD69 crosslinking was performed on eosinophils purified from the spleen of IL-5 transgenic mice to investigate CD69 signaling and its function in eosinophils. Then, qPCR, Western blot, enzyme-linked immunosorbent assay, and survival assay results were analyzed. RESULTS: Surface CD69 expression on lung tissue eosinophils in the asthma mice model was 2.91% ± 0.76%, whereas no expression was detected in the healthy group. CD69-expressed eosinophils intrinsically have an upregulation of IL-10 mRNA expression. Moreover, CD69 crosslinking induced further pronounced IL-10 production and apoptosis; these responses were mediated via the Erk1/2 and JNK pathways, respectively. CONCLUSIONS: Our results suggested that CD6

Association study of Interleukin 10 gene polymorphisms in Iraqi patients with multiple sclerosis

Multiple sclerosis (MS) is a type of autoimmune disease where immune cell attacks our cells mistakenly; its severity is measured by expanded disease status scale (EDSS). The study aims the investigation of −1082 polymorphism in interleukin 10 (IL-10) as one of the etiologies that develops the disease. This is a case-control study that allele-specific polymerase chain reaction (AS-PCR) were provided to compare 100 relapsing-remitting MS (RRMS) patients, which fulfills McDonald criteria with 100 healthy controls depending on the −1082 (G/A) polymorphism of the gene encoding IL-10. The A allele frequency of IL-10 gene has been considerably less in MS patients compare to healthy control (60.50 Vs. 81%). Genotype distributions of the single nucleotide polymorphism (SNP) -1082 fulfills Hardy-Weinberg equilibrium in cases (P = 0.155) but it doesn't in controls (P < 0.0001). In MS patients, Heterozygous (GA) genotypes were non-significantly associated with MS (OR = 0.834,95% CI = 0.6890 to 1.29, P 0.706) but homozygous (AA) were significantly associated with this condition (OR = 3.420, 95% CI = 1.450 to 8.065, P = 0.0037). To conclude, the genotype distribution of −1082 (G/A) polymorphism has been showed a significant difference in the case/control study recruited in Erbil province-Iraq, and EDSS is significantly higher in A allele's carrier genotypes. There was non-significance association AA genotypes and duration of the disease

Elevated Systemic IL-10 Levels Indicate Immunodepression Leading to Nosocomial Infections after Aneurysmal Subarachnoid Hemorrhage (SAH) in Patients

Background: Aneurysmal subarachnoid hemorrhage (SAH) is a highly complex disease with very high mortality and morbidity. About one-third of SAH patients suffer from systemic infections, predominantly pneumonia, that can contribute to excess mortality after SAH. Immunodepression is probably the most important mechanism leading to infections. Interleukin-10 (IL-10) is a master regulator of immunodepression, but it is still not clear if systemic IL-10 levels contribute to immunodepression, occurrence of infections and clinical outcome after SAH. Methods: This explorative study included 76 patients with SAH admitted to our neurointensive care unit within 24 h after ictus. A group of 24 patients without any known intracranial pathology were included as controls. Peripheral venous blood was withdrawn on day 1 and day 7 after SAH. Serum was isolated by centrifugation and stored at −80 °C until analysis. Serum IL-10 levels were determined by enzyme-linked immunoassay (ELISA). Patient characteristics, post-SAH complications and clinical outcome at discharge were retrieved from patients’ record files. Results: Serum IL-10 levels were significantly higher on day 1 and day 7 in SAH patients compared to controls. Serum IL-10 levels were significantly higher on day 7 in patients who developed any kind of infection, cerebral vasospasm (CVS) or chronic hydrocephalus. Serum IL-10 levels were significantly higher in SAH patients discharged with poor clinical outcome (modified Rankin Scale (mRS) 3–6 or Glasgow Outcome Scale (GOS) 1–3). Conclusion: Serum IL-10 might be an additional useful parameter along with other biomarkers to predict post-SAH infections

Analysis of the function of IL-10 in chickens using specific neutralising antibodies and a sensitive capture ELISA

AbstractIn mammals, the inducible cytokine interleukin 10 is a feedback negative regulator of inflammation. To determine the extent to which this function is conserved in birds, recombinant chicken IL-10 was expressed as a secreted human Ig Fc fusion protein (chIL-10-Fc) and used to immunise mice. Five monoclonal antibodies (mAb) which specifically recognise chicken IL-10 were generated and characterised. Two capture ELISA assays were developed which detected native chIL-10 secreted from chicken bone marrow-derived macrophages (chBMMs) stimulated with lipopolysaccharide (LPS). Three of the mAbs detected intracellular IL-10. This was detected in only a subset of the same LPS-stimulated chBMMs. The ELISA assay also detected massive increases in circulating IL-10 in chickens challenged with the coccidial parasite, Eimeria tenella. The same mAbs neutralised the bioactivity of recombinant chIL-10. The role of IL-10 in feedback control was tested in vitro. The neutralising antibodies prevented IL-10-induced inhibition of IFN-γ synthesis by mitogen-activated lymphocytes and increased nitric oxide production in LPS-stimulated chBMMs. The results confirm that IL-10 is an inducible feedback regulator of immune response in chickens, and could be the target for improved vaccine efficacy or breeding strategies