Release Of Pro-inflammatory Cytokines IL-6 Research Articles

Endoplasmic Reticulum Stress Differently Modulates the Release of IL-6 and IL-8 Cytokines in Human Glial Cells.

Endoplasmic reticulum (ER) stress is a significant player in the pathophysiology of various neurodegenerative and neuropsychiatric disorders. Despite the established link between ER stress and inflammatory pathways, there remains a need for deeper exploration of the specific cellular mechanisms underlying ER stress-mediated neuroinflammation. This study aimed to investigate how the severity of ER stress (triggered by different concentrations of tunicamycin) can impact the release of proinflammatory cytokines IL-6 and IL-8 from astrocytes and microglia, comparing the effects with those induced by well-known immunostimulants-tumor necrosis factor alpha (TNF-α) or lipopolysaccharide (LPS). Mild ER stress has a distinct effect on the cytokine release compared to more intense stress levels, i.e., diminished IL-6 production was accompanied by an increase in IL-8 level, which was significantly more pronounced in astrocytes than in microglia. On the contrary, prolonged or more severe ER stress induced inflammation in glial cells, leading to a time- and concentration-dependent buildup of proinflammatory IL-6, but unlike inflammatory agents, an ER stress inducer diminished IL-8 secretions by glial cells. The differences could hold importance in identifying ER stress markers as potential drug targets for the treatment of neurodegenerative diseases or mood disorders, yet this requires confirmation in more complex animal studies.

CD4+ T cell-associated cytokines induce a chronic pro-inflammatory phenotype in induced pluripotent stem cell-derived midbrain astrocytes.

Astrocytes are mediators of homeostasis but contribute to neuroinflammation in Parkinson's disease (PD). Mounting evidence suggests involvement of peripheral immune cells in PD pathogenesis. Therefore, this study aimed to determine the potential role of peripheral immune secreted cytokines in modulating midbrain astrocyte reactivity. Human iPSC-derived midbrain astrocytes were exposed to 5% and 10% CD4+ T cell conditioned media (CD4CM) for 24 h, 72 h, and 7 days to assess chronic exposure. Additionally, astrocytes were exposed to the Th17 cell cytokine, IL-17A (10 ng/mL), alone and in combination with TNF-α (0.3 ng/mL) to assess potential synergistic effects of both cytokines at 24 h, 72 h, and 7 days. CD4CM induced acute and chronic alterations in midbrain astrocytes. Increased NFκB translocation to the nucleus, increased expression of the pro-inflammatory genes, IL-1β, CXCL10 at 24 h, C3, LCN2, IL-6 at 24 and 48 h, as well as an increase in their release of pro-inflammatory cytokines IL-6 and CXCL10 at both these time points were observed. A synergistic response to the combination of IL-17A and TNF-α on increasing inflammatory gene expression and cytokine release occurred. IL-17A and TNF-α increased intensity of S100β at 24 h, decreased nuclear area and increased circularity of astrocytes at 72 h. A synergistic effect on γH2AX intensity at 72 h and an increase in LDH release at 7 days was observed. Our results demonstrate that IL-17A and TNF-α act synergistically, enhancing midbrain astrocyte reactivity to a similar degree as CD4CM. This highlights the importance of the peripheral immune secreted cytokines in increasing the reactivity status of midbrain astrocytes, implicating their role in PD.

Loss of stearoyl-CoA desaturase 2 disrupts inflammatory response in macrophages.

Macrophages are innate immune cells that patrol tissues and are the first responders to detect infection. They orchestrate the host immune response in eliminating invading pathogens and the subsequent transition from inflammation to tissue repair. Macrophage dysfunction contributes to age-related pathologies, including low-grade inflammation in advanced age that is termed "inflammaging." Our laboratory has previously identified that macrophage expression of a fatty acid desaturase, stearoyl-CoA desaturase 2 (SCD2), declines with age. Herein, we delineate the precise cellular effects of SCD2 deficiency in murine macrophages. We found that deletion of Scd2 from macrophages dysregulated basal and bacterial lipopolysaccharide (LPS)-stimulated transcription of numerous inflammation-associated genes. Specifically, deletion of Scd2 from macrophages decreased basal and LPS-induced expression of Il1b transcript that corresponded to decreased production of precursor IL1B protein and release of mature IL1B. Furthermore, we identified disruptions in autophagy and depletion of unsaturated cardiolipins in SCD2-deficient macrophages. To assess the functional relevance of SCD2 in the macrophage response to infection, we challenged SCD2-deficient macrophages with uropathogenic Escherichia coli and found that there was impaired clearance of intracellular bacteria. This increased burden of intracellular bacteria was accompanied by increased release of pro-inflammatory cytokines IL6 and TNF but decreased IL1B. Taken together, these results indicate that macrophage expression of Scd2 is necessary for maintaining the macrophage response to inflammatory stimuli. This link between fatty acid metabolism and fundamental macrophage effector functions may potentially be relevant to diverse age-related pathologies. IMPORTANCE Macrophages are immune cells that respond to infection, but their dysfunction is implicated in many age-related diseases. Recent evidence showed that macrophage expression of a fatty acid enzyme, stearoyl-CoA desaturase 2, declines in aged organisms. In this work, we characterize the effects when stearoyl-CoA desaturase 2 is deficient in macrophages. We identify aspects of the macrophage inflammatory response to infection that may be affected when expression of a key fatty acid enzyme is decreased, and these findings may provide cellular insight into how macrophages contribute to age-related diseases.

Metabolic profiling of parthenolide: Characterized by bioactive organosulfur compounds involving in hydrogen sulfide regulation

Parthenolide (PA) shows promising therapeutic effect on ulcerative colitis and colorectal cancer, and can ameliorate colon inflammation in a gut microbiota-dependent manner. In the present study, an effective UHPLC-QTOF-MS method aided by rat intestinal microbiota biotransformation was applied to characterize 34 metabolites (including a series of characteristic R-Sn-R type organosulfur compounds) of PA for the first time. Noteworthy, organosulfur metabolites PA-S2-PA and PA-S3-PA could react with cysteine to generate H2S via persulfide/polysulfide intermediates and significantly inhibit colonic cancer HT-29 and HCT116 cells viability and pro-inflammatory cytokine IL-6 release. These results indicated that PA and its organosulfur metabolites involved in H2S regulation in intestinal tract, which should be beneficial for a more in-depth understanding of the action mechanism of PA.

Molecular Mechanisms of the Anti-Inflammatory Effects of Epigallocatechin 3-Gallate (EGCG) in LPS-Activated BV-2 Microglia Cells.

Chronic neuroinflammation is associated with many neurodegenerative diseases, such as Alzheimer's. Microglia are the brain's primary immune cells, and when activated, they release various proinflammatory cytokines. Several natural compounds with anti-inflammatory and antioxidant properties, such as epigallocatechin 3-gallate (EGCG), may provide a promising strategy for inflammation-related neurodegenerative diseases involving activated microglia cells. The objective of the current study was to examine the molecular targets underlying the anti-inflammatory effects of EGCG in activated microglia cells. BV-2 microglia cells were grown, stimulated, and treated with EGCG. Cytotoxicity and nitric oxide (NO) production were evaluated. Immunoassay, PCR array, and WES™ Technology were utilized to evaluate inflammatory, neuroprotective modulators as well as signaling pathways involved in the mechanistic action of neuroinflammation. Our findings showed that EGCG significantly inhibited proinflammatory mediator NO production in LPS-stimulated BV-2 microglia cells. In addition, ELISA analysis revealed that EGCG significantly decreases the release of proinflammatory cytokine IL-6 while it increases the release of TNF-α. PCR array analysis showed that EGCG downregulated MIF, CCL-2, and CSF2. It also upregulated IL-3, IL-11, and TNFS10. Furthermore, the analysis of inflammatory signaling pathways showed that EGCG significantly downregulated mRNA expression of mTOR, NF-κB2, STAT1, Akt3, CCL5, and SMAD3 while significantly upregulating the expression of mRNA of Ins2, Pld2, A20/TNFAIP3, and GAB1. Additionally, EGCG reduced the relative protein expression of NF-κB2, mTOR, and Akt3. These findings suggest that EGCG may be used for its anti-inflammatory effects to prevent neurodegenerative diseases.

MTADV 5-MER peptide suppresses chronic inflammations as well as autoimmune pathologies and unveils a new potential target-Serum Amyloid A

Despite the existence of potent anti-inflammatory biological drugs e.g., anti-TNF and anti IL-6 receptor antibodies, for treating chronic inflammatory and autoimmune diseases, these are costly and not specific. Cheaper oral available drugs remain an unmet need. Expression of the acute phase protein Serum Amyloid A (SAA) is dependent on release of pro-inflammatory cytokines IL-1, IL-6 and TNF-α during inflammation. Conversely, SAA induces pro-inflammatory cytokine secretion, including Th17, leading to a pathogenic vicious cycle and chronic inflammation. 5- MER peptide (5-MP) MTADV (methionine-threonine-alanine-aspartic acid-valine), also called Amilo-5MER, was originally derived from a sequence of a pro-inflammatory CD44 variant isolated from synovial fluid of a Rheumatoid Arthritis (RA) patient. This human peptide displays an efficient anti-inflammatory effects to ameliorate pathology and clinical symptoms in mouse models of RA, Inflammatory Bowel Disease (IBD) and Multiple Sclerosis (MS). Bioinformatics and qRT-PCR revealed that 5-MP, administrated to encephalomyelytic mice, up-regulates genes contributing to chronic inflammation resistance. Mass spectrometry of proteins that were pulled down from an RA synovial cell extract with biotinylated 5-MP, showed that it binds SAA. 5-MP disrupted SAA assembly, which is correlated with its pro-inflammatory activity. The peptide MTADV (but not scrambled TMVAD) significantly inhibited the release of pro-inflammatory cytokines IL-6 and IL-1β from SAA-activated human fibroblasts, THP-1 monocytes and peripheral blood mononuclear cells. 5-MP suppresses the pro-inflammatory IL-6 release from SAA-activated cells, but not from non-activated cells. 5-MP could not display therapeutic activity in rats, which are SAA deficient, but does inhibit inflammations in animal models of IBD and MS, both are SAA-dependent, as shown by others in SAA knockout mice. In conclusion, 5-MP suppresses chronic inflammation in animal models of RA, IBD and MS, which are SAA-dependent, but not in animal models, which are SAA-independent.

Lipocalin 2 induces neuroinflammation and blood-brain barrier dysfunction through liver-brain axis in murine model of nonalcoholic steatohepatitis

BackgroundRecent clinical and basic research implicated a strong correlation between NAFLD/NASH phenotypes with ectopic manifestations including neuroinflammation and neurodegeneration, but the mediators and critical pathways involved are not well understood. Lipocalin 2 (Lcn2) is one of the important mediators exclusively produced in the liver and circulation during NASH pathology.MethodsUsing murine model of NASH, we studied the role of Lcn2 as a potent mediator of neuroinflammation and neurodegeneration in NASH pathology via the liver-brain axis.ResultsResults showed that high circulatory Lcn2 activated 24p3R (Lipocalin2 receptor) in the brain and induced the release of high mobility group box 1 (HMGB1) preferably from brain cells. Released HMGB1 acted as a preferential ligand to toll-like receptor 4 (TLR4) and induced oxidative stress by activation of NOX-2 signaling involving activated p65 protein of the NF-κB complex. Further, the HMGB1-derived downstream signaling cascade activated NLRP3 inflammasome and release of proinflammatory cytokines IL-6 and IL-1β from brain cells. In addition, to advance our present understanding, in vitro studies were performed in primary brain endothelial cells where results showed high circulatory Lcn2 influenced HMGB1 secretion. Mechanistically, we also showed that elevated Lcn2 level in underlying NASH might be a likely cause for induction of blood-brain barrier dysfunction since the adipokine decreased the expression of tight junction protein Claudin 5 and caused subsequent elevation of pro-inflammatory cytokines IL-6 and IL-1β.ConclusionIn conclusion, the NASH-induced brain pathology might be because of increased Lcn2-induced release of HMGB1 and accompanying neuroinflammation.

P0700FLAVONOID FISETIN ALLEVIETES RENAL DYSFUNCTION IN A SEVERE MOUSE MODEL OF HYPERURECEMIC NEPHROPATHY

Abstract Background and Aims Flavonoid Fisetin is a natural product that has a wide distribution in many fruits and vegetables and exerts anti-cancer, anti-oxidant, anti-inflammatory and anti-mitotic effects. The renal protective effects of fisetin have been confirmed in models of diabetic nephropathy, cisplatin-induced AKI, ischemia reperfusion kidney injury and lupus nephritis. The current study instigates the protective effect of fisetin in a severe mouse model of hyperuricemic nephropathy (HN) and explores its underlying mechanisms. Method Twenty-four C57BL6 mice were randomly divided into four groups: Control, Model, Allopurinol and fisetin. HN was established by oral administration of mixture of adenine (160 mg/kg/d)/potassium oxonate (2400 mg/kg/d) every other day for 28 days. Fisetin (100 mg/kg/d) and allopurinol (10 mg/kg/d) was given by gavage simultaneously. At the end of the treatment, serum uric acid, kidney functions (serum creatinine and BUN) kidney pathological changes, as well as underlying mechanisms were investigated by biochemical assay, histopathological score, RT-PCR, and western blotting analysis. Results Oral administration of fisetin to HN mice lowered serum uric acid and improved renal functions. Renal fibrosis and inflammation in HN mice were attenuated by fisetin treatment. fisetin alleviated extracellular matrix deposition and reduced the release of pro-inflammatory cytokine IL-6, IL-1β, TNF-α and in the kidneys of HN mice. Furthermore, fisetin inhibited the activation of TGF-β and STAT3 signaling pathways in the kidneys of HN mice Conclusion These data demonstrated that fiseitn exhibited renoprotective effect in a severe mouse model of hyperuricemic nephropathy (HN). Fisetin exerted anti-inflammatory and antifibrotic effects in HN mice, which might be associated with its inhibition on TGF-β /Smad3 and STAT3 pathways.

Apigenin reverses lung injury and immunotoxicity in paraquat-treated mice

Paraquat (PQ) induces acute lung injury (ALI) and immunotoxicity. Apigenin exerts anti-oxidant and anti-inflammatory properties. The purpose of this study was to investigate the possible protective effects of apigenin on PQ-induced ALI and immunotoxicity in mice. Female C57BL/6 mice received a single injection of PQ (50 mg/kg). Apigenin was given for 7 consecutive days starting 5 days before PQ exposure. The toxicity markers were evaluated in terms of weight loss, lung histopathology, oxidative stress, inflammation, and T cell functions after PQ exposure. Poisoned mice exhibited severe lung tissue lesions, inflammatory cell infiltration and the release of pro-inflammatory cytokines IL-6 and TNF-α. PQ administration increased the lung wet/dry ratios and lipid peroxidation by the increase of MDA levels and decreased anti-oxidase activity including SOD, GSH-PX, and CAT. While such effect on lung was reversed by apigenin. Importantly, PQ-induced immunotoxicity was also observed in a decrease of spleen weight, inhibition of T cell proliferation and T-cell secreting IL-2 from splenocytes. Further mechanism analysis found that PQ administration could decrease total splenocytes, CD4+ and CD8+ T cells, SOD, GSH-PX, and CAT activity, and increased the levels of MDA and the concentrations of pro-inflammatory cytokines IL-6 and TNF-α compared to control mice. However, apigenin treatment reversed PQ-induced immunotoxicity. In summary, all results suggest that apigenin has beneficial effects on PQ-induced ALI and immunotoxicity possibly, and it could be related, at least in part, to its ability in modulating inflammation and oxidative stress, although in-depth studies might be needed to fully understand the mechanism of action.

3-Dehydroandrographolide protects against lipopolysaccharide-induced inflammation through the cholinergic anti-inflammatory pathway

Acute lung injury (ALI) is a deadly disease without effective chemotherapy, so far. Traditional Chinese medicine andrographis herba is frequently used in the treatment of respiratory diseases. In searching for natural anti-ALI components from andrographis herba, the activities of 3-dehydroandrographolide (3-DA), a new natural andrographolide product from andrographis herba were evaluated. In this study, murine macrophage RAW 264.7 cells and BALB/c mice were treated with LPS (lipopolysaccharide, 100 ng/ml in vitro; 3 mg/kg, intratracheal) to establish inflammation models. 3-DA attenuated the release of pro-inflammatory cytokines IL-6 and TNF-α, inhibited the degradation and phosphorylation of IκBα, and suppressed the nuclear translocation of NF-κB p65 as well as the phosphorylation of Akt at Ser473 in LPS-stimulated RAW 264.7 macrophage cells. Furthermore, 3-DA increased α7nAchR expression level and bound with α7nAchR. More importantly, the anti-inflammatory effects of 3-DA were counteracted in the presence of α7nAchR siRNA or methyllycaconitine (MLA, a α7nAchR specific inhibitor), suggesting that α7nAchR is a potential target in the anti-inflammatory effects of 3-DA. Besides, 3-DA significantly inhibited inflammation in LPS-induced ALI mice, which was associated with the decrease of lung water content and inflammatory cytokines, the inhibition of neutrophil and macrophage infiltration, and activation of the NF-κB/Akt signaling pathway. Moreover, these protective effects were attenuated by the treatment of MLA. Taken together, 3-DA alleviates LPS-induced inflammation via the cholinergic anti-inflammatory pathway in vitro and in vivo. These findings provide a rationale for the role of the cholinergic anti-inflammatory pathway in inflammation and the promising clinical application of 3-DA to treat ALI.

Placental Pim-1 expression is increased in obesity and regulates cytokine- and toll-like receptor-mediated inflammation

IntroductionObesity is a growing epidemic, and as a consequence the number of obese pregnancies has also increased. Pregnancy is characterised by maternal and placental inflammation which is intensified with maternal obesity. The proviral integration site for Moloney murine leukemic virus (Pim)-1 protein is a serine/threonine kinase involved in a wide range of inflammatory diseases. In relation to obesity, however, its role has not been elucidated in human placenta. The aims were to determine the placental expression of Pim-1 with pre-existing maternal obesity and its role in regulating placental inflammation associated with obesity. MethodsHuman placenta was obtained at the time of term Caesarean section from lean and pre-existing obese pregnant women to determine the effect of maternal obesity on Pim-1 expression. To determine the effect of Pim-1 on the inflammatory response induced by bacterial endotoxin LPS and pro-inflammatory cytokines TNF-α or IL-1β, the chemical inhibitor SMI-4a and siRNA were used. ResultsPim-1 protein and mRNA expression was significantly increased in placenta of obese women. SMI-4a significantly suppressed the expression and release of pro-inflammatory cytokine IL-6 and chemokines GRO-α and MCP-1 when stimulated with LPS or TNF-α in placenta. Primary trophoblast cells transfected with Pim-1 siRNA had decreased expression and release of pro-inflammatory cytokines IL-1β, IL-6, chemokines GRO-α and MCP-1, when stimulated with LPS, TNF-α or IL-1β. DiscussionThe findings from this study implicate Pim-1 may contribute to placental inflammation in pregnancies complicated by maternal obesity. Thus, therapeutic targets for Pim-1 may improve fetal outcomes complicated by obese pregnancies.

Impaired autophagy activity is linked to elevated ER-stress and inflammation in aging adipose tissue.

Adipose tissue dysfunction in aging is associated with inflammation, metabolic syndrome and other diseases. We propose that impaired protein homeostasis due to compromised lysosomal degradation (micro-autophagy) might promote aberrant ER stress response and inflammation in aging adipose tissue. Using C57BL/6 mouse model, we demonstrate that adipose tissue-derived stromal vascular fraction (SVF) cells from old (18-20 months) mice have reduced expression of autophagy markers as compared to the younger (4-6 months) cohort. Elevated expressions of ER-stress marker CHOP and autophagy substrate SQSTM1/p62 are observed in old SVFs compared to young, when treated with either vehicle or with thapsigargin (Tg), an ER stress inducer. Treatment with bafilomycin A1 (Baf), a vacuolar-type H (+)-ATPase, or Tg elevated expressions of CHOP, and SQSTM1/p62 and LC-3-II, in 3T3-L1-preadipocytes. We also demonstrate impaired autophagy activity in old SVFs by analyzing increased accumulation of autophagy substrates LC3-II and p62. Compromised autophagy activity in old SVFs is correlated with enhanced release of pro-inflammatory cytokines IL-6 and MCP-1. Finally, SVFs from calorie restricted old mice (CR-O) have shown enhanced autophagy activity compared to ad libitum fed old mice (AL-O). Our results support the notion that diminished autophagy activity with aging contributes to increased adipose tissue ER stress and inflammation.

Changes in some pro-and anti-inflammatory cytokines produced by bovine peripheral blood mononuclear cells following foot and mouth disease vaccination

Interleukin (IL)-17 is exclusively produced by CD4 helper T-cells upon activation. It most often acts as a pro-inflammatory cytokine, which stimulates the release of pro-inflammatory cytokines IL-6, IL-8, TNF-α, and granulocyte-macrophage colony-stimulating factor (GM-CSF). In this study, we studied the in-vitro IL-17 response to specific antigens and a variety of mitogens and compared the IL-17 response to IL-2, IL-4, IL-5, IL-6, IL-10, and IFN-γ responses. We used a foot and mouth disease (FMD) vaccine as specific antigens and mitogens (phytohemagglutinin [PHA], pokeweed mitogen [PWM], and concanavalin A [Con A]) to stimulate peripheral blood mononuclear cells (PBMCs) of vaccinated calves. Cell culture supernatant was harvested and analyzed for cytokines, using commercially available bovine ELISA kits. The mitogens induced a significant increase in IL-17 production. IL-17 was produced at high levels in response to the T cell-stimulated mitogens, PHA, and Con A, and at low levels in response to PWM mitogens. In contrast, level of the produced IL-17 cytokines in response to the FMDV antigens was lower as compared to those produced by mitogens. The FMDV antigens and mitogens significantly increased IL-17 production. There was not a correlation between IL-17 production and type-1 cytokine, IFN-γ, and IL-2, while there was a correlation between type-2 cytokine, IL-4, and IL-5 at either cytokine level produced by PBMCs stimulated by FMDV antigens. Moreover, there was an interaction between IL-17 and IL-6, that is, as IL-6 cytokine level elevated or diminished, IL-17 cytokine level increased or decreased, as well.

Macrophage molecular signaling and inflammatory responses during ingestion of atherogenic lipoproteins are modulated by complement protein C1q

Background and aimsThis study investigated the effect of innate immune protein C1q on macrophage programmed responses during the ingestion of atherogenic lipoproteins. C1q plays a dual role in atherosclerosis where activation of complement by C1q is known to drive inflammation and promote disease progression. However, C1q is atheroprotective in early disease using mouse models. Our previous studies have highlighted a non-complement associated role for C1q in polarizing macrophages towards an M2-like anti-inflammatory phenotype during ingestion of targets such as atherogenic lipoproteins. This study aims to investigate the molecular mechanisms involved. MethodsWe investigated the molecular signaling mechanisms involved in macrophage polarization using an unbiased examination of gene expression profiles in human monocyte derived macrophages ingesting oxidized or acetylated low density lipoproteins in the presence or absence of C1q. ResultsExpression of genes involved in Janus kinase and signal transducer and activator of transcription (JAK-STAT) signaling, peroxisome proliferator activating receptor (PPAR) signaling and toll-like receptor (TLR) signaling were modulated by C1q in this screen. C1q was also shown to significantly suppress JAK-STAT pathway activation (a maximum 55% ± 13% reduction, p = 0.044) and increase transcriptional activation of PPARs (a maximum 229% ± 54% increase, p = 0.0002), consistent with an M2-like polarized response. These pathways were regulated in macrophages by C1q bound to different types of modified atherogenic lipoprotein and led to a reduction in the release of pro-inflammatory cytokine IL-6. ConclusionsThis study identifies potential molecular mechanisms for the beneficial role C1q plays in early atherosclerosis.

Resistance to cytotoxicity and sustained release of interleukin-6 and interleukin-8 in the presence of decreased interferon-γ after differentiation of glioblastoma by human natural killer cells.

Natural killer (NK) cells are functionally suppressed in the glioblastoma multiforme (GBM) tumor microenvironment. We have recently shown that survival and differentiation of cancer stem-like cells (CSCs)/poorly differentiated tumors are controlled through two distinct phenotypes of cytotoxic and non-cytotoxic/split anergized NK cells, respectively. In this paper, we studied the function of NK cells against brain CSCs/poorly differentiated GBM and their NK cell-differentiated counterparts. Brain CSCs/poorly differentiated GBM, differentiated by split anergized NK supernatants (supernatants from NK cells treated with IL-2+anti-CD16mAb) expressed higher levels of CD54, B7H1 and MHC-I and were killed less by the NK cells, whereas their CSCs/poorly differentiated counterparts were highly susceptible to NK cell lysis. Resistance to NK cells and differentiation of brain CSCs/poorly differentiated GBM by split anergized NK cells were mediated by interferon (IFN)-γ and tumor necrosis factor (TNF)-α. Brain CSCs/poorly differentiated GBM expressed low levels of TNFRs and IFN-γRs, and when differentiated and cultured with IL-2-treated NK cells, they induced increased secretion of pro-inflammatory cytokine interleukin (IL)-6 and chemokine IL-8 in the presence of decreased IFN-γ secretion. NK-induced differentiation of brain CSCs/poorly differentiated GBM cells was independent of the function of IL-6 and/or IL-8. The inability of NK cells to lyse GBM tumors and the presence of a sustained release of pro-inflammatory cytokines IL-6 and chemokine IL-8 in the presence of a decreased IFN-γ secretion may lead to the inadequacy of NK cells to differentiate GBM CSCs/poorly differentiated tumors, thus failing to control tumor growth.

Anti-Inflammatory Activities of Pentaherbs Formula, Berberine, Gallic Acid and Chlorogenic Acid in Atopic Dermatitis-Like Skin Inflammation.

Atopic dermatitis (AD) is a common allergic skin disease, characterized by dryness, itchiness, thickening and inflammation of the skin. Infiltration of eosinophils into the dermal layer and presence of edema are typical characteristics in the skin biopsy of AD patients. Previous in vitro and clinical studies showed that the Pentaherbs formula (PHF) consisting of five traditional Chinese herbal medicines, Flos Lonicerae, Herba Menthae, Cortex Phellodendri, Cortex Moutan and Rhizoma Atractylodis at w/w ratio of 2:1:2:2:2 exhibited therapeutic potential in treating AD. In this study, an in vivo murine model with oxazolone (OXA)-mediated dermatitis was used to elucidate the efficacy of PHF. Active ingredients of PHF water extract were also identified and quantified, and their in vitro anti-inflammatory activities on pruritogenic cytokine IL-31- and alarmin IL-33-activated human eosinophils and dermal fibroblasts were evaluated. Ear swelling, epidermis thickening and eosinophils infiltration in epidermal and dermal layers, and the release of serum IL-12 of the murine OXA-mediated dermatitis were significantly reduced upon oral or topical treatment with PHF (all p < 0.05). Gallic acid, chlorogenic acid and berberine contents (w/w) in PHF were found to be 0.479%, 1.201% and 0.022%, respectively. Gallic acid and chlorogenic acid could suppress the release of pro-inflammatory cytokine IL-6 and chemokine CCL7 and CXCL8, respectively, in IL-31- and IL-33-treated eosinophils-dermal fibroblasts co-culture; while berberine could suppress the release of IL-6, CXCL8, CCL2 and CCL7 in the eosinophil culture and eosinophils-dermal fibroblasts co-culture (all p < 0.05). These findings suggest that PHF can ameliorate allergic inflammation and attenuate the activation of eosinophils.

The association of interleukin-31 polymorphisms with interleukin-31 serum levels and risk of systemic lupus erythematosus.

Interleukin-31 (IL-31) is the most recently discovered member of the gp130/IL-6 cytokine family which is produced mainly by activated Th2 cells. IL-31 was proved to play a crucial role in autoimmune and inflammatory diseases such as atopic dermatitis, asthma, cutaneous T cell lymphomas, Kawasaki disease and allergic rhinitis. Previous studies have identified that IL-31 could significantly induce the release of proinflammatory cytokines IL-6. Moreover, a large number of studies have shown that IL-6 plays an important role in the pathogenesis of systemic lupus erythematosus (SLE). However, up to date, no study to data was reported on the relationship between IL-31 and SLE. Therefore, in the present study, we investigated the association between IL-31 polymorphisms and its serum levels with the risk of SLE in a Chinese population. We analyzed two single nucleotide polymorphisms of IL-31 gene rs7977932 C/G and rs4758680 G/T in 190 patients with SLE and 250 age- and sex-matched controls, using polymerase chain reaction-single base extension and DNA sequencing methods. Soluble IL-31 (sIL-31) levels were measured by ELISA. From this study, we found that there were significant differences in the genotype and allele frequencies of IL-31 gene rs7977932 C/G polymorphism between the group of patients with SLE and the control group (P<0.05). sIL-31 levels were increased in patients with SLE compared with controls (P<0.01). Moreover, genotypes carrying the IL-31 rs7977932 G variant allele were associated with increased IL-31 levels compared to the homozygous wild-type genotype in patients with SLE. The rs7977932 C/G polymorphism of IL-31 gene and its sIL-31 levels were associated with SLE in the Chinese population. Our data suggest that IL-31 gene may play a role in the development of SLE.

PPARγ regulated CIDEA affects pro-apoptotic responses in glioblastoma.

Refractoriness of glioblastoma multiforme (GBM) to current treatment paradigms has necessitated identification of new targets to better the existing therapeutic strategies. One such target is peroxisome proliferator-activated receptor gamma (PPARγ) – a transcription factor involved in regulation of lipid metabolism and inflammation. Expression of PPARγ, a known regulator of cell death-inducing DFFA-like effector (CIDEA), is modulated by hypoxia inducible factor (HIF-1α). While the involvement of CIDEA in lipid metabolism is known, its role in malignancies remains largely unknown. An elevated PPARγ and low CIDEA level was observed in GBM tumors as compared with surrounding non-neoplastic tissue. As reciprocal relation exists between PPAR and HIF-1α: and as HIF-1α is a key component in glioma progression, their role in regulating CIDEA expression in glioblastoma was investigated. Although HIF-1α inhibition had no effect on CIDEA expression, pharmacological inhibition of PPARγ elevated CIDEA levels. PPARγ mediated upregulation of CIDEA was accompanied by decreased recruitment of NFκB and SP1 to their predicted binding sites on CIDEA promoter. Ectopic expression of CIDEA triggered apoptosis, activated JNK, decreased HIF-1α activation and increased PPARγ levels in glioma cells. While CIDEA overexpression induced actin cytoskeletal disruption, cell cycle arrest, release of pro-inflammatory cytokine IL-6 in a JNK-dependent manner; CIDEA mediated apoptotic cell death, decreased STAT3 phosphorylation and increased p53 acetylation was JNK independent. This study highlights for the first time the existence of (i) PPARγ-CIDEA regulatory loop in glioma and (ii) novel function of CIDEA as regulator of glioma cell survival.

Genetic engineering strategies to prevent the effects of antibody and complement on xenogeneic chondrocytes.

Advances in animal transgenesis may allow using xenogeneic chondrocytes in tissue-engineering applications for clinical cartilage repair. Porcine cartilage is rejected by humoral and cellular mechanisms that could be overcome by identifying key molecules triggering rejection and developing effective genetic-engineering strategies. Accordingly, high expression of α1,2-fucosyltransferase (HT) in xenogeneic cartilage protects from galactose α1,3-galactose (Gal)-mediated antibody responses. Now, we studied whether expression of a complement inhibitor provides further protection. First, porcine articular chondrocytes (PAC) were isolated from non-transgenic, single and double transgenic pigs expressing HT and moderate levels of human CD59 (hCD59) and their response to human serum was assessed. High recombinant expression of human complement regulatory molecules hCD59 and hDAF was also attained by retroviral transduction of PAC for further analyses. Complement activation on PAC after exposure to 20 % human serum for 24 hours mainly triggered the release of pro-inflammatory cytokines IL-6 and IL-8. Transgenic expression of HT and hCD59 did not suffice to fully counteract this effect. Nevertheless, the combination of blocking anti-Gal antibodies (or C5a) and high hCD59 levels conferred very high protection. On the contrary, high hDAF expression attained the most dramatic reduction in IL-6/IL-8 secretion by a single strategy, but the additional inhibition of anti-Gal antibodies or C5a did not provide further improvement. Notably, we demonstrate that both hCD59 and hDAF inhibit anaphylatoxin release in this setting. In conclusion, our study identifies genetic-engineering approaches to prevent humoral rejection of xenogeneic chondrocytes for use in cartilage repair.

Protecting the BBB endothelium against cigarette smoke-induced oxidative stress using popular antioxidants: Are they really beneficial?

Blood Brain Barrier (BBB) exposed to realistic concentrations (comparable to a chronic heavy smoker) of Cigarette Smoke Extract (CSE) triggers a strong endothelial inflammatory response which can lead to the onset of neurological disorders. The involvement of Reactive Oxygen Species (ROS) in this inflammatory cascade is evident from the up-regulation of nuclear factor erythroid 2 related factor 2 (Nrf-2), a transcription factor involved in anti-oxidant response signaling in CSE exposed endothelial cells. We have shown that pre-treatment with α-tocopherol and/or ascorbic acid is highly protective for the BBB, thus suggesting that, prophylactic administration of antioxidants can reduce CSE and/or inflammatory-dependent BBB damage. We have assessed and ranked the protective effects of 5 popular OTC antioxidants (Coenzyme Q10, melatonin, glutathione, lipoic acid and resveratrol) against CSE-induced BBB endothelial damage using hCMEC/D3 cells. The analysis of pro-inflammatory cytokines release by ELISA revealed that resveratrol, lipoic acid melatonin and Co-Q10 inhibited the BBB endothelial release of pro-inflammatory cytokines IL-6 and IL-8, reduced (not Co-Q10) CSE-induced up-regulation of Platelet Cell Adhesion Molecule-1 (PECAM-1), Vascular Cell Adhesion Molecule-1 (VCAM-1) & E-selectin and inhibited monocytes-endothelial cell adhesion. The anti-inflammatory effects correlated with the anti-oxidative protection endowed by these compounds as evidenced by upregulation of NADPH: Quinone Oxidoreductase 1 (NQO1) and reduced cellular oxidative stress. CSE-induced release of Vascular Endothelial Growth Factor (VEGF) was inhibited by all tested compounds although the effect was not strictly dose-dependent. Further in vivo studies are required to validate our results and expand our current study to include combinatorial treatments.