Levels Of Cytokine Production Research Articles

Hyperglycemia Augmented Neuropathic Pain Involving the HMGB1/TLR4 Inflammatory Axis

<p>Peripheral neuropathy, accompanied by chronic neuropathic pain, is among the frequent and debilitating complications of diabetes. Hyperglycemia is a risk factor for neuropathic pain, but the pathological mechanisms are unclear. The high mobility group box 1 (HMGB1)/toll-like receptor 4 (TLR4) inflammatory axis is strongly implicated in the pathogenesis of neuropathic pain,so we examined how hyperglycemia influences the HMGB1/TLR4 axis and behavioural manifestations of hyperalgesia in the Sprague-Dawley rat sciatic nerve chronic constriction injury (CCI) model. Short-term hyperglycemia induced by streptozotocin significantly augmented mechanical allodynia and thermal hyperalgesia in CCI model rats compared to normoglycemic CCI model rats.These augmented nociceptive behaviours were associated with increased immune cell infiltration and activation, cytokine production and expression levels of TLR4 and HMGB1, as well as with reduced Sirt1 expression in spinal cord dorsal horn and sciatic nerve. High glucose also enhanced HMGB1/TLR4 inflammatory axis activity and cytokine production in lipopolysaccharide-stimulated RAW264.7 macrophages. Our results suggest that hyperglycemia predisposes individuals with peripheral nerve injury to neuropathic pain, in part through enhancement of Sirt1/HMGB1/TLR4 signalling and ensuing neuroinflammation.</p> <p> </p>

Caspase-4 and -5 Biology in the Pathogenesis of Inflammatory Bowel Disease.

Inflammatory bowel disease (IBD) is a chronic relapsing inflammatory disease of the gastrointestinal tract, associated with high levels of inflammatory cytokine production. Human caspases-4 and -5, and their murine ortholog caspase-11, are essential components of the innate immune pathway, capable of sensing and responding to intracellular lipopolysaccharide (LPS), a component of Gram-negative bacteria. Following their activation by LPS, these caspases initiate potent inflammation by causing pyroptosis, a lytic form of cell death. While this pathway is essential for host defence against bacterial infection, it is also negatively associated with inflammatory pathologies. Caspases-4/-5/-11 display increased intestinal expression during IBD and have been implicated in chronic IBD inflammation. This review discusses the current literature in this area, identifying links between inflammatory caspase activity and IBD in both human and murine models. Differences in the expression and functions of caspases-4, -5 and -11 are discussed, in addition to mechanisms of their activation, function and regulation, and how these mechanisms may contribute to the pathogenesis of IBD.

SARS-CoV-2 host-shutoff impacts innate NK cell functions, but antibody-dependent NK activity is strongly activated through non-spike antibodies.

The outcome of infection is dependent on the ability of viruses to manipulate the infected cell to evade immunity, and the ability of the immune response to overcome this evasion. Understanding this process is key to understanding pathogenesis, genetic risk factors, and both natural and vaccine-induced immunity. SARS-CoV-2 antagonises the innate interferon response, but whether it manipulates innate cellular immunity is unclear. An unbiased proteomic analysis determined how cell surface protein expression is altered on SARS-CoV-2-infected lung epithelial cells, showing downregulation of activating NK ligands B7-H6, MICA, ULBP2, and Nectin1, with minimal effects on MHC-I. This occurred at the level of protein synthesis, could be mediated by Nsp1 and Nsp14, and correlated with a reduction in NK cell activation. This identifies a novel mechanism by which SARS-CoV-2 host-shutoff antagonises innate immunity. Later in the disease process, strong antibody-dependent NK cell activation (ADNKA) developed. These responses were sustained for at least 6 months in most patients, and led to high levels of pro-inflammatory cytokine production. Depletion of spike-specific antibodies confirmed their dominant role in neutralisation, but these antibodies played only a minor role in ADNKA compared to antibodies to other proteins, including ORF3a, Membrane, and Nucleocapsid. In contrast, ADNKA induced following vaccination was focussed solely on spike, was weaker than ADNKA following natural infection, and was not boosted by the second dose. These insights have important implications for understanding disease progression, vaccine efficacy, and vaccine design.

Novel urea-thiourea hybrids bearing 1,4-naphthoquinone moiety: Anti-inflammatory activity on mammalian macrophages by regulating intracellular PI3K pathway, and molecular docking study

In this study, four novel urea-thiourea hybrids bearing 1,4-naphthoquinone moiety were synthesized by the reaction of 2,3-diaminonaphthalene-1,4-dione and various isothiocyanate derivatives in 75–88% yield, and their molecular structures were characterized by using 1H/13C NMR, FT-IR and HRMS techniques. Urea-thiourea hybrids have been known for their anti-inflammatory activities. Therefore, in our study, we aimed to decipher their biological activities on mammalian macrophages. Macrophages were stimulated by LPS and pro-inflammatory TNF, IL6, GMCSF and IL12p40 cytokine production levels were measured in the presence of these derivatives by ELISA. Our results suggest that these derivatives on stimulated mammalian macrophages had anti-inflammatory activities. In order to examine their intracellular mechanism of action, intracellular phosphorylated (activated) PI3K protein levels were measured in the presence of our novel derivatives by flow cytometry. Our results suggest that these novel derivatives had anti-inflammatory activities on mammalian macrophages by blocking the PI3K activation and decreasing the production of pro-inflammatory cytokines. Molecular docking calculations were also employed to elucidate the theoretical binding modes of our studied compounds with PI3Kγ. These derivatives have the potential to be utilized as anti-inflammatory drug agents against versatile inflammatory and autoimmune disorders.

Effect of resolvin D1 on experimental bacterial keratitis to prevent corneal scar.

The study aims to investigate the role of the lipid mediator resolvin D1 (RvD1) in bacterial keratitis in a murine model. The effect of RvD1 on Pseudomonas aeruginosa-stimulated human corneal epithelial cells (HCECs) and mouse macrophages and dendritic cells (DCs) was assessed. C57BL/6 mouse corneas were abraded and treated with RvD1 after stimulation with P. aeruginosa, following which cytokine production level in the cornea and drainage lymph nodes was compared with that in controls. Corneal opacity and thickness were assessed using anterior segment photographs, and optical coherence tomography and corneal infiltrates were analyzed using immunohistochemistry for neutrophils. RvD1 significantly inhibited pro-inflammatory cytokine production in HCECs, mouse macrophages, and DCs. Corneal opacity and corneal thickness were reduced, and the development of corneal infiltrates, specifically neutrophils, was also significantly inhibited by RvD1 in response to stimulation with P. aeruginosa. RvD1 inhibits P. aeruginosa-induced corneal inflammation. This finding supports a potential therapeutic approach for patients with bacterial keratitis.

Evaluation of equine xenogeneic mixed lymphocyte reactions using 5-ethynyl-2’-deoxyuridine (EdU)

Allogeneic solid organ transplantation is currently the only treatment option for end stage organ disease. The shortage of available donor organs has driven efforts to utilize xenogeneic organs for transplantation. In vitro methods for evaluating immune-compatibility are a quick and low cost means of screening novel tissue products prior to more involved, expensive, and invasive live animal studies. Recently, a new analog of the DNA base thymidine, 5-ethynyl-2’-deoxyuridine (EdU), was developed. It may be used in a fast, efficient and specific means of evaluating cell proliferation via flow cytometry. This study was designed to test and optimize this platform for assessing equine xenogeneic one-way mixed lymphocyte reaction (MLR) to porcine stimulator cells. Furthermore, it was hypothesized that an enriched T-lymphocyte (T-cell) population would generate a stronger proliferative response to stimulation, and higher levels of cytokine production when compared to unfractionated peripheral blood mononuclear cells (PBMCs). PBMCs and T-cells were isolated from 3 horses and 4 pigs. Equine xenogeneic MLRs were set up using porcine allogeneic MLRs as a reference for clinically acceptable levels of cell proliferation. Equine T-cells showed significantly greater EdU incorporation in one-way xenogeneic MLRs than equine PBMCs. However, there was no significant difference in cell proliferation between porcine T-cell and PBMC as responders in allogenic one-way MLRs. Given the results of this study, we consider that enriched equine T-cells should be used in preference to unfractionated PBMCs when attempting to evaluate the equine xenogeneic response using the EdU assay as an indicator of suitability for transplant in vivo.

Cytokine Production in Whole Blood Cells Culture of Patients with Alcohol Dependence and Autologous Plasma Oxidative Stress Markers

We studied spontaneous production of a spectrum of proinflammatory cytokines by cultured whole blood cells from men with alcohol dependence at the stage of withdrawal syndrome and oxidative stress markers (carbonylated proteins and TBA-reactive substances) in the plasma of these blood samples. Enhanced production of cytokines by blood cells and increased concentrations of oxidative stress markers in the autologous plasma were revealed in comparison with the corresponding parameters in the control (blood from healthy men). Direct correlations were found between the levels of spontaneous cytokine production by blood cells from subjects with alcohol dependence and the concentration of oxidized proteins and lipids in autologous plasma.

Regulation of the macrophage cellular response by Clinacanthus nutans extracts in J774.2 macrophages

Clinacanthus nutans is a medicinal plant from the Acanthaceae family, and it is well-known as 'Belalai gajah' and Sabah Snake Grass in Malaysia. C. nutans extracts have been reported to exhibit various bioactivities but reports about its immunomodulatory role are limited. Macrophage activation is lacking in certain macrophage-related immunodeficiency disorders, but it occurs constitutively in certain inflammatory diseases. Thus, the authors investigated the potential role of C. nutans extracts in regulating the macrophage cellular response in J774.2 mouse macrophages. J774.2 macrophages were treated with different concentrations (0–1000 µg/mL) of C. nutans ethanol, ethanol-aqueous, and aqueous extracts for 24, 48, and 72 h and the cytotoxicity effect was assayed using PrestoBlue assay. Then, the macrophages were incubated with 32 µg/mL of ethanol, 500 µg/mL of ethanol-aqueous, and 32 µg/mL of aqueous extracts of C. nutans for 48 h. Cytokine secretion, phagocytic activity, and expression of M1 and M2 activation markers were assessed using a multiplexed cytokine bead-based assay, fluorescence microscopy, and flow cytometry. All extracts did not show any significant cytotoxic effect towards J774.2 macrophages within the extract concentration range tested (cell viability >50%). The ethanol-aqueous extract showed the greatest potential for downregulating lipopolysaccharide-induced pro-inflammatory cytokine (IL-1β, IL-6, and IL-12p40) production and CD86 (M1 activation marker) expression level and for stimulating the phagocytic function of J774.2 macrophages.

Antidesma thwaitesianum Müll. Arg. fruit extract rich in 5-hydroxymethylfurfural exhibits anti-inflammatory effects in lipopolysaccharide-stimulated RAW264.7 macrophages

Introduction: Antidesma thwaitesianum Müll. Arg is a tropical fruit, which has been commonly used for healthy food and traditional herbal medicine. This study aimed to investigate the anti-inflammatory effects of A. thwaitesianum fruit extract (AFE) rich in 5-hydroxymethylfurfural (5-HMF) in lipopolysaccharide (LPS)-stimulated RAW264.7 macrophages. Methods: The chemical composition of AFE was analysed by gas chromatography/mass spectrometry (GC/MS). RAW264.7 cells were used as an in vitro inflammatory response model. RAW264.7 cells were pre-treated with various concentrations of AFE or 5-HMF and subsequently treated with LPS. Cell viability was assessed by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The production levels of pro-inflammatory cytokines and mediators, including nitric oxide (NO), interleukin (IL)-1β, and tumour necrosis factor (TNF)-α were determined by the Griess assay and enzyme-linked immunosorbent assay. The protein expression of inducible nitric oxide synthase (iNOS) was examined by western blot analysis. Results: AFE had a high content of 5-HMF (61.03% ± 0.49%). Pre-treatment with AFE and 5-HMF markedly reduced LPS-induced pro-inflammatory mediators and cytokines, namely NO, IL-1β, and TNF-α, in RAW264.7 cells; this reduction correlated with downregulation of iNOS expression. Conclusion: This study suggests that A. thwaitesianum fruit extract containing 5-HMF could modulate the LPS-induced inflammatory response by inhibiting NO, IL-1β, and TNF-α production and iNOS expression. A. thwaitesianum fruit extract rich in 5-HMF could be considered a potential therapeutic agent for the prevention of inflammation.

Functional Characteristics of Kombucha Fermented with Lactic Acid Bacteria, Yeast, and Acetic Acid Bacteria Derived from Korea Traditional Foods

In this study, to determine the importance of lactic acid bacteria (LAB) in Kombucha fermentation, biological functions, such as organic acid production and anti-inflammatory and antibacterial activities, of Kombucha, with or without LAB inoculation, were evaluated. Lactobacillus paracasei DK215, Saccharomyces cerevisiae C3, and Acetobacter pasteurianus P2 were selected as the inoculants. Organic acids were measured every 3 days from the end of fermentation using HPLC; the organic acid content of LAB-inoculated Kombucha was relatively high. Samples with or without LAB inoculation showed high antibacterial activity against Escherichia coli. The MTT assay results indicated no significant difference in concentration difference and cell death. In the NO production test, compared with the uninoculated Kombucha sample, the LAB-inoculated Kombucha sample exhibited a value similar to that of the group without LPS treatment. The levels of cytokine (IL-1α, IL-6, TNF-α) production were significantly lower than those of the LPS(+) group, indicating the anti-inflammatory activity potential of the Kombucha sample. This improvement in the biological function of the LAB-inoculated Kombucha further verifies the value of LAB in the fermented food and beverage industry.

Comparison of the Seven Interleukin-32 Isoforms' Biological Activities: IL-32θ Possesses the Most Dominant Biological Activity.

Cytokines are significantly associated with the homeostasis of immune responses in health and disease. Interleukin-32 (IL-32) is a cytokine originally discovered in natural killer cell transcript 4. IL-32 with different disorders has been described in terms of pathogenesis and the progression of diseases. Clinical studies have investigated IL-32 under various conditions, such as viral infection, autoimmune diseases, inflammatory diseases, certain types of cancer, vascular disease, and pulmonary diseases. The high expression of IL-32 was identified in different tissues with various diseases and found to have multiple transcripts of up to seven isoforms. However, the purification and biological activities of these isoforms have not been investigated yet. Therefore, in this study, we purified and compared the biological activity of recombinant IL-32 (rIL-32) isoforms. This is the first time for seven rIL-32 isoforms (α, β, δ, γ, ϵ, ζ, and θ) to be cloned and purified using an Escherichia coli expression system. Next, we evaluate the biological activities of these seven rIL-32 isoforms, which were used to treat different types of cells by assessing the levels of inflammatory cytokine production. The results revealed that rIL-32θ possessed the most dominant biological activity in both immune and non-immune cells.

The anti-inflammatory effects of Akkermansia muciniphila and its derivates in HFD/CCL4-induced murine model of liver injury

Inflammation plays a critical role in the promotion of hepatocyte damage and liver fibrosis. In recent years the protective role of Akkermansia muciniphila, a next-generation beneficial microbe, has been suggested for metabolic and inflammatory disorders. In this study, we aimed to evaluate the effects of live and pasteurized A. muciniphila and its extra cellular vesicles (EVs) on inflammatory markers involved in liver fibrosis in a mouse model of a high-fat diet (HFD)/carbon tetrachloride (CCl4)-induced liver injury. Firstly, the responses of hepatic stellate cells (HSCs) to live and pasteurized A. muciniphila and its EVs were examined in the quiescent and LPS-activated LX-2 cells. Next, the anti-inflammatory effects of different forms of A. muciniphila were examined in the mouse model of HFD/CCl4-induced liver injury. The gene expression of various inflammatory markers was evaluated in liver, colon, and white adipose tissues. The cytokine secretion in the liver and white adipose tissues was also measured by ELISA. The results showed that administration of live and pasteurized A. muciniphila and its EVs leads to amelioration in HSCs activation. Based on data obtained from the histopathological analysis, an improvement in gut health was observed through enhancing the epithelium and mucosal layer thickness and strengthening the intestinal integrity in all treatments. Moreover, live A. muciniphila and its EVs had inhibitory effects on liver inflammation and hepatocytes damage. In addition, the tissue cytokine production and inflammatory gene expression levels revealed that live A. muciniphila and its EVs had more pronounced anti-inflammatory effects on liver and adipose tissues. Furthermore, EVs had better effects on the modulation of gene expression related to TLRs, PPARs, and immune response in the liver. In conclusion, the present results showed that oral administration of A. muciniphila and its derivatives for four weeks could enhance the intestinal integrity and anti-inflammatory responses of the colon, adipose, and liver tissues and subsequently prevent liver injury in HFD/CCL4 mice.

Fatostatin ameliorates inflammation without affecting cell viability.

The mature form of sterol regulatory element‐binding protein (SREBP)1 is a transcription factor involved in lipid synthesis, which participates in toll like receptor 4‐triggered inflammatory pathways during the resolution phase of inflammation in macrophages. SREBP1 has thus attracted interest as a candidate target molecule for ameliorating inflammation. Fatostatin is a small molecule that inhibits the maturation and function of SREBP, and its role in regulating inflammation is poorly understood. To evaluate the anti‐inflammatory effect of fatostatin, we compared body weight, footpad and hock dimensions, and arthritis scores between K/BxN serum‐induced arthritis mice treated with fatostatin and those treated with dimethyl sulfoxide as the vehicle control. We performed hematoxylin and eosin staining of joints of distal paws to assess tissue inflammation. Moreover, inflammatory cytokine production levels and cell viability were measured in lipopolysaccharide‐responsive human embryonic kidney 293 cells (293/hTLR4A‐MD2‐CD14 cells) after fatostatin administration. In K/BxN serum‐induced arthritis mice, fatostatin treatment significantly reduced the arthritis scores and hyperplasia. In vitro analysis revealed that fatostatin significantly inhibited the secretion of inflammatory cytokines from cells activated with lipopolysaccharide, without affecting cell viability. This is the first study to demonstrate that fatostatin is an anti‐inflammatory agent that modulates the processing of lipid transcription factors without affecting cell viability. Accordingly, the study reveals the potential of anti‐inflammatory therapeutics that link lipid regulation and inflammation.

Interrelations of cytokine production levels in the mechanisms of inflammatory process regulation in patients with community-acquired pneumonia

The aim of study was to study of cytokine production levels and their relationship in the mechanisms of regulation of the inflammatory process in community-acquired pneumonia.
 Materials and methods. The study was carried out in the period 2017–2020 based on the therapeutic department of the Municipal non-profit enterprise “City Clinical Multidisciplinary Hospital No. 25” of Kharkiv City Council. The study involved 34 adult patients with CAP aged 18 to 80 years (mean age 36.5±10.3). The control group consisted of 20 apparently healthy individuals (mean age 39.5±12.5). Levels of IL-17, IL-1Ra, TGFβ1, visfatin, adiponectin were determined by enzyme-linked immunosorbent assay.
 Results. Levels of IL-1 Ra and IL-17 in the group of patients with САP were 3.77±0.24 and 33.08±0.10 pg/ml, respectively. In the PРI group, the level of these indicators was probably higher (p<0.05) and was 2.53±0.13 and 28.17±0.53 pg/ml. The level of TGFβ1 in the group of patients with CAP was 24.54±0.55 ng/ml and was slightly higher than in the group of AHI (26.33±0.62 ng/ml). No differences were found between adiponectin levels in the AHI group and patients with CAP, as in the case of visfatin levels. The disappearance of relationships in the system of normal regulation of the inflammatory process between one pair of cytokines and their appearance between other pairs indicates a violation of regulatory mechanisms in patients with community-acquired pneumonia.
 Conclusions. The identified characteristics of the cytokine profile reflect the end of the phase of active inflammation in the lungs and the beginning of the phase of compensatory reactions. The findings suggest the importance of studying the cytokine profile in patients with community-acquired pneumonia, which will help to develop new approaches to predict the course of pneumonia, ways to correct metabolic disorders that develop in this condition, and will help identify risk groups for this pathology

Clinical characteristics, cytokine profiles and plasma IgE in adults with asthma

Asthma is a disease with complicated network of inflammatory responses of cytokines and ImmunoglobulinE (IgE). The aim of this study was to explore the clinical characteristics, cytokine profile and plasma IgE in the Malaysian population. This is a cross-sectional study involving physician-diagnosed asthma patients (n = 287) recruited from the Chest Clinic, University of Malaya Medical Centre (UMMC). Blood (8 mL) was taken after consent was obtained. The peripheral blood leucocytes (PBL) were cultured in presence of a mitogen for 72 h to quantify cytokines [Interleukin-5(IL-5), Interleukin-9 (IL-9), Interleukin-12 Beta (IL-12ꞵ) and granulocyte-macrophage colony-stimulating factor (GM-CSF)] and plasma was used to quantify IgE levels with commercial ELISA kits. Results were compared against the same biomarkers in healthy subjects (n = 203). In addition, the amount of the biomarkers in the asthma patients were compared with their disease severity and clinical characteristics. Statistical tests in the SPSS software (Mann-Whitney U test and the Kruskal Wallis) were used to compare cytokine production and plasma IgE levels. The mean plasma IgE level was markedly higher (p < 0.0001) in asthmatics compared to controls. There were higher levels of IL-5, IL-9, IL-12ꞵ and GM-CSF (p < 0.0001) produced by cultured PBL from asthma patients compared to controls. However, our results did not expose a significant association between these cytokine levels and severity and clinical symptoms of asthma. However, there was a marked association between asthma severity and blood lymphocyte count [ꭓ2(2) = 6.745, p < 0.05]. These findings support the roles played by cytokines and IgE in the airway inflammation in asthma. The findings of this study provide new information about inflammatory cytokines in Malaysian asthma patients.

Identification of galangin as the bioactive compound from Alpinia calcarata (Haw.) Roscoe rhizomes to inhibit IRAK-1/ MAPK/ NF-κB p65 and JAK-1 signaling in LPS stimulated RAW 264.7 cells

Ethnopharmacological relevanceAlpinia calcarata (Haw.) Roscoe rhizomes are used to treat diabetes, rheumatism, gastrointestinal problems, inflammatory diseases, cough and respiratory problems in traditional practices. The primary objective of the study is to identify and isolate anti-inflammatory bioactive compounds from A.calcarata rhizomes and to assess its molecular mechanism. Materials and methodsThe bioassay-guided fractionation of methanolic extract of A. calcarata rhizomes yielded chloroform fraction as the effective fraction and galangin as the bioactive compound identified by NMR studies. The anti-inflammatory action of galangin was evaluated by determining NO and cytokine production in LPS stimulated RAW264.7 cells. Further, its mechanism was studied on the expression levels of mRNA and protein targets by qPCR and Western blot analysis. ResultsBased on the MTT assay, the concentration of 3.1–25 μM of galangin was selected for further studies. Galangin reduced the levels of NO and proinflammatory cytokines (TNF-α, IL-1β and IL-6) production in LPS induced RAW 264.7 cells in a dose-dependent manner. In addition, the qPCR analysis revealed a reduction in the mRNA expression levels of COX-2, IRAK 1 and JAK 1 in galangin treated LPS stimulated RAW 264.7 cells in a dose-dependent manner. Western blot analysis implicated that galangin has markedly reduced the protein expression levels of cell signaling regulators (JAK-1, IRAK-1, MyD88, MAPK (p38 and ERK) and NF-κB p65). ConclusionFrom the results, it is evident that the inhibition of these cell signaling regulators has contributed to the anti-inflammatory effects of galangin. To our knowledge, we are the first to report IRAK-1 and JAK-1 as therapeutic targets of galangin for its anti-inflammatory effect.

Anti-TIM1 suppresses airway inflammation and hyperresponsiveness via the STAT6 /STAT1 pathways in mice with allergic asthma.

T cell immunoglobulin and mucin domain-1 (TIM-1) is a transmembrane glycoprotein and has been reported as an molecular mechanism of allergic diseases. This study aimed to explore the effects of anti-TIM-1 monoclonal antibodies (anti-TIM1) on the development of allergic asthma. Female C57BL/6 mice were induced and challenged with ovalbumin (OVA) and received subsequent intranasal administration of anti-TIM1. The airway resistance of all mice was evaluated using a Buxco PFT system. Flow cytometry was used to detect the expression of TIM-1 in peripheral blood mononuclear cells. The level of cytokine production in the bronchial alveolar lavage fluid and serum was determined using ELISA. Mucous cells were observed using Alcian blue and periodic acid-Schiff staining. In addition, B-cell lymphoma gene 2(BCL2), T-box transcription factor (T-bet), GATA binding protein-3(GATA3), signal transducer and activator of transcription (STAT) 1, STAT6 were analyzed by western blot analysis. Their corresponding mRNA expression levels were determined by quantitative PCR. The mRNA expression level of Mucin 5AC in the lung tissues was also detected using quantitative RT-PCR. The results showed that the intranasal administration of anti-TIM1 ameliorated airway inflammation and hyperresponsiveness in an acute model of asthma. Following administration of anti-TIM1, both the mRNA and protein levels of T-bet were upregulated, while those of BCL2 and GATA3 were downregulated. Moreover, the phosphorylation levels of STAT1 and STAT6 were increased. Taken together, these findings demonstrated that intranasal administration of anti-TIM1 ameliorated allergic lung inflammation and remodeling in mouse models of asthma by repairing both the STAT1 and STAT6 pathways.

Lactiplantibacillus plantarum으로 발효한 황련해독탕 가미방의 기능성 화장품 조성물로서의 생리활성 증대에 관한 연구

Hwangryunhaedoktang is one of the prescriptions used in traditional medicine for skin diseases. In this study, Hwangryunhaedoktang-Gamibang (HG) was fermented with Lactiplantibacillus plantarum , a probiotic lactic acid bacterium, to evaluate its potential as a functional cosmetic composition. Strains with anti-inflammatory activity were selected by isolating lactic acid bacteria from kimchi, a traditional Korean fermented food. HG was inoculated with lactic acid bacteria and the viability was measured. The supernatant was obtained by centrifugation of fermented Hwangryunhaedoktang-Gamibang (HGF) and HG, and the filtered supernatant was freeze-dried and used in the experiment. By measuring DPPH and ABTS scavenging activity, it was confirmed that the antioxidant activity was increased. RAW264.7 cells were inoculated with HG and HGF to confirm anti-inflammatory activity through NO assay, and production levels of pro-inflammatory cytokines (IL-1β, IL-6, and TNF-α) were measured through ELISA assay. It was confirmed that HGF had a greater decrease in production than HG. Through lactobacilli fermentation, the beneficial probiotic properties and antioxidant and pro-inflammatory activities of lactic acid bacteria suggest potential clinical or technical applications.

Comparison of Overall Immunity Levels among Workers at Grape Orchard, Rose Greenhouse, and Open-Field Onion Farm

BackgroundOccupational hazards in crop farms vary diversely based on different field operations as soil management, harvesting processes, pesticide, or fertilizer application. We aimed at evaluating the immunological status of crop farmers, as limited systematic investigations on immune alteration involved with crop farming have been reported yet. MethodsImmunological parameters including plasma immunoglobulin level, major peripheral immune cells distribution, and level of cytokine production from activated T cell were conducted. Nineteen grape orchard, 48 onion open-field, and 21 rose greenhouse farmers were participated. ResultsSignificantly low proportion of natural killer (NK) cell, a core cell for innate immunity, was revealed in the grape farmers (19.8 ± 3.3%) in comparison to the onion farmers (26.4 ± 3.1%) and the rose farmers (26.9 ± 2.5%), whereas cytotoxic T lymphocyte proportion was lower in the grape and the onion farmers than the rose farmers. The proportion of NKT cell, an immune cell implicated with allergic response, was significantly higher in the grape (2.3 ± 0.3%) and the onion (1.6 ± 0.8%) farmers compared with the rose farmers (1.0 ± 0.4%). A significantly decreased interferon-gamma:interleukin-13 ratio was observed from ex vivo stimulated peripheral blood mononuclear cells of grape farmers compared with the other two groups. The grape farmers revealed the lowest levels of plasma IgG1 and IgG4, and their plasma IgE level was not significantly different from that of the onion or the rose farmers. ConclusionOur finding suggests the high vulnerability of workplace-mediated allergic immunity in grape orchard farmers followed by open-field onion farmers and then the rose greenhouse farmers.

Intracellular NAD+ Depletion Confers a Priming Signal for NLRP3 Inflammasome Activation.

Nicotinamide adenine dinucleotide (NAD+) is an important cofactor in many redox and non-redox NAD+-consuming enzyme reactions. Intracellular NAD+ level steadily declines with age, but its role in the innate immune potential of myeloid cells remains elusive. In this study, we explored whether NAD+ depletion by FK866, a highly specific inhibitor of the NAD salvage pathway, can affect pattern recognition receptor-mediated responses in macrophages. NAD+-depleted mouse bone marrow-derived macrophages (BMDMs) exhibited similar levels of proinflammatory cytokine production in response to LPS or poly (I:C) stimulation compared with untreated cells. Instead, FK866 facilitated robust caspase-1 activation in BMDMs in the presence of NLRP3-activating signals such as ATP and nigericin, a potassium ionophore. However, this FK866-mediated caspase-1 activation was completely abolished in Nlrp3-deficient macrophages. FK866 plus nigericin stimulation caused an NLRP3-dependent assembly of inflammasome complex. In contrast, restoration of NAD+ level by supplementation with nicotinamide mononucleotide abrogated the FK866-mediated caspase-1 cleavage. FK866 did not induce or increase the expression levels of NLRP3 and interleukin (IL)-1β but drove mitochondrial retrograde transport into the perinuclear region. FK866-nigericin-induced mitochondrial transport is critical for caspase-1 cleavage in macrophages. Consistent with the in vitro experiments, intradermal coinjection of FK866 and ATP resulted in robust IL-1β expression and caspase-1 activation in the skin of wild-type, but not Nlrp3-deficient mice. Collectively, our data suggest that NAD+ depletion provides a non-transcriptional priming signal for NLRP3 activation via mitochondrial perinuclear clustering, and aging-associated NAD+ decline can trigger NLRP3 inflammasome activation in ATP-rich environments.