IL-4 mRNA Expression Research Articles

IL1RAP expression and the enrichment of IL-33 activation signatures in severe neutrophilic asthma.

Interleukin (IL)-33 is an upstream regulator of type 2 (T2) eosinophilic inflammation and has been proposed as a key driver of some asthma phenotypes. To derive gene signatures from in vitro studies of IL-33-stimulated cells and use these to determine IL-33-associated enrichment patterns in asthma. Signatures downstream of IL-33 stimulation were derived from our in vitro study of human mast cells and from public datasets of in vitro stimulated human basophils, type 2 innate lymphoid cells (ILC2), regulatory T cells (Treg) and endothelial cells. Gene Set Variation Analysis (GSVA) was used to probe U-BIOPRED and ADEPT sputum transcriptomics to determine enrichment scores (ES) for each signature according to asthma severity, sputum granulocyte status and previously defined molecular phenotypes. IL-33-activated gene signatures were cell-specific with little gene overlap. Individual signatures, however, were associated with similar signalling pathways (TNF, NF-κB, IL-17 and JAK/STAT signalling) and immune cell differentiation pathways (Th17, Th1 and Th2 differentiation). ES for IL-33-activated gene signatures were significantly enriched in asthmatic sputum, particularly in patients with neutrophilic and mixed granulocytic phenotypes. IL-33 mRNA expression was not elevated in asthma whereas the expression of mRNA for IL1RL1, the IL-33 receptor, was up-regulated in the sputum of severe eosinophilic asthma. The mRNA expression for IL1RAP, the IL1RL1 co-receptor, was greatest in severe neutrophilic and mixed granulocytic asthma. IL-33-activated gene signatures are elevated in neutrophilic and mixed granulocytic asthma corresponding with IL1RAP co-receptor expression. This suggests incorporating T2-low asthma in anti-IL-33 trials.

LBMPL Vaccine Therapy Induces Progressive Organization of the Spleen Microarchitecture, Improved Th1 Adaptative Immune Response and Control of Parasitism in Leishmania infantum Naturally Infected Dogs.

The spleen plays a central role in human and canine visceral leishmaniasis, where the activation of the immune response occurs in one of the tissues where Leishmania infantum reproduces. Therefore, this organ is both a target to understand the mechanisms involved in the parasite control and a parameter for assessing the therapeutic response. In this sense, this study aimed to evaluate the main histological, immunological and parasitological aspects in the spleen of symptomatic dogs naturally infected by L. infantum treated with the therapeutic vaccine LBMPL. For this, dogs were divided into four groups: dogs uninfected and untreated (NI group); L. infantum-infected dogs that were not treated (INT group); L. infantum-infected dogs that received treatment only with monophosphoryl lipid A adjuvant (MPL group); and L. infantum-infected dogs that received treatment with the vaccine composed by L. braziliensis promastigote proteins associated with MPL adjuvant (LBMPL group). Ninety days after the therapeutics protocol, the dogs were euthanized and the spleen was collected for the proposed evaluations. Our results demonstrated a reduction of hyperplasia of red pulp and follicular area of white pulp, increased mRNA expression of IFN-γ, TNF-α, IL-12 and iNOS, and decreased IL-10 and TGF-β1, and intense reduction of splenic parasitism in dogs treated with the LBMPL vaccine. These results possibly suggest that the pro-inflammatory environment promoted the progressive organization of the splenic architecture favoring the cellular activation, with consequent parasite control. Along with previously obtained data, our results propose the LBMPL vaccine as a possible treatment strategy for canine visceral leishmaniasis (CVL).

Effect of Curcumin as Feed Supplement on Immune Response and Pathological Changes of Broilers Exposed to Aflatoxin B1.

In this study, we examined the protective effects of curcumin against the AFB1-induced immune response of and pathological changes in broilers. Histopathology examinations showed that at day 28, AFB1 (5 mg/kg) exposure leads to severe histological changes in the spleen, thymus and bursa of Fabricius with a decrease in the number and karyoplasmic area ratio of plasma cells. Curcumin alleviated the AFB1-induced immune organs’ damage as well as the changes in plasma cells in a dose-dependent manner. RT-PCR data showed that AFB1 significantly downregulated the IL-2 and IFN-γ mRNA expression levels in the thymus, spleen and bursa of Fabricius. However, curcumin supplementation improved the AFB1-induced immune organs’ damage via upregulated cytokines’ expression. Intriguingly, similar trends were noticed in abnormal morphological changes and the immune response at day 35 after the withdrawal of AFB1 and curcumin from the diet, suggesting the protective effects and immunomodulatory function against AFB1 in broilers. The current study provides a scientific experimental basis for the application of curcumin as a therapeutic drug or additive in animal husbandry productive practice.

2020 Research Grant. Mitochondrial targeted protection following incomplete spinal cord injury with noninvasive technology

BACKGROUND CONTEXT Most of the traumatic spinal cord injury (SCI) are treatable, but despite advances in rehabilitation strategies continued impairment in sensorimotor function persists and does not return to pre-injury levels. Following the initial trauma there is a secondary cascade of events characterized by damage to the vasculature of the spinal cord and, therefore, reducing local oxygen delivery to the mitochondria. The main source of ROS (reactive oxygen species; free radicals) production is the mitochondrial electron transport chain. During tissue injury, calcium is released, which activates mitochondrial proteins. The hyperactivate electron transport chain now generates a mitochondrial membrane potential that is above the optimal range. This, in turn, exponentially increases ROS production, which then initiates cellular death cascades causing further damage to the spinal cord, such as inhibiting neuronal differentiation. PURPOSE The purpose of this study was to use a noninvasive infrared red-light therapy (IRL) which reduces the mitochondrial hyperactivity show immediately after SCI. We hypothesized that IRL will reduce mitochondrial hyperactivity which will result in lower ROS production at the epicenter of SCI. METHODS A between-subjects design was used where male Sprague-Dawley rats (3 months old) were randomized into three groups (sham, SCI and SCI + IRL). A balloon catheter model of spinal cord compression was used to induce SCI. The IRL treatment was for 2 hours under anesthesia. RESULTS We found that IRL therapy significantly (p < 0.03) reduced ROS production at the epicenter of SCI. In addition, we found that IL-8 and IL-12 mRNA expression were significantly lower in the SCI + IRL relative to the SCI group, respectively. CONCLUSIONS The results of this pilot study indicated that IRL treatment reduced mitochondrial hyperactivity associated with the initial 24 hours following SCI. FDA DEVICE/DRUG STATUS This abstract does not discuss or include any applicable devices or drugs. Most of the traumatic spinal cord injury (SCI) are treatable, but despite advances in rehabilitation strategies continued impairment in sensorimotor function persists and does not return to pre-injury levels. Following the initial trauma there is a secondary cascade of events characterized by damage to the vasculature of the spinal cord and, therefore, reducing local oxygen delivery to the mitochondria. The main source of ROS (reactive oxygen species; free radicals) production is the mitochondrial electron transport chain. During tissue injury, calcium is released, which activates mitochondrial proteins. The hyperactivate electron transport chain now generates a mitochondrial membrane potential that is above the optimal range. This, in turn, exponentially increases ROS production, which then initiates cellular death cascades causing further damage to the spinal cord, such as inhibiting neuronal differentiation. The purpose of this study was to use a noninvasive infrared red-light therapy (IRL) which reduces the mitochondrial hyperactivity show immediately after SCI. We hypothesized that IRL will reduce mitochondrial hyperactivity which will result in lower ROS production at the epicenter of SCI. A between-subjects design was used where male Sprague-Dawley rats (3 months old) were randomized into three groups (sham, SCI and SCI + IRL). A balloon catheter model of spinal cord compression was used to induce SCI. The IRL treatment was for 2 hours under anesthesia. We found that IRL therapy significantly (p < 0.03) reduced ROS production at the epicenter of SCI. In addition, we found that IL-8 and IL-12 mRNA expression were significantly lower in the SCI + IRL relative to the SCI group, respectively. The results of this pilot study indicated that IRL treatment reduced mitochondrial hyperactivity associated with the initial 24 hours following SCI.

M2 Macrophages promote IL-33 expression, ILC2 expansion and mucous metaplasia in response to early life rhinovirus infections.

Wheezing-associated rhinovirus (RV) infections are associated with asthma development. We have shown that infection of immature mice with RV induces type 2 cytokine production and mucous metaplasia which is dependent on IL-33 and type 2 innate lymphoid cells (ILC2s) and intensified by a second heterologous RV infection. We hypothesize that M2a macrophages are required for the exaggerated inflammation and mucous metaplasia in response to heterologous RV infection. Wild-type C57Bl/6J mice and LysMCre IL4Rα KO mice lacking M2a macrophages were treated as follows: (1) sham infection on day 6 of life plus sham on day 13 of life, (2) RV-A1B on day 6 plus sham on day 13, (3) sham on day 6 and RV-A2 on day 13, or (4) RV-A1B on day 6 and RV-A2 on day 13. Lungs were harvested one or seven days after the second infection. Wild-type mice infected with RV-A1B at day 6 showed an increased number of Arg1- and Retnla-expressing lung macrophages, indicative of M2a polarization. Compared to wild-type mice infected with RV on day 6 and 13 of life, the lungs of LysMCre IL4Rα KO mice undergoing heterologous RV infection showed decreased protein abundance of the epithelial-derived innate cytokines IL-33, IL-25 and TSLP, decreased ILC2s, decreased mRNA expression of IL-13 and IL-5, and decreased PAS staining. Finally, mRNA analysis and immunofluorescence microscopy of double-infected LysMCre IL4Rα KO mice showed reduced airway epithelial cell IL-33 expression, and treatment with IL-33 restored the exaggerated muco-inflammatory phenotype.ConclusionEarly-life RV infection alters the macrophage response to subsequent heterologous infection, permitting enhanced IL-33 expression, ILC2 expansion and intensified airway inflammation and mucous metaplasia.

Effects of niacin on intestinal epithelial Barrier, intestinal Immunity, and microbial community in weaned piglets challenged by PDCoV

The objective was to evaluate effects of niacin on the intestinal epithelial barrier, intestinal immunity, and microbial community in weaned piglets challenged by Porcine Deltacoronavirus (PDCoV). In this study, fifteen weaned piglets were randomly assigned to 1 of 3 groups, (1) control group, normal diet; (2) PDCoV group, infected with 1 × 107 TCID50 of the PDCoV CHN-HN-17 strain by oral administration; (3) NA + PDCoV group, infected with 1 × 107 TCID50 of the PDCoV CHN-HN-17 strain by oral administration following administration of 40 mg of niacin for three days. The results showed that PDCoV infection induced diarrhea and other clinical symptoms with intestinal villi shedding and atrophy in weaned piglets. Niacin alleviated the symptoms of diarrhea and intestinal damage of PDCoV-infected weaned piglets. Additionally, PDCoV increased (P < 0.05) the mRNA expression of tight junction proteins [zonula occludens-1 (ZO-1) and Claudin] and antimicrobial peptides [porcine β defensin 1 (pBD1), pBD2, proline-arginine rich 39-amino acid peptide (PR39) and protegrin 1–5 (PG1-5) in the jejunum and ileum of weaned piglets, while niacin increased (P < 0.05) the expression of PG1-5 compared with PDCoV. PDCoV increased (P < 0.05) the contents of serum interleukin-1β (IL-1β), IL-8 and intestinal IL-8, and up-regulated the mRNA expression of tumor necrosis factor-α (TNF-α), IL-1β, IL-6, IL-10, IL-12, and IL-18 in ileum of weaned piglets compared with control. However, niacin decreased (P < 0.05) the contents of serum IL-1β, IL-6 and intestinal IL-10 and IL-8, and also reduced (P < 0.05) the mRNA expression of ileal TNF-α, IL-10 and IL-12 in the PDCoV-infected piglets. Compared with control, PDCoV up-regulated (P < 0.05) the mRNA expression of key genes related to innate immune and antiviral molecules [toll-like receptor 4 (TLR4), NOD1, NOD2, DDX58, CCL2, STAT2, Mx1, IFN-γ, and protein kinase R (PKR) in the ileum of weaned piglets. Niacin decreased (P < 0.05) the mRNA expression of NOD1, NOD2, STAT2, IFN-γ, and PKR in PDCoV-infected weaned piglets. Moreover, the mRNA expression of IL-6 decreased (P < 0.05) and 2′-5′-oligoadenylate synthetase (OAS), IFN-α, and PKR increased (P < 0.05) in PDCoV-infected IPEC-J2 cells treated with niacin in vitro. Furthermore, niacin decreased (P < 0.05) the elevation of protein expression including inducible NOS (iNOS), nuclear factor-κB (NF-κB p65), inhibitor kappa B (IKKβ), histone deacetylase [Sirtuin 1 (SIRT1) and histone deacetylase 7 (HDAC7) and phosphorylation of histone H3 at serine s10 (pH3s10) in the ileum of PDCoV-infected piglets, and increased (P < 0.05) the expression of G protein-coupled receptor (GPR109A). PDCoV disrupted the composition and structure of microflora in the colon of weaned piglets, and reduced the relative abundance of the beneficial bacteria Spirobacterium, but niacin could improve the intestinal microbial flora of the PDCoV-infected piglets associated with increasing the relative abundance of Lactobacillus. Overall, niacin could alleviate diarrhea, intestinal barrier damages, intestinal immune response and colonic microflora disfunction in PDCoV-infected weaned piglets.

Psychological Stress Exacerbates Inflammation of the Ileum via the Corticotropin-Releasing Hormone-Mast Cell Axis in a Mouse Model of Eosinophilic Enteritis.

The effects of psychological stress on eosinophilic gastrointestinal disorders have not been elucidated. This study investigated the effects of psychological stress in a mouse model of eosinophilic enteritis (EoN). BALB/c mice were treated with ovalbumin (OVA) to create an EoN model and subjected to either water avoidance stress (WAS) or sham stress (SS). Microscopic inflammation, eosinophil and mast cell counts, mRNA expression, and protein levels of type 2 helper T cell (Th2) cytokines in the ileum were compared between groups. We evaluated ex vivo intestinal permeability using an Ussing chamber. A corticotropin-releasing hormone type 1 receptor (CRH-R1) antagonist was administered before WAS, and its effects were analyzed. WAS significantly increased diarrhea occurrence and, eosinophil and mast cell counts, and decreased the villus/crypt ratio compared to those in the SS group. The mRNA expression of CRH, interleukin IL-4, IL-5, IL-13, eotaxin-1, and mast cell tryptase β2 significantly increased, and the protein levels of IL-5, IL-13, and OVA-specific immunoglobulin E (IgE) also significantly increased in the WAS group. Moreover, WAS significantly increased the intestinal permeability. The CRH-R1 antagonist significantly inhibited all changes induced by WAS. Psychological stress exacerbated ileal inflammation via the CRH-mast cell axis in an EoN mouse model.

Dipeptidyl Peptidase 4 Inhibitor‒Associated Bullous Pemphigoid Is Characterized by an Altered Expression of Cytokines in the Skin

Dipeptidyl peptidase 4 inhibitors (DPP4is), commonly used drugs for treatment of type 2 diabetes, increase the risk for bullous pemphigoid (BP). Currently, the mechanism leading to the loss of immunological tolerance of the cutaneous adhesion molecule BP180 as well as similarities and differences in disease progression between DPP4i-associated BP (DPP4i-BP) and DPP4i-independent regular BP are largely unknown. We analyzed the expression of 32 cytokines and two proteases by Luminex and ELISA assays in samples taken from lesional and nonlesional skin of patients with regular BP or DPP4i-BP and healthy controls. Cytokines mediating B-cell survival and targeting such as BAFF, CCL4, CXCL12, and IL-6 were expressed at a higher level in the lesional regular BP skin than the levels in the lesional DPP4i-BP skin. The DPP4i-BP samples had increased levels of eosinophilic cytokines CCL1, CCL17, CCL26, and IL-5, which correlated with the serum level of anti-BP180 NC16A IgG autoantibodies. The mRNA expression of BAFF, IL6, CCL1, CCL17, CCL26, and IL5 measured by qPCR correlated with the protein levels. Taken together, the cutaneous cytokine profiles were found to provide distinctive molecular fingerprints between regular BP and DPP4i-BP.

Humulus japonicus attenuates LPS-and scopolamine-induced cognitive impairment in mice

BackgroundNeuroinflammation plays an important role in cognitive decline and memory impairment in neurodegenerative disorders. Previously, we demonstrated that Humulus japonicus (HJ) has anti-inflammatory effects in rodent models of Alzheimer’s disease and Parkinson’s disease. The present study aimed to examine the protective potential of HJ extracts against lipopolysaccharide (LPS)-induced cognitive impairment and scopolamine-induced amnesia in mouse models. Cognitive improvement of mice was investigated by novel object recognition test. For analyzing effects on neuroinflammation, immunohistochemistry and quantitative real-time polymerase chain reaction (qRT-PCR) assays were performed.ResultsWe found that the oral administration of HJ significantly improved cognitive dysfunction induced by LPS in a novel object recognition test. The LPS-induced activation of microglia was notably decreased by HJ treatment in the cortex and hippocampus. HJ administration with LPS also significantly increased the mRNA expression of interleukin (IL)-10 and decreased the mRNA expression of IL-12 in the parietal cortex of mice. The increased expression of LPS-induced complement C1q B chain (C1bq) and triggering receptor expressed on myeloid cells 2 (Trem2) genes was significantly suppressed by HJ treatment. In addition, HJ administration significantly improved novel object recognition in a scopolamine-induced amnesia mouse model.ConclusionsThese findings revealed that HJ has a beneficial effect on cognitive impairment and neuroinflammation induced by systemic inflammation and on amnesia induced by scopolamine in mice.

Schistosoma mansoni infection decreases IL-33-mRNA expression and increases CXCL9 and CXCL10 production by peripheral blood cells.

Schistosoma mansoni infections, particularly egg antigens, induce Th2-dominant granulomatous responses accompanied by remarkable immunoregulatory mechanisms that avoid intense fibrosis. Interleukin (IL)-33 is a cytokine that stimulates the early activation of Th2 responses, and its soluble ST2 receptor (sST2) avoids granulomatous response, as well as CXCL9 and CXCL10 chemokines that have antifibrotic activity. However, in schistosomiasis, these molecules have not been suitably studied. Therefore, this study aimed to measure IL-33 and sST2 RNA, cytokines, and chemokines in peripheral blood cultures from individuals living in schistosomiasis-endemic areas. Peripheral blood cells from individuals with S. mansoni (n = 34) and non-infected individuals (n = 31) were cultured under mitogen stimulation. Supernatant chemokines and cytokines were evaluated using a cytometric bead array, and IL-33 and sST2 mRNA expression was measured using qPCR. Infected individuals showed higher levels of CXCL8, CXCL9, CXCL10, IFN-γ, TNF-α, IL-6, IL-2, IL-4, and IL-10; there was a lower expression of IL-33 mRNA and similar expression of sST2mRNA in infected than non-infected individuals. In conclusion, for the first time, we demonstrated lower IL-33mRNA expression and high levels of the antifibrotic chemokines CXCL9 and CXCL10 in schistosomiasis mansoni, which could control exacerbations of the disease in individuals from endemic areas.

Kaempferol-3-O-Rhamnoside Inhibits the Proliferation of Jurkat Cells Through Jun Amino-Terminal Kinase Signaling

Background: Indonesian herbal medicine has become target of new drugs against diseases, including cancer. The high incidence and mortality rate of cancer, anticancer resistance, and side effects of chemotherapy contribute to the urgency of researching novel anticancer drugs. A natural product from Schima wallichii Korth., an Indonesian herbal medicine empirically used for many diseases, has shown anticancer activity in MCF-7 and LNCaP cells. Objective: In this study, we investigated the antiproliferative mechanism of the active compound of S. wallichii, kaempferol-3-O-rhamnoside, against Jurkat cells. Methods: Treated cells were analyzed using a proliferation assay and real time-reverse transcriptase polymerase chain reaction for IL-2 mRNA measurement. The mechanism of antiproliferative activity was assesed by western blotting analysis for Mitogen Activated Protein Kinases (MAPKs). Results: Kaempferol-3-O-rhamnoside has an antiproliferative activity at IC50 of 76.3 μM and slightly inhibited IL-2 mRNA expression. The mechanism to inhibit Jurkat cells proliferation was through the stimulation of phosphorylated Jun amino-terminal kinase. Conclusions: The present study observed the molecular mechanism of antiproliferative activity of kaempferol-3-O-rhamnoside.

The Effect of Oral Administration of the Antibacterial Peptide MPX on Intestinal Inflammation of Mice in Experimental Infection with Escherichia Coli Strain O157: H7

Escherichia coli is a gram-negative bacterium, an intestinal pathogen that can cause intestinal inflammation. Antimicrobial peptides are a class of small molecule peptides, which has good antibacterial activity against a variety of gram-positive and negative bacteria. In this regard, the authors aimed to study the effect of the antimicrobial peptide MPX, which was administered orally, on the intestinal wall of mice infected with the intestinal barrier function, which were infected with E. coli. Synthesis and purification of the antimicrobial peptide MPX (H-INWKGIAAMAKKLL-NH2) was performed by Jier Sheng Hua (Shanghai, China). Mass spectrometry and liquid chromatography (HPLC) were used for this purpose. Escherichia Coli (O157: H7 ATCC43889) was isolated from human faeces in which haemolytic uremic syndrome was reported. Solid LB agar (Solarbio, China) was used to isolate enterohemorrhagic E. coli. The culture was obtained from the Chinese Institute of Veterinary Drug Control (Beijing, China). The results of necropsy found that using of an orally administered MPX could alleviate the damage of E. coli to the liver, spleen, and the lungs were less affected. According to H&amp;E results in case of an orally administered MPX group considerably relieved duodenum and organs on day 7 and day 28. qRT-PCR results showed orally administered MPX could reduce the inflammation-related factors in the mRNA expression of IL-2 and IL-6 and TNF-α on day 7 and day 28. In addition, orally administered MPX could significantly increase them RNA expression of tight junction proteins Occludin and Zo-1 on day 7 and day 28. The results of immune histochemistry further showed that an orally administered MPX could increase the mRNA expression of MUC2 in jejunum. The above results showed that orally administered MPX could alleviate the attack of E. coli on the intestinal tract of mice, relieve intestinal inflammation, and improve the intestinal barrier function. This study lays a theoretical foundation for adding antimicrobial peptides to food. In orally administered MPX authors can see reducing the mRNA expression of inflammation-related factors, thereby alleviating the intestinal inflammation caused by E. coli infection in mice. Authors can add that orally administered MPX shows an increase in mRNA expression of tight junction protein in intestines and improves the intestinal barrier function. This study lays the foundation for adding antimicrobial peptides to food to relieve inflammation and improve barrier function in clinical practices

Serum IL-5, POSTN and IL-33 levels in chronic rhinosinusitis with nasal polyposis correlate with clinical severity

BackgroundChronic rhinosinusitis (CRS) is a group of heterogeneous diseases characterized by epithelial inflammation and tissue eosinophilic infiltration. IL-5, POSTN, and IL-33 are important factors that act as chemoattractants for eosinophils, and a tissue-remodeling protein positively correlated with eosinophils in blood and mediators of eosinophilic infiltration. The aim of the study was to determine the expression of IL-5, POSTN and IL-33, at the gene and protein levels, in eosinophilic CRS with nasal polyps (CRSwNP) and without nasal polyps (CRSsNP), and to correlate this expression with clinical severity.Materials and methodsThe study included 40 CRSwNP patients and 53 CRSsNP patients and 40 control subjects. The expression of IL-5, POSTN and IL-33 mRNA was determined in sinonasal mucosal samples and in nasal polyp tissue by real-time PCR. Protein levels in the serum of CRSwNP patients were measured by ELISA. Computed tomography was evaluated according to Lund–Mackay scores, and visual analog scale scores were assessed.ResultsNP tissue demonstrated significantly higher IL-5 and POSTN mRNA expression than the sinonasal tissue in the CRSsNP and CRSwNP groups. CRS groups demonstrated elevated IL-33 mRNA expression in comparison to controls irrespective of the presence of NP. No correlation was found between IL-5, POSTN and IL-33 mRNA expression and disease severity. CRSwNP group demonstrated significantly higher serum IL-5, POSTN and IL-33 protein levels than controls, and this corresponds to disease severity.ConclusionSerum IL-5, POSTN and IL-33 levels may be important markers for classification of eosinophilic CRSwNP patients, along with disease severity.

Possible association of rotavirus IgG with cytokine expression levels and dyslipidemia in rotavirus-infected type 1 diabetic children

BackgroundRotavirus (RV) has been postulated as a viral trigger for the onset of autoimmune disorders, such as type 1 diabetes (T1D). This study aimed to examine the conceivable association of RV IgG with cytokine levels and dyslipidemia in the pathogenesis of pediatric T1D.MethodsThis study included 30 healthy controls and 80 children with T1D who were divided into two groups based on the time since their T1D diagnosis: newly diagnosed (ND ≤ 1 year; n = 30) and previously diagnosed (PD > 1 year; n = 50). ND and PD patients were also separated into negative and positive according to IgG detection (RV IgG−, ND−, and PD−; RV IgG+, ND+, and PD+).ResultsPositive polymerase chain reaction for RVs was evidenced in 7.5% of children with T1D. Anti-RV IgG was 30% and 36% in ND and PD, respectively, compared to healthy controls (2 of 30, 6.6%; P < 0.05). Fasting blood sugar and hemoglobin A1c significantly increased in PD+ compared to PD−. Interferon-γ and interleukin (IL)-15 levels significantly increased. IL-12 and IL-22 mRNA expression was upregulated in ND+ patients compared to that in ND− patients. IL-37 mRNA expression was significantly downregulated in ND− and ND+ patients compared to that in healthy controls. Total cholesterol and high- and low-density lipoprotein-cholesterol levels were significantly lower in PD+ than in PD−; whereas triglyceride levels were higher than those in healthy controls.ConclusionsThis study suggested that anti-RV IgG may have a role in the pathogenesis, development, and progression of T1D, and RV infections are implicated in dyslipidemia and inflammation status.

Tim-3/Galectin-9 signaling pathway is involved in the cytokine changes in mice with alveolar echinococcosis.

Tim-3/Galectin-9 is involved in the immune escape of many pathogens. However, the role of Tim-3/Galectin-9 in persistent infection of Echinococcus multilocularis (Em), which is related to immune escape, is still unclear. To investigate the role of Tim-3/Galectin-9 and related cytokines in mice with persistent infection of Em. Em infection model was established by injecting the protoscoleces. Serum was collected at days 2, 8, 30, 60, 90, 180 and 270 after infection. Lymphocytes were isolated from liver tissue samples with Ficoll. Tim-3 + CD4 + T percentage was analyzed by flow cytometry. CD4 + T cells were isolated from liver tissues of Em infected mice and cultured in vitro. The mRNA levels of Tim-3, Galectin-9, IFN-γ and IL-4 were detected by qRT-PCR. Cytokine levels in serum and culture supernatant (IFN-γ and IL-4) were analyzed by cytometric bead array. The expression of Tim-3 and Galectin-9 mRNA significantly increased after 30 days of infection, reached peak on day 90, and then decreased slightly on days 180-270. The expression of IFN-γ mRNA, increased on day 2 and 8 after infection, slightly decreased on days 30-60, and obvious decreased on days 90-270, but were still higher than those of the control group. The expression of IL-4 mRNA gradually increased along with the time of infection. In serum of Em infected mice, level of IFN-γ peaked at day 30 and then gradually decreased; whereas IL-4 level peaked at day 90 and then gradually decreased. In vitro experiment found that Tim-3/Galectin-9 directly caused the changes in the levels of IFN-γ and IL-4. Tim-3/Galectin-9 signaling pathway may be involved in the development of persistent infection of Em by regulating the production of Th1 and Th2 cytokines.

Expression of genes encoding interleukin 15 and its receptor subunits in the duodenal and colonic mucosae of dogs with chronic enteropathy

A pro-inflammatory role of interleukin (IL)-15 and IL-15 receptor (R) in chronic intestinal inflammation, such as inflammatory bowel disease, has been reported in humans. However, the contribution of IL-15 signaling in the pathogenesis of canine chronic enteropathy (CE) remains unclear. Therefore, as a first step in elucidating the importance of IL-15 signaling in canine CE, we measured the mRNA expression of IL-15 and IL-15R subunits, including IL-15Rα, IL-15Rβ, and IL-15Rγ, in the duodenal and colonic mucosae of healthy dogs and those with CE, including food-responsive enteropathy (FRE), antibiotic-responsive enteropathy (ARE), and immunosuppressant-responsive enteropathy (IRE). Real-time PCR analysis revealed significantly lower IL-15Rα mRNA expression levels in the duodenal mucosa of dogs with IRE compared to healthy dogs. In contrast, the mRNA expression levels of IL-15, IL-15Rβ, and IL-15Rγ in the duodenal mucosa and IL-15, IL-15Rα, IL-15Rβ, and IL-15Rγ in the colonic mucosa did not differ among healthy dogs and those with FRE, ARE, or IRE. These findings suggest that decreased mRNA expression of IL-15Rα might be involved in the pathogenesis of duodenitis in dogs with IRE. Moreover, even in canine CE, IL-15 signaling appears to play different roles in duodenitis and colitis in dogs with FRE, ARE, and IRE. However, there were no correlations between the gene expression levels of IL-15Rα and clinical severity or histopathological scores in the duodenum of dogs with IRE. Further studies are necessary to investigate the IL-15Rα protein localization and to determine how impaired IL-15Rα expression contributes to the development of duodenitis in dogs with IRE.

Effects of high-grain diet feeding on mucosa-associated bacterial community and gene expression of tight junction proteins and inflammatory cytokines in the small intestine of dairy cattle

The objective of this study was to investigate changes in the composition of mucosa-associated bacterial community, the morphology of the small intestinal epithelia, and the gene expressions of junction proteins and inflammatory cytokines in the small intestines of dairy cattle fed a high-grain (HG) diet. A total of 12 ruminally cannulated Holstein cows in mid-lactation were randomly fed either a conventional (CON) diet (40% concentrate, dry matter basis) or an HG diet (60% concentrate, dry matter basis) for 4 wk. At the end of the feeding trial, all the cows were slaughtered and then examined for changes in the small intestinal mucosa-associated bacterial communities using 16S full-length amplicon sequencing. Furthermore, the gene expression of tight junction proteins and inflammatory cytokines in the small intestinal epithelium were studied using real-time quantitative PCR. The results of nonmetric multidimensional scaling plots showed that an HG diet altered the composition of mucosa-associated bacterial communities in the jejunum and ileum. The HG feeding only increased the numbers of operational taxonomic units in the mucosa-associated bacterial community in the jejunum. At the genus level, the HG diet increased the abundance of uncultured Succinivibrionaceae and Lachnospiraceae incertae sedis in the duodenal mucosa, whereas the proportions of Veillonella and Selenomonas increased in the jejunal mucosa. Compared with the CON group, the proportions of Acetitomaculum in both the jejunal and the ileal mucosa were higher in the HG group. Analysis via PICRUSt2 (version 2.2.0-b) suggested that the HG diet increased the abundance of genes related to biodegradation of xenobiotics in the jejunal mucosa and the abundance of genes related to immune disease in the ileal mucosa. Additionally, the group fed an HG diet had higher concentrations of lipopolysaccharides in the jejunal and ileal digesta. The HG feeding caused a downregulation of the mRNA expression of occludin and ZO-1 in the jejunal epithelium, as well as of claudin-1, claudin-4, and ZO-1 in the ileal epithelium. Moreover, the HG diet caused an increase in the mRNA expression of IL-1β, IL-2, and IFN-γ in the jejunal epithelium, but a higher expression of IL-2 and IFN-γ in the ileal epithelium. Correlation analysis revealed that the alteration of lipopolysaccharide levels and mucosa-associated bacterial community might partly contribute to changes in the expression of the epithelial cytokines in the jejunum and ileum during HG feeding. These findings suggest that microbiota residing in the small intestine provide essential health benefits to host dairy cattle.

Enhanced production of IL-2 from anti-CD3 antibody-stimulated mouse spleen cells by artepillin C, a major component of Brazilian green propolis

ObjectivesIn this report, we attempt to clarify the immune modulatory effects of Brazilian green propolis (BGP) and its major component, artepillin C, on the cytokine production of anti-CD3 antibody-stimulated mouse spleen cells. We also estimate the physiological mechanism affecting artepillin C's upon the cells. MethodsMale C3H/HeN mouse spleen cells stimulated by antiCD3 monoclonal antibody were co-cultured with BGP, artepillin C, and HC030031, a transient receptor potential ankyrin 1 (TRPA1) Ca2+ channel antagonist. The synthesis of interferon (IFN)-γ, interleukin (IL)-6, IL-17, IL-4, IL-10, and IL-2 was assayed by enzyme-linked immunoassay. The expression of IL-2 mRNA and the protein product were examined by reverse transcription-quantitative polymerase chain reaction and Western blot analyses, respectively. ResultsThe production of IL-2 was markedly enhanced, while that of IL-4 and IL-10 was not significantly affected; by contrast, the production of IFN-γ, IL-6, and IL-17 was significantly reduced in the antibody-stimulated spleen cells treated with BGP at a non-cytostatic concentration. These effects were reproduced in the cells treated with artepillin C. The expression of IL-2 mRNA was unaffected; however, that of the protein was significantly enhanced in the artepillin C-treated cells compared to untreated control cells. The enhancement of protein expression and the production of IL-2 by artepillin C was significantly alleviated by adding HC030031. ConclusionsArtepillin C is an important regulator of cytokine synthesis from activated spleen cells. The agent specifically augmented the expression of IL-2 via the Ca2+-permeable cation channel, TRPA1, at least in part, at the translational or secretion levels.

Sex-specific association between placental inflammatory cytokine mRNA expression and preschoolers’ behavioral development: The Ma’anshan birth cohort study

BackgroundPlacental inflammation may contribute to brain abnormalities and childhood neuropsychiatric disorders, but limited knowledge is available on the association of placental inflammatory cytokine levels and offspring’s behavioral development. This study aimed to examine the sex-specific association between placental inflammatory cytokine mRNA expression and preschoolers’ behavioral development. Methods3474 pregnant women were recruited as the initial study population in the Ma’anshan birth cohort (MABC) study. Placentas (n = 2519) were collected during childbirth, and the mRNA expression of IL-8, IL-1β, CRP, TNF-α, IL-6, IL-10, and IL-4 was assessed. The Child Behavior Checklist 1.5–5 (CBCL 1.5–5) was used to assess children’s behavioral development at 4 years old. A T-score ≥ 60 on summary scales or a score ≥ 65 on syndrome scales was regarded as the borderline clinical range. Multiple linear regression models and binary logistic regression models were applied to explore the sex-specific associations between placental inflammatory cytokines mRNA transcript levels and preschoolers’ behavioral development. ResultsSex-specific associations between placental inflammatory cytokines mRNA expression and preschoolers’ behavioral development were observed. There was a positive association between IL-8 and CBCL scores for boys on anxious/depressed problems, aggressive behaviors, externalizing problems and total problems. Logistic regression models showed that high levels of IL-8 were associated with a higher risk of girls’ emotionally reactive problems and sleep problems compared to low/medium levels. High TNF-α was correlated with increased sleep problem scores in boys, and medium TNF-α (vs. low levels) was associated with an increased risk of girls’ externalizing problems. Medium levels of CRP, IL-1β, and IL-6 were found to be associated with a decreased risk of girls’ behavioral problems compared to low/high levels. For anti-inflammatory cytokines, medium IL-10 and IL-4 (vs. low levels) were observed to be associated with a lower risk of internalizing problems in boys and externalizing problems in girls, respectively. High IL-10 was correlated with decreased attention problem scores in boys. ConclusionThis study indicates that placental inflammatory cytokine mRNA expression of IL-8, CRP, TNF-α, IL-1β, IL-4 and IL-10 may be associated with preschoolers’ behavioral development in a sex-specific manner.

Multi-omic and spatial dissection of immunotherapy response groups in non–small cell lung cancer (NSCLC).

8544 Background: Immune checkpoint inhibitors (ICI) have shown durable benefit in a subset of non-small cell lung cancer (NSCLC) patients. The composition of the tumour microenvironment (TME) is becomingly increasingly recognised as an important factor to predict response to therapy. Methods: Here, we applied digital spatial profiling of the tumour and stromal compartments from a 2nd line NSCLC ICI-treated cohort (n = 41 patient) and standard of care (SOC), platinum treated NSCLC cohort (n = 47), to identify tissue-based signatures of response to therapy. Results: We demonstrate by mIHC that the interaction of CD68+ macrophages with PD1+, FoxP3+ cells is significantly enriched in ICI refractory tumours (p = 0.012). Patients sensitive to ICI therapy expressed higher levels of IL2 receptor alpha (CD25, p = 0.028) within the tumour compartments, which corresponded with the increased expression of IL2 mRNA (p = 0.001) within their stroma. Immuno-inhibitory markers CTLA-4 (p = 0.021) and IDO-1 (p = 0.023) were supressed in ICI-responsive patients. Tumour CD44 (p = 0.02) was depleted in the response group and corresponded inversely with significantly higher stromal expression of one of its ligands, SPP1 (osteopontin, p = 0.008). Analysis of dysregulated transcripts indicated the potential inhibition of stromal interferon-gamma (IFNγ) activity, estrogen-receptor and Wnt-1 signalling activity within the tumour cells of ICI responsive patients. Cox survival analysis indicated tumour CD44 expression was associated with poorer prognosis (HR = 1.61, p = 0.01), consistent with its depletion in ICI sensitive patients. Similarly, stromal CTLA-4 (HR = 1.78, p = 0.003) and MDSC/M2 macrophage marker ARG1 (HR = 2.37, p = 0.01) were associated with poorer outcome while BAD (HR = 0.5, p = 0.01) appeared protective. The SOC cohort paralleled similar roles for immune checkpoints and pro-apoptotic markers, with LAG3 (HR = 3.81, p = 0.04) indicating poorer outcome, and BIM (HR = 0.16, p = 0.014) with improved outcome. Interestingly, stromal mRNA for E-selectin (HR = 652, p = 0.001), CCL17 (HR = 70, p = 0.006) and MTOR (HR = 1065, p = 0.008) were highly associated with poorer outcome in ICI treated patients, indicating pro-tumourigenic features in the tumour microenvironment that may facilitate ICI resistance. Conclusions: Through multi-modal approaches, we have dissected the characteristics of NSCLC treatment groups and provide evidence for the role of several markers including IL2, CD25, CD44 and SPP1 in the efficacy of current generations of ICI therapy.