Proinflammatory Cytokine Gene Expression Research Articles

Pseudomonas quinolone signal induces organelle stress and dysregulates inflammation in human macrophages

Pseudomonas quinolone signal (PQS) is a quorum-sensing molecule associated with Pseudomonas aeruginosa that regulates quorum sensing, extracellular vesicle biogenesis, iron acquisition, and the secretion of virulence factors. PQS has been shown to have immunomodulatory effects on the host. It induces oxidative stress, modulates cytokine levels, and activates regulated cell death in the host. In this study, we investigated the effects of PQS (10 μM) on host organelle dynamics and dysfunction in human macrophages at the interphase of endoplasmic reticulum (ER), mitochondria, and lysosome. This study showed that PQS increases cytosolic Ca+2 levels and elevates ER stress, as evidenced by increased expression of BiP and activation of the PERK-CHOP axis of unfolded protein response (UPR). Moreover, PQS also negatively affects mitochondria by disrupting mitochondrial membrane potential and increasing mitochondrial ROS generation (mROS). Additionally, PQS stimulation decreased the number of acridine orange-positive lysosomes, indicating lysosomal destabilization. Furthermore, PQS-induced lysosomal destabilization also induces overexpression of the lysosomal stress-responsive gene TFEB. Besides organelle dysfunction, PQS dysregulates inflammation-related genes by upregulating NLRC4, TMS1, and Caspase 1 while downregulating NLRP3 and IL-1β. Also, PQS increases gene expression of pro-inflammatory cytokines (IL-6, TNF-α, and IFN-γ). In conclusion, our findings suggest that PQS negatively affects human macrophages by interfering with organelle function and dysregulating inflammatory response. Consequently, this study provides crucial insight into PQS-driven macrophage dysfunction and may contribute to a better understanding of Pseudomonas aeruginosa-associated infections.

Porphyromonas gingivalis outer membrane vesicles in cerebral ventricles activate microglia in mice.

Porphyromonas gingivalis (Pg) is thought to be involved in the progression of Alzheimer's disease (AD). Whether Pg or its contents can reach the brain and directly affect neuropathology is, however, unknown. Here, we investigated whether outer membrane vesicles (OMVs) of Pg translocate to the brain and induce the pathogenic features of AD. Pg OMVs were injected into the abdominal cavity of mice for 12 weeks. Pg OMV translocation to the brain was detected by immunohistochemistry using an anti-gingipain antibody. Tau protein and microglial activation in the mouse brain were examined by western blotting and immunohistochemistry. The effect of gingipains on inflammation was assessed by real-time polymerase chain reaction using human microglial HMC3 cells. Gingipains were detected in the region around cerebral ventricles, choroid plexus, and ventricular ependymal cells in Pg OMV-administered mice. Tau and phosphorylated Tau protein increased and microglia were activated. Pg OMVs also increased the gene expression of proinflammatory cytokines in HMC3 cells in a gingipain-dependent manner. Pg OMVs, including gingipains, can reach the cerebral ventricle and induce neuroinflammation by activating microglia. Pg OMVs may provide a better understanding of the implications of periodontal diseases in neurodegenerative conditions such as AD.

In vitro phenotypic effects of Lipoxin A4 on M1 and M2 polarized macrophages derived from THP-1.

Lipoxin A4 (LXA4) is a specialized pro-resolving mediator involved in the resolution phase of inflammation that is crucial for the return of tissues to homeostasis, healing, and regenerative processes. LXA4 can modify the microenvironment via its receptor, formyl peptide receptor 2 (FPR2) and thus modulate the inflammatory response. However, the effect of exogeneous LXA4 application on polarized macrophages remains unstudied. The objective of this study was to assess the effect of LXA4 on macrophage activity and on the phenotype modulation of polarized M1 and M2 macrophages derived from THP-1 monocytes. Once differentiated, human macrophages were incubated with interleukin 4 (IL-4) and IL-13 to obtain M2-polarized macrophages or with interferon gamma and lipopolysaccharide for classical macrophage activation. The mRNA and protein expression of M1 and M2 markers confirmed the polarization of THP-1-derived macrophages. LXA4 (0-100nM) did not affect the viability of M1 and M2 macrophages or the phagocytic activity of these cells. Gene expression of FPR2, referred as a receptor for the LXA4, was higher in M1 compared with M2, and was not modified by the LXA4 at the doses used. Moreover, LXA4 exhibited anti-inflammatory properties illustrated by the decreasing in the gene expression of pro-inflammatory cytokines (IL-6, tumor necrosis factor alpha, IL-1β) in M1 and by the increase in the expression of anti-inflammatory cytokines (IL-10) in M2 macrophages. These results provide new insights regarding the potential of LXA4 to regulate the polarization state of macrophages.

Effects of enzymatically hydrolyzed yeast supplementation on blood attributes, antioxidant status and gene expression of cytokines in vaccinated dairy cows

This study was done to assess the effects of hydrolyzed yeast on blood cells counts, antioxidant status and gene expression of pro-inflammatory and anti-inflammatory cytokines of vaccinated dairy cows. Twenty Holstein lactating dairy cows were randomly divided into four groups received basal diet without (control) and with 10, 20 and 30 g/d/head of hydrolyzed yeast. Cows were vaccinated against foot and mouth disease virus at day 10 and blood samples were taken 10 days after. Cows received hydrolyzed yeast at doses of 20 and 30 g/d/head had the lowest and those in the control group had the highest malondialdehyde concentration. Cows received hydrolyzed yeast at dose of 20 g/d/head had the highest lymphocyte and the lowest neutrophil/lymphocyte ratio. IL-1β and TNF-α downregulated and INF-γ up-regulated in cows received different doses of hydrolyzed yeast. The lowest expression of IL-1β and TNF-α and the highest expression of INF-γ was for cows received hydrolyzed yeast at dose of 20 g/d/head. The highest titers against foot and mouth disease virus were for cows received hydrolyzed yeast at dose of 20 g/d/head and the lowest titer was for the control group. Supplementation of 20 g/d/head hydrolyzed yeast recommended in vaccinated cows.

Single cell analysis of docosahexaenoic acid suppression of sequential LPS-induced proinflammatory and interferon-regulated gene expression in the macrophage.

Preclinical and clinical studies suggest that consumption of long chain omega-3 polyunsaturated fatty acids (PUFAs) reduces severity of chronic inflammatory and autoimmune diseases. While these ameliorative effects are conventionally associated with downregulated expression of proinflammatory cytokine and chemokine genes, our laboratory has recently identified Type 1 interferon (IFN1)-regulated gene expression to be another key target of omega-3 PUFAs. Here we used single cell RNA sequencing (scRNAseq) to gain new mechanistic perspectives on how the omega-3 PUFA docosahexaenoic acid (DHA) influences TLR4-driven proinflammatory and IFN1-regulated gene expression in a novel self-renewing murine fetal liver-derived macrophage (FLM) model. FLMs were cultured with 25 µM DHA or vehicle for 24h, treated with modest concentration of LPS (20 ng/ml) for 1 and 4h, and then subjected to scRNAseq using the 10X Chromium System. At 0h (i.e., in the absence of LPS), DHA increased expression of genes associated with the NRF2 antioxidant response (e.g. Sqstm1, Hmox1, Chchd10) and metal homeostasis (e.g.Mt1, Mt2, Ftl1, Fth1), both of which are consistent with DHA-induced polarization of FLMs to a more anti-inflammatory phenotype. At 1h post-LPS treatment, DHA inhibited LPS-induced cholesterol synthesis genes (e.g. Scd1, Scd2, Pmvk, Cyp51, Hmgcs1, and Fdps) which potentially could contribute to interference with TLR4-mediated inflammatory signaling. At 4h post-LPS treatment, LPS-treated FLMs reflected a more robust inflammatory response including upregulation of proinflammatory cytokine (e.g. Il1a, Il1b, Tnf) and chemokine (e.g.Ccl2, Ccl3, Ccl4, Ccl7) genes as well as IFN1-regulated genes (e.g. Irf7, Mx1, Oasl1, Ifit1), many of which were suppressed by DHA. Using single-cell regulatory network inference and clustering (SCENIC) to identify gene expression networks, we found DHA modestly downregulated LPS-induced expression of NF-κB-target genes. Importantly, LPS induced a subset of FLMs simultaneously expressing NF-κB- and IRF7/STAT1/STAT2-target genes that were conspicuously absent in DHA-pretreated FLMs. Thus, DHA potently targeted both the NF-κB and the IFN1 responses. Altogether, scRNAseq generated a valuable dataset that provides new insights into multiple overlapping mechanisms by which DHA may transcriptionally or post-transcriptionally regulate LPS-induced proinflammatory and IFN1-driven responses in macrophages.

Aucubin administration suppresses STING signaling and mitigated high-fat diet-induced atherosclerosis and steatohepatosis in LDL receptor deficient mice.

The rising obesity epidemic in developed countries is associated with many chronic inflammatory diseases including atherosclerosis and nonalcoholic steatohepatitis (NASH). Consuming aucubin may benefit health by suppressing inflammation. Herein, we studied the effects of aucubin consumption on atherosclerosis and NASH progression induced by high-fat diet (HFD) in LDL receptor deficient (LDLr-/-) mice. Adult LDLr-/- mice were fed with HFD for 12 weeks and received oral administration of aucubin for the last 6 weeks. Aucubin did not alter body weight or dyslipidemia, but lowered hyperglycemia and mitigated HFD-induced atherosclerosis and hepatic impairments in LDLr-/- mice. Aucubin administration inhibited HFD-induced inflammation and downregulated mRNA and protein expression of stimulator of IFN genes (STING) in both aortas and livers of LDLr-/- mice. In vitro, aucubin suppressed mitochondrial DNA (mtDNA)-induced activation of STING/NFκB pathway and downregulated gene expression of pro-inflammatory cytokines in cultured bone marrow-derived macrophages (BMDM). Furthermore, aucubin enhanced microRNA-181a-5p (miR-181a-5p) levels in both aortas and livers of LDLr-/- mice. Importantly, miR-181a-5p mimicked the inhibitory effect of aucubin on STING/NFκB pathway and inflammation in BMDM. In conclusion, aucubin consumption attenuated HFD-induced atherosclerosis and NASH progression in LDLr-/- mice, possibly through modulating miR-181a-5p/STING and inhibiting inflammation.

ODP254 WFS1 Loss of Function Causes ER Stress-Mediated Inflammation in Pancreatic β-Cells

Abstract Wolfram syndrome is a rare genetic disorder characterized by juvenile-onset diabetes mellitus, optic nerve atrophy, hearing loss, diabetes insipidus, and progressive neurodegeneration. Pathogenic variants in the WFS1 gene are the main causes of Wolfram syndrome. WFS1 encodes a transmembrane protein localized to the endoplasmic reticulum (ER) and regulates the unfolded protein response (UPR). Loss of function of WFS1 leads to dysregulation of insulin production and secretion, ER calcium depletion, and cytosolic calpains activation, resulting in the activation of apoptotic cascades. Although the terminal UPR is well known to yield an inflammatory response called "sterilized inflammation" that accelerates β-cell dysfunction and death in diabetes, the contribution of β-cell inflammation to the development of diabetes in Wolfram syndrome has not been fully understood. Here we show that WFS1-deficiency enhances the gene expression of pro-inflammatory cytokines and chemokines, leading to cytokine-induced ER-stress and cell death in pancreatic β-cells. Under thechronic[BL1] high-glucose condition, CRISPR/Cas9-mediated Wfs1-knockout (KO) INS-1 832/13 cells, the pancreatic β-cell model of Wolfram syndrome, showed enhanced gene expression levels of ER stress markers (Chop, sXbp1, Bip, and Txnip). Moreover, the gene expression levels of pro-inflammatory cytokines (Il-1β, Il-6), chemokine (Ccl2), and vascular endothelial growth factor A (VegfA) were higher in Wfs1- KO INS-1 832/13 cells when compared to Wfs1-wild type INS-1 832/13 cells. [BL2] The high glucose-induced pro-inflammatory cytokine genes in Wfs1-KO INS-1 832/13 cells were mediated by the PKR-like ER-resident kinase (PERK) and inositol-requiring enzyme 1α (IRE1α)pathways. Furthermore, the IFN-γ and IL-1β treatment enhanced ER stress-mediated cell death and upregulated the pro-inflammatory cytokine gene expression in Wfs1-KO INS-1 832/13 cells. To confirm these findings, we investigated whether inflammation occurs in a mouse model of Wolfram syndrome. Compared to wild-type mice, 10-month-old whole body Wfs1-KO mouse islets showed a significantly higher density of Iba1, which is a marker for macrophage expression. The islet-infiltrated macrophages in these Wfs1-KO mice expressed the M1-macrophage marker (CD68). There was also strong fibrosis and a high density of endothelial cell marker (CD31) in the Wfs1-KO mouse islets. These results demonstrate that WFS1 loss-of-function enhances the inflammatory responses in pancreatic β-cells. The results in our study identified sterile inflammation as a new pathological mechanism in the development of diabetes in Wolfram syndrome. In Wolfram syndrome model mice, islets are highly infiltrated with macrophages, suggesting that the vicious cycle of ER stress and sterile inflammation accelerates the progression of diabetes in a cell-autonomous and cell-nonautonomous manner. A deeper understanding of the pathophysiology of Wolfram syndrome is essential for developing novel therapeutic targets for the disease. Presentation: No date and time listed

The targeted anti-Salmonella bacteriophage attenuated the inflammatory response of laying hens challenged with Salmonella Gallinarum

Fowl typhoid is a severe disease caused by Salmonella Gallinarum with considerable mortality and morbidity in laying hen farms. The current study has focused on controlling the infection in laying hens using anti-Salmonella spp. bacteriophage. The treatments included, PC, without challenge; NC, S. Gallinarum challenged (SGC); B5, 5 mg bacteriophage/kg+SGC; B10, 10 mg bacteriophage/kg+SGC. The Salmonella shedding, inflammatory responses, and gene expression of pro-inflammatory cytokines, toll-like receptor (TLR), and heat shock protein (HSP) in the jejunum, liver, and thigh muscle were tested in laying hens. Supplementation of bacteriophage reduced the abundance of S. Gallinarum in the excreta at d 3, 7, and 14. The abundance of S. Gallinarum was lower in the B10 than the B5 at d 7. Supplementation of bacteriophage decreased the abundance of S. Gallinarum in the oviduct, spleen, and cecum at d 14. The laying hens in the NC group showed an increased relative spleen weight compared with the PC and B10 treatments. Among the SGC treatments, the NC treatment showed higher gene expressions of IL-4 compared with the B5, higher gene expressions of interferon (IFNγ), TLR4, and tumor necrosis factor-α (TNF-α) compared with the B5 and B10, and higher gene expressions of HSP27 compared with the B10 in the jejunum. Dietary supplementation of B10 decreased the mRNA expressions of TLR4 and TNF-α compared with the B5 treatment in the jejunum. The NC treatment showed the highest gene expressions of HSP27, TLR4, and TNF-α in the liver. Dietary supplementation of B10 showed lower mRNA expressions of HSP27 compared with the B5 treatment in the liver. Moreover, the IFNγ and HSP27 were upregulated in the NC treatment compared with the B5 and B10 in the muscle. In conclusion, it can be suggested that bacteriophage is an effective supplement to control S. Gallinarum infection in laying hens and possibly lower horizontal contaminations in laying hen flocks.

A class of anti-inflammatory lipids decrease with aging in the central nervous system.

Lipids contribute to the structure, development, and function of healthy brains. Dysregulated lipid metabolism is linked to aging and diseased brains. However, our understanding of lipid metabolism in aging brains remains limited. Here we examined the brain lipidome of mice across their lifespan using untargeted lipidomics. Co-expression network analysis highlighted a progressive decrease in 3-sulfogalactosyl diacylglycerols (SGDGs) and SGDG pathway members, including the potential degradation products lyso-SGDGs. SGDGs show an age-related decline specifically in the central nervous system and are associated with myelination. We also found that an SGDG dramatically suppresses LPS-induced gene expression and release of pro-inflammatory cytokines from macrophages and microglia by acting on the NF-κB pathway. The detection of SGDGs in human and macaque brains establishes their evolutionary conservation. This work enhances interest in SGDGs regarding their roles in aging and inflammatory diseases and highlights the complexity of the brain lipidome and potential biological functions in aging.

Lactobacillus paracaseiN1115 attenuates obesity in high-fat diet-induced obese mice.

Disruption of the microbial structure of intestinal bacteria due to a high-fat diet (HFD) is closely associated with metabolic disorders, such as obesity and type 2 diabetes. Probiotics are known to modulate the gut microbiota; therefore, we demonstrated the capability of Lactobacillus paracasei N1115 (LC-N1115) to attenuate obesity. Four-week-old male C57BL/6J mice were fed a HFD for 12 weeks to induce obesity and were then randomized to supplemented placebo or LC-N1115 treatment group for another 12 weeks. LC-N1115 treatment reduced weight gain and liver fat accumulation as well as triglyceride, total cholesterol, and low-density lipoprotein cholesterol levels. The administration of LC-N1115 suppressed the expression of fatty acid synthase, interleukin-1 β, and toll-like receptor 4. Notably, the operational taxonomic units that negatively and positively correlated with the obesity phenotypes were enriched and reduced, respectively, in the LC-N1115 treatment group. These results indicate that LC-N1115 attenuates obesity by modulating the gut microbiota and the expression of lipid synthesis and proinflammatory cytokine genes.

Predicting the efficacy of tofacitinib therapy based on gene expression of proinflammatory cytokines and proteases in cultured blood cells of patients with rheumatoid arthritis

The effectiveness of personalized therapy for rheumatoid arthritis (RA) is associated with the correct choice of the drug and the ability to predict its effect before starting the treatment.Objective: to study in patients with RA the relationship between results of therapy and initial expression of genes responsible for bone and articular cartilage resorption (matrix metalloproteinase 9 – MMP9, – cathepsin K) and inflammation (tumor necrosis factor α – TNFα – and interleukin 1β – IL1β) in mononuclear cells of peripheral blood (PBMC), cultured with tofacitinib (TOFA).Patients and methods. We examined 12 patients with RA who had not previously received TOFA. The average age of the patients was 51 years, the average duration of the disease was 37.6 months. After 3 months of TOFA therapy, 6 patients achieved remission, while the rest had high and moderate disease activity. PBMC were isolated before therapy using a Ficoll density gradient and cultured in the presence of 100 nM TOFA for 48 h. Total RNA obtained from these cells was used to analyze the expression of MMP9, cathepsin K, IL1β, and TNFα genes using a real-time quantitative reverse transcription polymerase chain reaction.Results and discussion. TOFA is able to modify gene expression in cultured PBMC from RA patients compared to control cells. The initial expression of all the studied genes was significantly increased in cultured with TOFA cells of patients with persistent high and moderate disease activity during therapy, while TNFα gene expression was significantly reduced in patients who achieved remission.Conclusion. In patients with RA who have not previously received TOFA, a decrease in TNFα gene expression in blood cells cultured with this drug before the start of therapy may be a prognostic biomarker for achieving remission during TOFA therapy.

Genomic Basis for Individual Differences in Susceptibility to the Neurotoxic Effects of Diesel Exhaust.

Air pollution is a known environmental health hazard. A major source of air pollution includes diesel exhaust (DE). Initially, research on DE focused on respiratory morbidities; however, more recently, exposures to DE have been associated with neurological developmental disorders and neurodegeneration. In this study, we investigated the effects of sub-chronic inhalation exposure to DE on neuroinflammatory markers in two inbred mouse strains and both sexes, including whole transcriptome examination of the medial prefrontal cortex. We exposed aged male and female C57BL/6J (B6) and DBA/2J (D2) mice to DE, which was cooled and diluted with HEPA-filtered compressed air for 2 h per day, 5 days a week, for 4 weeks. Control animals were exposed to HEPA-filtered air on the same schedule as DE-exposed animals. The prefrontal cortex was harvested and analyzed for proinflammatory cytokine gene expression (Il1β, Il6, Tnfα) and transcriptome-wide response by RNA-seq. We observed differential cytokine gene expression between strains and sexes in the DE-exposed vs. control-exposed groups for Il1β, Tnfα, and Il6. For RNA-seq, we identified 150 differentially expressed genes between air and DE treatment related to natural killer cell-mediated cytotoxicity per Kyoto Encyclopedia of Genes and Genomes pathways. Overall, our data show differential strain-related effects of DE on neuroinflammation and neurotoxicity and demonstrate that B6 are more susceptible than D2 to gene expression changes due to DE exposures than D2. These results are important because B6 mice are often used as the default mouse model for DE studies and strain-related effects of DE neurotoxicity warrant expanded studies.

Effects of exogenous taurine supplementation on the growth, antioxidant capacity, intestine immunity, and resistance against Streptococcus agalactiae in juvenile golden pompano (Trachinotus ovatus) fed with a low-fishmeal diet.

Taurine has various biological functions in fish, playing an essential role in growth, resistance to oxidative stress, and intestine immunity. Here, we evaluated the effects of exogenous taurine added to low-fishmeal diets on the growth, anti-oxidative stress, intestine immunity, and Streptococcus agalactiae resistance in juvenile golden pompano (Trachinotus ovatus). Our study showed that exogenous taurine supplementation of 1.2% (T3 group) greatly enhanced the weight gain rate and specific growth rate (SGR) of juvenile golden pompano, significantly upregulating growth-related factor expression in the brain and liver, as well as the levels of growth-related parameters in the serum. Polynomial regression analysis using SGR estimated the optimal dietary taurine level for golden pompano at 1.18%. Moderate exogenous taurine also increased the muscular thickness and villus length within the intestine, maintained intestinal physical barrier stability, activated the Nrf2/Keap-1/HO-1 signaling pathway, increased intestinal antioxidant enzyme gene expression and antioxidant enzyme activity in the serum, and upregulated immunoglobulin and complement levels in parallel with declining reactive oxygen species (ROS) levels in the serum. Antioxidant factor expression was also upregulated in the intestine. Furthermore, supplementation suppressed NF-κB signaling and intestinal pro-inflammatory cytokine gene expression, increased anti-inflammatory cytokine gene expression, and improved intestine immunity. Finally, taurine supplementation improved the survival rate of golden pompano challenged with S. agalactiae. Overall, our findings provide additional information and support for the rational use of taurine in healthy aquatic animal farming.

Inhibition of Pseudomonas aeruginosa LPS-Induced airway inflammation by RIPK3 in human airway.

Although the physiological function of receptor-interacting protein kinase (RIPK) 3 has emerged as a critical mediator of programmed necrosis/necroptosis, the intracellular role it plays as an attenuator in human lungs and human bronchial epithelia remains unclear. Here, we show that the expression of RIPK3 dramatically decreased in the inflamed tissues of human lungs, and moved from the nucleus to the cytoplasm. The overexpression of RIPK3 dramatically increased F-actin formation and decreased the expression of genes for pro-inflammatory cytokines (IL-6 and IL-1β), but not siRNA-RIPK3. Interestingly, whereas RIPK3 was bound to histone 1b without LPS stimulation, the interaction between them was disrupted after 15 min of LPS treatment. Histone methylation could not maintain the binding of RIPK3 and activated movement towards the cytoplasm. In the cytoplasm, overexpressed RIPK3 continuously attenuated pro-inflammatory cytokine gene expression by inhibiting NF-κB activation, preventing the progression of inflammation during Pseudomonas aeruginosa infection. Our data indicated that RIPK3 is critical for the regulation of the LPS-induced inflammatory microenvironment. Therefore, we suggest that RIPK3 is a potential therapeutic candidate for bacterial infection-induced pulmonary inflammation.

Antimicrobial peptide hepcidin contributes to restoration of the intestinal flora after Aeromonas hydrophila infection in Acrossocheilus fasciatus

Hepcidin is a cysteine-rich antimicrobial peptide that serves an important role in the immunity system of fishes. It exhibits antibacterial, antifungal, antiviral, and antitumor activities. However, the exact role of fish hepcidin in the regulation of the intestinal flora still remains a mystery. In our study, we sequenced and characterized hepcidin from the liver of Acrossocheilus fasciatus. Phylogenetic tree analysis showed that A. fasciatus hepcidin and Gobiocypris rarus hepcidin were the most closely related, and both belonged to the fish HAMP1 cluster. Studies conducted on in vivo tissue distribution showed that the expression of hepcidin was highest in healthy A. fasciatus liver. Aeromonas hydrophila infection was confirmed by the increased expression of pro-inflammatory cytokine genes and bacterial loads in A. fasciatus tissues. After A. hydrophila infection, hepcidin expression significantly increased in the liver, spleen, and head kidney. In vitro antibacterial assays showed that hepcidin exhibits strong broad spectrum antibacterial activity. Furthermore, we examined the regulatory effect of hepcidin on the intestinal flora and found that A. fasciatus hepcidin restored the reduced diversity and compositional changes in intestinal flora caused by A. hydrophila infection. Our results suggest that hepcidin could regulate the intestinal flora in fishes; however, the underlying mechanisms need to be explored in greater detail.

Analysis of local expression of inflammatory cytokines genes in patients with gastroesophageal reflux disease and its association with types and number of gastroesophageal refluxes

The aim of the study was to evaluate gene expression of proinflammatory cytokines in esophageal mucosa in patients with erosive esophagitis (EE) and non-erosive form (NERD) of gastroesophageal reflux disease (GERD) and perform correlation analysis between the genes expression and types and number of gastroesophageal refluxes. Methods. This was a single-center prospective study. Esophageal mucosa samples were taken from lower part of the esophagus during endoscopy. Expression of IL-1b, IL-10, IL-18, TNFA, TLR4, GATA3, CD68 and beta-2 macroglobulin genes was assessed with ImmunoQuantex assays. Multichannel intraluminal esophageal pH-impedance studies were performed with Ohmega recorder and 2pH-6impedance catheters. Results. Data of 60 patients (females - 34; mean age (M±SD) 54.6 years, 20 - with erosive esophagitis) were available for the analysis. In those with EE, there was higher expression of IL-18 (5.89±0.4 vs 5.28±1.1, p=0.04), GATA3 (2.92±0.86 vs 2.23±0.96, p=0.03) genes, and inflammatory index (92.12±32.41% vs 70.1±30.8%, p=0.02) compared to NERD. Direct correlation was found between acid exposure time and expression of IL-1b (Spearman rank R = 0.29), IL-18 (R=0.31), TNFA (R=0.35), GATA3 (R=0.34), TLR4 (R=0.29), CD68 (R=0.37) genes and inflammatory index (R=0.3). Indirect correlation was revealed between mean рН and expressions of IL-18 (R= -0.28), TNFA (R= -0.33), GATA3 (R= -0.28), TLR4 (R= -0.28), CD68 (R= -0.39) genes and inflammatory index (R= -0.17). No correlation was found between the genes’ expression and number of gastroesophageal refluxes. Conclusions: Expression of proinflammatory cytokines’ genes differ in patients with EE and NERD. There is a correlation between expression of studied genes and esophageal acidity.

Silver-Silica nanoparticles induced dose-dependent modulation of histopathological, immunohistochemical, ultrastructural, proinflammatory, and immune status of broiler chickens

Silver nanoparticles (AgNPs) are a powerful disinfectant, but little information is available on their potential use as a growth promoter and the safety margin of this. In this study, 480 1-day-old Cobb chicks were assigned to one control and three treated groups. The treated groups were supplemented with silver-doped silica nanoparticles (SiO2@AgNPs) at three dietary levels (8, 16, and 20 mg/kg diet) for 35 days. The results revealed no significant changes in the growth performance and oxidative parameters, and in most of the hematological and biochemical parameters among the control and treated groups. In contrast, dose-dependent adverse effects were exerted on the histopathological structure and immunohistochemical expression of CD45 in liver, kidneys, and lymphoid organs (spleen, bursa, and thymus). In addition, the relative weight of lymphoid organs and the serum levels of immunoglobulins M and G were significantly diminished. Moreover, the gene expression of proinflammatory cytokines (IL-β1 and TNF-α) and the ultrastructural morphology in breast muscle showed significant dose-dependent alterations. It could be concluded that the dietary supplementation of SiO2@AgNPs at a level of 8 mg/kg diet or more has dose-dependent proinflammatory and immunosuppressive effects on broiler chickens.

Antagonism of CGRP Receptor: Central and Peripheral Mechanisms and Mediators in an Animal Model of Chronic Migraine.

Calcitonin-gene-related peptide (CGRP) plays a key role in migraine pathophysiology and more specifically in the mechanisms underlying peripheral and central sensitization. Here, we explored the interaction of CGRP with other pain mediators relevant for neuronal sensitization in an animal model of chronic migraine. Male Sprague-Dawley rats were exposed to nitroglycerin (NTG, 5 mg/kg, i.p.) or vehicle co-administered with the CGRP receptor antagonist olcegepant (2 mg/kg i.p.), or its vehicle, every other day over a 9-day period. Twenty-four hours after the last injection of NTG (or vehicle), behavioral test and ex vivo analysis were performed. Olcegepant attenuated NTG-induced trigeminal hyperalgesia in the second phase of the orofacial formalin test. Interestingly, it also reduced gene expression and protein levels of CGRP, pro-inflammatory cytokines, inflammatory-associated miRNAs (miR-155-5p, miR-382-5p, and miR-34a-5p), and transient receptor potential ankyrin channels in the medulla–pons area, cervical spinal cord, and trigeminal ganglia. Similarly, olcegepant reduced the NTG-induced increase in CGRP and inflammatory cytokines in serum. The findings show that the activation of the CGRP pathway in a migraine animal model was associated to the persistent activation of inflammatory pathways, which was paralleled by a condition of hyperalgesia. These molecular events are relevant for informing us about the mechanisms underlying chronic migraine.

Probiotics suppress nonalcoholic steatohepatitis and carcinogenesis progression in hepatocyte-specific PTEN knockout mice

Nonalcoholic fatty liver disease (NAFLD), a hepatic characteristic of metabolic syndrome, received significant attention in clinical settings. The multiple-hit theory is one of the proposed mechanisms of NAFLD, and gut dysbiosis is considered a hit. Thus, controlling gut microbiota is a potential target in the management of NAFLD, and probiotics can be used as a treatment agent for NAFLD. The current study aimed to investigate the efficacy of probiotics against nonalcoholic steatohepatitis in a hepatocyte-specific PTEN knockout mouse model that mimics the characteristics of human NAFLD. Probiotics were administered to male knockout mice for 8 or 40 weeks. Next, we assessed hepatic inflammation, fibrosis, carcinogenesis, and oxidative stress. Probiotics were found to reduce serum transaminase levels, NAFLD activity score, and the gene expression of pro-inflammatory cytokines. In addition, they decreased liver fibrosis grade, which was examined via Sirius red staining, gene expression of fibrotic markers, and hydroxyproline. Furthermore, probiotics suppressed the number of liver tumors, particular in HCC. Probiotics reduced oxidative stresses, including glutathione levels, and anti-oxidative stress marker, which may be an underlying mechanism for their beneficial effects. In conclusion, probiotics treatment had beneficial effects against NAFLD and carcinogenesis in hepatocyte-specific PTEN knockout mice.

Effects of Different Titanium Surfaces Created by 3D Printing Methods, Particle Sizes, and Acid Etching on Protein Adsorption and Cell Adhesion, Proliferation, and Differentiation.

The surfaces of 3D printed titanium prostheses have major impacts on the clinical performance of the prostheses. To investigate the surface effects of the products generated by 3D printed titanium on osseointegration, six surface types of titanium discs produced by the direct metal laser sintering (DMLS) and electron beam melting (EBM) methods, with two sizes of titanium particles and post-printing acid etching, were used to examine the surface topography and to explore the protein adsorption, pro-inflammatory cytokine gene expressions, and MC3T3-E1 cell adhesion, proliferation, and differentiation. The EBM-printed disc showed a stripy and smooth surface without evidence of the particles used, while the DMLS surface contained many particles. After acid etching, small particles on the DMLS surface were removed, whereas the large particles were left. Moreover, distinct proteins with low molecular weights were attached to the 3D printed titanium discs but not to the pre-printing titanium particles. The small titanium particles stimulated the highest TNF-α and IL-6 gene expressions at 24 h. The alizarin red content and osteocalcin gene expression at day 21 were the highest in the groups of acid-etched discs printed by DMLS with the small particles and by EBM. Therefore, the acid-treated surfaces without particles favor osteogenic differentiation. The surface design of 3D printed titanium prostheses should be based on their clinical applications.