IL-6 mRNA Expression Research Articles

Effects of Yacon Leaf Extract on MCP-1 and IL-10 Expressions and Macrophage Phenotypes in CKD Mouse Model

Macrophages are essential in tissue homeostasis and immunity, but also contribute to disease development and progression. Chronic kidney disease (CKD) is characterized by interstitial infiltration of macrophages, the density of which correlates inversely with kidney survival. Studies have shown that yacon (Smallanthus sonchifolius) has beneficial effects on CKD. Therefore, this study aimed to investigate the effects of yacon leaf extract on mice with subtotal nephrectomy by evaluating the M1 and M2 macrophage counts and mRNA expressions of monocyte chemoattractant protein-1 (MCP-1 and IL-10. The mice were randomly divided into five groups: SO (negative control: underwent sham operation), SN (positive control: underwent subtotal nephrectomy), and yacon-treated groups: YK1, YK2, and YK3 (underwent subtotal nephrectomy, given peroral yacon leaf extract for 14 days with doses of 24,5 mg/kgBW/day, 49 mg/kgBW/day, and 98 mg/kgBW/day, respectively). The macrophage subtypes were assessed using immunohistochemistry anti-CD68 for M1 and anti-Arginase I for M2. MCP-1 and IL-10 mRNA expressions were analyzed using semi-quantitative PCR. Results showed that yacon leaf extract could significantly lower the M2 macrophage count (p<0.001) and the mRNA expressions of MCP-1 and IL-10 in all yacon-treated groups when compared to the SN group. However, the M1 macrophage count was only lower in the YK2 group (p=0.009). In conclusion, the administration of yacon leaf extract could attenuate kidney injury by lowering the macrophage count and the expression of MCP-1 and IL-10.

The placenta in fetal death: molecular evidence of dysregulation of inflammatory, proliferative, and fetal protective pathways

BackgroundIt is estimated that over 2 million cases of fetal death occur worldwide every year, but, despite the high incidence, several basic and clinical characteristics of this disorder are still unclear. Placenta is suggested to play a central role in fetal death. Placenta produces hormones, cytokines and growth factors that modulate functions of the placental-maternal unit. Fetal death has been correlated with impaired secretion of some of these regulatory factors. Objective(s)The aim of the present study was to evaluate, in placentas collected from fetal death, the gene expression of inflammatory, proliferative and protective factors. Study designCases of fetal death in singleton pregnancy were retrospectively selected, excluding pregnancies complicated by fetal anomalies, gestational diabetes, intrauterine growth restriction and moderate to severe maternal diseases. A group of placentas collected from healthy singleton term pregnancies were used as controls. Groups were compared regarding maternal and gestational age, fetal sex and birth weight. Placental mRNA expression of inflammatory (IL-6), proliferative (Activin A, TGF-β1) and regulatory (VEGF, VEGFR2, ATP-binding cassette (ABC) transporters ABCB1 and ABCG2, sphingosine 1-phosphate (S1P) signaling pathway) markers was conducted using real-time PCR. Statistical analysis and graphical representation of the data were performed using the GraphPad Prism 5 software. For the statistical analysis, Student’s t-test was used, and P values < 0.05 were considered significant. ResultsPlacental mRNA expression of IL-6 and VEGFR2 resulted significantly higher in the fetal death group compared to controls (P<0.01), while activin A, ABCB1 and ABCG2 expression resulted significantly lower (P<0.01). A significant alteration in the S1P signaling pathway was found in the fetal death group, with an increased expression of the specific receptor isoforms sphingosine 1-phosphate receptor 1, 3 and 4 (S1P1, S1P3, S1P4) and of sphingosine kinase 2 (SK2), one of the enzyme isoforms responsible for S1P synthesis (P<0.01). Conclusion(s): The present study confirmed a significantly increased expression of placental IL-6 and VEGFR2 mRNA, and for the first time showed an increased expression of S1P receptors and SK2 as well as a decreased expression of activin A and of selected ATP-binding cassette transporters, suggesting that multiple inflammatory and protective factors are deranged in placenta of fetal death.

Co-exposure of decabromodiphenyl ethane and polystyrene nanoplastics damages grass carp (Ctenopharyngodon idella) hepatocytes: Focus on the role of oxidative stress, ferroptosis, and inflammatory reaction

Decabromodiphenyl ethane (DBDPE) and polystyrene nanoplastics (PS-NPs) are emerging pollutants that seriously threaten the ecological safety of the aquatic environment. However, the hepatotoxicity effect of their combined exposure on aquatic organisms has not been reported to date. In, this study, the effects of single or co-exposure of DBDPE and PS-NPs on grass carp hepatocytes were explored and biomarkers related to oxidative stress, ferroptosis, and inflammatory cytokines were evaluated. The results show that both single and co-exposure to DBDPE and PS-NPs caused oxidative stress. Oxidative stress was induced by increasing the contents of pro-oxidation factors (ROS, MDA, and LPO), inhibiting the activity of antioxidant enzymes (CAT, GPX, T-SOD, GSH, and T-AOC), and downregulating the mRNA expressions of antioxidant genes (GPX1, GSTO1, SOD1, and CAT); the effects of combined exposure were stronger overall. Both single and co-exposure to DBDPE and PS-NPs also elevated Fe2+ content, promoted the expressions of TFR1, STEAP3, and NCOA4, and inhibited the expressions of FTH1, SLC7A11, GCLC, GSS, and GPX4; these effects resulted in iron overload-induced ferroptosis, where co-exposure had stronger adverse effects on ferroptosis-related biomarkers than single exposure. Moreover, single or co-exposure enhanced inflammatory cytokine levels, as evidenced by increased mRNA expressions of IL-6, IL-12, IL-17, IL-18, IL-1β, TNF-α, IFN-γ, and MPO. Co-exposure exhibited higher expression of pro-inflammatory cytokines compared to single exposure. Interestingly, the ferroptosis inhibitor ferrostatin-1 intervention diminished the above changes. In brief, the results suggest that DBDPE and PS-NPs trigger elevated levels of inflammatory cytokines in grass crap hepatocytes. This elevation is achieved via oxidative stress and iron overload-mediated ferroptosis, where cytotoxicity was stronger under co-exposure compared to single exposure. Overall, the findings contribute to elucidating the potential hepatotoxicity mechanisms in aquatic organisms caused by co-exposure to DBDPE and PS-NPs.

White blood cells, TNF-α, and interleukin-6 in subjects with infantile colic treated with Lacticaseibacillus rhamnosus GG (ATCC 53103): a randomised prospective study.

Recent metanalysis reported that certain probiotic strains, such as Limosilactobacillus reuteri and Lacticaseibacillus rhamnosus (LGG), seem effective for treatment of infantile colic of exclusively breastfed infants; some reports have also linked probiotics to have an immunological effect, however further investigation are needed to fully understand the exact mechanism. The objective of this study was to assay white blood cells, tumour necrosis factor (TNF)-α and interleukin (IL)-6 values in peripheral blood in subjects treated in a randomised, double-blind, placebo-controlled trial for infantile colic with LGG. Fifty-eight infants were enrolled and followed for a study period of 28 days. Parent were asked to record daily crying time using a structured cry diary. Peripheral white blood cells was assessed and RNA (mRNA) expression of TNF-α and IL-6 was measured using TaqMan real-time PCR-maternal amplification. Infants with colic treated with LGG showed a reduction in daily crying duration after 28 days of treatment and a reduction in values of IL-6 ( P < 0.005) and TNF-α ( P < 0.05); we observe also a significantly decreasing of IL-6 in the placebo group while decrease of TNF-α was not significant in this group. A significant decreased values of monocytes ( P < 0.05) was observed in infants treated with LGG. Our data therefore showed, in addition to crying time reduction, a significant decrease of TNF-α and a significant reduction of monocytes cells in colicky infants treated with LGG, compared to placebo group. This observation supports the hypothesis that probiotics may have anti-inflammatory properties. Further studies are needed to better understand the influence of probiotic on immunity cells.

Sacubitril/valsartan ameliorates cardiac function and ventricular remodeling in CHF rats via the inhibition of the tryptophan/kynurenine metabolism and inflammation

Sacubitril/valsartan has been highly recognized as a treatment for Chronic heart failure (CHF). Its potential cardioprotective benefits and mechanisms, however, remain to be explored. Metabolomics can be used to identify the metabolic characteristics and related markers, as well as the influence of drugs, thereby opening up the new mechanism for sacubitril/valsartan therapy in CHF disease. In this study, the ligation of left anterior descending and exhaustive swimming were used to induce a rat model of CHF after myocardial infarction. The efficacy was appraised with echocardiography, serum NT-proBNP, and histopathologica. UPLC-Q/TOF–MS combined with multivariate statistical analysis approach were used to analyze the effect of sacubitril/valsartan on CHF rats. RT-qPCR and western blot were performed to investigate the tryptophan/kynurenine metabolism pathway. Accordingly, the basal cardiac function were increased, while the serum NT-proBNP and collagen volume fraction decreased in CHF rats with sacubitril/valsartan. Sacubitril/valsartan regulated the expression of kynurenine et.al 8 metabolomic biomarkers in CHF rats serum, and it contributed to the cardioprotective effects through tryptophan metabolism pathway. In addition, the mRNA and protein expression of the indoleamine 2,3-dioxygenase (IDO) in the myocardial tissue of CHF rats, were down-regulated by sacubitril/valsartan, which was the same with the IL-1β, IFN-γ, TNF-α, COX-2, and IL-6 mRNA expression, and IL-1β, IFN-γ, and TNF-α expression in serum. In conclusion, sacubitril/valsartan can ameliorate cardiac function and ventricular remodeling in CHF rats, at least in part through inhibition of tryptophan/kynurenine metabolism.

Effect of Dietary Supplementation of Enteric Avian-Origin Lactobacillus casei-Fermented Soybean Meal on the Growth Performance and Intestinal Health of Broiler Chickens

The bacterial strain is key to fermentation, and the intestinal tract in livestock and poultry is a resource bank of good natural strains. The objective of this study was to evaluate the effect of soybean meal fermented using Lactobacillus casei, isolated from healthy broiler intestines with excellent organic acid production, on the intestinal health and growth performance of broilers. A total of 120 Arbor Acre male broiler chickens aged 21 days were fed until 42 days of age. These chickens were randomly divided into four groups with five replicates per group. Each replicate contained six broiler chickens. The specific groups were the control group (basal diet), the low-dose fermented soybean meal (FSBM) additive group (FSBML, basal diet + 0.2 kg/t FSBM), the middle-dose FSBM additive group (FSBMM, basal diet + 2 kg/t FSBM), and the high-dose FSBM additive group (FSBMH, basal diet + 5 kg/t FSBM). The results demonstrated a significant increase in the average daily feed intake (ADFI) and average daily gain (ADG) of the FSBMH group (p &lt; 0.05). The FSBMH group displayed a significantly increased villus height (VH) to crypt depth (CD) ratio (VH/CD) for the duodenum (p &lt; 0.05) and rectum (p &lt; 0.05). The examination of the ileal mucosa showed that the FSBMH group (p &lt; 0.05) had significantly higher levels of glutathione (GSH) activity, as well as higher relative mRNA expression of ZO-1, ZO-2, Occludin, IL-4, IL-6, MCP-1, TNF-α, IFN-α, IFN-β, and IFN-γ. However, the activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) were significantly lower in the FSBMH group (p &lt; 0.05). The FSBMH group also showed higher levels of Nitriliruptoraceae and Ruminococcaceae. In conclusion, the addition of 5 kg/t FSBM to diets had an ameliorative effect on broiler growth performance and intestinal health.

Effects of idebenone and coenzyme Q10 on NLRP3/caspase-1/IL-1β pathway regulation on ethanol-induced hepatotoxicity in rats

Chronic and excessive alcohol consumption leads to liver toxicity. There is a need to investigate effective therapeutic strategies to alleviate alcohol-induced liver injury, which remains the leading cause of liver-related morbidity and mortality worldwide. Therefore here, we looked into and evaluated how ethanol-induced hepatotoxicity was affected by coenzyme Q10 (CoQ10) and its analog, idebenone (IDE), on the NLRP3/caspase-1/IL-1 pathway. Hepatotoxicity induced in rats through the oral administration of gradually increasing dosages of ethanol (from 2 to 6 g/kg/day) over 30 days and the effect of CoQ10 (10 or 20 mg/kg) and IDE (50 or 100 mg/kg) were evaluated. Serum hepatotoxicity markers (ALT, AST, GGT, ALP, and TBIL), tissue oxidative stress markers and the mRNA expressions of IL-1β, IL-18, TGF-β, NF-κB, NLRP3, and caspase-1 were evaluated. Masson’s trichrome staining was also used to visualize fibrosis in the liver tissue. The results indicated that ethanol exposure led to hepatotoxicity as well as considerable NLRP3/caspase-1/IL-1β pathway activation. Moreover, CoQ10 or IDE treatment reduced measured parameters in a dosage-dependent manner. Thus, by inhibiting the NLRP3/caspase-1/IL-1 pathway, CoQ10 and IDE can prevent the hepatotoxicity caused by ethanol, although CoQ10 is more effective than IDE. This study will provide insight into new therapeutic avenues that take advantage of the anti-inflammatory and antioxidant properties of CoQ10 and IDE in ethanol-induced liver diseases.

Biofilm-derived membrane vesicles exhibit potent immunomodulatory activity in Pseudomonas aeruginosa PAO1.

Pathogenic bacteria form biofilms on epithelial cells, and most bacterial biofilms show increased production of membrane vesicles (MVs), also known as outer membrane vesiclesin Gram-negative bacteria. Numerous studies have investigated the MVs released under planktonic conditions; however, the impact of MVs released from biofilms on immune responses remains unclear. This study aimed to investigate the characteristics and immunomodulatory activity of MVs obtained from both planktonic and biofilm cultures of Pseudomonas aeruginosa PAO1. The innate immune responses of macrophages to planktonic-derived MVs (p-MVs) and biofilm-derived MVs (b-MVs) were investigated by measuring the mRNA expression of proinflammatory cytokines. Our results showed that b-MVs induced a higher expression of inflammatory cytokines, including Il1b, Il6, and Il12p40, than p-MVs. The mRNA expression levels of Toll-like receptor 4 (Tlr4) differed between the two types of MVs, but not Tlr2. Polymyxin B significantly neutralized b-MV-mediated cytokine induction, suggesting that lipopolysaccharideof native b-MVs is the origin of the immune response. In addition, heat-treated or homogenized b-MVs induced the mRNA expression of cytokines, including Tnfa, Il1b, Il6, and Il12p40. Heat treatment of MVs led to increased expression of Tlr2 but not Tlr4, suggesting that TLR2 ligands play a role in detecting the pathogen-associated molecular patternsin lysed MVs. Taken together, our data indicate that potent immunomodulatory MVs are produced in P. aeruginosa biofilms and that this behavior could be a strategy for the bacteria to infect host cells. Furthermore, our findings would contribute to developing novel vaccines using MVs.

Distinct Effects of Respiratory Viral Infection Models on miR-149-5p, IL-6 and p63 Expression in BEAS-2B and A549 Epithelial Cells.

Respiratory viruses cause airway inflammation, resulting in epithelial injury and repair. miRNAs, including miR-149-5p, regulate different pathological conditions. We aimed to determine how miR-149-5p functions in regulating pro-inflammatory IL-6 and p63, key regulators of airway epithelial wound repair, in response to viral proteins in bronchial (BEAS-2B) and alveolar (A549) epithelial cells. BEAS-2B or A549 cells were incubated with poly (I:C, 0.5 µg/mL) for 48 h or SARS-CoV-2 spike protein-1 or 2 subunit (S1 or S2, 1 μg/mL) for 24 h. miR-149-5p was suppressed in BEAS-2B challenged with poly (I:C), correlating with IL-6 and p63 upregulation. miR-149-5p was down-regulated in A549 stimulated with poly (I:C); IL-6 expression increased, but p63 protein levels were undetectable. miR-149-5p remained unchanged in cells exposed to S1 or S2, while S1 transfection increased IL-6 expression in BEAS-2B cells. Ectopic over-expression of miR-149-5p in BEAS-2B cells suppressed IL-6 and p63 mRNA levels and inhibited poly (I:C)-induced IL-6 and p63 mRNA expressions. miR-149-5p directly suppressed IL-6 mRNA in BEAS-2B cells. Hence, BEAS-2B cells respond differently to poly (I:C), S1 or S2 compared to A549 cells. Thus, miR-149-5p dysregulation may be involved in poly (I:C)-stimulated but not S1- or S2-stimulated increased IL-6 production and p63 expression in BEAS-2B cells.

Rosmarinic acid alleviates fungal keratitis caused by Aspergillus fumigatus by inducing macrophage autophagy

Fungal keratitis (FK) is an infectious keratopathy can cause serious damage to vision. Its severity is related to the virulence of fungus and response of inflammatory. Rosmarinic acid (RA) extracted from Rosmarinus officinalis exhibits antioxidant, anti-inflammatory and anti-viral properties. The aim of this study was to investigate the effect of RA on macrophage autophagy and its therapeutic effect on FK. In this study, we demonstrated that RA reduced expression of proinflammatory cytokine, lessened the recruitment of inflammatory cells in FK. The relative contents of autophagy markers, such as LC3 and Beclin-1, were significantly up-regulated in RAW 264.7 cells and FK. In addition, RA restored mitochondrial membrane potential (MMP) of macrophage to normal level. RA not only reduced the production of intracellular reactive oxygen species (ROS) but also mitochondria ROS (mtROS) in macrophage. At the same time, RA induced macrophage to M2 phenotype and down-regulated the mRNA expression of IL-6, IL-1β, TNF-α. All the above effects could be offset by the autophagy inhibitor 3-Methyladenine (3-MA). Besides, RA promote phagocytosis of RAW 264.7 cells and inhibits spore germination, biofilm formation and conidial adherence, suggesting a potential therapeutic role for RA in FK.

Study on the bioactive ingredients and mechanism of Huangqi against diabetic retinopathy based on network pharmacology and experimental verification.

Diabetic retinopathy (DR) is one of the most well-known microvascular complications of diabetes mellitus. As a traditional Chinese medicine, Huangqi (HQ), has been used for treating DR for a long time. However, its anti-DR active ingredients and mechanism are still unknown. Therefore, we designed this study to explore the active components and mechanism of HQ against DR via network pharmacology analysis. The ingredients of HQ, and potential targets of HQ and DR were obtained from public databases. We used the protein-protein interaction (PPI) network, Kyoto Encyclopedia of Genes and Genomes (KEGGs) pathway enrichment, and Gene Ontology (GO) analysis to identify core targets and pathways of HQ against DR. Finally, molecular docking and vitro experiments were applied to validate our results. A total of 34 potential targets of HQ against DR were obtained. Based on PPI network, VEGFA, PTGS2, Interleukin-6 (IL-6), and CCL2 were considered as core targets. GO analysis involved 692 biological processes, 21 cellular components, and 35 molecular functions. KEGG enrichment analysis manifested that the anti-DR effect of HQ was mainly mediated via the AGE-RAGE signaling pathway in diabetic complications. The molecular docking results indicated that kaempferol had higher affinity with CCL2, IL-6, VEGFA, and PTGS2. The vitro experiments showed that the mRNA expressions of CCL2, IL-6, VEGFA, and PTGS2 in ARPE-19 cells were differentially decreased after kaempferol treatment. This study preliminarily unveiled that the therapeutic efficacy of HQ against DR might be attributed to the reduced expression of CCL2, IL-6, VEGFA, and PTGS2.

#2009 FGF23 in patients with diabetic kidney disease and atherosclerosis

Abstract Background and Aims Diabetes mellitus (DM) and chronic kidney disease (CKD) are risk factors for vascular atherosclerosis, a pathological process with an important inflammatory component. The coexistence of these conditions in patients with diabetic kidney disease (DKD) has additive effects with worst cardiovascular mortality and morbidity rates when compared to non-diabetic CKD patient on cardiovascular outcomes. Accumulating research has shown that increased levels of bone-derived fibroblast growth factor 23 (FGF23), the major phosphate-regulating hormone, constitute a novel predictor of cardiovascular and all-cause mortality in CKD patients, with potential effects on disorders of glucose metabolism as well. The objective of this comparative study is to determine the differences in FGF23 levels in two groups of patients with CKD, with and without DM, matched by age, sex and renal function, all of them with clinical atherosclerotic disease. We also evaluated the relationships of FGF23 with several inflammatory markers and parameters of glucose metabolism. Method Sixty-six patients with CKD stages 3-4 (33 of them with DM) and diagnosed with peripheral vascular disease were included in the study. Whole blood and serum samples were obtained from all participants. CRP values, neutrophil/lymphocyte (N/L) ratio, HbA1c (%), fasting glucose, and Triglycerides and glucose (TyG) index, among other parameters, were obtained using standardized routine methods. Circulating levels of the intact form of FGF23, tumor necrosis factor (TNF) α, interleukin (IL) 6, and IL10 were determined by ELISA immunoassays in both groups. We also performed gene expression analysis for these three cytokines in peripheral blood cells (PBCs) using qPCR. Results The mean values of eGFR, ACR and phosphate were 37.6 ± 11.2 ml/min/1.73 m2; 294.1 ± 271.2 mg/g; 3.8 ± 0.82 mg/dL; respectively without differences between patients with and without DM The median circulating FGF23 values were higher in the DM group (51.59 [37.73-89.68] vs. 38.1 [33-54.14] pg/mL; p&amp;lt;0.001). Likewise, patients with DM presented higher levels of TNFα both in serum concentration (1.78 [0.72-2.47] vs. 1.28 [1.01-1.88] pg/mL; p&amp;lt;0.01) as in the expression of mRNA in PBCs (2.1 ± 1.26 vs. 1.87 ± 1.16 a.u.; p&amp;lt;0.05). No differences were observed for IL6 and IL10 levels, nor for CRP concentration or N/L ratio. As expected, fasting glucose, HbA1c and TyG index were higher in the group of subjects with DM. FGF23 was inversely correlated with eGFR (r=−0.435, p&amp;lt;0.0001), while it showed a positive association with ACR (r=0.416, p&amp;lt;0.001) and, interestingly, with proinflammatory cytokines including circulating TNFα (r=0.44, p&amp;lt;0.001), circulating IL6 (r=0.27, p=0.04), and TNF mRNA expression in PBCs (r=0.245, p=0.047). Furthermore, circulating FGF23 was also direct and significantly correlated with HbA1c (r=0.34, p&amp;lt;0.01). Conclusion These results suggest that FGF23 levels are higher in DKD than in non-DM CKD patients with similar renal function. DKD patients also had higher levels of the proinflammatory cytokine TNFα. Our results also show positive correlations of FGF23 with the inflammatory cytokines TNFα and IL6, as well as with HbA1c. Previous studies, shown that elevated TNFα and glucose promote FGF23 production, which may help to explain the higher levels of FGF23 in patients with DM Future studies should clarify whether FGF23 is simply a marker of atherosclerosis disease severity or a contributing factor to this complication in CKD.

Aspergillus awamori: potential antioxidant, anti-inflammatory, and anti-apoptotic activities in acetic acid-induced ulcerative colitis in rats

Ulcerative colitis (UC) is a chronic colonic inflammation with a significant health hazard. Aspergillus awamori (A. awamori) is a microorganism with various bioactive compounds with natural antioxidant and anti-inflammatory properties. The present work aimed to elucidate the protective and therapeutic effects of varying concentrations of A. awamori against acetic acid (AA)-induced ulcerative colitis (UC) in rats. Nine groups of albino male rats were established: a control negative group (G1), a control positive group (G2,AA), and preventive protocol groups (including G3A, G4A, and G5A) that received 100 mg, 50 mg, and 25 mg/kg b.w, respectively, of A. awamori orally and daily from the 1st day of the experiment and for 7 consecutive days. Then, they were subjected to one dose of AA intrarectally on day 8th. G3B, G4B, and G5B were termed as curative protocol groups that received one dose of AA on day 8th and then administered 100 mg, 50 mg, and 25 mg/kg b.w. of A. awamori, respectively, on day 9th and continued receiving these doses daily until day 16th. Rats in the AA group exhibited marked histopathological alterations of the distal colon, with an exaggeration of the DAI. In addition, a remarkable increase in oxidative stress was represented by the elevation of MDA and NO levels with a decline in SOD and GPx activities. In addition, upregulation of TNF-α, IL-6, and IL-1β mRNA expressions and downregulation of Muc2 and Nrf2 levels were detected. Unambiguously, a remarkable anti-inflammatory effect was noticed either in A. awamori prevented or treated groups expounded by reducing and regulating TNF-α, IL-6, and IL-1β with improved pathological lesion scoring. The Muc2, Nrf2, and bcl-2 gene levels were upregulated and restored also. In summary, the findings in this work reveal that A. awamori supplementation successfully alleviated the UC induced by AA, which had a better effect when administered before colitis induction.

Liquiritin Alleviates Inflammation in Lipopolysaccharide-Induced Human Corneal Epithelial Cells

Purpose This research was designed to elucidate the anti-inflammatory impacts of liquiritin on lipopolysaccharide (LPS)-activated human corneal epithelial cells (HCECs). Methods The Cell Counting kit-8 (CCK-8) assay was adopted to assess cell viability. The enzyme-linked immunosorbent assay (ELISA) was used to detect the secretion levels of the proinflammatory cytokines IL-6, IL-8, and TNF-α. Transcriptome analysis was conducted to identify the genes that exhibited differential expression between different treatment. The model group included cells treated with LPS (10 µg/mL), the treatment group comprised cells treated with liquiritin (80 µM) and LPS (10 µg/mL), and the control group consisted of untreated cells. To further validate the expression levels of the selected genes, including CSF2, CXCL1, CXCL2, CXCL8, IL1A, IL1B, IL24, IL6, and LTB, quantitative real-time PCR was performed. The expression of proteins related to the Akt/NF-κB signaling pathway was assessed through western blot analysis. NF-κB nuclear translocation was evaluated through immunofluorescence staining. Results The secretion of IL-6, IL-8, and TNF-α in LPS-induced HCECs was significantly downregulated by liquiritin. Based on the transcriptome analysis, the mRNA expression of pro-inflammatory cytokines, namely IL-6, IL-8, IL-1β, IL-24, TNF-α, and IL-1α was overproduced by LPS stimulation, and suppressed after liquiritin treatment. Furthermore, the Western blot results revealed a remarkable reduction in the phosphorylation degrees of NF-κB p65, IκB, and Akt upon treatment with liquiritin. Additionally, immunofluorescence analysis confirmed liquiritin’s inhibition of LPS-induced p65 nuclear translocation. Conclusions Collectively, these findings imply that liquiritin suppresses the expression of proinflammatory cytokines, and the anti-inflammatory impacts of liquiritin may be caused by its repression of the Akt/NF-κB signaling pathway in LPS-induced HCECs. These data indicate that liquiritin could provide a potential therapeutic application for inflammation-associated corneal diseases.

No-observed adverse effect levels of deoxynivalenol and aflatoxin B1 in combination induced immune inhibition and apoptosis in vivo and in vitro

Mycotoxins are toxic metabolites produced by fungal species, commonly exist in animal feeds, and pose a serious risk to human as well as animal health. But limited studies have focused on combined effects of no-observed adverse effect levels. In vivo study, 6 weeks old twenty-four mice were individually exposed to Deoxynivalenol (DON) at 0.1 mg/kg BW, Aflatoxin B1 (AFB1) at 0.01 mg/kg BW, and mixture of DON and AFB1 (0.1 mg/kg BW and 0.01 mg/kg BW, respectively) for 28 days. Then, DON at 0.5 μg/mL, AFB1 at 0.04 μg/mL, and mixtures of DON and AFB1 (0.5 μg/mL, 0.04 μg/mL, respectively) were applied to porcine alveolar macrophages (PAMs) in vitro study. Our in vivo results revealed that the combined no-observed adverse effect levels of DON and AFB1 administration decreased IgA and IgG levels in the serum, the splenic TNF-α, IFN-γ, IL-2 and IL-6 mRNA expression and T-lymphocyte subset levels (CD4+ and CD8+) in the spleen. Additionally, the combined administration increased caspase-3, caspase-9, Bax, Cyt-c, and decreased Bcl-2 protein expression. Taken together, the combined no-observed adverse effect levels of DON and AFB1 could induce immunosuppression, which may be related to apoptosis. This study provides new insights into the combined immune toxicity (DON and AFB1).

House dust mite allergen Der f 2 drives IL-6 and GM–CSF expression in airway epithelial cells via p38 MAPK/NF-κB signaling

Objective Der f 2, a major allergen derived from Dermatophagoides farinae, is a leading cause of allergic asthma. IL-6 and GM–CSF play essential roles in the exacerbation of asthma. However, the mechanical act by which Der f 2 mediates the expression of IL-6, IL-8, and GM–CSF in airway epithelial cells remains incompletely elucidated. Herein, we aimed to explore the effect of Der f 2 on IL-6 and GM–CSF expression in the human airway epithelial cell BEAS-2B and A549. Methods Recombinant Der f 2 (rDf2) was acquired using Pichia pastoris. BEAS-2B and A549 cells were used as cell model. The expression of genes and proteins and the involvement of the signaling cascade were assessed using RT–PCR, quantitative real-time PCR (qPCR), Western blotting, and ELISA, respectively. Results Our findings showed that rDf2 significantly induced mRNA expression and protein production of IL-6 and GM-CSF in BEAS-2B and A549 cells. In contrast, rDf2 did not influence IL-8 expression or production in both cells. Mechanistic studies revealed that rDf2 triggered activation of the p38 MAPK and JNK. Inhibition of p38, but not JNK, significantly attenuated rDf2-induced IL-6 and GM–CSF expression and production. Conclusion This study demonstrates that Der f 2 promotes the expression and production of the pro-inflammatory cytokines IL-6 and GM–CSF in airway epithelial cells via activation of the p38 signaling pathway. These findings provide insights into the molecular mechanisms that Der f 2 may exacerbate airway inflammation.

Invivo evaluation of Andrographis paniculata and Boesenbergia rotunda extract activity against SARS-CoV-2 Delta variant in Golden Syrian hamsters: Potential herbal alternative for COVID-19 treatment

The ongoing COVID-19 pandemic has triggered extensive research, mainly focused on identifying effective therapeutic agents, specifically those targeting highly pathogenic SARS-CoV-2 variants. This study aimed to investigate the in vivo antiviral efficacy and anti-inflammatory activity of herbal extracts derived from Andrographis paniculata and Boesenbergia rotunda, using a Golden Syrian hamster model infected with Delta, a representative variant associated with severe COVID-19. Hamsters were intranasally inoculated with the SARS-CoV-2 Delta variant and orally administered either vehicle control, B. rotunda, or A.paniculata extract at a dosage of 1000 mg/kg/day. Euthanasia was conducted on days 1, 3, and 7 post-inoculation, with 4 animals per group. The results demonstrated that oral administration of A. paniculata extract significantly alleviated both lethality and infection severity compared with the vehicle control and B. rotunda extract. However, neither extract exhibited direct antiviral activity in terms of reducing viral load in the lungs. Nonetheless, A. paniculata extract treatment significantly reduced IL-6 protein levels in the lung tissue (7278 ± 868.4 pg/g tissue) compared to the control (12,495 ± 1118 pg/g tissue), indicating there was a decrease in local inflammation. This finding is evidenced by the ability of A. paniculata extract to reduce histological lesions in the lungs of infected hamsters. Furthermore, both extracts significantly decreased IL-6 and IP-10 mRNA expression in peripheral blood mononuclear cells of infected hamsters compared to the control group, suggesting systemic anti-inflammatory effects occurred. In conclusion, A. paniculata extract's potential therapeutic application for SARS-CoV-2 arises from its observed capacity to lessen inflammatory cytokine concentrations and mitigate lung pathology.

Expression of cytokines at baseline correlate/predict in the disc the outcome of surgery after disc degeneration: A 12-month follow-up study.

Low back pain (LBP) is a highly prevalent condition that comprise a large portion of outpatient practice, challenging the diagnosis and treatment. However, the diagnostic tools are limited to clinical history, physical examination and imaging. Degenerative disc disease (DDD) is a significant cause of LBP, and emerging literature confirms the elevated levels of biomarkers in the discs. These biomarkers may serve as a tool for diagnosis, but may also be useful in predicting the treatment outcome. Here, we examine the expression of various cytokines on 1-year recovery from patients with LBP. Patient-reported outcome (PRO) in terms of pain intensity (VAS), disability (ODI), and quality of life (Eq-5D) is collected from 44 patients at baseline and 12 months after surgery to study the influence of baseline TNF-α, IL-1β, and IL-6 mRNA expression in both annulus fibrosus (AF) and nucleus pulposus (NP). Between baseline and follow-up, our cohort showed improvement in VAS back pain (p < 0.001), VAS leg pain (p < 0.001), ODI (p = 0.02), and Eq-5D (p = 0.01). Baseline levels of IL-1 β was positively correlated with VAS back pain scores in AF (p = 0.05) and NP (p = 0.01) at 1-year follow-up. TNF-α expression at baseline was also positively correlated to ODI scores (p = 0.01) at follow-up and inversely correlated to improvements in ODI score between baseline and follow-up, suggesting that high TNF-α expression at baseline is associated with poor outcomes from surgery. The results from our study support that TNF-α expression at baseline can serve as a very important predictor of treatment response from lumbar fusion surgery.

Comparison of the recovery characteristics for canine corneal ulcer treated with corneoconjunctival transposition or conjunctival autografts

Corneal ulceration induced by Staphylococcus pseudintermedius (S. pseudintermedius) is a common clinical eye disease. Antibiotics combined with corneoconjunctival transposition (CCT) or conjunctival autografts (CA) are often used, but the recovery characteristics are still unknown. In this experiment, canine corneal ulcer models induced by S. pseudintermedius and treated with levofloxacin eye drops (LED) were created. The models were used to compare the recovery characteristics of CCT and CA, combined with LED, by clinical observation, histopathology, and cytokine expression detected by qRT-PCR analysis. The results showed that the ulcerative cornea with only LED treatment perforated after 48 h. The mRNA expression of TLR2, IL-1β, IL-6, IL-8, and TNF-α genes was significantly elevated on 14, 28, and 35 days after the surgery compared to normal (p < 0.01). On day 42, the inflammatory damage had resolved, but the corneal transparency and arrangement of collagen fibrils in the CCT group were higher than those in the CA group. The mRNA expression of EGF, FGF, TGF-β1 and VEGF genes increased significantly (p < 0.01), mostly until day 42, proving that CCT and CA surgery contributed to the corneal recovery, and relieved the inflammatory reaction, with the elimination of corneal cicatrices needing a period of reconstruction. Therefore, this study has provided, for the first time, the method for establishing a canine corneal ulcer model induced by S. pseudintermedius. More importantly, the recovery of canine ulcerative corneas with CCT or CA surgery is reported for the first time.

Regulation of electroacupuncture on non-canonical pathway of hepatocellular pyroptosis in rats with nonalcoholic fatty liver disease

To observe the effect and mechanism of electroacupuncture (EA) on non-canonical pathway of hepatocellular pyroptosis in nonalcoholic fatty liver disease (NAFLD). Sixty male SD rats were randomly divided into a normal diet group (n=15) and a high fat modeling group (n=45). The rats in the high fat modeling group were fed with customized high fat diet for 8 weeks to establish NAFLD model. Thirty successfully modeled rats were selected and randomly divided into a model group (n=10), an EA group (n=10) and a non-acupoint with shallow needling group (n=10), and 10 rats were randomly selected from the normal diet group as the control group additionally. In the EA group, EA was applied at bilateral "Fenglong" (ST 40) and "Ganshu" (BL 18), with disperse-dense wave, in frequency of 4 Hz/20 Hz and in intensity of 3 mA. In the non-acupoint with shallow needling group, shallow needling was delivered at points 5 mm from bilateral "Fenglong" (ST 40) and "Ganshu" (BL 18), the EA stimulation parameters were same as the EA group. The intervention was given once a day, 20 min a time, 5 days a week for 4 weeks in the two groups. After intervention, the liver morphology was observed by oil red "O" staining, the serum levels of lipopolysaccharide (LPS), interleukin (IL)-1β, IL-18 and tumor necrosis factor-α (TNF-α) were detected by ELISA, the protein expression of gasdermin D (GSDMD), GSDMD-N, cysteine aspartic acid specific protease-11 (Caspase-11), IL-1β, IL-18 and TNF-α in liver tissue were detected by Western blot, the mRNA expression of GSDMD, Caspase-11, IL-1β, IL-18 and TNF-α in liver tissue was detected by real-time PCR in rats of each group. In the model group, vacuoles in different size were found in the hepatocellular cytoplasm, and the fat droplets were in schistose accumulation. Compared with the model group, the hepatocellular fat droplets and the degree of hepatic steatosis were reduced in the EA group and the non-acupoint with shallow needling group. Compared with the control group, the serum levels of LPS, IL-1β, IL-18 and TNF-α were increased (P<0.01), the protein and mRNA expression of GSDMD, Caspase-11, IL-1β, IL-18, TNF-α as well as the protein expression of GSDMD-N in the liver tissue were increased (P<0.01) in the model group. Compared with the model group, the serum levels of LPS, IL-1β, IL-18 and TNF-α were decreased (P<0.01), the protein and mRNA expression of GSDMD, IL-1β, IL-18 and TNF-α in the liver tissue were decreased (P<0.01), the protein expression of GSDMD-N and the mRNA expression of Caspase-11 in the liver tissue were decreased (P<0.01) in the EA group and the non-acupoint with shallow needling group. Compared with the model group, the protein expression of Caspase-11 in the liver tissue was decreased (P<0.01) in the EA group. Compared with the non-acupoint with shallow needling group, the serum levels of LPS, IL-1β, IL-18 and TNF-α were decreased (P<0.01), the protein and mRNA expression of GSDMD, Caspase-11, IL-1β and IL-18 in the liver tissue were decreased (P<0.01), the protein expression of GSDMD-N and the mRNA expression of TNF-α in the liver tissue were decreased (P<0.01) in the EA group. EA can inhibit hepatocellular pyroptosis in NAFLD rats, and its mechanism may be related to reducing the serum level of LPS, and down-regulating the expression of the non-canonical pathway related factors i.e. GSDMD, GSDMD-N, Caspase-11, IL-1β, IL-18 and TNF-α.