NF-κB Protein Levels Research Articles

Pre-Exposure to Stress-Inducing Agents Increase the Anticancer Efficacy of Focused Ultrasound against Aggressive Prostate Cancer Cells.

Despite the initial success in treatment of localized prostate cancer (PCa) using surgery, radiation or hormonal therapy, recurrence of aggressive tumors dictates morbidity and mortality. Focused ultrasound (FUS) is being tested as a targeted, noninvasive approach to eliminate the localized PCa foci, and strategies to enhance the anticancer potential of FUS have a high translational value. Since aggressive cancer cells utilize oxidative stress (Ox-stress) and endoplasmic reticulum stress (ER-stress) pathways for their survival and recurrence, we hypothesized that pre-treatment with drugs that disrupt stress-signaling pathways in tumor cells may increase FUS efficacy. Using four different PCa cell lines, i.e., LNCaP, C4-2B, 22Rv1 and DU145, we tested the in vitro effects of FUS, alone and in combination with two clinically tested drugs that increase Ox-stress (i.e., CDDO-me) or ER-stress (i.e., nelfinavir). As compared to standalone FUS, significant (p < 0.05) suppressions in both survival and recurrence of PCa cells were observed following pre-sensitization with low-dose CDDO-me (100 nM) and/or nelfinavir (2 µM). In drug pre-sensitized cells, significant anticancer effects were evident at a FUS intensity of as low as 0.7 kW/cm2. This combined mechanochemical disruption (MCD) approach decreased cell proliferation, migration and clonogenic ability and increased apoptosis/necrosis and reactive oxygen species (ROS) production. Furthermore, although activated in cells that survived standalone FUS, pre-sensitization with CDDO-me and/or nelfinavir suppressed both total and activated (phosphorylated) NF-κB and Akt protein levels. Thus, a combined MCD therapy may be a safe and effective approach towards the targeted elimination of aggressive PCa cells.

Upregulation of hsa-miR-625-5p Inhibits Invasion of Acute Myeloid Leukemia Cancer Cells through ILK/AKT Pathway.

Objective Acute myeloid leukemia (AML) is characterized by abnormalities of differentiation and growth of primary hematopoietic stem cells (HSCs) in the blood and bone marrow. In many studies, miR-625-5p has been shown to inhibit downstream pathways from affecting the metastasis and invasion of the integrin-linked kinase (ILK) signaling pathway. It has been proved that the expression of miR-625-5p decreases in AML cell lines. This study aimed to investigate the effect of miR-625-5p upregulation on the invasion of KG1 ell line in vitro. Materials and Methods In this experimental study, we investigated the impact of upregulation of miR-625-5p on invasion via the ILK/AKT pathway in the KG1 cell line. After transfection using the viral method, the cellular invasion was assessed by invasion assay and the levels of miR-625-5p genes and protein were evaluated by quantitative polymerase chain reaction (qPCR) and western blotting. Moreover, CXCR4 level was assessed by flow cytometry. Results The invasion significantly reduced in MiR-625-5p-transfected KG1 cells (P<0.01) that was concomitant with remarkably decreasing in the expression levels of ILK, NF-κB, and COX2 genes compare with the control group (P<0.01). Incontrast, MMP9, AP1, and AKT significantly increased (P<0.01, P<0.001 and P<0.01, respectively) and GSK3β did not change significantly in MiR-625-5p-transfected KG1 cells. The protein level of NF-κB decreased (P<0.01) and MMP9 increased, however it was not significant. Moreoever, the expression of CXCR4 was significantly lower (P<0.01) in comparison with the control group. ConclusionmiR-625-5p leads to a reduction in cell invasion in the AML cell line through ILK pathway. Therefore, it could be a breakthrough in future AML-related research. However, further studies are needed to support this argument.

Protective effect of glucagon-like peptide-1 mediated by ultrasound microbubbles on myocardial injury in rats with diabetic cardiomyopathy

ABSTRACT Current study was performed to investigate the therapeutic effects of UTMD combined with Semaglutide, a GLP-1 R agonist, on DCM in GK rats. After 6-week intervention, all rats were examined by conventional echocardiography, and the indicators of papillary muscle horizontal wall were measured. At the end of experiment, the DCM rats were sacrificed, and then blood samples were taken to detect blood lipids and the activities of oxidative stress-related factors in myocardial tissues. Moreover, the myocardial tissues were also taken to observe histomorphological changes and myocardial fibrosis via H&E and Masson staining, respectively. Furthermore, the mRNA and protein levels of PPAR-γ and NF-κB in myocardial tissues were detected. As a result, the glycometabolism and the cardiac functions of DCM rats received combination therapy of GLP-1 R agonist and UTMD were improved. Significantly increased SOD and GSH-Px as well as significantly decreased MDA levels were also observed in rats of combined group compared with others. In addition, both mRNA and protein levels of PPAR-γ and NF-κB were significantly lower than those of the model control ones and Semaglutide microbubbles alone (all P < 0.001). Further pathological histology analysis demonstrated that combination therapy effectively ameliorated fibrosis and myocardial morphological changes of DCM rats. In summary, UTMD combined with Semaglutide can effectively protect cardiac function in rats with DCM by significantly alleviating myocardial fibrosis and oxidative stress. Abbreviations Ultrasound-targeted microbubble destruction, UTMD; glucagon-like peptide-1 receptor, GLP-1R; diabetic cardiomyopathy, DCM; Goto-Kakizaki, GK; velocity vector imaging, VVI; left ventricular end-diastolic diameter LVIDd; left ventricular end-systolic diameter, LvIDs; left ventricular end-diastolic pressure, LVEDP; fractional shortening, LVFs; left ventricular ejection fraction, LVEF; mean peak radial velocity, Vs; radial strain, Sr; radial strain rate, SRr; superoxide dismutase, SOD; malondialdehyde, MDA; glutathione peroxidase, GSH-Px; peroxisome proliferator-activated receptor-γ, PPAR-γ; nuclear transcription factor κB, NF-κB; insulin resistance, IR; total cholesterol, TC; total triglycerides, TG; creatine kinase, CK; lactate dehydrogenase, LDH; cardiac troponin I, cTnI; collagen volume fraction, CVF; Hematoxylin eosin, H&E.

Aspirin Attenuates Hyperoxia-Induced Acute Respiratory Distress Syndrome (ARDS) by Suppressing Pulmonary Inflammation via the NF-κB Signaling Pathway.

Acute respiratory distress syndrome (ARDS) is a common destructive syndrome with high morbidity and mortality rates. Currently, few effective therapeutic interventions for ARDS are available. Clinical trials have shown that the effectiveness of aspirin is inconsistent. The contribution of platelets to the inflammatory response leading to the development of ARDS is increasingly recognized. The antiplatelet agent aspirin reportedly exerts a protective effect on acid- and hyperoxia-induced lung injury in murine models. Our previous study showed that pretreatment with aspirin exerts protective effects on hyperoxia-induced lung injury in mice. However, the mechanisms and therapeutic efficacy of aspirin in the posttreatment of hyperoxia-induced acute lung injury (ALI) remain unclear. In this study, we used a homozygous NF-κB-luciferase+/+ transgenic mouse model and treated mice with low-dose (25 μg/g) or high-dose (50 μg/g) aspirin at 0, 24, and 48 h after exposure to hyperoxia (inspired oxygen fraction (FiO2) > 95%). Hyperoxia-induced lung injury significantly increased the activation of NF-κB in the lung and increased the levels of macrophages infiltrating the lung and reactive oxygen species (ROS), increased the HO-1, NF-κB, TNF-α, IL-1β, and IL-4 protein levels, and reduced the CC10, SPC, eNOS, Nrp-1, and IκBα protein levels in the lung tissue. Pulmonary edema and alveolar infiltration of neutrophils were also observed in the lung tissue of mice exposed to hyperoxia. However, in vivo imaging revealed that posttreatment with aspirin reduced luciferase expression, suggesting that aspirin might reduce NF-κB activation. Posttreatment with aspirin also reduced hyperoxia-induced increases in the numbers of lung macrophages, intracellular ROS levels, and the expression of TNF-α, IL-1β, and IL-4; it also increased CC10, SPC and Nrp-1 levels compared with hyperoxia exposure alone. Lung histopathology also indicated that the aspirin posttreatment significantly reduced neutrophil infiltration and lung edema compared with hyperoxia exposure alone. Aspirin effectively induces an anti-inflammatory response in a model of hyperoxia-induced lung injury. Thus, aspirin may have potential as a novel treatment for hyperoxia-induced ALI.

Dietary Resveratrol Alleviates AFB1-Induced Ileum Damage in Ducks via the Nrf2 and NF-κB/NLRP3 Signaling Pathways and CYP1A1/2 Expressions

The aim of this study was to explore the mechanism underlying the protective effects of resveratrol against Aflatoxin B1-induced ileum injury in ducks. A corn–soybean meal-basal diet and two test diets (500 mg/kg resveratrol +0.2 mg Aflatoxin B1/kg, 0.2 mg AFB1/kg) were used in a 10-wk design trial (n = 15 ducks/group). These results showed that the toxicity of Aflatoxin B1 significantly reduced the antioxidant capacity of duck ileum and induced inflammation, oxidative stress, mitochondrial dysfunction and DNA damage in ducks. The expression of genes, including CYP1A2, CYP2A6, and CYP3A4, at the mRNA level was significantly upregulated (p &lt; 0.05) by AFB1. The level of Nrf2 was suppressed (p &lt; 0.05) and the mRNA and protein level of NF-κB was activated (p &lt; 0.05) in the AFB1 group. However, supplementation with 500 mg/kg dietary resveratrol in Aflatoxin B1-induced ducks significantly ameliorated these alterations and decreased the mRNA expression of CYP1A1 and CYP1A2 (p &lt; 0.05) and the production of AFB1-DNA adducts (p &lt; 0.05). The results proved that resveratrol alleviated ileum injury induced by AFB1, decreased the production of AFB1-DNA adducts by downregulating the expression of CYP1A1 and CYP1A2, and reduced DNA damage and oxidative stress via the Nrf2/ Keap1 and NF-κB/NLRP3 signaling pathways.

Ursolic acid protects against anoxic injury in cardiac microvascular endothelial cells by regulating intercellular adhesion molecule-1 and toll-like receptor 4/MyD88/NF-κB pathway.

Cardiac microvascular endothelial cells (CMECs) are rapidly damaged after myocardial ischemia or hypoxia. In this study, we intend to explore whether ursolic acid (UA) can protect CMECs against hypoxia/reoxygenation (H/R) injury and to detect related molecular mechanism. CMECs were subjected to H/R condition in the absence or presence of UA. Cell behaviors were measured by Cell Counting Kit-8, transwell, ELISA and western blot assays. siRNA was applied to reduce ICAM1 expression, then the effect of co-treatment of UA and si-ICAM1 on CMECs has been detected by biological experiments. Under H/R stimulation, the proliferation and migration of CMECs were inhibited, as well as the inflammation and oxidative stress were enhanced. UA treatment obviously reversed these H/R-induced injuries and reduced ICAM1 expression. Moreover, knockdown of ICAM1 could alleviate the H/R-induced injuries and strengthen the protective effect of UA on CMECs under H/R condition. Additionally, the protein levels of TLR4, MyD88 and p-P65 NF-κB were obviously increased after H/R stimulation, whereas the addition of UA could alter the phenomena by reducing TLR4, MyD88, and p-P65 NF-κB expression. Our results insinuated that UA could alleviate H/R-induced injuries in CMECs by regulating ICAM1 and TLR4/MyD88/NF-κB pathway.

Astragalus polysaccharide attenuates overexercise-induce myocardial injury via activating AMPK signaling pathway to suppress inflammation and oxidative stress.

Excessive exercise leads to myocardial injury or even sudden exercise death. For the vast sports population, appropriate physiological state is a necessary condition for exercise. The present study aims to investigate the cardioprotective effects and potent mechanism of astragalus polysaccharide (APS) treatment against the exercise-induced myocardial injury via in vitro cell-based assay and in vivo model rat. Efficacies of APS incubation on the inflammatory response and oxidative stress induced by LPS were both explored in H9c2 cells by using CCK-8 and western blotting method, respectively. Normal SD rats were randomly divided into saline-treated overexercise rat group, and APS-treated overexercise rat groups with three doses. Then long-term swimming training load cycle (8 week) were performed on these rats. Finally, the changes on body weight, myocardial morphological and injury indicators, as well as the inflammation-related proteins in overexercise-induced model rats were all assessed. Three concentrations of APS all significantly increased cell viability, and decreased the apoptosis of cardiomyocytes in LPS-treated H9c2 cells. Moreover, chronic treatment of APS at all three doses also could obviously decreased myocardial injury-related indicators. Furthermore, the histopathologic examination exhibited that the APS successfully attenuated the changes of myocardial tissues, reduced the lipid accumulation and the protein levels of IL-1β, TNF-α and NF-κB. Furthermore, the APS could activate the AMPK signaling pathway, enhance the autophagy and suppress the production of ROS. On conclusions, APS exerted the protective efficacies on overexercise-induced myocardial injury by activating the AMPK signaling pathway to increase autophagy and suppress the inflammation response, oxidative stress, apoptosis of myocardial cells.

The Inhibitory Effects of Vanillin on the Growth of Melanoma by Reducing Nuclear Factor-κB Activation.

Melanoma is skin cancer, and the treatments are not efficient enough. Therefore, finding new drugs seems to be an essential need. Vanillin, which is extracted from vanilla seed, has anti-cancer effects by reducing nuclear factor-κB (NF). We explored the anti-tumor effects of vanillin in the melanoma model and its possible mechanism. In the MTT assay, mice melanoma cells (B16F10) were treated with vanillin (1, 2, 3, 4, 5 μg/mL) for 24 and 48 h. In an animal model, B16F10 was subcutaneously injected into C57BL/6 mice. After the development of tumors, the mice were treated with 50 and 100 mg/kg/day of vanillin for 10 days. The tumor size and expression level of NF-κB protein were measured. In the MTT assay, vanillin in all concentrations significantly decreased B16F10 cell viability after 24 h incubation. The size of melanoma tumors was reduced in both doses 50 and 100 mg/kg/day in mice. NF-κB protein expression was decreased in the 100 mg/kg/day group in comparison with the control group. We found that vanillin by reducing NF-κB expression may have anti-tumor effects and reduced melanoma tumor size and cell viability.

Dexmedetomidine mitigates myocardial ischemiareperfusion injury via regulation of HMGB1-TLR4-NF-κB signaling axis

Purpose: To study the effect of dexmedetomidine (Dex) on myocardial ischemia-reperfusion injury (MIRI), and the associated mechanism of action.Methods: Sixty Sprague-Dawley (SD) rats were assigned to sham, ischemia-reperfusion (I/R), Dex, and MD groups (methyllycaconitine prior to injection with Dex), with 15 rats in each group. Pathological changes in myocardial tissues were determined in all groups. Protein expression levels of HMGB1, TLR4, NF-κB and myeloid differentiation protein 88 (MyD88) in serum and myocardial tissues were assayed and compared.Results: Protein levels of HMGB1, TLR4, MyD88 and NF-κB were significantly higher in heart muscle I/R rats than those in sham group, but lower in heart muscle of rats in Dex group than in heart muscle of I/R rats (p &lt; 0.05). However, they were significantly up-regulated in MD group, relative to Dex group (p&lt; 0.05).Conclusion: Dex exerts a protective effect against ischemia/reperfusion-induced myocardial damage via HMGB1-TLR4-NF-κB signal axis via CAP, and thus, is a potential agent for the management of myocardial disease.

Molecular Mechanism of Biofilm Locator Protein Kinase Dbf2p-related kinase 1 in Regulating Innate Immune Response to Interleukin 17-induced Viral Pneumonia

ABSTRACT It focused on the antiviral immune regulation of biofilm-localized protein kinase Dbf2p-related kinase 1 (NDR1) in viral pneumonia. Mouse alveolar monocyte RAW264.7 was used as blank control, and viral pneumonia cell model was prepared by infecting cells with respiratory syncytial virus (RSV). NDR1 overexpression vector and siRNA interference sequences were synthesized, and overexpression/silence NDR1 cell model was fabricated. About 50 ng/mL interleukin 17 (IL-17) was given to stimulate. Enzyme-linked immunosorbent assay (ELISA), quantitative reverse transcription PCR (RT-qRCR), and Western blot were performed to detect cytokines and chemokines, mRNA of inflammatory factors, and signal molecule protein expression. Notably, RSV infection increased RSV-F mRNA in RAW264.7 cells and reduced NDR1 mRNA and protein. Secretion levels of IL-6, interferon β (IFN-β), chemokine (C-X-C motif) ligand 2 (CXCL2), and chemokine (C-C motif) ligand 2 (CCL20) increased in the model group versus blank control (P< 0.05). IL-6, IFN-β, tumor necrosis factor α (TNF-α), and toll-like receptor 3 (TLR3) mRNA were up-regulated (P < 0.05). Extracellular signal-regulated kinase (ERK1/2), p38 protein phosphorylation, human recombinant 1 (TBK1), and nuclear factor kappa-B (NF-κB) protein levels increased (P < 0.05). After overexpression of NDR1, the secretion levels of cytokines and chemokines, inflammatory factors mRNA, and signal molecule protein increased significantly. After NDR1 was silenced, cytokines and chemokines, inflammatory factors mRNA, and signal molecule protein were not significantly different versus blank control group (P > 0.05). In short, NDR1 regulated innate immune response to viral pneumonia induced by IL-17, which can be used as a new target for the treatment of IL-17-induced inflammatory response and autoimmune diseases.

ILC2s induce adaptive Th2-type immunity in different stages of tuberculosis through the Notch-GATA3 pathway

The study is to investigate the roles of group 2 innate lymphoid cells (ILC2s) in different courses of tuberculosis (TB). Serum and PBMCs were respectively isolated from the TST negative, LTBI (latent TB infection), ATB (active TB) and RTB (recurrent TB) patients. Flow cytometry was used to detect Th1, Th2 and ILC2s in the peripheral blood. The mRNA and protein levels of GATA3, RORα, CRTH2, Hes1, Notch1, NF-κB, and ID2 were detected by qRT-PCR and Western blotting. ILC2 cells from ATB and RTB patients were stimulated with rJagged2 or DAPT in vitro, and co-cultured with CD4+ T cells from TST negative group. ELISA was used to detect cytokine levels. The results showed that compared with the TST negative or LTBI group, Th2 cells and serum IL-4 in ATB group increased dramatically, accompanied by an increase of Th2/Th1 ratio in ATB patients, especially in RTB group. However, ILC2s in the ATB and RTB group increased significantly, along with increased GATA3, RORα, and CRTH2 levels. After rJagged2 stimulation in vitro, the levels of Hes1, Notch1, NF-κB, RORα, GATA3 and ID2 and those of IL-4, IL-5 and IL-13 were significantly increased. These effects were abrogated by DAPT treatment. Then, ILC2s, especially those from RTB patients, induced Th2-type immune response after co-culturing with CD4+ T cells. In conclusion, our results suggest that ILC2s may promote Th2-type immune response in different stages of TB via the Notch-GATA3 pathway.

Novel roles of lipopolysaccharide and TLR4/NF-κB signaling pathway in inflammatory response to liver injury in Budd-Chiari syndrome.

BACKGROUNDBudd-Chiari syndrome (BCS) is an uncommon disorder characterized by obstruction of hepatic venous outflow. To date, the exact mechanism underlying hepatic injury derived from the hepatic venous outflow obstruction in BCS remains largely unknown.AIMTo assess the role of NF-κB-mediated inflammation in BCS-induced liver injury in humans and rats.METHODSA total of 180 rats were randomly assigned into nine groups, including four BCS model groups (1, 3, 6 and 12 wk), four sham-operated groups (1, 3, 6 and 12 wk), and a control group. Lipopolysaccharide (LPS) levels in each group were detected by the Tachypleus Amebocyte Lysate assay. The mRNA and protein levels of TLR4, NF-κB, tumor necrosis factor (TNF)-α, interleukin (IL)-2 and interferon (IFN)-γ were quantified. In addition, 60 patients with BCS and 30 healthy controls were enrolled, and their blood samples were analyzed.RESULTSHepatic and plasma LPS levels were significantly increased in rats. The mRNA and protein expression levels of TLR4, NF-κB and inflammatory cytokines (TNF-α, IL-2 and IFN-γ) in liver tissues were significantly higher in the BCS model groups compared with the other two groups. In addition, the model groups (1, 3, 6 and 12 wk after BCS induction) showed significant differences in the levels of LPS, TLR4, NF-κB, TNF-α, IL-2 and IFN-γ. Notably, there was a significant correlation between the LPS concentrations and mRNA and protein levels of TLR4, NF-κB and inflammatory cytokines. Importantly, it was revealed that the levels of LPS, TLR4, NF-κB and inflammatory cytokines were significantly greater in chronic BCS patients than healthy controls and acute BCS patients.CONCLUSIONLPS level is markedly elevated in BCS, in turn activating the TLR4/NF-κB signaling pathway, leading to induction of inflammatory cytokines (TNF-α, IL-2 and IFN-γ) in response to BCS-induced liver injury.

The molecular mechanisms of signal pathway activating effect of E2F-1/NF-κB/GSK-3β on cognitive dysfunction of Alzheimer rats

ABSTRACT Alzheimer disease (AD) seriously harms human health and its onset is insidious. Therefore, it is of great significance to find out the pathogenesis of AD disease for improving the prevention and treatment effect of the disease. The study drew attention to the influence of E2F-1/NF-κB/GSK-3β signaling pathway on cognitive dysfunction of Alzheimer rats. 60 specific pathogen-free (SPF) SD rats were selected as research subjects. The, the AD model was created by injecting Aβ1-42 into hippocampus CA1 region of AD rats using a microscopic syringe. Besides, Morris water maze test and Western blot were performed to detect the cognitive function, the levels of destination protein and active oxidation products in the brain of rats. Compared to the Sham group, the escape latency and the distance of the model group significantly increased (P < 0.05), and the number of times to pass the target quadrant was significantly reduced (P < 0.05); the expression levels of E2F-1 and NF-κB protein in the hippocampus and the phosphorylation levels of Tau231, Tau262, Tau396, Tau404 and T216-GSK-3β protein of the model group were significantly increased (P < 0.05); the ROS/RNS value in the hippocampus of the model group significantly increased (P < 0.05). AD model rats exhibit obvious cognitive dysfunction, which is associated with the activation of E2F-1/NF-κB/GSK-3β signaling pathway and the heightened Tau protein phosphorylation level.

Neuroprotective effects of liraglutide against inflammation through the AMPK/NF-κB pathway in a mouse model of Parkinson's disease.

Parkinson's disease (PD) is a neurodegenerative disease with increasing incidence in aged populations, second only to Alzheimer's disease. Increasing evidence has shown that inflammation plays an important role in the occurrence and development of Parkinson's disease. Growing evidence has shown that AMP-activated protein kinase (AMPK) and NF-κB are closely related to inflammation. Glucagon-like peptide 1 (GLP-1) is a hormone that is primarily secreted by intestinal endocrine L cells, and it has a variety of physiology through binding to GLP-1 receptor. GLP-1can be used for treatment of type 2 diabetes. In addition, GLP-1 also has anti-neuroinflammation activity. However, the exact mechanism behind how GLP-1 regulates neuroinflammation remains unclear. This study was designed to examine the effect of liraglutide on 1-methyl-4-phenyl-1,2,3,6- tetrahydropyridine (MPTP)-induced injury in mice and its potential mechanism of action. Results showed that liraglutide dose-dependently ameliorated mouse behavior including swimming time and locomotor activity, increased the number of tyrosine hydroxylase (TH)-positive neurons and protein level, and reduced Iba1 and GFAP expression in the substantia nigra (SN). Liraglutide treatment also increased p-AMPK expression and reduced NF-κB protein level. Applying the AMPK inhibitor Dorsomorphin (Compound C) reversed the effect of liraglutide-reducing p-AMPK and increasing NF-κB expression. Finally, GFAP protein level increased, along with a decrease in TH expression. In conclusion, these results suggest that liraglutide can suppress neuroinflammation. Moreover, this effect is mediated through the AMPK/NF-κB signaling pathway.

Sophocarpine Alleviates Injury-Induced Intima Hyperplasia of Carotid Arteries by Suppressing Inflammation in a Rat Model.

Introduction: Balloon angioplasty is a commonly applied procedure for treating atherosclerotic vascular diseases. However, the maintenance of long-term lumen patency is relatively difficult due to the occurrence of restenosis. Previous research has shown that the occurrence of vascular wall inflammation is associated with higher rates of restenosis. Sophocarpine (SPC) can exert various therapeutic effects such as anti-oxidation, anti-inflammation, anti-tumor, antivirus and immune regulation. This study aimed to investigate whether SPC can alleviate intimal hyperplasia following balloon injury in a rat carotid artery model. Methods: Twenty Sprague–Dawley rats were randomly assigned to four groups: (i) control, (ii) balloon injury, (iii) balloon injury followed by saline injection, and (iv) balloon injury followed by SPC administration. Each group contained five rats. A high-pressure balloon of 3 mm × 20 mm was placed in the carotid artery. The balloon was inflated to a pressure of 8 atmospheres to carry out rat carotid artery balloon injury model. The areas of neointimal and media were determined by Verhoeff_Van Gieson staining, and the intima-to-media (I:M) ratios were subsequently evaluated. After that, the protein levels of IL-6, IL-1β, MCP-1, NF-κB, TNF-α, VCAM-1, ICAM-1 and eNOS were measured. Results: The ratio of I:M was remarkably higher in the balloon injury group than in the control group (p < 0.01). SPC could significantly decrease the ratio of I:M compared with the balloon injury group (p < 0.01). Besides, the protein levels of IL-6, IL-1β, MCP-1, NF-κB, TNF-α, ICAM-1 and VCAM-1 were increased in rat carotid arteries exposed to balloon injury (p < 0.01), and treatment with SPC could attenuate these effects (p < 0.05). Furthermore, balloon injury inhibited the protein expression of eNOS (p < 0.01), and SPC could elevate its level (p < 0.05). Conclusions: SPC could alleviate an intimal hyperplasia in balloon-injured carotid artery, and the mechanisms underlying this protective effect might be due to its inhibitory potency against inflammation signals. Our study also implies the potential applicability of SPC in treating restenosis after balloon angioplasty.

Syringaresinol Resisted Sepsis-Induced Acute Lung Injury by Suppressing Pyroptosis Via the Oestrogen Receptor-β Signalling Pathway.

Acute lung injury (ALI) is a common lung disease characterized by severe acute inflammatory lung injury in patients with sepsis. Syringaresinol (SYR) has been reported to have anti-apoptotic and anti-inflammatory effects, but whether it could prevent pyroptosis to improve sepsis-induced ALI remains unclear. The purpose of this work was to examine the impact of SYR on sepsis-induced ALI and investigate the underlying mechanisms. The ALI model was induced by caecal ligation and puncture (CLP) in C57BL/6 mice, structural damage in the lung tissues was determined using haematoxylin and eosin (HE) staining, and the levels of related inflammatory cytokines and macrophage polarization were examined by enzyme-linked immunosorbent assays (ELISAs) and flow cytometry, respectively. The activation of the NLRP3 inflammasome and the protein levels of TLR4, NF-κB and MAPKs was measured by western blotting. The results demonstrated that SYR pretreatment significantly reduced lung tissue histological damage, inhibited the production of proinflammatory cytokines and albumin in bronchoalveolar lavage fluid (BALF), and decreased myeloperoxidase (MPO) levels, thereby alleviating lung tissue injury. Meanwhile, septic mice treated with SYR displayed a higher survival rate and lower percentage of M1 macrophages in the BALF and spleen than septic mice. In addition, lung tissues from the CLP + SYR group exhibited downregulated protein expression of NLRP3, ASC, GSDMD caspase-1 p20 and TLR4, along with decreased phosphorylated levels of NF-κB, ERK, JNK and P38, indicating that SYR administration effectively prevented CLP-induced pyroptosis in the lung. SYR also suppressed LPS-induced pyroptosis in RAW 264.7 cells by inhibiting the activation of the NLRP3 inflammasome, which was abolished by an oestrogen receptor-β (ERβ) antagonist (PHTPP). In conclusion, SYR exerted protective effects on CLP-induced ALI via the oestrogen receptor-β signalling pathway.

Neuroprotective effects of sonochemical- synthesized SiO2 nanoparticles in vivo models of ischemic/reperfusion injury in stroke

Cerebral ischemic injury is one of the debilitating diseases that showed inflammation plays an essential role in aggravating ischemic damage. After synthesizing silica nanoparticles (SiO 2 NPs) by sonochemical method, serum parameters in the presence of different concentrations of SiO 2 NPs are measured for toxicity assay. Rats were separated randomly into control, ischemia/reperfusion, and ischemia/reperfusion + SiO 2 NPs groups. Transient forebrain ischemia induced with bilateral occlusion of both common carotid arteries followed by 60minuts of reperfusion. SiO 2 NPs were administered (500 mg/kg/day p.o.) 21 days before ischemia/reperfusion time. Animals sacrificed and frontal cortex and hippocampal tissues used to determine malondialdehyde (MDA) level, nitric oxide (NO), glutathione (GSH) levels, an essential antioxidant, superoxide dismutase (SOD), alterations in the level of cytokines, TNFα, IL-1β, MCP-1, and phosphor Ik-кB. We also revealed the involvement of NF-κB downregulation by using western blotting. We reported on a histological investigation. The results showed that SiO 2 NPs with a diameter of around 50 nm in dose of 500 mg/kg didn't change the level of liver enzyme (including ALT, AST and ALP) and hematological parameters. 500 mg/kg SiO 2 NPs showed significant effects on remission of behavioral impairment. Ischemia/reperfusion oxidative injury in the rat hippocampus demonstrated a significant increase in MDA, TNFα, MCP-1, IL-1β, phosphor Ik-кB, NO levels, and a significant decrease in GSH contents and SOD activities in the hippocampal tissue compared to the control group. Pretreatment of ischemic rats with SiO 2 NPs decreased the elevated levels of MDA, TNFα, MCP-1, IL-1β, phosphor Ik-кB, and NO levels. A significant alteration observed in SOD activities and GSH content results between treated and untreated ischemia/reperfusion brains in rats. Decreased protein level of NF-κB also measured in SiO 2 NPs-treated animals. Untreated ischemia/reperfusion brains had significantly decreased in number of cells in CA1 hippocampus, nevertheless SiO 2 NPs increase the normal cell and decrease the neurodegeneration in hippocampus but it was not significant alteration. SiO 2 NPs reduced the damage caused by cerebral ischemia/reperfusion in rats and its molecular mechanism attributed to the downregulation of NF-κB signaling pathway.

Effect of electroacupuncture stimulation of different acupoint groups on lipid metabolism and liver TLR4/NF-κB signaling in obese rats

To observe the effect of electroacupuncture (EA) of different acupoint combinations on lipid metabolism and liver Toll-like receptor 4 (TLR4)/nuclear factor kappa B (NF-κB) signaling in obese rats, so as to explore the specificity of regulatory effects of different acupoints for obesity. Male SD rats were randomly divided into normal control, model, hindlimb acupoint EA ［"Fenglong" (ST40)-"Zusanli" (ST36), hindlimb EA］, abdominal acupoint EA ［("Zhongwan" (CV12)-"Tianshu" (ST25), "Guanyuan" (CV4) - the opposite ST25, abdominal EA］, BiaoBen-acupoint EA ［ST40-ST36, CV12 -CV4, ST25, BiaoBen EA］ groups, with 10 rats in each group. The obesity model was induced by feeding the rats with high fat diet. EA (2 Hz, 1 mA) was applied to the acupoint groups mentioned above for 10 min (per acupoint group), 3 times a week for 8 weeks. After the treatment, the body weight and food intake in each group were recorded. The contents of serum total cholesterol (TC), triglyceride (TG), non-esterified fatty acid (NEFA) were detected by using an automatic biochemical analyzer. The expression levels of TLR4 and NF-κB p65 mRNAs and proteins in the liver tissue were detected by quantitative real-time PCR and Western blot, separately. Compared with the normal group, the body weight and food intake, serum TC, TG and NEFA contents, hepatic TLR4 and NF-κB p65 protein and gene expression were significantly increased in the model group (P<0.05, P<0.01). After the treatment, compared with the model group, the body weight, food intake, TC, TG, NEFA, TLR4 and NF-κB p65 protein and mRNA expression were significantly decreased in the hindlimb EA, abdominal EA and BiaoBen EA groups (P<0.05, P<0.01). Comparison among the 3 treatment groups showed that the serum TC, TG and NEFA contents were significantly lower in the hindlimb EA and BiaoBen EA groups than in the abdominal EA group (P<0.01，P<0.05). The expression levels of TLR4 and NF-κB p65 mRNAs and proteins were significantly lower in the BiaoBen EA group than in the hindlimb EA and abdominal EA groups (P<0.01, P<0.05). BiaoBen EA is superior to abdominal EA in improving lipid metabolism in obesity rats, which may be related to its stronger effectiveness in down-regulating hepatic TLR4/NF-κB p65 signaling.

Quercetin attenuates lipopolysaccharide-mediated inflammatory injury in human nasal epithelial cells via regulating miR-21/DMBT1/NF-κB axis

Background Quercetin (Qu) belongs to a flavonoid polyphenolic compound present in fruits and vegetables which has been confirmed to exert anti-inflammatory properties. Our study aimed to explore the impacts of quercetin on lipopolysaccharide (LPS)-induced inflammatory injury and signal transduction of miR-21/DMBT1/NF-κB axis in human nasal epithelial cells (HNEpC). Methods HNEpCs were cultured and treated with 1 μg/mL of LPS and a gradient concentration (10, 100, and 200 μM) of quercetin for 24 h. Cell viability, apoptosis, and cytokines were detected to assess the inflammatory injury in LPS-exposed HNEpCs. The expressions of miR-21, DMBT1, and NF-κB mRNA were detected by quantitative real-time polymerase chain reaction (qRT-PCR). The levels of DMBT1 and NF-κB protein were measured by western blotting. Results LPS treatment reduced cell viability, promoted cell apoptosis and inflammatory response, down-regulated miR-21 expression and up-regulated DMBT1, and NF-κB in HNEpC cells. Quercetin exerted the opposite effects to attenuate LPS-induced inflammatory injury in HNEpC cells at a concentration-dependent way. Additionally, miR-21 directly targeted DMBT1 to reduce its expression and further inducing cell viability via inhibiting cell apoptosis and inflammatory response. MiR-21 inhibition or DMBT1 over-expression weakened the protective effects of quercetin against LPS-induced inflammatory injury in HNEpC cells. Conclusions Quercetin could protect HNEpC cells against LPS-induced inflammatory injury via inducing miR-21/DMBT1/NF-κB axis. Therefore, quercetin could be utilized as a potential compound to treat for allergic rhinitis.

Activation of TLR2 heterodimers-mediated NF-κB, MAPK, AKT signaling pathways is responsible for Vibrio alginolyticus triggered inflammatory response in vitro

Vibrio alginolyticus is an important zoonotic marine pathogenic bacterium. Previous studies on the mechanism of innate immune against V. alginolyticus infection have been limited to aquatic animals, however, how V. alginolyticus activates mammalian immune cells has not been fully clarified. Here, ELISA combined RT-qPCR assays were used to detect the secretion and transcription level of pro-inflammatory cytokines and TLRs during V. alginolyticus infection of mice peritoneal macrophages (PMϕs). Western blotting was used to explore the phosphorylation levels of p38, JNK, ERK, AKT and NF-κB protein. Immunofluorescence assay was used to determine the location of NF-κB protein. Inhibition assay was used to study the role of up-regulated TLR in activated signaling pathways and the role of these pathways in the release of pro-inflammatory cytokines. Our data showed that V. alginolyticus can up-regulate the expression levels of IL-1β, IL-6, IL-12 and TNF-α in PMϕs. In addition, V. alginolyticus stimulation activated the phosphorylation of p38, JNK and ERK were TLR2 heterodimers-dependent, whereas inhibitors of SB203580 (p38), SCH772984 (ERK) and SP600125 (JNK) significantly reduced IL-1β, IL-6, IL-12 and TNF-α production. We further revealed that V. alginolyticus activated the signaling pathways of AKT via TLR2 heterodimers. The inhibitor of MK-2206 2HCl (AKT) negatively regulated the IL-1β, IL-6 and TNF-α release levels. Moreover, V. alginolyticus infection of PMϕs resulted in TLR2 heterodimers-mediated activation of NF-κB and induced translocation of phosphorylated NF-κB protein from the cytoplasm into the nucleus via IκBα degradation. V. alginolyticus induced IL-1β, IL-6, IL-12 and TNF-α release were blocked by the specific NF-κB inhibitor, BAY 11–7082. Taken together, our results suggested that activation of the TLR2 heterodimers-mediated downstream signaling pathways NF-κB, MAPK and AKT is responsible for inflammatory response during Vibrio alginolyticus infection in vitro.