NF-κB P65 DNA Binding Activity Research Articles

444 Lactate and its induced EGR1 are novel key factors that turn hot tumors into cold tumors

Tumors with an inflamed tumor microenvironment are considered “hot tumors” and show good responses to immune checkpoint inhibitors (ICIs). Conversely, non-inflamed tumors, so-called “cold tumors,” have no tumor-suppressive cytokines and rarely respond to ICIs. Therefore, turning non-inflamed (cold) tumors into inflamed (hot) tumors is important for maximizing the effect of ICIs. We showed that lactate, a product of the Warburg effect, inhibited the efficacy of ICIs and suppressed IL-12 p40 expression in dendritic cells (DCs) through reducing NF-κB p65, p50 and c-Rel DNA-binding activity to the IL-12 p40 promoter.

Dapagliflozin Protects H9c2 Cells Against Injury Induced by Lipopolysaccharide via Suppression of CX3CL1/CX3CR1 Axis and NF-κB Activity.

Dapagliflozin, a selective Sodium-glucose cotransporter-2 (SGLT2) inhibitor, has been shown to play a key role in the control and management of metabolic and cardiac diseases. The current study aims to address the effects of dapagliflozin on the expression of fractalkine (FKN), known as CX3CL1, and its receptors CX3CR1, Nuclear factor-kappa B(NF-κB) p65 activity, Reactive oxygen species (ROS), and inflammation in LPS-treated H9c2 cell line. H9c2 cells were cultured with lipopolysaccharide (LPS) to establish a model of LPS-induced damage, and then, subsequently were treated with dapagliflozin for 72 h. Our work included measurement of cell viability (MTT), Malondialdehyde (MDA), intracellular ROS, tumor necrosis factor-α (TNF-α), NF-κB activity, and expression of CX3CL1/CX3CR1. The results showed that LPS-induced reduction of cell viability was successfully rescued by dapagliflozin treatment. The cellular levels of MDA, ROS, and TNF-α, as an indication of cellular oxidative stress and inflammation, were significantly elevated in H9c2 cells compared to the control group. Furthermore, dapagliflozin ameliorated inflammation and oxidative stress through the modulation of the levels of MDA, TNF-α, and ROS. Correspondingly, dapagliflozin reduced the expression of CX3CL1/CX3CR1, NF-κB p65 DNA binding activity, and it also attenuated nuclear acetylated NF-κB p65 in LPS-induced injury in H9c2 cells compared to untreated cells. These findings shed light on the novel pharmacological potential of dapagliflozin in the alleviation of LPS-induced CX3CL1/CX3CR1-mediated injury in inflammatory conditions such as sepsis-induced cardiomyopathy.

Topical application of metformin accelerates cutaneous wound healing in streptozotocin-induced diabetic rats.

Diabetic chronic wound, which is one of the diabetic complications caused by hyperglycemia, characterized by prolonged inflammation has become one of the most serious challenges in the clinic. Hyperglycemia during diabetes not only causes prolonged inflammation and delayed wound healing but also modulates the activation of nuclear factor-kappa B (NF-κB) and the expression of matrix metalloproteinases (MMPs). Although metformin is the oldest oral antihyperglycemic drug commonly used for treating type 2 diabetes, few studies have explored the molecular mechanism of its topical effect on wound healing. Therefore, we aimed to investigate the molecular effects of topical metformin application on delayed wound healing, which's common in diabetes. In this context, we created a full-thickness excisional wound model in Wistar albino rats and, investigated NF-κB p65 DNA-binding activity and expression levels of RELA (p65), MMP2 and MMP9 in wound samples taken on days 0, 3, 7, and 14 from diabetic/non-diabetic rats treated with metformin and saline. As a result of our study, we showed that topically applied metformin accelerates wound healing by suppressing NF-κB p65 activity and diminishing the expression of MMP2 and MMP9. Diabetic wounds treated with metformin healed even faster than those in the control group that mimicked standard wound healing.

A mechanism of cooling hot tumors: Lactate attenuates inflammation in dendritic cells

SummaryTurning non-inflamed (cold) tumors into inflamed (hot) tumors is important for maximizing the effect of immune checkpoint inhibitors (ICIs) against malignancies. We showed that lactate, a product of the Warburg effect, inhibited the efficacy of ICIs and suppressed IL-12 p40 expression in dendritic cells (DCs) through reducing NF-κB p65, p50, and c-Rel DNA-binding activity to the IL-12 p40 promoter. Additionally, lactate promoted the expression of early growth response protein 1 (EGR1), whose expression was increased in human invasive melanoma compared with non-invasive melanoma. We also found that EGR1 interacts with serum response factor (SRF) and represses the expression of CD80 in DCs. These findings suggest that lactate and its induced EGR1 are key factors that turn hot tumors into cold tumors and may represent targets in cancer treatment with ICIs.

Triptolide dose-dependently improves LPS-induced alveolar hypercoagulation and fibrinolysis inhibition through NF-κB inactivation in ARDS mice

BackgroundAlveolar hypercoagulation and fibrinolysis inhibition were associated with the refractory hypoxemia and the high mortality in patient with acute respiratory distress syndrome (ARDS), and NF-κB pathway was confirmed to contribute to the process. Triptolide (TP) significantly inhibited NF-κB pathway and thus depressed accessive inflammatory response in ARDS. We speculate that TP could improve alveolar hypercoagulation and fibrinolytic inhibition in LPS-induced ARDS via NF-κB inactivation. PurposeThe aim of this experiment was to explore the efficacy and potential mechanism of TP on alveolar hypercoagulation and fibrinolysis inhibition in LPS-induced ARDS in mice. Methods50 μl of LPS (5 mg/ml) was inhalationally given to C57BL/6 mice to set up ARDS model. Male mice were randomly accepted with LPS, LPS + TP (1 μg/kg, 10 μg/kg, 50 μg/kg respectively), or with NEMO Binding domain peptide (NBD), an inhibitor of NF-κB. TP (1 μg/kg, 10 μg/kg, 50 μg/kg) were intraperitoneally injected or 10 μg/50 μl of NBD solution were inhaled 30 min before LPS inhalation. A same volume of normal saline (NS) substituted for TP in mice in control. The endpoint of experiment was at 8 hours after LPS stimulation. Pulmonary tissues were taken for hematoxylin-eosin (HE) staining, wet / dry ratio and for lung injury scores (LIS). Tissue factor (TF) and plasminogen activator inhibitor (PAI)-1 in lung tissue were detected by Western-blotting and by quantitative Real-time PCR(qPCR) respectively. Concentrations of TF, PAI-1, thrombin-antithrombin complex (TAT), procollagen peptide type Ⅲ (PⅢP) and activated protein C (APC) in bronchoalveolar lavage fluid (BALF) were measured by ELISA. NF-κB activation and p65-DNA binding activity in pulmonary tissue were simultaneously determined. ResultsLPS stimulation resulted in pulmonary edema, neutrophils infiltration, obvious alveolar collapse, interstitial congestion, with high LIS, which were all dose-dependently ameliorated by Triptolide. LPS also dramatically promoted the expressions of TF and PAI-1 either in mRNA or in protein in lung tissue, and significantly stimulated the secretions of TF, PAI-1, TAT, PⅢP but inhibited APC production in BALF, which were all reversed by triptolide treatment in dose-dependent manner. TP dose-dependently inhibited the activation of NF-κB pathway induced by LPS, indicated by the changes of phosphorylations of p65 (p-p65), p-IKKα/β and p-IκBα, and weakened p65-DNA binding activity. TP and NBD had same efficacies either on alveolar hypercoagulation and fibrinolysis inhibition or on NF-κB signalling pathway in ARDS mice. ConclusionsTP dose-dependently improves alveolar hypercoagulation and fibrinolysis inhibition in ARDS mice through inhibiting NF-κB signaling pathway. Our data demonstrate that TP is expected to be an effective selection in ARDS.

Protective effect of toll-interacting protein overexpression against paraquat-induced lung injury in mice and A549 cells through inhibiting oxidative stress, inflammation, and NF-κB signaling pathway

Toll-interacting protein (Tollip) is a pivotal negative regulator of inflammatory response. In the present study, the effects of Tollip overexpression on paraquat (PQ)-induced lung injury were explored through in vivo and in vitro investigations. Upon stimulation with PQ in mice, the expression of Tollip was down-regulated. Histopathological analysis revealed that the overexpression of Tollip significantly decreased inflammatory cell infiltration. Similarly, the levels of myeloperoxidase (MPO) and interleukin-1β (IL-1β) were lowered by Tollip overexpression in PQ-administrated mice. Besides, the overexpression of Tollip reduced reactive oxygen species (ROS) generation and malondialdehyde (MDA) level but enhanced superoxide dismutase (SOD) activity in PQ-treated A549 cells. Meanwhile, Tollip overexpression lowered the level of IL-1β and decreased the protein expressions of p-p65 in the cytoplasm and nuclear p65. Importantly, inhibition of NF-κB signaling pathway probably by decreasing NF-κB p65-DNA binding activity was induced by Tollip overexpression. Taken together, Tollip overexpression attenuated PQ-initiated lung injury possibly via reduction of oxidative stress and inflammation and suppression of NF-κB signaling pathway activation, which provided some novel ideas for the treatment of lung damage mediated by PQ.

Human Amnion Membrane Proteins Prevent Doxorubicin-Induced Oxidative Stress Injury and Apoptosis in Rat H9c2 Cardiomyocytes.

Doxorubicin (DOX) is widely used as an effective chemotherapy agent in cancer treatment. Cardiac toxicity in cancer treatment with DOX demand urgent attention and no effective treatment has been established for DOX-induced cardiomyopathy. It has been well documented that human amniotic membrane proteins (AMPs), extracted from amnion membrane (AM), have antioxidant, anti-apoptotic, and cytoprotective properties. Therefore, in this study, we aimed to investigate the protective effects of AMPs against cardiotoxicity induced by DOX in cultured rat cardiomyocyte cells (H9c2). DOX-induced cell injury was evaluated using multi-parametric assay including thiazolyl blue tetrazolium bromide (MTT), the release of lactic dehydrogenase (LDH), intracellular Ca2+ , reactive oxygen species (ROS) levels, cellular antioxidant status, mitochondrial membrane potential (ΔΨm), malondialdehyde (MDA), and NF-κB p65 DNA-binding activity. Moreover, expression profiling of apoptosis-related genes (P53, Bcl-2, and Bax) and Annexin V by flow cytometry were used for cell apoptosis detection. It was shown that AMPs pretreatment inhibited the cell toxicity induced by DOX. AMPs effectively attenuated the increased levels of LDH, Ca2+ , ROS, and MDA and also simultaneously elevated the ΔΨm and antioxidant status such as superoxide dismutase (SOD) and Catalase (CAT) in pretreated H9c2 cardiomyocytes. Besides, the activity of NF-kB p65 was reduced and the p53 and Bax protein levels were inhibited in these myocardial cells subjected to DOX. These findings provide the first evidence that AMPs potently suppressed DOX-induced toxicity in cardiomyocytes through inhibition of oxidative stress and apoptosis. Thus, AMPs can be a potential therapeutic agent against DOX cardiotoxicity.

Ethanol extract of Elephantopus scaber Linn. Attenuates inflammatory response via the inhibition of NF-κB signaling by dampening p65-DNA binding activity in lipopolysaccharide-activated macrophages.

Elephantopus scaber Linn. (E.scaber) is a widely-used traditional herb whose use has been documented for various inflammatory diseases such as fever, sore throat, dysentery, carbuncle and so on. However, the effect and mechanism of E.scaber in LPS-activated macrophages remain unclear. This study aims to investigate the anti-inflammatory mechanism of the ethanol extract of E.scaber (ESE) in lipopolysaccharide (LPS)-induced inflammatory models. Griess reagent was used to determine NO production, and the levels of TNF-α, IL-6, MCP-1 and IL-1β were determined by ELISA kits. The molecular mechanism research was performed by RT-PCR, Western blot, and electrophoretic mobility shift assay (EMSA). LPS-induced endotoxemia mouse model was used for evaluating the in vivo anti-inflammatory action of ESE. ESE suppressed LPS-induced iNOS, TNF-α, IL-6, MCP-1 and IL-1β transcription as well as supernatant NO, TNF-α, IL-6, MCP-1 and IL-1β production in macrophages. Although ESE inhibited NF-κB activation, it did not affect the IκBα phosphorylation and degradation and the NF-κB p65 nuclear translocation. The result of EMSA revealed that ESE inhibited the NF-κB p65-DNA binding activity. Additionally, ESE also decreased the proinflammatory cytokines in serum and peritoneal lavage fluid of LPS-induced endotoxemic mice. ESE has a potently anti-inflammatory effect through inhibiting the NF-κB p65-DNA binding activity in LPS-activated macrophages.

Cyclic Stretch Force Induces Periodontal Ligament Cells to Secrete Exosomes That Suppress IL-1β Production Through the Inhibition of the NF-κB Signaling Pathway in Macrophages.

In the oral mechanical environment, periodontal ligament cells (PDL cells) contribute to maintaining periodontal tissue homeostasis. Recent studies showed that exosomes, which are small vesicles secreted by various types of cells, play a pivotal role in cell-to-cell communication in biological processes. We examined the secretion of exosomes from PDL cells stimulated with cyclic stretch and their role in the inflammatory response of macrophages using the human macrophage cell line THP-1 and human primary monocytes/macrophages. We prepared supernatants from human PDL cells (PDL-sup) stimulated with cyclic stretch. The treatment of macrophages with PDL-sup, but not PDL-sup from unstimulated PDL cells, inhibited the production of IL-1β in LPS/nigericin-stimulated macrophages. The pretreatment of PDL cells with GW4869, an inhibitor of exosome secretion, or siRNA for Rab27B, which controls exosome secretion, abrogated the inhibitory effects of PDL-sup. A transmission electron microscopy analysis demonstrated the existence of exosomes with diameters ranging between 30 and 100 nm in PDL-sup, suggesting that exosomes in PDL-sup contribute to this inhibition. An immunofluorescence microscopy analysis revealed that exosomes labeled with PKH67, a fluorescent dye, were incorporated by macrophages as early as 2 h after the addition of exosomes. Purified exosomes inhibited IL-1β production in LPS/nigericin-stimulated macrophages and the nuclear translocation of NF-κB as well as NF-κB p65 DNA-binding activity in LPS-stimulated macrophages, suggesting that exosomes suppress IL-1β production by inhibiting the NF-κB signaling pathway. Our results indicate that PDL cells in mechanical environments contribute to the maintenance of periodontal immune/inflammatory homeostasis by releasing exosomes.

Effect of Astragalus membranaceus in Ovalbumin-Induced Allergic Rhinitis Mouse Model.

Background Astragalus membranaceus (AM), a traditional Chinese medicine, has been used to treat allergic diseases, but the mechanism for treating allergic rhinitis (AR) remains unclear. Objective The purpose of this study was to look at the anti-inflammatory effect of AM on AR and the mechanism of anti-allergy. Methods The mouse model of AR was induced by ovalbumin. Allergic symptoms, number of eosinophils in nasal mucosa, and levels of inflammatory cells in nasal lavage fluid were analyzed. We explored the serum immunoglobulin E (IgE), interleukin-4 (IL-4), IL-5, IL-13, interferon-γ (IFN-γ), and IL-10. Besides, the relative mRNA of IL-4, IL-5, and IL-13 was also detected in nasal mucosa tissue. The proportion of CD4+ CD25+ Foxp3+ T cells in the spleen and nasal lymphoid tissue were analyzed. The mRNA levels of nuclear factor-kappa B p65 (NF-κB p65) and inhibitory kappa B alpha (IκBα), as well as NF-κB p65 DNA binding activity, were tested. We also measured the protein levels of NF-κB p65 and p-NF-κB p65 in nasal mucosa. Results AM could reduce the number of eosinophils in the nasal mucosa and decrease the levels of inflammatory cells in nasal lavage fluid. The serum IgE, IL-4, IL-5, and IL-13 were also decreased, while levels of IFN-γ and IL-10 were increased. The relative mRNA of IL-4, IL-5, and IL-13 was decreased by AM. AM increased the proportion of CD4+ CD25+ Foxp3+ T cells in the spleen and nasal lymphoid tissue. In addition, AM could reduce the activity of NF-kB by inhibiting the mRNA expression and DNA binding activity of NF-κB p65. However, AM had no significant effect on mRNA of IκBα. Above all, AM could reduce the p-NF-κB p65 protein expression of nasal mucosa. Conclusions AM could reduce the secretion of inflammatory cytokines by increasing the level of CD4+ CD25+ Foxp3+ T cells and inhibiting the activation of the NF-κB.

Curcumin Attenuates Asthmatic Airway Inflammation and Mucus Hypersecretion Involving a PPARγ-Dependent NF-κB Signaling Pathway In Vivo and In Vitro.

Asthma is characterized by airway inflammation and mucus hypersecretion. Curcumin possessed a potent anti-inflammatory property involved in the PPARγ-dependent NF-κB signaling pathway. Then, the aim of the current study was to explore the value of curcumin in asthmatic airway inflammation and mucus secretion and its underlying mechanism. In vivo, mice were sensitized and challenged by ovalbumin (OVA) to induce chronic asthma. Airway inflammation and mucus secretion were analyzed. In vitro, BEAS-2B cells were obtained. MCP-1, MUC5AC, and PPARγ expression and the phosphorylation of NF-κB p65 and NF-κB p65 DNA-binding activity were measured in both the lungs and BEAS-2B cells. shRNA-PPARγ was used to knock down PPARγ expression. We found that OVA-induced airway inflammation and mucus hypersecretion in mice, OVA and IL-4-induced upregulation of MCP-1 and MUC5AC, suppression of PPARγ, and activation and translocation of NF-κB p65 were notably improved by curcumin both in vivo and in vitro. Our data also showed that these effects of curcumin were significantly abrogated by shRNA-PPARγ. Taken together, our results indicate that curcumin attenuated OVA-induced airway inflammation and mucus hypersecretion in mice and suppressed OVA- and IL-4-induced upregulation of MCP-1 and MUC5AC both in vivo and in vitro, most likely through a PPARγ-dependent NF-κB signaling pathway.

β-Casomorphin-7 Ameliorates Sepsis-Induced Acute Kidney Injury by Targeting NF-κB Pathway.

BackgroundThe aim of this study was to investigate the protective effect of β-casomorphin-7 (β-CM-7) and its possible mechanisms on acute kidney injury (AKI).Material/MethodsRats were randomly divided into a sham group, a cecal ligation and puncture (CLP) group, and a CLP+β-CM-7 group. Kidney index, kidney function, and histopathology changes were assessed. The expression of neutrophil gelatinase-associated lipocalin (NGAL), kidney injury molecule-1 (Kim-1), nuclear factor of kappa light polypeptide gene enhancer in B-cells inhibitor alpha (IκBα), and p-IκBα in kidney tissues were detected by Western blotting. Inflammatory and oxidative stress factors were detected by ELISA kits.ResultsThe results showed that treatment with β-CM-7 reduced the levels of creatinine (Cre), blood urea nitrogen (BUN), NGAL, and Kim-1 induced by CLP, weakening the pathological damage. In the CLP + β-CM-7 group, the tumor necrosis factor-α (TNF-α) level and the DNA-binding activity of NF-κB p65 were significantly reduced and the interleukin-10 (IL-10) level was significantly increased compared with the CLP group. b-CM-7 decreased the expression of p-IκBα/IκBα. In addition, β-CM-7 increased the activity of superoxide dismutase (SOD) and decreased the level of malondialdehyde (MDA) in kidney tissue.Conclusionsβ-CM-7 attenuated sepsis-induced AKI through reducing inflammation and oxidative stress and by inhibition of nuclear factor (NF)-κB activities. This study provides a new therapeutic agent for attenuating sepsis-induced kidney injury.

Natural Compound Oridonin Inhibits Endotoxin-Induced Inflammatory Response of Activated Hepatic Stellate Cells.

Hepatic stellate cells (HSCs) play an important role in hepatic fibrogenesis and inflammatory modulation. Endotoxin is dramatically increased in portal venous blood after serious injury and can contribute to liver damage. However, the mechanism underlying endotoxin's effects on HSCs remains largely unknown. Oridonin is a bioactive diterpenoid isolated from Rabdosia rubescens that exhibits anti-inflammatory properties in different tissues. In the present study, we determined the effects of oridonin on endotoxin-induced inflammatory response and signaling pathways in vitro. The production of proinflammatory cytokines in activated human HSCs line LX-2 was measured by ELISA and Western blots. Immunofluorescence and nuclear fractionation assay were used to determine NF-κB activity. Oridonin treatment significantly inhibited LPS-induced proinflammatory cytokines IL-1β, IL-6, and MCP-1 production as well as cell adhesion molecules ICAM-1 and VCAM-1. Additionally, oridonin blocked LPS-induced NF-κB p65 nuclear translocation and DNA binding activity. Oridonin prevented LPS-stimulated NF-κB regulator IKKα/β and IκBα phosphorylation and IκBα degradation. Combined treatment of oridonin and an Hsp70 substrate binding inhibitor synergistically suppressed LPS-stimulated proinflammatory cytokines and NF-κB pathway activation. Therefore, oridonin inhibits LPS-stimulated proinflammatory mediators through IKK/IκBα/NF-κB pathway. Oridonin could be a promising agent for a hepatic anti-inflammatory.

The metastasis suppressor, NDRG1, attenuates oncogenic TGF-β and NF-κB signaling to enhance membrane E-cadherin expression in pancreatic cancer cells.

The metastasis suppressor, N-myc downstream-regulated gene-1 (NDRG1), plays multifaceted roles in inhibiting oncogenic signaling and can suppress the epithelial mesenchymal transition (EMT), a key step in metastasis. In this investigation, NDRG1 inhibited the oncogenic effects of transforming growth factor-β (TGF-β) in PANC-1 pancreatic cancer cells, promoting expression and co-localization of E-cadherin and β-catenin at the cell membrane. A similar effect of NDRG1 at supporting E-cadherin and β-catenin co-localization at the cell membrane was also demonstrated for HT-29 colon and CFPAC-1 pancreatic cancer cells. The increase in E-cadherin in PANC-1 cells in response to NDRG1 was mediated by the reduction of three transcriptional repressors of E-cadherin, namely SNAIL, SLUG and ZEB1. To dissect the mechanisms how NDRG1 inhibits nuclear SNAIL, SLUG and ZEB1, we assessed involvement of the nuclear factor-κB (NF-κB) pathway, as its aberrant activation contributes to the EMT. Interestingly, NDRG1 comprehensively inhibited oncogenic NF-κB signaling at multiple sites in this pathway, suppressing NEMO, Iĸĸα and IĸBα expression, as well as reducing the activating phosphorylation of Iĸĸα/β and IĸBα. NDRG1 also reduced the levels, nuclear co-localization and DNA-binding activity of NF-κB p65. Further, Iĸĸα, which integrates NF-κB and TGF-β signaling to upregulate ZEB1, SNAIL and SLUG, was identified as an NDRG1 target. Considering this, therapies targeting NDRG1 could be a new strategy to inhibit metastasis, and as such, we examined novel anticancer agents, namely di-2-pyridylketone thiosemicarbazones, which upregulate NDRG1. These agents downregulated SNAIL, SLUG and ZEB1 in vitro and in vivo using a PANC-1 tumor xenograft model, demonstrating their marked potential.

Antifibrosis Effect of Novel Oridonin Analog CYD0618 Via Suppression of the NF-κB Pathway

BackgroundLiver fibrosis is characterized as excessive deposition of the extracellular matrix proteins, primarily by activated hepatic stellate cells (HSCs). NF-κB has been reported as one of the major mediators of HSC activation. Previously, our team reported that oridonin exhibited antihepatic fibrogenetic activity in vitro. In this study, we examined the effects of its novel derivative CYD0618 on HSC viability, apoptosis, and NF-κB signaling. MethodsCell proliferation of activated human and rat HSC lines LX-2 and HSC-T6 was measured using Alamar Blue Assay. Apoptosis was measured by a Cell Death Detection ELISA kit. Cellular proteins were determined by Western blots and immunofluorescence. ResultsCYD0618 significantly inhibited LX-2 and HSC-T6 cell proliferation in a dose-dependent manner. CYD0618 induced cell apoptosis in both cell lines. CYD0618 treatment increased cell cycle inhibitory protein p21, p27, and induced apoptosis marker cleaved poly (ADP-ribose) polymerase, while suppressing the expression of Collagen type 1. CYD0618 blocked lipopolysaccharide (LPS)-induced NF-κB p65 nuclear translocation and DNA binding activity and prevented LPS-induced NF-κB inhibitory protein IκBα phosphorylation and degradation. LPS-stimulated NF-κB downstream target cytokines IL-6 and MCP-1 were attenuated by CYD0618. Endogenous and LPS-stimulated NF-κB p65 S536 phosphorylation was inhibited by CYD0618 treatment. ConclusionsThe potent antihepatic fibrogenetic effect of CYD0618 may be mediated via suppression of the NF-κB pathway.

Caffeic acid phenethyl ester inhibits the progression of ovarian cancer by regulating NF-κB signaling

Ovarian cancer is one of the most common malignancies in women and characterized by a rapid progression to metastasis, which restricts effective treatment options. Caffeic acid phenethyl estre (CAPE), a natural honeybee product, exhibits a variety of biological activities, anti-tumor is included. Our study aims to explore whether CAPE could inhibit the progression of ovarian cancer and the underlying molecular mechanism. The establishment of ovarian cancer model was set up in mice through caudal vein injection of SKOV-3 cells. Results indicated that CAPE treatment remarkably decreased the viability, migration and invasion of SKOV-3 cells. Besides that, the apoptosis of SKOV-3 cells was significantly promoted by CAPE treatment. Moreover, the growth of ovarian cancer was tremendously inhibited by CAPE in vivo and its action was accompanied by the obstructed Ki67 and PCNA expression. Furthermore, nuclear factor kappa b (NF-κB) pathway was dramatically suppressed by CAPE through the inhibition of IκB phosphorylation, nuclear translocation of p65 and NF-κB p65 DNA binding activity. By contrast, cells transfected with p65 siRNA exhibited decreased cell viability, migration and invasion along with increased cell apoptosis in SKOV-3 cells. However, CAPE treatment could enhance these alters induced by p65 siRNA in KOV-3 cells. Taken together, these findings suggested that CAPE could restrain the progression of ovarian cancer via inactivating NF-κB signaling, and may provide novel therapeutic regimens for ovarian cancer.

Modulation of nuclear factor-kappa B activation by the endoplasmic reticulum stress sensor PERK to mediate estrogen-induced apoptosis in breast cancer cells

Stress responses are critical for estrogen (E2)-induced apoptosis in E2-deprived breast cancer cells. Nuclear factor-kappa B (NF-κB) is an important therapeutic target to prevent stress responses in chronic inflammatory diseases including cancer. However, whether E2 activates NF-κB to participate in stress-associated apoptosis in E2-deprived breast cancer cells is unknown. Here, we demonstrated that E2 differentially modulates NF-κB activity according to treatment time. E2 initially has significant potential to suppress NF-κB activation; it completely blocks tumor necrosis factor alpha (TNFα)-induced activation of NF-κB. We found that E2 preferentially and constantly enhances the expression of the adipogenic transcription factor CCAAT/enhancer binding protein beta (C/EBPβ), which is responsible for the suppression of NF-κB activation by E2 in MCF-7:5C cells. Interestingly, NF-κB p65 DNA-binding activity is increased when E2 is administered for 48 h, leading to the induction of TNFα and associated apoptosis. Blocking the nuclear translocation of NF-κB can completely prevent the induction of TNFα and apoptosis induced by E2. Further examination revealed that protein kinase RNA-like endoplasmic reticulum kinase (PERK), a stress sensor of unfolded protein response (UPR), plays an essential role in the late activation of NF-κB by E2. This modulation between PERK and NF-κB is mainly mediated by a stress responsive transcription factor, transducer and activator of transcription 3 (STAT3), independently of the classic canonical IκBα signaling pathway. Thus, inhibition of PERK kinase activity completely blocks the DNA binding of both STAT3 and NF-κB, thereby preventing induction of NF-κB-dependent genes and E2-induced apoptosis. All of these findings suggest that PERK is a key regulator to convey stress signals from the endoplasmic reticulum to the nucleus and illustrate a crucial role for the novel PERK/STAT3/NF-κB/TNFα axis in E2-induced apoptosis in E2-deprived breast cancer cells.

Protective effect of icaritin on focal cerebral ischemic–reperfusion mice

ObjectiveTo investigate the protective effects of icaritin (ICT), one of the active ingredients in Epimedii Folium, on mouse model of cerebral ischemia-reperfusion (I/R) in vivo. MethodsICR mice were subjected to an 1 h transient middle cerebral artery occlusion (MCAO) and followed by 24 h of reperfusion. Neurological deficits, infarct volume, brain edema and survive rate were measured, respectively. The levels of brain IL-1β, TNF-α, ROS and DNA-binding activity of NF-κB p65 were measured by ELISA kits. The levels of malondialdehyde (MDA) and activities of superoxide dismutase (SOD) were detected by spectrophotometry, and the release of nitric oxide (NO) were detected by Griess kit. ResultsICT markedly reduced the neurological deficit scores, brain edema, infarct volume and increased the survival rate of the cerebral I/R mice. The expression of IL-1β, TNF-α, NO, MDA and DNA-binding activity of NF-κB p65 were significantly inhibited by ICT, while the activity of SOD were up-regulated at the same time. ConclusionICT possessed significant neuroprotective effects in cerebral I/R mice, which might be related to prevent neuroinflammatory and oxidative damage.

Resolvin D1 attenuates imiquimod-induced mice psoriasiform dermatitis through MAPKs and NF-κB pathways

BackgroundResolvin D1 (RvD1), a pro-resolution lipid mediator derived from docosahexaenoic acid (DHA), has been described to promote several kinds of inflammatory resolution. However, the effects and anti-inflammatory mechanisms of RvD1 on psoriasis have not been previously reported. ObjectiveThe present study aimed to determine the protective effects and the underlying mechanisms of RvD1 on imiquimod (IMQ)-induced psoriasiform dermatitis. MethodsMice were topically treated with IMQ to develop psoriasiform dermatitis on their shaved back, pretreated intraperitoneally (i.p.) with or without RvD1 or tert-butoxycarbonyl Met-Leu-Phe peptide (Boc), a lipoxin A4 (ALX) receptor antagonist. The severity was monitored and graded using a modified human scoring system, the Psoriasis Area and Severity Index (PASI), histopathology, and the signature cytokines of psoriasis (IL-23, IL-17, IL-22 and TNF-α). The mRNA and protein levels of inflammatory cytokines were quantified by quantitative real-time PCR (QRT-PCR) and ELISA. The expressions of signaling proteins MAPKs and NF-κB p65 were analyzed using western blotting. Electrophoretic mobility shift assay (EMSA) was used to check NF-κB p65 DNA binding activity. ResultsOur study showed that RvD1 alleviated IMQ-induced psoriasiform dermatitis and improved skin pathological changes. RvD1 markedly inhibited IMQ-induced activation of ERK1/2, p38, JNK (c-Jun N-terminal protein kinase, a subfamily of MAPKs), and NF-κB. Furthermore, pretreatment with Boc, would not exacerbate skin inflammation of IMQ-induced mice, but significantly reversed the beneficial effects of RvD1 on IMQ-induced psoriasiform inflammation. ConclusionRvD1 can obviously improve skin inflammation in IMQ-induced mice psoriasiform dermatitis. The protective mechanisms might be related to its selective reaction with lipoxin A4 receptor/Formyl-peptide receptor 2 (ALX/FPR2), by downregulating relevant cytokines of the IL-23/IL-17 axis expression, the inhibition of MAPKs and NF-κB signaling transduction pathways. Thus, these results show that RvD1 could be a possible candidate for psoriasis therapy.

Berberine suppresses LPS-induced inflammation through modulating Sirt1/NF-κB signaling pathway in RAW264.7 cells

Chronic inflammation is a major contributing factor in the pathogenesis of many diseases. Natural product berberine (BBR) exhibits potent anti-inflammatory effect in vitro and in vivo, while the underlying mechanisms remain elusive. Sirt1, a NAD+-dependent protein deacetylase, was recently found to play an important role in modulating the development and progression of inflammation. Thus, we speculate that Sirt1 might mediate the inhibitory effect of BBR on inflammation. In LPS-stimulated RAW264.7 macrophages, BBR treatment significantly downregulated the expression of proinflammatory cytokines such as MCP-1, IL-6 and TNF-α. Importantly, BBR potently reversed LPS-induced down-regulation of Sirt1. Consistently, the inhibitory effects of BBR on proinflammatory cytokines expression was largely abrogated by Sirt1 inhibition either by EX527, a Sirt1 inhibitor or Sirt1 siRNA. Further mechanistic studies revealed that BBR-induced inhibition of NF-κB is Sirt1-dependent, as either pharmacologically or genetically inactivating Sirt1 enhanced the IκΒα degradation, IKK phosphorylation, NF-κB p65 acetylation and DNA-binding activity. Taken together, our results provide the first evidence that BBR potently suppressed inflammatory responses in macrophages through inhibition of NF-κB signaling via Sirt1-dependent mechanisms.