ABSTRACT
Neuroinflammation has been shown to exacerbate ischemic brain injury, and is considered as a prime target for the development of stroke therapies. Clinacanthus nutans Lindau (C. nutans) is widely used in traditional medicine for treating insect bites, viral infection and cancer, due largely to its anti-oxidative and anti-inflammatory properties. Recently, we reported that an ethanol extract from the leaf of C. nutans could protect the brain against ischemia-triggered neuronal death and infarction. In order to further understand the molecular mechanism(s) for its beneficial effects, two experimental paradigms, namely, in vitro primary cortical neurons subjected to oxygen-glucose deprivation (OGD) and in vivo rat middle cerebral artery (MCA) occlusion, were used to dissect the anti-inflammatory effects of C. nutans extract. Using promoter assays, immunofluorescence staining, and loss-of-function (siRNA) approaches, we demonstrated that transient OGD led to marked induction of IL-1ß, IL-6 and TNFα, while pretreatment with C. nutans suppressed production of inflammatory cytokines in primary neurons. C. nutans inhibited IL-1ß transcription via preventing NF-κB/p65 nuclear translocation, and siRNA knockdown of either p65 or IL-1ß mitigated OGD-mediated neuronal death. Correspondingly, post-ischemic treatment of C. nutans attenuated IκBα degradation and decreased IL-1ß, IL-6 and TNFα production in the ischemic brain. Furthermore, IL-1ß siRNA post-ischemic treatment reduced cerebral infarct, thus mimicking the beneficial effects of C. nutans. In summary, our findings demonstrated the ability for C. nutans to suppress NF-κB nuclear translocation and inhibit IL-1ß transcription in ischemic models. Results further suggest the possibility for using C. nutans to prevent and treat stroke patients.
Subject(s)
Acanthaceae/chemistry , Anti-Inflammatory Agents/therapeutic use , Brain Ischemia/drug therapy , Infarction, Middle Cerebral Artery/drug therapy , Interleukin-1beta/biosynthesis , NF-kappa B/metabolism , Neurons/drug effects , Plant Extracts/pharmacology , Plant Leaves/chemistry , Plants, Medicinal/chemistry , Animals , Anti-Inflammatory Agents/pharmacology , Cell Death/drug effects , Cells, Cultured , Cerebral Infarction/pathology , Drug Evaluation, Preclinical , Glucose/pharmacology , Interleukin-1beta/genetics , Male , NF-KappaB Inhibitor alpha/metabolism , Oxygen/pharmacology , Phytotherapy , Promoter Regions, Genetic , Protein Transport/drug effects , RNA Interference , RNA, Small Interfering/genetics , Rats , Rats, Long-Evans , Transcription Factor RelA/antagonists & inhibitors , Transcription Factor RelA/genetics , Transcription, Genetic/drug effects , Tumor Necrosis Factor-alpha/biosynthesis , Tumor Necrosis Factor-alpha/geneticsABSTRACT
Nuclear factor-κB (NF-κB) is an important transcription factor involved in various biological functions, including tumorigenesis. Hence, NF-κB has attracted attention as a target factor for cancer treatment, leading to the development of several inhibitors. However, existing NF-κB inhibitors do not discriminate between its subunits, namely, RelA, RelB, cRel, p50, and p52. Conventional methods used to evaluate interactions between transcription factors and DNA, such as electrophoretic mobility shift assay and luciferase assays, are unsuitable for high-throughput screening (HTS) and cannot distinguish NF-κB subunits. We developed a HTS method named DNA strand exchange fluorescence resonance energy transfer (DSE-FRET). This assay is suitable for HTS and can discriminate a NF-κB subunit. Using DSE-FRET, we searched for RelA-specific inhibitors and verified RelA inhibition for 32,955 compounds. The compound A55 (2-(3-carbamoyl-6-hydroxy-4-methyl-2-oxopyridin-1(2H)-yl) acetic acid) selectively inhibited RelA-DNA binding. We propose that A55 is a seed compound for RelA-specific inhibition and could be used in clinical applications.
Subject(s)
Drug Evaluation, Preclinical/methods , Fluorescence Resonance Energy Transfer/methods , Transcription Factor RelA/antagonists & inhibitors , Transcription Factor RelA/chemistry , Binding Sites , Cell Line, Tumor , DNA/chemistry , DNA/metabolism , High-Throughput Screening Assays , Humans , Models, Molecular , Molecular Conformation , Molecular Docking Simulation , Molecular Dynamics Simulation , Molecular Structure , Protein Binding , Structure-Activity RelationshipABSTRACT
M1 macrophages serve one edge as proinflammatory and M2 macrophages serve the other edge as an anti-inflammatory macrophage. It appears that a related "switch" in macrophage morphology may also happen in the course of atherosclerosis, which has not yet been elucidated. An atherogenic diet (AD) was given to rats, and induction of macrophage differentiation and the nuclear localization of nuclear factor-kappa B (NFκB) were investigated by Western blot and immunofluorescence. Chemokines were analyzed using an antibody array with 32 target proteins. M2 macrophage transformation was confirmed in diosgenin-treated aorta by immunofluorescence and was validated in vitro using THP-1 cells. MAC387 (macrophage marker) and NFκBp65 (inflammatory hub) were upregulated in oxidatively-modified low-density lipoprotein (OxyLDL) and AD-induced condition. Macrophage differentiation, which induced the formation of inflammatory mediators, was not significantly suppressed by the inhibition of NFκB using dexamethasone. M1 macrophage polarization was identified in OxyLDL-induced monocytes, which are proinflammatory in nature, whereas M2 macrophage polarization was noticed in diosgenin-treated monocytes, which exhibit anti-inflammatory properties. M1-and M2-specific chemokines were analyzed using chemokine antibody array. Furthermore, the expression of proinflammatory macrophage (M1) was noticed in AD-induced aorta and anti-inflammatory macrophage (M2) was observed in diosgenin-treated aorta. This is the first report where, unifying the mechanism of diosgenin as aan nti-atherosclerotic and the expression of M1 and M2 specific chemokines is shown by downregulating NFκB and not by preventing the differentiation of monocyte into a macrophage, but by allowing macrophage to differentiate into M2, which aids in preventing the atherosclerotic progression.
Subject(s)
Aorta/metabolism , Atherosclerosis/metabolism , Cell Polarity , Cytokines/metabolism , Diosgenin/pharmacology , Macrophages/metabolism , Plant Extracts/pharmacology , Transcription Factor RelA/metabolism , Animals , Antigens, CD/genetics , Antigens, CD/metabolism , Antigens, Differentiation, Myelomonocytic/genetics , Antigens, Differentiation, Myelomonocytic/metabolism , Atherosclerosis/drug therapy , Atherosclerosis/prevention & control , CD36 Antigens/genetics , CD36 Antigens/metabolism , Cell Differentiation/drug effects , Cyclooxygenase 2/genetics , Cyclooxygenase 2/metabolism , Dexamethasone/pharmacology , Diet, Atherogenic/adverse effects , Dioscorea/chemistry , Diosgenin/therapeutic use , Humans , Lipoproteins, LDL/pharmacology , Male , Monocytes/metabolism , Plant Extracts/therapeutic use , Rats , Signal Transduction/drug effects , THP-1 Cells , Transcription Factor RelA/antagonists & inhibitors , Transcription Factor RelA/geneticsABSTRACT
BACKGROUND: Melanoma is a lethal cancer. NF-κB has been validated as a molecular target for melanoma treatment. Current therapies for melanoma have limitations. Novel targeted therapeutics are needed. Arnicolide D (Ar-D), a sesquiterpene lactone isolated from the dried whole plant of Centipeda minima (L.) A. Br. et Aschers., has been reported to inhibit NF-κB activity in colorectal cancer cells. PURPOSE: To investigate the anti-melanoma effects of Ar-D in vitro and in vivo; and to determine whether Ar-D inhibits the NF-κB pathway in melanoma cells. METHODS: A B16F10 allograft mouse model and two melanoma cell lines (A375 and B16F10) were used to investigate the anti-melanoma effects of Ar-D in vivo and in vitro. Dacarbazine was used as a positive control. Cell viability was assessed by MTT and crystal violet staining assays. Cell cycle arrest and apoptosis were analyzed by flow cytometry. Protein levels were determined by Immunoblotting. RESULTS: In vivo assays showed that the average tumor weight in Ar-D-treated group (4â¯mg/kg, i.p, 15 days) was reduced by 53.7%, when compared with the control group. In vitro studies demonstrated that Ar-D reduced cell viability, induced G2/M cell cycle arrest and apoptosis, elevated levels of cell cycle regulatory proteins p53 and p21, and lowered levels of G2/M checkpoint proteins Cdc2 and Cyclin B1 in melanoma cells. Mechanistically, Ar-D inhibited the activity of IKKα/ß, the degradation of IκBα, and the phosphorylation and expression of NF-κB p65 in melanoma cells. CONCLUSION: Ar-D has anti-melanoma effects, and inhibition of the IKK/IκBα/NF-κB p65 pathway is involved in the effects.
Subject(s)
Antineoplastic Agents/pharmacology , Apoptosis/drug effects , Lactones/pharmacology , Melanoma/drug therapy , Sesquiterpenes/pharmacology , Signal Transduction/drug effects , Transcription Factor RelA/antagonists & inhibitors , Animals , Cell Cycle Checkpoints/drug effects , Cell Line, Tumor , Cell Survival/drug effects , Disease Models, Animal , Humans , Male , Mice , Mice, Inbred C57BL , Phosphorylation , Transcription Factor RelA/metabolismABSTRACT
Hepatitis and hepatocellular carcinoma are serious human diseases. Here, we examined the in vivo and in vitro inhibitory effect of extracts of Qizhu decoction (a traditional Chinese medicine) on hepatitis caused by diethylnitrosamine or hepatitis B virus and on diethylnitrosamine-induced hepatocellular carcinoma. The results showed that both the aqueous and ethanol extracts (QC and QS, respectively) of Qizhu decoction significantly inhibited hepatic inflammation and liver cancer induced by diethylnitrosamine or hepatitis B virus by suppressing NF-κB signaling and decreasing the levels of TNF-α and IL-1ß. Both QC and QS inhibited the proliferation and migration of primary cancer hepatocytes by reducing cyclin B1, cyclin D1 and N-cadherin expression and increasing E-cadherin expression. QC and QS also promoted the apoptosis of primary cancer hepatocytes by upregulating caspase-3 and downregulating BCL-2 expression. The knockdown of p65 in NF-κB signaling inhibited the ability of QC and QS to significantly reduce the colony formation ability of liver cancer cells. Additionally, QC and QS might significantly inhibit the DNA replication of hepatitis B virus in vivo and in vitro, and we found that corilagin and polydatin were the active compounds of QC and QS. Taken together, our in vitro findings and our results in C57BL/6 mice showed that extracts of Qizhu decoction might inhibit hepatitis and hepatocellular carcinoma by suppressing NF-κB signaling.
Subject(s)
Carcinoma, Hepatocellular/drug therapy , Drugs, Chinese Herbal/therapeutic use , Hepatitis B virus/genetics , Hepatitis B, Chronic/drug therapy , Hepatitis, Animal/drug therapy , Liver Neoplasms/drug therapy , Transcription Factor RelA/metabolism , Animals , Carcinoma, Hepatocellular/chemically induced , Carcinoma, Hepatocellular/pathology , Diethylnitrosamine/pharmacology , Disease Models, Animal , Drugs, Chinese Herbal/pharmacology , Gene Knockdown Techniques , Hep G2 Cells , Hepatitis B, Chronic/virology , Hepatitis, Animal/chemically induced , Humans , Liver Neoplasms/chemically induced , Liver Neoplasms/pathology , Male , Medicine, Chinese Traditional/methods , Mice , Mice, Inbred C57BL , Mice, Transgenic , Signal Transduction/drug effects , Signal Transduction/genetics , Transcription Factor RelA/antagonists & inhibitors , Transcription Factor RelA/genetics , TransfectionABSTRACT
BACKGROUND AND AIM: ß-Arrestins (ß-arrs) are regulators and mediators of G protein-coupled receptor signaling that are functionally involved in inflammation. Nuclear factor-κB p65 (NF-κBp65) activation has been observed early in the onset of pancreatitis. However, the effect of ß-arrs in acute pancreatitis (AP) is unclear. The aim of this study is to investigate whether ß-arrs are involved in AP through activation of NF-κBp65. METHODS: Acute pancreatitis was induced by either caerulein injection or choline-deficient supplemented with ethionine diet (CDE). ß-arr1 wild-type and ß-arr1 knockout mice were used in the experiment. The survival rate was calculated in the CDE model mice. Histological and western blot analyses were performed in the caerulein model. Inflammatory mediators were detected by real-time polymerase chain reaction in the caerulein-induced AP mice. Furthermore, AR42J and PANC-1 cell lines were used to further study the effects of ß-arr1 in caerulein-induced pancreatic cells. RESULTS: ß-Arr1 but not ß-arr2 is significantly downregulated in caerulein-induced AP in mice. Targeted deletion of ß-arr1 notably upregulated expression of the pancreatic inflammatory mediators including tumor necrosis factor α and interleukin 1ß as well as interleukin 6 and aggravated AP in caerulein-induced mice. ß-Arr1 deficiency increased mortality in mice with CDE-induced AP. Further, ß-arr1 deficiency enhanced caerulein-induced phosphorylation of NF-κBp65 both in vivo and in vitro. CONCLUSION: ß-Arr1 alleviates AP via repression of NF-κBp65 activation, and it is a potentially therapeutic target for AP.
Subject(s)
Pancreatitis/genetics , Pancreatitis/metabolism , Transcription Factor RelA/antagonists & inhibitors , Transcription Factor RelA/metabolism , beta-Arrestin 1/genetics , beta-Arrestin 1/metabolism , Acute Disease , Animals , Cell Line, Tumor , Ceruletide , Choline Deficiency/complications , Disease Models, Animal , Down-Regulation , Ethionine , Female , Humans , Interleukin-1beta/metabolism , Interleukin-6/metabolism , Mice, Knockout , Pancreatitis/chemically induced , Pancreatitis/pathology , Phosphorylation , Survival Rate , Tumor Necrosis Factor-alpha/metabolismABSTRACT
BACKGROUND: Artemisia scoparia Waldst and Kit is a famous traditional Chinese medicine widely distributed in Xinjiang, China. Flavonoids extracted from it exhibits inhibitory activities against several influenza virus strains. Despite this fact, the antiviral properties of CST, one of such flavonoids, against the influenza virus has not been reported. Thus, the aim of this study is to investigate the anti-influenza virus efficacy and antiviral mechanism of CST. METHODS: The inhibitory activity of CST against influenza viruses was assessed by using viral titers and performing Western blot, qRT-PCR, and immunofluorescence assays in Madin-Darby canine kidney (MDCK) cells and a human monocytic cell line (THP-1). The mechanism of CST against influenza virus was analyzed by hemagglutination inhibition (HI) assay, neuraminidase (NA) inhibition assay, and Western blot. RESULTS: CST reduced viral titers and influenza A virus (IAV) RNA and protein synthesis in a dose-dependent manner. Mechanistically, CST had no inhibitory effect on the attachment and release processes of the viral life cycle, as indicated by the HI and NA assays. Conversely, the CST-mediated inhibition of IAV is possibly linked to the inactivation of the NF-κB/p65 signal pathway. CST also suppressed the activation of JNK MAPK and P38 MAPK in vitro. In line with NF-κB/p65 inhibition, the expression levels of proinflammatory cytokines (TNF-α, IL-1ß, IL-8, and IL-10) and the inflammation-related protein COX-2 were downregulated by CST. CONCLUSIONS: CST inhibited IAV replication by downregulating the NF-κB signal transduction pathway. CST may be a potential agent or supplement against IAV infection.
Subject(s)
Antiviral Agents/pharmacology , Artemisia/chemistry , Flavones/pharmacology , Influenza A Virus, H1N1 Subtype/drug effects , Influenza A Virus, H3N2 Subtype/drug effects , Transcription Factor RelA/genetics , Virus Replication/drug effects , Animals , Antiviral Agents/isolation & purification , Cyclooxygenase 2/genetics , Cyclooxygenase 2/metabolism , Dogs , Dose-Response Relationship, Drug , Flavones/isolation & purification , Gene Expression Regulation , Hemagglutination Inhibition Tests , Host-Pathogen Interactions , Humans , Influenza A Virus, H1N1 Subtype/genetics , Influenza A Virus, H1N1 Subtype/growth & development , Influenza A Virus, H1N1 Subtype/metabolism , Influenza A Virus, H3N2 Subtype/genetics , Influenza A Virus, H3N2 Subtype/growth & development , Influenza A Virus, H3N2 Subtype/metabolism , Interleukin-10/genetics , Interleukin-10/metabolism , Interleukin-1beta/genetics , Interleukin-1beta/metabolism , Interleukin-8/genetics , Interleukin-8/metabolism , Madin Darby Canine Kidney Cells , Neuraminidase/antagonists & inhibitors , Neuraminidase/genetics , Neuraminidase/metabolism , Plant Extracts/chemistry , Signal Transduction , THP-1 Cells , Transcription Factor RelA/antagonists & inhibitors , Transcription Factor RelA/metabolism , Tumor Necrosis Factor-alpha/genetics , Tumor Necrosis Factor-alpha/metabolism , Viral Load/drug effects , Viral Proteins/antagonists & inhibitors , Viral Proteins/genetics , Viral Proteins/metabolismABSTRACT
Oxyresveratrol and its glycoside are important natural active materials. As an effective tyrosine kinase inhibitor, oxyresveratrol may prevent herpes virus infection, inflammation and oxidative stress, as well as protect nerves. In addition, it is known to inhibit cell apoptosis following cerebral ischemia. In recent years, oxyresveratrol and its glycoside have been widely investigated, and their useful biological activities have been explored, indicating that they may be worthy of further comprehensive research. The aim of the present study was to evaluate the photoprotective effects of oxyresveratrol and its ability to abrogate inflammation and oxidative stress in a rat model of spinal cord injury (SCI). The authors identified that oxyresveratrol significantly reversed the SCIinduced inhibition of Basso, Beattie, and Bresnahan scores, inhibited the SCImediated increase in spinal cord water content, significantly suppressed SCIinduced nuclear factorκB/p65, tumor necrosis factorα, interleukin (IL)1ß and IL6 activities and reversed the malondialdehyde, superoxide dismutase, glutathione (GSH) and GSH peroxidase activities in SCI rats. SCIinduced granulocytemacrophage colonystimulating factor (GMCSF), inducible nitric oxide synthase (iNOS) and cyclooxygenase2 (COX2) protein expression was significantly suppressed by oxyresveratrol, and SCImediated inhibition of nuclear factor (erythroidderived 2)like 2 (Nrf2) protein expression was significantly increased by oxyresveratrol. In conclusion, these results suggest that the effects of oxyresveratrol restores SCI, and abrogates inflammation and oxidative stress in rat model of SCI via the GMCSF, iNOS, COX2 and Nrf2 signaling pathway.
Subject(s)
Antioxidants/pharmacology , Locomotion/drug effects , Neuroprotective Agents/pharmacology , Plant Extracts/pharmacology , Spinal Cord Injuries/drug therapy , Spinal Cord/drug effects , Stilbenes/pharmacology , Animals , Disease Models, Animal , Drug Administration Schedule , Female , Gene Expression Regulation , Glutathione Peroxidase/genetics , Glutathione Peroxidase/metabolism , Granulocyte-Macrophage Colony-Stimulating Factor/genetics , Granulocyte-Macrophage Colony-Stimulating Factor/metabolism , Inflammation , Injections, Intraperitoneal , Interleukin-1beta/antagonists & inhibitors , Interleukin-1beta/genetics , Interleukin-1beta/metabolism , Interleukin-6/antagonists & inhibitors , Interleukin-6/genetics , Interleukin-6/metabolism , Malondialdehyde/metabolism , Nitric Oxide Synthase Type II/genetics , Nitric Oxide Synthase Type II/metabolism , Oxidative Stress , Rats , Rats, Sprague-Dawley , Spinal Cord/metabolism , Spinal Cord/pathology , Spinal Cord Injuries/genetics , Spinal Cord Injuries/metabolism , Spinal Cord Injuries/pathology , Superoxide Dismutase/genetics , Superoxide Dismutase/metabolism , Transcription Factor RelA/antagonists & inhibitors , Transcription Factor RelA/genetics , Transcription Factor RelA/metabolism , Tumor Necrosis Factor-alpha/antagonists & inhibitors , Tumor Necrosis Factor-alpha/genetics , Tumor Necrosis Factor-alpha/metabolismABSTRACT
To investigate the mechanism and effect of Psoralea corylifolia(PC) in the treatment of NAFLD in juvenal mice. The NAFLD model in juvenal mice was established by feeding high-fat diet. Then PC herbal granules (at low and high dose) were administered for 5 weeks. Blood glucose (FBG, PG-1 h/2 h), blood lipid (TC, TG, HDL-C, LDL-C), fasting insulin, liver function (ALT, AST) were examined. HOMA-IR was calculated. Hepatic histological changes were observed. The content of TG, inflammatory factor (TNF-α, IL-8) and protein expressions of CD44, NF-κB p65, p-NF-κB p65 in hepatic tissues were determined. The ratio of p-NF-κB p65 to NF-κB p65 (p-p65/p65) was calculated. The result showed that compared with the model group, both PC treatment groups showed reduction in hepatic steatosis, inflammatory cell infiltration and fibroplasia in portal area. HOMA-IR, ALT, AST, FBG, PG-2 h, TC, TG, LDL-C concentrations and hepatic TG content were also significantly decreased, with the reduction of TNF-α, IL-8 contents, CD44 expression and p-p65/p65 ratio in hepatic tissues (P<0.01). High-dose PC group had a better effect than low-dose group (P<0.01, P<0.05). In conclusion, PC is effective in treating hepatic injury, glucolipid metabolism disturbances and fibrosis in juvenal NAFLD mice. The mechanism may be related to inhibition of inflammation and down-regulation of the activation of hepatic NF-κB.
Subject(s)
Drugs, Chinese Herbal/pharmacology , Non-alcoholic Fatty Liver Disease/drug therapy , Psoralea/chemistry , Transcription Factor RelA/antagonists & inhibitors , Animals , Diet, High-Fat , Interleukin-8/metabolism , Liver/drug effects , Mice , Tumor Necrosis Factor-alpha/metabolismABSTRACT
Viral infections may predispose the airways to secondary bacterial infections that can lead to unfavorable progression of principally self-limiting illnesses. Such complicated respiratory infections include pneumonia, bronchitis, sinusitis, acute otitis media, and sepsis, which cause high morbidity and lethality. Some of the pathogenic consequences of viral infections, like the expression of bacterial adhesion receptors and the disturbance of physical barrier integrity due to inflammation, may create permissive conditions for co-infections. Influenza virus A (H3N2) is a major pathogen that causes secondary bacterial infections and inflammation that lead to pneumonia. The herbal medicine Echinacea purpurea, on the other hand, has been widely used to prevent and treat viral respiratory infections, and recent clinical data suggest that it may prevent secondary infection complications as well. We investigated the role of standardized E. purpurea (Echinaforce® extract or EF) on H3N2-induced adhesion of live nontypeable Haemophilus influenzae (NTHi) and Staphylococcus aureus, along with the expression of bacterial receptors, intracellular adhesion molecule-1 (ICAM-1), fibronectin, and platelet activating factor receptor (PAFr), by BEAS-2B cells. Inflammatory processes were investigated by determining the cellular expression of IL-6 and IL-8 and the involvement of Toll-like receptor (TLR-4) and NFκB p65. We found that influenza virus A infection increased the adhesion of H. influenzae and S. aureus to bronchial epithelial cells via upregulated expression of the ICAM-1 receptor and, to some extent, of fibronectin and PAFr. Echinaforce (EF) significantly reduced the expression of ICAM-1, fibronectin, and PAFr and consequently the adhesion of both bacterial strains. EF also effectively prevented the super-expression of inflammatory cytokines by suppressing the expression of NFκB and possibly TLR-4. These results indicate that E. purpurea has the potential to reduce the risk of respiratory complications by preventing virus-induced bacterial adhesion and through the inhibition of inflammation super-stimulation (cytokine storms).
Subject(s)
Anti-Infective Agents/pharmacology , Echinacea/chemistry , Epithelial Cells/drug effects , Superinfection/prevention & control , Toll-Like Receptor 4/antagonists & inhibitors , Transcription Factor RelA/antagonists & inhibitors , Cell Line , Coinfection , Epithelial Cells/cytology , Epithelial Cells/microbiology , Epithelial Cells/virology , Fibronectins/genetics , Fibronectins/immunology , Gene Expression Regulation , Haemophilus influenzae/drug effects , Haemophilus influenzae/growth & development , Haemophilus influenzae/pathogenicity , Host-Pathogen Interactions , Humans , Influenza A Virus, H3N2 Subtype/drug effects , Influenza A Virus, H3N2 Subtype/growth & development , Influenza A Virus, H3N2 Subtype/pathogenicity , Intercellular Adhesion Molecule-1/genetics , Intercellular Adhesion Molecule-1/immunology , Interleukin-6/genetics , Interleukin-6/immunology , Interleukin-8/genetics , Interleukin-8/immunology , Lung/cytology , Lung/drug effects , Lung/microbiology , Lung/virology , Plant Extracts/pharmacology , Platelet Membrane Glycoproteins/genetics , Platelet Membrane Glycoproteins/immunology , Receptors, G-Protein-Coupled/genetics , Receptors, G-Protein-Coupled/immunology , Signal Transduction , Staphylococcus aureus/drug effects , Staphylococcus aureus/growth & development , Staphylococcus aureus/pathogenicity , Superinfection/microbiology , Superinfection/virology , Toll-Like Receptor 4/genetics , Toll-Like Receptor 4/immunology , Transcription Factor RelA/genetics , Transcription Factor RelA/immunologyABSTRACT
The human intracellular enzyme AKR1B1 belongs to the aldo-keto reductase superfamily. The AKR1B1-catalyzed reduction of aldehydes is part of the intracellular inflammatory pathway leading to the activation of NF-κB and the expression of pro-inflammatory genes. The present study is aimed at determining the inhibition of AKR1B1 brought about by an extract of artichoke leaves (bracts), and the effects of this extract and three participating compounds on the expression of AKR1B1, COX-2, and MMP-2 proteins in THP-1 cells. It seeks to identify the ability of the test substances to modulate the lipopolysaccharide (LPS)-induced activation of NF-κB in cells and the intracellular oxidant effect of test substances after incubation with LPS. Low concentrations of the extract inhibit the enzyme AKR1B1. After stimulation by LPS, the extract attenuated the activity of NF-κB in THP-1 cells, but no changes in the expression of AKR1B1 were recorded. The extract diminished the expression of the inflammation-related enzymes COX-2 and MMP-2, probably by inhibiting the activity of NF-κB. The extract significantly diminished the intracellular reactive oxygen species after a brief LPS incubation, which may also have reduced intracellular inflammation. The diminished activity of NF-κB in the cells could be linked to the inhibition of the activity of AKR1B1. Copyright © 2017 John Wiley & Sons, Ltd.
Subject(s)
Aldehyde Reductase/antagonists & inhibitors , Cynara scolymus/chemistry , Leukemia/pathology , Plant Extracts/pharmacology , Plant Leaves/chemistry , Aldehyde Reductase/genetics , Aldehyde Reductase/metabolism , Animals , Cells, Cultured , Cyclooxygenase 2/metabolism , Down-Regulation/drug effects , Gene Expression Regulation/drug effects , Humans , Inflammation/chemically induced , Inflammation/genetics , Inflammation/metabolism , Leukemia/genetics , Leukemia/metabolism , Lipopolysaccharides , Matrix Metalloproteinase 2/genetics , Matrix Metalloproteinase 2/metabolism , Rats , Signal Transduction/drug effects , Signal Transduction/genetics , Transcription Factor RelA/antagonists & inhibitors , Transcription Factor RelA/metabolismABSTRACT
NFκB-mediated inflammation contributes to liver injury during nonalcoholic steatohepatitis (NASH). We hypothesized that antiinflammatory activities of green tea extract (GTE) during NASH would lower tumor necrosis factor receptor-1 (TNFR1)- and Toll-like receptor-4 (TLR4)-mediated NFκB activation. Male C57BL6/J mice (6 weeks old) were fed a low-fat (LF) or high-fat (HF) diet for 12 weeks to induce NASH. They were then randomized to continue on these diets supplemented with 0 or 2% GTE (n=10/group) for an additional 8 weeks prior to evaluating NASH, NFκB inflammation and TNFR1 and TLR4 receptor complexes and their respective ligands, TNFα and endotoxin. HF feeding increased (P<.05) serum alanine aminotransferase (ALT) activity and histological evidence of NASH compared with LF controls. HF-mediated increases in NFκB p65 phosphorylation were also accompanied by increased serum TNFα and endotoxin concentrations, mRNA expression of hepatic TNFR1 and TLR4 and MyD88 protein levels. GTE in LF mice had no effect (P>.05) on liver histology or inflammatory responses. However, GTE in HF mice decreased biochemical and histological parameters of NASH and lowered hepatic p65 phosphorylation in association with decreased serum TNFα, mRNA expression of TNFR1 and TLR4 and MyD88 protein. GTE in HF-fed mice also lowered serum endotoxin and up-regulated mRNA expression of duodenal occludin and zonula occluden-1 and ileal occludin and claudin-1 that were otherwise lowered in expression by HF feeding. These data suggest that dietary GTE treatment reduces hepatic inflammation in NASH by decreasing proinflammatory signaling through TNFR1 and TLR4 that otherwise increases NFκB activation and liver injury.
Subject(s)
Anti-Inflammatory Agents, Non-Steroidal/therapeutic use , Camellia sinensis/chemistry , Dietary Supplements , Non-alcoholic Fatty Liver Disease/diet therapy , Plant Extracts/therapeutic use , Receptors, Tumor Necrosis Factor, Type I/antagonists & inhibitors , Toll-Like Receptor 4/antagonists & inhibitors , Animals , Anti-Inflammatory Agents, Non-Steroidal/adverse effects , Diet, High-Fat/adverse effects , Food Handling , Gene Expression Regulation , Ligands , Liver/immunology , Liver/metabolism , Male , Mice, Inbred C57BL , Non-alcoholic Fatty Liver Disease/etiology , Non-alcoholic Fatty Liver Disease/immunology , Non-alcoholic Fatty Liver Disease/metabolism , Oxidation-Reduction , Phosphorylation , Plant Extracts/adverse effects , Plant Leaves/chemistry , Protein Processing, Post-Translational , Random Allocation , Receptors, Tumor Necrosis Factor, Type I/genetics , Receptors, Tumor Necrosis Factor, Type I/metabolism , Toll-Like Receptor 4/genetics , Toll-Like Receptor 4/metabolism , Transcription Factor RelA/antagonists & inhibitors , Transcription Factor RelA/metabolismABSTRACT
Zerumbone ameliorates retinal damage by blocking advanced glycation end products and their receptor system in streptozotocin-diabetic rats. Because of the multiple factors involved in diabetic retinopathy (DR) etiology, the mechanisms of zerumbone that are mainly responsible for its ameliorative effect on DR need to be further clarified. In the present study, zerumbone (20 mg or 40 mg/kg) or fenofibric acid (100 mg/kg) was orally administered to diabetic rats by intragastric gavage once daily for three consecutive months. Zerumbone displayed similar characteristics to fenofibric acid in reducing retinal vascular permeability and leukostasis in diabetic rats. Fundus photographs showed that large retinal vessel diameters were decreased in zerumbone-treated diabetic rats. Zerumbone not only down-regulated the gene expression of retinal angiogenic parameters, but also reduced the expression of inflammatory cytokines and chemokines in the retina of diabetic rats. Moreover, zerumbone reduced the p38 MAPK phosphorylation and abrogated the nuclear translocation of NF-κB p65 in the retina of diabetic rats. In conclusion, treatment of diabetic rats with zerumbone attenuates the severity of retinal inflammation and angiogenesis, via inhibition of p38 MAPK and NF-κB signaling pathways. These benefits of zerumbone for DR appear to be linked to its antihyperglycemic and antihyperlipidemic effects.
Subject(s)
Diabetes Mellitus, Experimental/pathology , Diabetic Retinopathy/drug therapy , Diabetic Retinopathy/prevention & control , Microvessels/drug effects , Retinal Vessels/drug effects , Sesquiterpenes/therapeutic use , Transcription Factor RelA/antagonists & inhibitors , p38 Mitogen-Activated Protein Kinases/antagonists & inhibitors , Animals , Capillary Permeability/drug effects , Fenofibrate/analogs & derivatives , Fenofibrate/therapeutic use , Glycation End Products, Advanced/antagonists & inhibitors , Male , Microvessels/injuries , Rats , Rats, Wistar , Retina/drug effects , Retinal Vessels/injuries , Signal Transduction/drug effects , Streptozocin , Transcription Factor RelA/metabolism , p38 Mitogen-Activated Protein Kinases/metabolismABSTRACT
Gardeniae Fructus (GF, Zhi Zi in China), a fruit of Gardenia jasminoides Ellis, has been used in traditional medicine to reduce inflammation and headache and to treat hepatic disorders, hypertension, and icterus. In recent studies, extract of raw or stir-baked GF was shown to have pharmacological activities for viral infection, thrombosis, hyperlipidemia, convulsion, inflammation, oxidative stress, and others. In addition, baked GF extract suppressed the proteolytic activities and altered the cellular morphology of tumor cells. However, the effects of ethanol extract of baked GF (EBGF) on the metastatic and angiogenic capacities of malignant tumor cells and its detailed mechanism of action have not been reported. In this study, we found that EBGF significantly inhibited phorbol 12-myristate 13-acetate (PMA)-induced MMP-9 and -13 and uPA expression via suppression of PMA-induced nuclear translocation of NF-κBp65. Metastatic potential, including migration, invasion, and colonization, was substantially reduced by EBGF with no cytotoxicity. In addition, EBGF attenuated tumor-induced angiogenesis, including microvessel sprouting, migration of endothelial cells (ECs), and tube formation of ECs, by inhibiting the release of pro-angiogenic factors from tumor cells. In C57BL/6 mice, we observed that daily administration of EBGF at 50 and 100 mg/kg suppressed metastatic colonization of B16F10 melanoma cells in the lungs. Furthermore, EBGF administration did not cause adverse effects, suggesting that EBGF is safe and may be a potential herbal medicine capable of controlling metastatic malignant cancers.
Subject(s)
Angiogenesis Inhibitors/pharmacology , Antineoplastic Agents/pharmacology , Gardenia/metabolism , Hypoxia-Inducible Factor 1, alpha Subunit/antagonists & inhibitors , Melanoma, Experimental/pathology , Plant Extracts/pharmacology , Transcription Factor RelA/antagonists & inhibitors , Animals , Cell Line, Tumor , Cell Movement/drug effects , Human Umbilical Vein Endothelial Cells , Lung Neoplasms/drug therapy , Lung Neoplasms/secondary , Matrix Metalloproteinase 13/biosynthesis , Matrix Metalloproteinase 9/biosynthesis , Melanoma, Experimental/drug therapy , Mice , Mice, Inbred C57BL , Neoplasm Metastasis/pathology , Neovascularization, Pathologic/prevention & control , Tetradecanoylphorbol Acetate/toxicityABSTRACT
Dihydromyricetin (DMY) a flavonoid derived from medicinal plant Ampelopsis grossedentata, possesses anti-oxidative and anti-inflammatory effects in vitro, however, the in vivo anti-inflammatory action of DMY remains unknown. In the current study, carrageenan-induced paw edema in rat, an acute inflammation model, and RAW264.7 macrophages activated by LPS were employed to evaluate the anti-inflammatory potency of DMY in vivo and in vitro. Results showed that DMY significantly attenuated rat paw edema induced by carrageenan. Also, DMY markedly inhibited NO secretion, iNOS, and COX-2 protein expression, as well as p65 phosphorylation via suppression of IKKß activity and IKKα/ß phosphorylation in RAW264.7 cells. And using high resolution Atomic Force Microscope (AFM), we also proved that DMY prevented morphological change and membrane alterations of RAW 264.7 macrophages caused by LPS stimulation. As activation of macrophages is one of major factors in carrageenan-induced paw edema of rats, the anti-inflammatory action of DMY is suggested to be closely associated with suppression of macrophage activation. These findings indicate that DMY is valuable of being further investigated as a candidate new agent for treating inflammatory conditions, and suggest that AFM could be a powerful nanotool for anti-inflammatory investigations. SCANNING 38:901-912, 2016. © 2016 Wiley Periodicals, Inc.
Subject(s)
Anti-Inflammatory Agents/pharmacology , Flavonols/pharmacology , I-kappa B Kinase/antagonists & inhibitors , Macrophages/drug effects , Animals , Female , HEK293 Cells , Humans , Macrophages/immunology , Mice , Microscopy, Atomic Force , Nitric Oxide/biosynthesis , Nitric Oxide Synthase Type II/genetics , RAW 264.7 Cells , Rats , Rats, Wistar , Transcription Factor RelA/antagonists & inhibitorsABSTRACT
Aberrant activation of oncogenic signaling pathways plays a pivotal role in tumor initiation and progression. The purpose of the present study was to investigate the chemopreventive and therapeutic efficacy of blueberry in the hamster buccal pouch (HBP) carcinogenesis model based on its ability to target TGF-ß, PI3K/Akt, MAPK and NF-κB signaling and its impact on invasion and angiogenesis. Squamous cell carcinomas were induced in the HBP by 7,12-dimethylbenz[a]anthracene (DMBA). The effect of blueberry on the oncogenic signaling pathways and downstream events was analyzed by quantitative real-time PCR and immunoblotting. Experiments with the ECV304 cell line were performed to explore the mechanism by which blueberry regulates angiogenesis. Blueberry supplementation inhibited the development and progression of HBP carcinomas by abrogating TGF-ß and PI3K/Akt pathways. Although blueberry failed to influence MAPK, it suppressed NF-κB activation by preventing nuclear translocation of NF-κB p65. Blueberry also modulated the expression of the oncomiR miR-21 and the tumor suppressor let-7. Collectively, these changes induced a shift to an anti-invasive and anti-angiogenic phenotype as evidenced by downregulating matrix metalloproteinases and vascular endothelial growth factor. Blueberry also inhibited angiogenesis in ECV304 cells by suppressing migration and tube formation. The results of the present study suggest that targeting oncogenic signaling pathways that influence acquisition of cancer hallmarks is an effective strategy for chemointervention. Identification of modulatory effects on phosphorylation, intracellular localization of oncogenic transcription factors and microRNAs unraveled by the present study as key mechanisms of action of blueberry is critical from a therapeutic perspective.
Subject(s)
Benz(a)Anthracenes/toxicity , Blueberry Plants/chemistry , Dietary Supplements , Fruit/chemistry , Mouth Neoplasms/prevention & control , Neoplasms, Squamous Cell/prevention & control , Neovascularization, Pathologic/prevention & control , Signal Transduction , 9,10-Dimethyl-1,2-benzanthracene , Active Transport, Cell Nucleus/drug effects , Animals , Carcinogens/toxicity , Cell Line, Transformed , Freeze Drying , Gene Expression Regulation, Neoplastic/drug effects , Humans , Male , Mesocricetus , Mouth Neoplasms/blood supply , Mouth Neoplasms/metabolism , Mouth Neoplasms/pathology , Neoplasm Invasiveness , Neoplasm Proteins/antagonists & inhibitors , Neoplasm Proteins/genetics , Neoplasm Proteins/metabolism , Neoplasms, Squamous Cell/blood supply , Neoplasms, Squamous Cell/metabolism , Neoplasms, Squamous Cell/pathology , Neovascularization, Pathologic/chemically induced , Neovascularization, Pathologic/metabolism , Neovascularization, Pathologic/pathology , Random Allocation , Signal Transduction/drug effects , Transcription Factor RelA/antagonists & inhibitors , Transcription Factor RelA/metabolism , Transforming Growth Factor beta/antagonists & inhibitors , Transforming Growth Factor beta/metabolismABSTRACT
Lipopolysaccharide (LPS) is the major structural component of Gram-negative bacteria cell wall and a highly pro-inflammatory toxin. Naringenin is found in Citrus fruits and exhibits antioxidant and anti-inflammatory properties through inhibition of NF-κB activation but its effects in LPS-induced inflammatory pain and leukocyte recruitment were not investigated yet. We investigated the effects of naringenin in mechanical hyperalgesia, thermal hyperalgesia and leukocyte recruitment induced by intraplantar injection of LPS in mice. We found that naringenin reduced hyperalgesia to mechanical and thermal stimuli, myeloperoxidase (MPO, a neutrophil and macrophage marker) and N-acetyl-ß-D-glucosaminidase (NAG, a macrophage marker) activities, oxidative stress and cytokine (TNF-α, IL-1ß, IL-6, and IL-12) production in the paw skin. In the peritoneal cavity, naringenin reduced neutrophil and mononuclear cell recruitment, and abrogated MPO and NAG activity, cytokine and superoxide anion production, and lipid peroxidation. In vitro, pre-treatment with naringenin inhibited superoxide anion and cytokine (TNF-α, IL-1ß, IL-6, and IL-12) production by LPS-stimulated RAW 264.7 macrophages. Finally, we demonstrated that naringenin inhibited NF-κB activation in vitro and in vivo. Therefore, naringenin is a promising compound to treat LPS-induced inflammatory pain and leukocyte recruitment.
Subject(s)
Anti-Inflammatory Agents, Non-Steroidal/therapeutic use , Dietary Supplements , Flavanones/therapeutic use , Hyperalgesia/prevention & control , Leukocytes, Mononuclear/immunology , Neutrophils/immunology , Transcription Factor RelA/antagonists & inhibitors , Animals , Anti-Inflammatory Agents, Non-Steroidal/metabolism , Antioxidants/therapeutic use , Behavior, Animal , Biomarkers/metabolism , Flavanones/metabolism , Hot Temperature/adverse effects , Hyperalgesia/immunology , Hyperalgesia/metabolism , Leukocytes, Mononuclear/drug effects , Leukocytes, Mononuclear/metabolism , Lipopolysaccharides/toxicity , Macrophage Activation/drug effects , Macrophages/drug effects , Macrophages/immunology , Macrophages/metabolism , Male , Mice , Neutrophil Infiltration/drug effects , Neutrophils/drug effects , Neutrophils/metabolism , Oxidative Stress/drug effects , RAW 264.7 Cells , Skin/drug effects , Skin/immunology , Skin/metabolism , Stress, Mechanical , Transcription Factor RelA/metabolismABSTRACT
The novel compounds NSC745885 and NSC757963 developed at our laboratory were tested against a panel of 60 cancer cell lines at the National Cancer Institute, USA, and a panel of 39 cancer cell lines at the Japanese Foundation of Cancer Research. Both compounds demonstrated selective unique multi-log differential patterns of activity, with GI50 values in the sub-micro molar range against cancer cells rather than normal cardiac cells. NSC757963 showed high selectivity towards the leukemia subpanel. Activities of both compounds strongly correlated to expression of NFKB1 and CSNK2B genes, implying that they may inhibit the NF-κB pathway. Immunocytochemical microscopy of OVCAR-3 cells showed clear cytosolic accumulation of the NF-κB p65 subunit following treatment. Western blotting showed dose dependent inhibition of the nuclear expression of the NF-κB p65 subunit with subsequent accumulation in the cytosol following treatment. Docking experiments showed binding of both compounds to the NF-κB activator IKKß subunit preventing its translocation to the nucleus. Collectively, these results confirm the ability of our compounds to inhibit the constitutively active NF-κB pathway of OVCAR-3 cells. Furthermore, COMPARE analysis indicated that the activity of NSC757963 is similar to the antituberculosis agent rifamycin SV, this was confirmed by testing the antimycobacterial activity of NSC757963 against Mycobacterium tuberculosis, results revealed potent activity suitable for use in clinical practice. Molecular properties and Lipinski's parameters predicted acceptable bioavailability properties with no indication of mutagenicity, tumorigenicity, irritability and reproductive effects. Oral absorption experiments using the human Caco-2 model showed high intestinal absorption of NSC745885 by passive transport mechanism with no intestinal efflux or active transport mechanisms. The unique molecular characterization as well as the illustrated anticancer spectra of activity and bioavailability properties warrant further development of our compounds and present a foundation brick in the pre-clinical investigations to implement such compounds in clinical practice.
Subject(s)
Antineoplastic Agents/pharmacology , Antitubercular Agents/pharmacology , Gene Expression Regulation, Neoplastic , Thiadiazoles/pharmacology , Transcription Factor RelA/antagonists & inhibitors , Antineoplastic Agents/chemical synthesis , Antitubercular Agents/chemical synthesis , Biological Availability , Cell Line, Tumor , Drug Evaluation, Preclinical , Gene Expression Profiling , Humans , I-kappa B Kinase/antagonists & inhibitors , I-kappa B Kinase/genetics , I-kappa B Kinase/metabolism , Intestinal Absorption/drug effects , Microbial Sensitivity Tests , Models, Biological , Molecular Docking Simulation , Mycobacterium tuberculosis/drug effects , Mycobacterium tuberculosis/growth & development , Signal Transduction , Thiadiazoles/chemical synthesis , Transcription Factor RelA/genetics , Transcription Factor RelA/metabolismABSTRACT
OBJECTIVES: Although kaffir lime has been reported to exhibit antioxidant and antileukemic activity, little is known about the antimicrobial effect of kaffir lime extract. Because Streptococcus mutans has been known to cause biofilm formation, it has been considered the most important causative pathogen of dental caries. Thus, the effective control of its effects on the oral biofilm is the key to the prevention of dental caries. The aims of the present study were to investigate the effect of kaffir lime leaves extract on biofilm formation and its antibacterial activity on S. mutans. METHODS: We examined the effect of kaffir lime leaves extract on growth and biofilm formation of S. mutans. For the investigation we used a kaffir lime extract with high phenolic content. The minimum inhibitory concentration of the extract was determined by broth microdilution assay. The inhibitory effect of the test substances on biofilm formation was also investigated by biofilm formation assay and qRT-PCR of biofilm formation-associated genes. RESULTS: Kaffir lime leaves extract inhibits the growth of S. mutans, corresponding to the activity of an antibiotic, ampicillin. Formation of biofilm by S. mutans was also inhibited by the extract. These results were confirmed by the down-regulation of genes associated with the biofilm formation. CONCLUSIONS: The findings highlight the ability of kaffir lime leaves extract to inhibit S. mutans activity, which may be beneficial in the prevention of biofilm formation on dental surface, reducing dental plaque and decreasing the chance of dental carries.
Subject(s)
Biofilms/drug effects , Citrus/chemistry , Plant Extracts/pharmacology , Plant Leaves/chemistry , Streptococcus mutans/drug effects , Anti-Bacterial Agents/pharmacology , Bacterial Proteins/antagonists & inhibitors , Bacterial Proteins/genetics , Bacterial Proteins/metabolism , Dental Caries/drug therapy , Down-Regulation , Microbial Sensitivity Tests , Phenols/analysis , Phenols/pharmacology , RNA, Bacterial/genetics , Transcription Factor RelA/antagonists & inhibitors , Transcription Factor RelA/genetics , Transcription Factor RelA/metabolismABSTRACT
We recently reported that enzyme-treated asparagus extract (ETAS) attenuates hydrogen peroxide (H(2)0(2))-stimulated matrix metalloproteinase-9 expression in skin fibroblast L929 cells. To further elucidate the anti-aging effects of ETAS on skin, we examined whether ETAS has preventive effects on H202-induced pro-inflammatory responses of skin fibroblasts. H(2)0(2) induced Ser536 phosphorylation and nuclear accumulation of nuclear factor-κB (NF-κB) p65, and increased the mRNA levels .of interleukin-12α (IL-12α)-and inducible nitric oxide synthase (iNOS) in L929 cells. Pretreatment of the cells with JSH-23, an inhibitor of NF-κB nuclear translocation, abolished the H(2)(0(2)-induced expression of IL-12α and iNOS, indicating that the increased transcription is regulated by p65. The H(2)0(2)-stimulated nuclear accumulation of p65 and-induction of IL12a and iNOS mRNA were significantly attenuated after pretreatment with ETAS for 3 h, and these responses were completely abolished when the duration was extended to 24 h. However, ETAS did not affect the H(2)0(2)-stimulated degradation of IκBα and phosphorylation of p65. On the other hand, ETAS treatment for 24 h resulted in decreased protein levels of importin-α. These results suggest that ETAS prevents pro-inflammatory responses by suppressing the p65 nuclear translocation in skin fibroblasts induced by H202.