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1.
Braz. j. med. biol. res ; 53(6): e9346, 2020. graf
Article in English | LILACS, ColecionaSUS | ID: biblio-1132516

ABSTRACT

Atherosclerosis (AS) is a common vascular disease, which can cause apoptosis of vascular endothelial cells. Notoginsenoside R1 (NGR1) is considered an anti-AS drug. MicroRNAs (miRNAs) are believed to play a vital role in cell apoptosis and angiogenesis. This study aimed to explore the mechanism of NGR1 for treating AS through miRNAs. Flow cytometry was used to detect the apoptosis rate. The levels of inflammatory cytokines interleukin (IL)-6 and IL-1β were detected using ELISA. Reactive oxygen species (ROS) and malondialdehyde (MDA) levels were measured using corresponding assay kits. Quantitative real-time polymerase chain reaction (qRT-PCR) assay was performed to detect miR-221-3p expression. Dual-luciferase reporter and RNA immunoprecipitation assays were carried out to examine the relationship between miR-221-3p and toll-like receptors 4 (TLR4). Also, western blot analysis was performed to determine the levels of TLR4 and nuclear factor kappa B (NF-κB) signaling pathway-related proteins. Oxidized low-density lipoprotein (ox-LDL) induced human umbilical vein endothelial cells (HUVECs) apoptosis, inflammation, and oxidative stress. NGR1 alleviated the negative effect of ox-LDL through promoting the expression of miR-221-3p in HUVECs. TLR4 was a target of miR-221-3p, and its overexpression could reverse the inhibition effects of miR-221-3p on apoptosis, inflammation, and oxidative stress. NGR1 improved miR-221-3p expression to inhibit the activation of the TLR4/NF-κB pathway in ox-LDL-treated HUVECs. NGR1 decreased ox-LDL-induced HUVECs apoptosis, inflammation, and oxidative stress through increasing miR-221-3p expression, thereby inhibiting the activation of the TLR4/NF-κB pathway. This study of the mechanism of NGR1 provided a more theoretical basis for the treatment of AS.


Subject(s)
Humans , Apoptosis/drug effects , Oxidative Stress/drug effects , Ginsenosides/pharmacology , MicroRNAs/adverse effects , Human Umbilical Vein Endothelial Cells/drug effects , Inflammation/metabolism , Lipoproteins, LDL/metabolism , Enzyme-Linked Immunosorbent Assay , Signal Transduction , Transcriptional Activation , Up-Regulation , Blotting, Western , NF-kappa B/antagonists & inhibitors , Reactive Oxygen Species , MicroRNAs/metabolism , Immunoprecipitation , Toll-Like Receptor 4/antagonists & inhibitors , Human Umbilical Vein Endothelial Cells/metabolism , Real-Time Polymerase Chain Reaction
2.
Acta cir. bras ; 34(2): e201900209, 2019. graf
Article in English | LILACS | ID: biblio-989056

ABSTRACT

Abstract Purpose: To explore the effect of milk fat globule-epidermal growth factor 8 (MFG-E8) on sepsis-induced acute kidney injury (SAKI). Methods: Male C57BL/6 mice were randomized to control, sham, CLP, CLP+PBS, and CLP+rmMFG-E8 groups. SAKI was induced by cecal ligation and puncture (CLP). Recombinant mouse MFG-E8 (rmMFG-E8) (20 μg/kg) or PBS (vehicle) was administered intraperitoneally. Blood, urine and renal tissue were collected at 24 h after CLP. Blood samples were tested for serum kidney injury biomarker and cytokines. Urine samples were collected to detect KIM-1, and NGAL. Real-time PCR was tested for Bax and Bcl-2. TUNEL staining was used to determine renal apoptosis. Western blot was used to detect the expression of Bax, Bcl-2, and proteins in the NF-κB pathway. Results: MFG-E8 alleviated SAKI by decreasing serum Cre, BUN, urine KIM-1 and NGAL and by mitigating renal pathological changes significant (p < 0.05). IL-1β, IL-6, TNF-α were significantly inhibited by MFG-E8 (p < 0.05). Apoptosis induced by SAKI was markedly suppressed by MFG-E8. Finally, MFG-E8 attenuated the activation of the NF-��B signaling pathway in SAKI. Conclusion: MFG-E8 has beneficial effects on SAKI, which may be achieved by inhibiting the NF-κB pathway.


Subject(s)
Animals , Male , Rats , NF-kappa B/antagonists & inhibitors , Sepsis/complications , Protective Agents/therapeutic use , Acute Kidney Injury/prevention & control , Milk Proteins/therapeutic use , Antigens, Surface/therapeutic use , Signal Transduction/drug effects , Acute Kidney Injury/etiology , Mice, Inbred C57BL
3.
Biol. Res ; 51: 9, 2018. graf
Article in English | LILACS | ID: biblio-950895

ABSTRACT

BACKGROUND: Diabetic nephropathy (DN) is the leading cause of end-stage renal failure, contributing to severe morbidity and mortality in diabetic patients. Berberine (BBR) has been well characterized to exert renoprotective effects in DN progression. However, the action mechanism of BBR in DN remains to be fully understood. METHODS: The DN rat model was generated by intraperitoneal injection of streptozotocin (STZ, 65 mg/kg body weight) while 30 mM high glucose (HG)-treated podocytes were used as an in vitro DN model. The fasting blood glucose level and ratio of kidney weight to body weight were measured after BBR treatment (50, 100, or 200 mg/kg) in STZ-induced DN rats. The renal injury parameters including 24-h urinary protein, blood urea nitrogen and serum creatinine were assessed. qRT-PCR was performed to detect the transcript amounts of inflammatory factors. The concentrations of inflammatory factors were evaluated by ELISA kits. Western blot analysis was conducted to measure the amounts of TLR4/NF-κB-related proteins. The apoptotic rate of podocytes was analyzed by flow cytometry using Annexin V/propidium iodide. RESULTS: Berberine reduced renal injury in STZ-induced DN rat model, as evidenced by the decrease in fasting blood glucose, ratio of kidney weight to body weight, 24-h urinary protein, serum creatinine, and blood urine nitrogen. BBR attenuated the systemic and renal cortex inflammatory response and inhibited TLR4/NF-κB pathway in STZ-induced DN rats and HG-induced podocytes. Also, HG-induced apoptosis of podocytes was lowered by BBR administration. Furthermore, blockade of TLR4/NF-κB pathway by resatorvid (TAK-242) or pyrrolidine dithiocarbamate aggravated the inhibitory effect of BBR on HG-induced inflammatory response and apoptosis in podocytes. CONCLUSIONS: Berberine ameliorated DN through relieving STZ-induced renal injury, inflammatory response, and podocyte HG-induced apoptosis via inactivating TLR4/NF-κB pathway.


Subject(s)
Animals , Male , Rats , Berberine/pharmacology , Signal Transduction/drug effects , NF-kappa B/antagonists & inhibitors , Diabetes Mellitus, Experimental/drug therapy , Diabetic Nephropathies/drug therapy , Toll-Like Receptor 4/antagonists & inhibitors , Rats, Sprague-Dawley
4.
Braz. j. med. biol. res ; 50(4): e5861, 2017. tab, graf
Article in English | LILACS | ID: biblio-839274

ABSTRACT

Myocardial ischemia is a major cause of death and remains a disease with extremely deficient clinical therapies and a major problem worldwide. Cold inducible RNA-binding protein (CIRBP) is reported to be involved in multiple pathological processes, including myocardial ischemia. However, the molecular mechanisms of myocardial ischemia remain elusive. Here, we first overexpressed CIRBP by transfection of pc-CIRBP (pcDNA3.1 containing coding sequenced for CIRBP) and silenced CIRBP by transfection of small interfering RNA targeting CIRBP (siCIRBP). pcDNA3.1 and the negative control of siCIRBP (siNC) were transfected into H9C2 cells to act as controls. We then constructed a cell model of myocardial ischemia through culturing cells in serum-free medium with hypoxia in H9C2 cells. Subsequently, AlamarBlue assay, flow cytometry and western blot analysis were used, respectively, to assess cell viability, reactive oxygen species (ROS) level and apoptosis, and expression levels of IκBα, p65 and Bcl-3. We demonstrated that CIRBP overexpression promoted cell proliferation (P<0.001), inhibited cell apoptosis (P<0.05), reduced ROS level (P<0.001), down-regulated phosphorylated levels of IκBα and p65 (P<0.01 or P<0.001), and up-regulated expression of Bcl-3 (P<0.001) in H9C2 cells with myocardial ischemia. The influence of CIRBP knockdown yielded opposite results. Our study revealed that CIRBP could protect H9C2 cells against myocardial ischemia through inhibition of NF-κB pathway.


Subject(s)
Animals , Rats , Myocardial Ischemia/metabolism , Myocardial Ischemia/prevention & control , NF-kappa B/antagonists & inhibitors , Protective Agents/pharmacology , RNA-Binding Proteins/pharmacology , Apoptosis/drug effects , Blotting, Western , Cell Proliferation/drug effects , Cell Survival , Cells, Cultured , Flow Cytometry , Gene Expression Regulation , NF-kappa B/metabolism , Reactive Oxygen Species/analysis , Reproducibility of Results , Reverse Transcriptase Polymerase Chain Reaction , RNA, Small Interfering , RNA-Binding Proteins/genetics , RNA-Binding Proteins/metabolism , Time Factors , Transfection/methods
5.
Yonsei Medical Journal ; : 647-651, 2016.
Article in English | WPRIM | ID: wpr-21850

ABSTRACT

PURPOSE: In the gastric mucosa of Helicobacter pylori (H. pylori)-infected patients with gastritis or adenocarcinoma, proliferation of gastric epithelial cells is increased. Hyperproliferation is related to induction of oncogenes, such as β-catenin and c-myc. Even though transcription factors NF-κB and AP-1 are activated in H. pylori-infected cells, whether NF-κB or AP-1 regulates the expression of β-catenein or c-myc in H. pylori-infected cells has not been clarified. The present study was undertaken to investigate whether H. pylori-induced activation of NF-κB and AP-1 mediates the expression of oncogenes and hyperproliferation of gastric epithelial cells. MATERIALS AND METHODS: Gastric epithelial AGS cells were transiently transfected with mutant genes for IκBα (MAD3) and c-Jun (TAM67) or treated with a specific NF-κB inhibitor caffeic acid phenethyl ester (CAPE) or a selective AP-1 inhibitor SR-11302 to suppress activation of NF-κB or AP-1, respecively. As reference cells, the control vector pcDNA was transfected to the cells. Wild-type cells or transfected cells were cultured with or without H. pylori. RESULTS: H. pylori induced activation of NF-κB and AP-1, cell proliferation, and expression of oncogenes (β-catenein, c-myc) in AGS cells, which was inhibited by transfection of MAD3 and TAM67. Wild-type cells and the cells transfected with pcDNA showed similar activities of NF-κB and AP-1, proliferation, and oncogene expression regardless of treatment with H. pylori. Both CAPE and SR-11302 inhibited cell proliferation and expression of oncogenes in H. pylori-infected cells. CONCLUSION: H. pylori-induced activation of NF-κB and AP-1 regulates transcription of oncogenes and mediates hyperproliferation in gastric epithelial cells.


Subject(s)
Blotting, Western , Caffeic Acids , Cell Line, Tumor , Cell Proliferation , DNA, Bacterial/analysis , DNA-Binding Proteins/metabolism , Epithelial Cells/metabolism , Gastric Mucosa/metabolism , Gastritis/pathology , Gene Expression Regulation, Bacterial , Helicobacter Infections/metabolism , Helicobacter pylori/pathogenicity , Humans , NF-kappa B/antagonists & inhibitors , Peptide Fragments , Phenylethyl Alcohol/analogs & derivatives , Proto-Oncogene Proteins c-jun , Repressor Proteins , Transcription Factor AP-1/biosynthesis , Transcription Factors/metabolism , beta Catenin/metabolism
7.
Article in English | WPRIM | ID: wpr-51698

ABSTRACT

We evaluated the effect of cobalt chloride (CoCl2) on TNF-alpha and IFN-gamma-induced-inflammation and reactive oxygen species (ROS) in renal tubular epithelial cells (HK-2 cells). We treated HK-2 cells with CoCl2 before the administration of TNF-alpha/IFN-gamma. To regulate hemeoxygenase-1 (HO-1) expression, the cells were treated CoCl2 or HO-1 siRNA. CoCl2 reduced the generation of ROS induced by TNF-alpha/IFN-gamma. TNF-alpha/IFN-gamma-treated-cells showed an increase in the nuclear translocation of phosphorylated NF-kappaBp65 protein, the DNA-binding activity of NF-kappaBp50 and NF-kappaB transcriptional activity and a decrease in IkappaBalpha protein expression. These changes were restored by CoCl2. We noted an intense increase in monocyte chemoattractant protein-1 (MCP-1) and regulated on activation normal T cell expressed and secreted (RANTES) production in TNF-alpha/IFN-gamma-treated cells. We demonstrated that this effect was mediated through NF-kappaB signaling because an NF-kappaB inhibitor significantly reduced MCP-1 and RANTES production. CoCl2 effectively reduced MCP-1 and RANTES production. The expression of HO-1 was increased by CoCl2 and decreased by HO-1 siRNA. However, knockdown of HO-1 by RNA interference did not affect MCP-1 or RANTES production. We suggest that CoCl2 has a protective effect on TNF-alpha/IFN-gamma-induced inflammation through the inhibition of NF-kappaB and ROS in HK-2 cells. However, CoCl2 appears to act in an HO-1-independent manner.


Subject(s)
Cell Line , Chemokine CCL2/metabolism , Chemokine CCL5/metabolism , Cobalt/pharmacology , Epithelial Cells/cytology , Heme Oxygenase-1/antagonists & inhibitors , Humans , Inflammation , Interferon-gamma/pharmacology , Kidney Tubules, Proximal/cytology , NF-kappa B/antagonists & inhibitors , NF-kappa B p50 Subunit/genetics , Oxidative Stress/drug effects , Phosphorylation , Protein Binding , RNA Interference , RNA, Small Interfering/metabolism , Transcription Factor RelA/metabolism , Tumor Necrosis Factor-alpha/pharmacology
8.
Indian J Exp Biol ; 2013 Apr; 51(4): 313-321
Article in English | IMSEAR | ID: sea-147597

ABSTRACT

Osteoarthritis (OA), which is also called degenerative arthritis, is the leading cause of disabilities in the old people. The Chinese traditional herb Epimedium grandiflorum had long been found to attenuate osteoarthritis process, but the detailed mechanism was not clear. To study the mechanisms of E. grandiflorum in the treatment of osteoarthritis, rabbit osteoarthritis model combined with D-galactose was used. After different treatments for 10 weeks, cartilage sections were analyzed by immunohistochemistry for uPA, uPAR and PAI expression level. E. grandiflorum could significantly attenuate OA condition and decrease uPA, uPAR and PAI expression. The extract of E. grandiflorum, icariin also had a similar effect when compared with E. grandiflorum treatment alone. Rabbit chondrocytes were further isolated to be stimulated by TNFα combined with different reagents treatment. Here, icariin treatment significantly reduced nuclear factor kappa B NF-B (P65) activity, decreased uPA expression level and increased IBα protein level. The results indicated that E. grandiflorum and its extract icariin could attenuate OA condition, reduce the expression of uPA and uPAR and increase PAI in experimental rabbit model and this effect may be conducted by suppressing NF-kB activity by increasing IkBα level.


Subject(s)
Animals , Cartilage/metabolism , Chondrocytes/cytology , Disease Models, Animal , Drugs, Chinese Herbal/therapeutic use , Epimedium/metabolism , Female , Flavonoids/therapeutic use , Galactose/metabolism , I-kappa B Proteins/metabolism , Immunohistochemistry , Male , Medicine, Chinese Traditional , NF-kappa B/antagonists & inhibitors , NF-kappa B/metabolism , Osteoarthritis/drug therapy , Plasminogen Activator Inhibitor 1/metabolism , Rabbits , Receptors, Urokinase Plasminogen Activator/metabolism , Tumor Necrosis Factor-alpha/metabolism , Urokinase-Type Plasminogen Activator/metabolism
9.
Article in English | WPRIM | ID: wpr-210924

ABSTRACT

The balance between tissue-type plasminogen activator (t-PA) and plasminogen activator inhibitor type 1 (PAI-1) regulates fibrinolysis. PAI-1 expression increases in atherosclerotic arteries and vascular smooth muscle cells (VSMCs) are one of major constituents of atheroma. We investigated the impact of lysophosphatidylcholine (lysoPC), an active component of oxidized low-density lipoprotein, on the plasminogen activator system of the rat VSMCs. The lysoPC stimulated the protein and gene expressions of PAI-1 but did not affect the protein expression of t-PA. Fibrin overlay zymography revealed that lysoPC increased the activity of PAI-1 in the conditioned media, while concurrently decreasing that of free t-PA. Vitamin E inhibited the lysoPC-induced PAI-1 expression. Further, lysoPC increased the intracellular reactive oxygen species (ROS) formation. Caffeic acid phenethyl ester, an inhibitor of NF-kappaB, blocked this lysoPC effect. Indeed, lysoPC induced the NF-kappaB-mediated transcriptional activity as measured by luciferase reporter assay. In addition, genistein, an inhibitor of protein-tyrosine kinase (PTK), diminished the lysoPC effect, while 7,12-dimethylbenz[a]anthracene, a stimulator of PTK, stimulated PAI-1 production. In conclusion, lysoPC does not affect t-PA expression but induces PAI-1 expression in the VSMC by mediating NF-kappaB and the genistein-sensitive PTK signaling pathways via oxidative stress. Importantly, lysoPC stimulates the enzyme activity of PAI-1 and suppresses that of t-PA.


Subject(s)
Animals , Benz(a)Anthracenes/pharmacology , Caffeic Acids/pharmacology , Cells, Cultured , Genistein/pharmacology , Lipoproteins, LDL/metabolism , Lysophosphatidylcholines/pharmacology , Muscle, Smooth, Vascular/cytology , NF-kappa B/antagonists & inhibitors , Oxidative Stress/drug effects , Phenylethyl Alcohol/analogs & derivatives , Plasminogen Activator Inhibitor 1/agonists , Protein Kinase Inhibitors/pharmacology , Protein-Tyrosine Kinases/antagonists & inhibitors , Rats , Rats, Sprague-Dawley , Reactive Oxygen Species/metabolism , Signal Transduction/drug effects , Tissue Plasminogen Activator/metabolism , Transcription, Genetic/drug effects , Up-Regulation/drug effects , Vitamin E/pharmacology
10.
Article in English | WPRIM | ID: wpr-149763

ABSTRACT

This study was designed to investigate the effects of the prenylated flavonoid kurarinone on TNF-related apoptosis inducing ligand (TRAIL)-induced apoptosis and its underlying mechanism. A low dose of kurarinone had no significant effect on apoptosis, but this compound markedly promoted tumor cell death through elevation of Bid cleavage, cytochrome c release and caspase activation in HeLa cells treated with TRAIL. Caspase inhibitors inhibited kurarinone-mediated cell death, which indicates that the cytotoxic effect of this compound is mediated by caspase-dependent apoptosis. The cytotoxic effect of kurarinone was not associated with expression levels of Bcl-2 and IAP family proteins, such as Bcl-2, Bcl-xL, Bid, Bad, Bax, XIAP, cIAP-1 and cIAP-2. In addition, this compound did not regulate the death-inducing receptors DR4 and DR5. On the other hand, kurarinone significantly inhibited TRAIL-induced IKK activation, IkappaB degradation and nuclear translocation of NF-kappaB, as well as effectively suppressed cellular FLICE-inhibitory protein long form (cFLIPL) expression. The synergistic effects of kurarinone on TRAIL-induced apoptosis were mimicked when kurarinone was replaced by the NF-kappaB inhibitor withaferin A or following siRNA-mediated knockdown of cFLIPL. Moreover, cFLIP overexpression effectively antagonized kurarinone-mediated TRAIL sensitization. These data suggest that kurarinone sensitizes TRAIL-induced tumor cell apoptosis via suppression of NF-kappaB-dependent cFLIP expression, indicating that this compound can be used as an anti-tumor agent in combination with TRAIL.


Subject(s)
Antineoplastic Agents/pharmacology , Apoptosis/drug effects , CASP8 and FADD-Like Apoptosis Regulating Protein/genetics , Caspase 3/metabolism , Caspase 8/metabolism , Drug Synergism , Enzyme Activation/drug effects , Flavonoids/pharmacology , Gene Expression/drug effects , Gene Knockdown Techniques , HeLa Cells , Humans , NF-kappa B/antagonists & inhibitors , Protein Transport/drug effects , RNA, Small Interfering/genetics , Signal Transduction , TNF-Related Apoptosis-Inducing Ligand/physiology , Up-Regulation/drug effects
11.
Article in English | WPRIM | ID: wpr-57563

ABSTRACT

Microglial cells are the resident innate immune cells that sense pathogens and tissue injury in the central nervous system (CNS). Microglial activation is critical for neuroinflammatory responses. The synthetic compound 2-hydroxy-3',5,5'-trimethoxychalcone (DK-139) is a novel chalcone-derived compound. In this study, we investigated the effects of DK-139 on Toll-like receptor 4 (TLR4)-mediated inflammatory responses in BV2 microglial cells. DK-139 inhibited lipopolysaccharide (LPS)-induced TLR4 activity, as determined using a cell-based assay. DK-139 blocked LPS-induced phosphorylation of IkappaB and p65/RelA NF-kappaB, resulting in inhibition of the nuclear translocation and trans-acting activity of NF-kappaB in BV2 microglial cells. We also found that DK-139 reduced the expression of NF-kappaB target genes, such as those for COX-2, iNOS, and IL-1beta, in LPS-stimulated BV2 microglial cells. Interestingly, DK-139 blocked LPS-induced Akt phosphorylation. Inhibition of Akt abrogated LPS-induced phosphorylation of p65/RelA, while overexpression of dominant-active p110CAAX enhanced p65/RelA phosphorylation as well as iNOS and COX2 expression. These results suggest that DK-139 exerts an anti-inflammatory effect on microglial cells by inhibiting the Akt/IkappaB kinase (IKK)/NF-kappaB signaling pathway.


Subject(s)
Animals , Binding Sites , Cell Line , Chalcones/chemistry , Cyclooxygenase 2/metabolism , I-kappa B Kinase/metabolism , Inflammation/drug therapy , Interleukin-1beta/metabolism , Lipopolysaccharides/immunology , Microglia/drug effects , Molecular Dynamics Simulation , NF-kappa B/antagonists & inhibitors , Nitric Oxide Synthase Type II/metabolism , Phosphorylation/drug effects , Protein Binding , Proto-Oncogene Proteins c-akt/antagonists & inhibitors , Rats , Signal Transduction , Toll-Like Receptor 4/antagonists & inhibitors , Transcription Factor RelA/metabolism
13.
Braz. j. infect. dis ; 15(3): 189-194, May-June 2011. ilus
Article in English | LILACS | ID: lil-589946

ABSTRACT

OBJECTIVE: Activation of nuclear factor kappaB by diverse bacteria regulates the secretion of chemokines and cytokines. Staphylococcus aureus (S. aureus)-infected osteoblasts can significantly increase the secretion of interleukin-6 and monocyte chemoattractant protein-1. The aim of this study was to investigate whether S. aureus can activate nuclear factor kappaB in human osteoblasts, and whether the activation of nuclear factor kappaB by S. aureus regulates the secretion of interleukin-6 and monocyte chemoattractant protein-1. METHODS: Immunoblot and electrophoretic mobility shift assay were used to detect the degradation of IκBa and activation of nuclear factor kappaB in human osteoblasts in response to S. aureus, respectively. Enzyme-linked immunosorbent assay was used to measure the secretion of interleukin-6 and monocyte chemoattractant protein-1 in the supernatants. Lastly, carbobenzoxyl-l-leucinyl-l-leucinyl-l-leucinal, an inhibitor of the nuclear factor kappaB, was used to determine if activation of nuclear factor kappaB by S. aureus in human osteoblasts regulates the secretions of interleukin-6 and monocyte chemoattractant protein-1. RESULTS: Our results for the first time demonstrated that S. aureus can induce the degradation of IκBa and activation of nuclear factor kappaB in human osteoblasts in a time and dose-dependent manner. In addition, inhibition of nuclear factor kappaB by carbobenzoxyl-l-leucinyl-l-leucinyl-l-leucinal suppressed the secretion of interleukin-6 and monocyte chemoattractant protein-1 in the supernatants of S. aureus-infected human osteoblasts in a dose-dependent manner. CONCLUSION: These findings suggest that S. aureus can activate nuclear factor kappaB in human osteoblasts, and subsequently regulate the secretion of interleukin-6 and monocyte chemoattractant protein-1. The nuclear factor kappaB transcription factor regulates a number of genes involved in a wide variety of biological processes. Further study of the effects of nuclear factor kappaB activation on S. aureus-infected human osteoblast may provide us new insights into discovery of the immune mechanisms in osteomyelitis.


Subject(s)
Humans , NF-kappa B/metabolism , Osteoblasts/microbiology , Signal Transduction/physiology , Staphylococcus aureus/physiology , Blotting, Western , Electrophoretic Mobility Shift Assay , Enzyme-Linked Immunosorbent Assay , Leupeptins/pharmacology , NF-kappa B/antagonists & inhibitors
14.
Article in English | WPRIM | ID: wpr-190966

ABSTRACT

Recent epidemiologic studies clearly showed that early intensive glucose control has a legacy effect for preventing diabetic macrovascular complications. However, the cellular and molecular processes by which high glucose leads to macrovascular complications are poorly understood. Vascular smooth muscle cell (VSMC) dysfunction due to high glucose is a characteristic of diabetic vascular complications. Activation of nuclear factor-kappaB (NF-kappaB) may play a key role in the regulation of inflammation and proliferation of VSMCs. We examined whether VSMC proliferation and plasminogen activator inhibitor-1 (PAI-1) expression induced by high glucose were mediated by NF-kappaB activation. Also, we determined whether selective inhibition of NF-kappaB would inhibit proliferation and PAI-1 expression in VSMCs. VSMCs of the aorta of male SD rats were treated with various concentrations of glucose (5.6, 11.1, 16.7, and 22.2 mM) with or without an inhibitor of NF-kappaB or expression of a recombinant adenovirus vector encoding an IkappaB-alpha mutant (Ad-IkappaBalphaM). VSMC proliferation was examined using an MTT assay. PAI-1 expression was assayed by real-time PCR and PAI-1 protein in the media was measured by ELISA. NF-kappaB activation was determined by immunohistochemical staining, NF-kappaB reporter assay, and immunoblotting. We found that glucose stimulated VSMC proliferation and PAI-1 expression in a dose-dependent manner up to 22.2 mM. High glucose (22.2 mM) alone induced an increase in NF-kappaB activity. Treatment with inhibitors of NF-kappaB such as MG132, PDTC or expression of Ad-IkappaB-alphaM in VSMCs prevented VSMC proliferation and PAI-1 expression induced by high glucose. In conclusion, inhibition of NF-kappaB activity prevented high glucose-induced VSMC proliferation and PAI-1 expression.


Subject(s)
Animals , Aorta/cytology , Cardiovascular Diseases/prevention & control , Cell Proliferation/drug effects , Cells, Cultured , Diabetes Complications/prevention & control , Gene Expression Regulation/drug effects , Glucose/immunology , Leupeptins/pharmacology , Male , Muscle, Smooth, Vascular/cytology , Myocytes, Smooth Muscle/cytology , NF-kappa B/antagonists & inhibitors , Plasminogen Activator Inhibitor 1/genetics , Proline/analogs & derivatives , Rats , Rats, Sprague-Dawley , Thiocarbamates/pharmacology
15.
Article in English | WPRIM | ID: wpr-44286

ABSTRACT

The massive reorganization of microtubule network involves in transcriptional regulation of several genes by controlling transcriptional factor, nuclear factor-kappa B (NF-kappaB) activity. The exact molecular mechanism by which microtubule rearrangement leads to NF-kappaB activation largely remains to be identified. However microtubule disrupting agents may possibly act in synergy or antagonism against apoptotic cell death in response to conventional chemotherapy targeting DNA damage such as adriamycin or comptothecin in cancer cells. Interestingly pretreatment of microtubule disrupting agents (colchicine, vinblastine and nocodazole) was observed to lead to paradoxical suppression of DNA damage-induced NF-kappaB binding activity, even though these could enhance NF-kappaB signaling in the absence of other stimuli. Moreover this suppressed NF-kappaB binding activity subsequently resulted in synergic apoptotic response, as evident by the combination with Adr and low doses of microtubule disrupting agents was able to potentiate the cytotoxic action through caspase-dependent pathway. Taken together, these results suggested that inhibition of microtubule network chemosensitizes the cancer cells to die by apoptosis through suppressing NF-kappaB DNA binding activity. Therefore, our study provided a possible anti-cancer mechanism of microtubule disrupting agent to overcome resistance against to chemotherapy such as DNA damaging agent.


Subject(s)
Animals , Antibiotics, Antineoplastic/therapeutic use , Apoptosis , Caspases/metabolism , Cell Line , Colchicine/pharmacology , DNA/metabolism , DNA Damage , Doxorubicin/therapeutic use , Humans , Mice , Microtubules/chemistry , NF-kappa B/antagonists & inhibitors , Neoplasms/drug therapy , Nocodazole/pharmacology , Protein Binding , Signal Transduction , Tubulin Modulators/pharmacology , Vinblastine/pharmacology
16.
Rev. argent. microbiol ; 41(3): 134-140, jul.-sep. 2009. graf
Article in Spanish | LILACS | ID: lil-634627

ABSTRACT

El virus de la Lengua azul (VLA) es un ARN virus de doble cadena que induce apoptosis tanto en cultivos celulares como en tejidos blanco. Con el fin de dilucidar el mecanismo de apoptosis en la infección por el VLA, en el presente trabajo examinamos en detalle, por la técnica de Western blot, las señales celulares de caspasas, Bax, citocromo c, Smac/DIABLO y factor nuclear kappa B (NF-kB) que se activan en la infección viral. Hemos comprobado que luego de la infección in vitro con el VLA, se detectó la activación de la caspasa 8 y con ello el mecanismo extrínseco de la apoptosis. También detectamos por primera vez no sólo la activación de miembros de la familia Bcl-2 (Bax), sino también la liberación del citocromo c y la proteína Smac/DIABLO, confirmando que en la infección por el VLA está involucrado el mecanismo secuencial intrínseco de la apoptosis. Asimismo, demostramos que la infección por el VLA activa el NF-kB y que la apoptosis es sustancialmente reducida mediante la inhibición del mismo. La activación de las señales celulares tales como Bax, citocromo c, Smac/DIABLO y NF-kB presentados en este trabajo, esclarecen los mecanismos apoptóticos durante la infección por el VLA para una mayor comprensión del papel primario que juega la apoptosis en la patogénesis del virus.


Bluetongue (BTV) is a double-stranded RNA virus that induces apoptosis both in mammalian cell cultures and in target tissues. To elucidate the apoptosis pathways in BTV infection, we have examined in detail the apoptosis mechanism by examination of caspases, Bax, cytochrome c, Smac/DIABLO and NF-kB signalling pathways. In this report, after cell infection with BTV, the activation of caspase 8 was detected, proving the extrinsic receptor binding apoptotic pathway. Apoptosis followed a sequential pathway involving the detection of activated Bcl-2 family members. Furthermore, its translocation to the mitochondria, as well as the release of cytochrome c and Smac/Diablo confirmed that BTV apoptosis involves the sequential intrinsic pathway. In addition, we demonstrated that NF-kB was activated following BTV infection and cell treatment with an inhibitor peptide before BTV infection, prevented NF-kB activation and substantially reduced cellular apoptosis. Our accumulating data concerning the activation of Bax, cytochrome c, Smac/DIABLO and NF-kB clarify the mechanism of apoptosis during BTV infection, and confer a better understanding of the primary role of apoptosis in BTV pathogenesis.


Subject(s)
Humans , Apoptosis/physiology , Bluetongue virus/physiology , Signal Transduction/physiology , Cytopathogenic Effect, Viral , Caspases/metabolism , Cytochromes c/metabolism , Enzyme Activation , HeLa Cells/virology , Intracellular Signaling Peptides and Proteins/metabolism , Mitochondria/physiology , Mitochondrial Proteins/metabolism , NF-kappa B/antagonists & inhibitors , NF-kappa B/metabolism , Peptides/pharmacology
17.
Article in English | WPRIM | ID: wpr-34745

ABSTRACT

Cordycepin (3'-deoxyadenosine) has been shown to exhibit many pharmacological activities, including anti-cancer, anti-inflammatory, and anti-infection activities. However, the anti-skin photoaging effects of cordycepin have not yet been reported. In the present study, we investigated the inhibitory effects of cordycepin on matrix metalloproteinase-1 (MMP-1) and -3 expressions of the human dermal fibroblast cells. Western blot analysis and real-time PCR revealed cordycepin inhibited UVB-induced MMP-1 and -3 expressions in a dose-dependent manner. UVB strongly activated NF-kappa B activity, which was determined by I kappa B alpha degradation, nuclear localization of p50 and p65 subunit, and NF-kappa B binding activity. However, UVB-induced NF-kappa B activation and MMP expression were completely blocked by cordycepin pretreatment. These findings suggest that cordycepin could prevent UVB-induced MMPs expressions through inhibition of NF-kappa B activation. In conclusion, cordycepin might be used as a potential agent for the prevention and treatment of skin photoaging.


Subject(s)
Aging/physiology , Cells, Cultured , Deoxyadenosines/pharmacology , Dermis/cytology , Dose-Response Relationship, Drug , Enzyme Induction/drug effects , Fibroblasts/drug effects , Gene Expression Regulation, Enzymologic , Humans , Infant, Newborn , Male , Matrix Metalloproteinase 1/antagonists & inhibitors , Matrix Metalloproteinase 3/antagonists & inhibitors , NF-kappa B/antagonists & inhibitors , Skin/physiopathology , Ultraviolet Rays
18.
Braz. j. med. biol. res ; 41(12): 1037-1046, Dec. 2008. ilus
Article in English | LILACS | ID: lil-502154

ABSTRACT

Stress is triggered by numerous unexpected environmental, social or pathological stimuli occurring during the life of animals, including humans, which determine changes in all of their systems. Although acute stress is essential for survival, chronic, long-lasting stress can be detrimental. In this review, we present data supporting the hypothesis that stress-related events are characterized by modifications of oxidative/nitrosative pathways in the brain in response to the activation of inflammatory mediators. Recent findings indicate a key role for nitric oxide (NO) and an excess of pro-oxidants in various brain areas as responsible for both neuronal functional impairment and structural damage. Similarly, cyclooxygenase-2 (COX-2), another known source of oxidants, may account for stress-induced brain damage. Interestingly, some of the COX-2-derived mediators, such as the prostaglandin 15d-PGJ2 and its peroxisome proliferator-activated nuclear receptor PPARγ, are activated in the brain in response to stress, constituting a possible endogenous anti-inflammatory mechanism of defense against excessive inflammation. The stress-induced activation of both biochemical pathways depends on the activation of the N-methyl-D-aspartate (NMDA) glutamate receptor and on the activation of the transcription factor nuclear factor kappa B (NFκB). In the case of inducible NO synthase (iNOS), release of the cytokine TNF-α also accounts for its expression. Different pharmacological strategies directed towards different sites in iNOS or COX-2 pathways have been shown to be neuroprotective in stress-induced brain damage: NMDA receptor blockers, inhibitors of TNF-α activation and release, inhibitors of NFκB, specific inhibitors of iNOS and COX-2 activities and PPARγ agonists. This article reviews recent contributions to this area addressing possible new pharmacological targets for the treatment of stress-induced neuropsychiatric disorders.


Subject(s)
Animals , Humans , Encephalitis , Inflammation Mediators/metabolism , Stress, Psychological/complications , /therapeutic use , Encephalitis/drug therapy , Encephalitis/etiology , Encephalitis/metabolism , Interleukin-1/metabolism , NF-kappa B/antagonists & inhibitors , Nitric Oxide Synthase/metabolism , Nitric Oxide/metabolism , Nitrosation/physiology , Oxidation-Reduction , PPAR gamma/agonists , Stress, Psychological/metabolism , Tumor Necrosis Factor-alpha/antagonists & inhibitors
19.
Rev. chil. reumatol ; 24(4): 197-199, 2008. graf
Article in Spanish | LILACS | ID: lil-532984

ABSTRACT

El factor de transcripción NFkB tiene una participación muy importante en el desarrollo y mantención de una serie de patologías humanas, principalmente aquellas con un componente inflamatorio, como la artritis reumatoide (AR). Al mismo tiempo participa en procesos tan diversos como la regulación de la respuesta inmune y el desarrollo embrionario. Una mejor comprensión de los mecanismos y funciones de NFkB permitiría el desarrollo de drogas específicas y efectivas para el tratamiento de patologías inflamatorias y autoinmunes, tratando de no interferir con las funciones normales de este sistema.


Transcription factor NFkB has an important role in development and maintenance of a lot of human pathologies, mainly those with an inflammatory component, for example rheumatoid arthritis (RA). At the same time, it participates in processes as diverse as development and immune response. A better understanding of NFkB mechanisms and functions will allow the development of more specific and effective drugs for the treatment of inflammatory and autoimmune disorders, without interfering with normal functions of this system.


Subject(s)
Humans , Arthritis, Rheumatoid/metabolism , Autoimmune Diseases/metabolism , NF-kappa B/physiology , Inflammation/metabolism , Arthritis, Rheumatoid/immunology , Autoimmune Diseases/drug therapy , NF-kappa B/antagonists & inhibitors , NF-kappa B/immunology , Inflammation/immunology , Inflammation/drug therapy
20.
Article in English | WPRIM | ID: wpr-205424

ABSTRACT

Reactive oxygen species (ROS) play a crucial role in acute lung injury. Tissue inflammation, the increased vascular permeability, and plasma exudation are cardinal features of acute lung injury. Angiopoietin-1 (Ang1) has potential therapeutic applications in preventing vascular leakage and also has beneficial effects in several inflammatory disorders. Recently developed COMP-Ang1 is more potent than native Ang1 in phosphorylating tyrosine kinase with immunoglobulin and EGF homology domain 2 receptor in endothelial cells. However, there are no data on effects and related molecular mechanisms of COMP- Ang1 on ROS-induced acute lung injury. We used hydrogen peroxide (H2O2)-inhaled mice to evaluate the effect of COMP-Ang1 on pulmonary inflammation, bronchial hyper-responsiveness, and vascular leakage in acute lung injury. The results have revealed that VEGF expression, the levels of IL-4, TNF-alpha, IL-1 beta, intercellular adhesion molecule-1, and vascular cell adhesion molecule-1 in lungs, the levels of hypoxia-inducible factor-1alpha (HIF-1 alpha) and NF-kappa B in nuclear protein extracts, phosphorylation of Akt, and vascular permeability were increased after inhalation of H2O2 and that the administration of COMP-Ang1 markedly reduced airway hyper-responsiveness, pulmonary inflammation, plasma extravasation, and the increases of cytokines, adhesion molecules, and VEGF in lungs treated with H2O2. We have also found that the activation of HIF-1a and NF-kappa B and the increase of phosphoinositide 3-kinase activity in lung tissues after H2O2 inhalation were decreased by the administration of COMP-Ang1. These results suggest that COMP-Ang1 ameliorates ROS-induced acute lung injury through attenuating vascular leakage and modulating inflammatory mediators.


Subject(s)
Acute Lung Injury/chemically induced , Administration, Inhalation , Airway Resistance/drug effects , Animals , Bronchial Hyperreactivity/drug therapy , Bronchoalveolar Lavage Fluid , Capillary Permeability/drug effects , Cytokines/antagonists & inhibitors , Female , Hydrogen Peroxide/adverse effects , Hypoxia-Inducible Factor 1, alpha Subunit/antagonists & inhibitors , Intercellular Adhesion Molecule-1/metabolism , Mice , Mice, Inbred BALB C , NF-kappa B/antagonists & inhibitors , Pneumonia/drug therapy , Recombinant Fusion Proteins/administration & dosage , Vascular Cell Adhesion Molecule-1/metabolism
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