RESUMO
Six transmembrane protein of prostate 2 (STAMP2) is a critical modulator of inflammation and metabolism in adipose tissue. There are no data on the expression of STAMP2 in chronic kidney disease, which is an inflammatory disease related to metabolic disorders. This study aimed to investigate STAMP2 expression in the kidney and heart in 5/6 nephrectomy (Nx) rats, and the effect of omega-3 fatty acid (FA) on STAMP2 expression. Male Sprague Dawley rats were divided into three groups: sham control (0.9% saline), 5/6 Nx (0.9% saline), and 5/6 Nx treated with omega-3 FA (300 mg per kg per day by gastric gavage). The expression of STAMP2 in the kidney and heart were examined by western blotting. Serum creatinine levels were higher in 5/6 Nx rats than in controls. Compared with sham controls, the expression of IκB, NF-κB, NOX4, SREBP-1, and LXR were upregulated and STAMP2 and phosphorylated-AMPK expression were downregulated in the kidney and heart of 5/6 Nx rats. Omega-3 FA supplementation prevented these changes in biomarkers related to inflammation and metabolic lipid disorders. Omega 3-FA supplementation induced the upregulation of STAMP2 protein in 5/6 Nx rats, which was associated with an attenuation of inflammation- and metabolic disease-related markers.
Assuntos
Ácidos Graxos Ômega-3/farmacologia , Falência Renal Crônica/metabolismo , Rim/metabolismo , Proteínas de Membrana/biossíntese , Miocárdio/metabolismo , Quinases Proteína-Quinases Ativadas por AMP , Animais , Creatinina/sangue , Modelos Animais de Doenças , Proteínas I-kappa B/biossíntese , Rim/patologia , Rim/cirurgia , Falência Renal Crônica/etiologia , Falência Renal Crônica/patologia , Receptores X do Fígado/biossíntese , Masculino , Miocárdio/patologia , NADPH Oxidase 4/biossíntese , NF-kappa B/biossíntese , Nefrectomia , Proteínas Quinases/biossíntese , Ratos , Ratos Sprague-Dawley , Proteína de Ligação a Elemento Regulador de Esterol 1/biossínteseRESUMO
Sourdough fermentation of cereal foods is an excellent source of obtaining peptides due to the ability of lactic acid bacteria to activate cereal proteases and produce strain-specific peptidases. With the aim of identifying the lactic acid bacterial strains potentially most effective in producing bioactive peptides, 131 lactobacilli isolates from Italian sourdoughs, used in baking technology, have been screened for proteolytic and peptidase activity. Of these, 23 strains were selected and singly inoculated in liquid sourdoughs from which a Low Molecular Weight fraction containing peptides was obtained. Evaluation of the antioxidant and anti-inflammatory activities of the extracts was performed on cultured cells (RAW 264.7 murine macrophage, murine H-end endothelium cells and Human intestinal Caco-2 cells) by assaying Reactive Oxygen Species (ROS) content, NFkB/IkB expression level and Interleukin-1ß production. As a result, three lactobacilli strains showed a high antioxidant and anti-inflammatory ability enabling the development of model sourdoughs that will potentially increase the nutritional benefits of bread.
Assuntos
Anti-Inflamatórios/metabolismo , Antioxidantes/metabolismo , Pão/microbiologia , Lactobacillus/metabolismo , Peptídeo Hidrolases/metabolismo , Animais , Células CACO-2 , Linhagem Celular , Fermentação , Farinha/microbiologia , Humanos , Proteínas I-kappa B/biossíntese , Interleucina-1beta/biossíntese , Itália , Lactobacillus/classificação , Lactobacillus/isolamento & purificação , Camundongos , Peptídeos/metabolismo , Proteólise , Células RAW 264.7 , Espécies Reativas de Oxigênio/análiseRESUMO
Sodium azide (NaN3) is a chemical compound with multiple toxic effects on vascular and neuronal systems, causing hypotension and neurotoxicity, respectively. In order to test its effects on the immune system, human and mouse macrophage-like cell lines were treated with nontoxic doses of NaN3 and the changes in LPS-induced inflammatory activation was measured. Interestingly, the LPS-induced expression of monocyte chemoattractant protein (MCP)-1 was suppressed by NaN3 without affecting the expression of IL-8 and TNF-α. Further analysis of cellular signaling mediators involved in the expression of these cytokines revealed that NaN3 suppressed the LPS-induced activation of signal transducers and activator of transcription (STAT)1 and inhibitor of κB (IκB) ς, which are involved in the LPS-induced expression of MCP-1, while the LPS-induced activation of nuclear factor κ-light-chain-enhancer of activated B cells (NF-κB) was not affected. The LPS-induced expression of MCP-2 and CXCL10, which are also regulated by STAT1, was suppressed by NaN3. Similarly, the LPS-induced expression of IL-6, which is regulated by IκBζ, was suppressed by NaN3. These results demonstrate that NaN3 selectively suppresses the LPS-induced expression of pro-inflammatory mediators through the suppression of STAT1 and IκBζ activation. These new findings about the activity of NaN3 may contribute to the development of specific regulators of macrophage activity during acute and chronic inflammation.
Assuntos
Quimiocina CCL2/biossíntese , Regulação da Expressão Gênica/efeitos dos fármacos , Proteínas I-kappa B/antagonistas & inibidores , Lipopolissacarídeos/farmacologia , Macrófagos/efeitos dos fármacos , Proteínas Nucleares/antagonistas & inibidores , Fator de Transcrição STAT1/antagonistas & inibidores , Azida Sódica/farmacologia , Proteínas Adaptadoras de Transdução de Sinal/antagonistas & inibidores , Animais , Linhagem Celular Tumoral , Quimiocina CCL2/genética , Proteínas I-kappa B/biossíntese , Proteínas I-kappa B/genética , Mediadores da Inflamação/metabolismo , Interleucina-8/biossíntese , Interleucina-8/genética , Leucemia Monocítica Aguda/patologia , Macrófagos/metabolismo , Camundongos , NF-kappa B/metabolismo , Proteínas Nucleares/biossíntese , Proteínas Nucleares/genética , Fosforilação/efeitos dos fármacos , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , Células RAW 264.7 , Fator de Transcrição STAT1/metabolismo , Fator de Necrose Tumoral alfa/biossíntese , Fator de Necrose Tumoral alfa/genéticaRESUMO
Oleanolic acid (OA) and its several derivatives possess chemopreventive and chemotherapeutic functions against a series of cancer types. Many chemotherapeutic compounds are effective in improving the quality of life and prolonging the survival of patients with gastric cancer, therefore progress in the treatment of gastric cancer, especially the anticancer effects of OA derivatives must be achieved. The inhibitory effect of SZC017, a newly synthesized derivative of OA, on cell viability was determined by MTT assay. Furthermore, flow cytometry, transmission electron microscopy, and western blot analysis revealed that the inhibition of cell viability by OA was mediated by triggering the intrinsic apoptosis of gastric cancer cells, and inducing S phase arrest of SGC7901 cells. Mechanistically, SZC017 was effective against gastric cancer cells via inhibiting Akt/NFκB signaling and topoisomerase I and IIα proteins. Taken together, our data indicate that SZC017 may be a potential chemotherapeutic agent against gastric cancer cells.
Assuntos
Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Ácido Oleanólico/análogos & derivados , Ácido Oleanólico/farmacologia , Piperidinas/farmacologia , Neoplasias Gástricas/patologia , Inibidores da Topoisomerase/farmacologia , Antineoplásicos/síntese química , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , DNA Topoisomerases Tipo I/biossíntese , DNA Topoisomerases Tipo I/genética , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Proteínas I-kappa B/biossíntese , Proteínas I-kappa B/genética , Inibidor de NF-kappaB alfa , NF-kappa B/antagonistas & inibidores , Proteínas de Neoplasias/antagonistas & inibidores , Proteínas de Neoplasias/biossíntese , Proteínas de Neoplasias/genética , Ácido Oleanólico/síntese química , Piperidinas/síntese química , Proteínas Proto-Oncogênicas c-akt/biossíntese , Proteínas Proto-Oncogênicas c-akt/genética , Transdução de Sinais/efeitos dos fármacos , Inibidores da Topoisomerase/síntese químicaRESUMO
UNLABELLED: TNF is highly expressed in synovial tissue of rheumatoid arthritis (RA) patients, where it induces proinflammatory cytokine secretion. However, in other cases, TNF will cause cell death. Considering the abnormal proliferation and activation of rheumatoid arthritis synovioblasts, the proper rate of synovioblast apoptosis could possibly relieve arthritis. However, the mechanism mediating TNF-induced synovioblast survival versus cell death in RA is not fully understood. Our objective was to study the role of CD147 in TNF downstream pathway preference in RA synovioblasts. We found that overexpressing TNF in synovial tissue did not increase the apoptotic level and, in vitro, TNF-induced mild synovioblast apoptosis and promoted IL-6 secretion. CD147, which was highly expressed in rheumatoid arthritis synovial fibroblasts (RASFs), increased the resistance of synovioblasts to apoptosis under TNF stimulation. Downregulating CD147 both increased the apoptotic rate and inhibited IκB kinase (IKK)/IκB/NF-κB pathway-dependent proinflammatory cytokine secretion. Further, we determined that it was the extracellular portion of CD147 and not the intracellular portion that was responsible for synovioblast apoptosis resistance. CD147 monoclonal antibody inhibited TNF-induced proinflammatory cytokine production but had no effect on apoptotic rates. Thus, our study indicates that CD147 is resistant to TNF-induced apoptosis by promoting IKK/IκB/NF-κB pathway, and the extracellular portion of CD147 is the functional region. KEY MESSAGES: CD147 inhibits TNF-stimulated RASF apoptosis. CD147 knockdown decreases IKK expression and inhibits NF-κB-related cytokine secretion. CD147's extracellular portion is responsible for apoptosis resistance. CD147 antibody inhibits TNF-related cytokine secretion without additional apoptosis.
Assuntos
Apoptose/fisiologia , Artrite Reumatoide/patologia , Basigina/metabolismo , Quinase I-kappa B/metabolismo , Proteínas I-kappa B/metabolismo , NF-kappa B/metabolismo , Membrana Sinovial/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Adulto , Basigina/genética , Células Cultivadas , Citocinas/metabolismo , Feminino , Fibroblastos/metabolismo , Humanos , Quinase I-kappa B/biossíntese , Proteínas I-kappa B/biossíntese , Masculino , Pessoa de Meia-Idade , Osteoartrite/patologia , Interferência de RNA , RNA Interferente Pequeno , Membrana Sinovial/citologiaRESUMO
Oleanolic acid (OA) possesses various pharmacological activities, such as antitumor and anti-inflammation; however, its clinical applications are limited by its relatively weak activities and low bioavailability. In this study, we evaluated the cytotoxic activity of seven novel OA derivatives, one of which, SZC014 [2-(pyrrolidine-1-yl) methyl-3-oxo-olean-12-en-28-oic acid], exhibited the strongest antitumor activity; its anticancer effect on gastric cancer cells and action mechanisms were investigated. The viability of OA and seven synthesized derivatives treating gastric cancer cells was detected using tetrazolium (MTT). Among them, SZC014 exhibited the strongest cytotoxic activity against gastric cancer cells (SGC7901, MGC803, and MKN-45). The effect of SZC014 on cell cycle was identified by propidium iodide (PI) staining assay. The cellular apoptosis induced by SZC014 was tested by annexin V/PI. The cellular morphological changes and ultrastructural structures affected by SZC014 were observed and imaged through inverted phase contrast microscope and transmission electron microscopy. Western blotting was performed to explore the expression of proteins associated with apoptosis (caspase 3, caspase 9, Bax, Bcl-2, and Bcl-xL), autophagy (Beclin 1 and ATG 5), and nuclear factor-κB (NF-κB) signal pathway, respectively. The cytotoxic activities of all the seven synthesized OA derivatives were stronger than that of OA against gastric cancer cells. SZC014 exhibited stronger cytotoxic activity than other OA derivatives, inhibited the proliferation of gastric cancer cells, besides, induced G2/M phase cell cycle arrest in SGC7901 cells. Both apoptosis and autophagy were found simultaneously in SZC014-treated SGC7901 cells. Caspase-dependent apoptosis induced by SZC014 was confirmed to be associated with upregulation of Bax and downregulation of Bcl-2 and Bcl-xL, while upregulation of Beclin 1 and ATG 5 was inferred to be involved in SZC014-induced autophagy. Moreover, treating cells with SZC014 resulted in a decrease in phosphorylation of IκBα and NF-κB/p65 and NF-κB/p65 nuclear translocation. The cytotoxic activities of seven OA derivatives were generally stronger than that of OA, among which, SZC014 possessed the most potent anticancer activity in SGC7901 cells and would be a promising chemotherapic agent for the treatment of gastric cancer.
Assuntos
Proteínas I-kappa B/genética , NF-kappa B/genética , Ácido Oleanólico/análogos & derivados , Ácido Oleanólico/administração & dosagem , Pirrolidinas/administração & dosagem , Neoplasias Gástricas/tratamento farmacológico , Fator de Transcrição RelA/genética , Apoptose/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Proteínas I-kappa B/biossíntese , Inibidor de NF-kappaB alfa , NF-kappa B/biossíntese , Proteínas de Neoplasias/biossíntese , Proteínas de Neoplasias/genética , Ácido Oleanólico/química , Fosforilação , Transdução de Sinais/efeitos dos fármacos , Neoplasias Gástricas/genética , Neoplasias Gástricas/patologia , Fator de Transcrição RelA/biossínteseRESUMO
Glomerular hypertrophy and extracellular matrix accumulation are early features of diabetic nephropathy (DN). High glucose-induced oxidative stress is implicated in the etiology of DN. This study aims to investigate the effect of eleutheroside E (EE) on high glucose mediated rat mesangial cells (MCs) proliferation and monocyte chemoattractant protein-1 (MCP-1) expression and the underlying mechanism. MCs proliferation was assessed by MTT assay. Reactive oxygen species (ROS) level and MCP-1 expression were evaluated by ELISA kit. The protein expression of p47, NF-κB p65, p-NF-κB p65, IκBα, p-IκBα, IKKß and p-IKKß were determined by Western blot. The results showed that treatment with EE markedly attenuated high glucose induced MCs proliferation and in a dose-dependent manner. Intervention with EE also significantly blocked high glucose induced intracellular ROS production by decreasing NADPH oxidase activity. Meanwhile, EE administration could effectively alleviate the high glucose-stimulated activation of NF-κB, the degradation of IκBα and the expression of MCP-1. These results demonstrate that high glucose enhances MCs proliferation and MCP-1 expression by activating the ROS/NF-κB pathway and can be inhibited by EE. Our findings provide a new perspective for the clinical treatment of DN.
Assuntos
Quimiocina CCL2/biossíntese , Nefropatias Diabéticas/tratamento farmacológico , Glucosídeos/administração & dosagem , Lignanas/administração & dosagem , Células Mesangiais/efeitos dos fármacos , NF-kappa B/biossíntese , Animais , Proliferação de Células/efeitos dos fármacos , Quimiocina CCL2/genética , Nefropatias Diabéticas/genética , Nefropatias Diabéticas/patologia , Modelos Animais de Doenças , Regulação da Expressão Gênica/efeitos dos fármacos , Glucose/administração & dosagem , Humanos , Proteínas I-kappa B/biossíntese , Proteínas I-kappa B/genética , Células Mesangiais/metabolismo , Células Mesangiais/patologia , NADPH Oxidases/biossíntese , NADPH Oxidases/genética , Inibidor de NF-kappaB alfa , NF-kappa B/genética , Ratos , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos , Fator de Transcrição RelA/biossíntese , Fator de Transcrição RelA/genéticaRESUMO
The allyl sulfides, including diallyl sulfide (DAS), diallyl disulfide (DAD), and diallyl trisulfide (DAT), contained in garlic and members of the Allium family, have a variety of pharmacological activities. Therefore, allyl sulfides have been evaluated as potential novel chemotherapeutic agents. Here, we found that DAT inhibited nuclear factor-κB (NF-κB) signaling and induced apoptosis in primary effusion lymphoma (PEL), a subtype of non-Hodgkin's B-cell lymphoma caused by Kaposi's sarcoma-associated herpesvirus (KSHV). We examined the cytotoxic effects of DAS, DAD and DAT on PEL cells. DAT significantly reduced the viability of PEL cells compared with uninfected B-lymphoma cells, and induced the apoptosis of PEL cells by activating caspase-9. DAT induced stabilization of IκBα, and suppressed NF-κB transcriptional activity in PEL cells. We examined the mechanism underlying DAT-mediated IκBα stabilization. The results indicated that DAT stabilized IκBα by inhibiting the phosphorylation of IκBα by the IκB kinase (IKK) complex. Furthermore, DAT induced proteasomal degradation of TRAF6, and DAT suppressed IKKß-phosphorylation through downregulation of TRAF6. It is known that activation of NF-κB is essential for survival of PEL cells. In fact, the NF-κB inhibitor BAY11-7082 induced apoptosis in PEL cells. In addition, DAT suppressed the production of progeny virus from PEL cells. The administration of DAT suppressed the development of PEL cells and ascites in SCID mice xenografted with PEL cells. These findings provide evidence that DAT has antitumor activity against PEL cells in vitro and in vivo, suggesting it to be a novel therapeutic agent for the treatment of PEL.
Assuntos
Compostos Alílicos/administração & dosagem , Quinase I-kappa B/metabolismo , Proteínas I-kappa B/biossíntese , Peptídeos e Proteínas de Sinalização Intracelular/biossíntese , Linfoma de Efusão Primária/tratamento farmacológico , NF-kappa B/genética , Proteínas de Neoplasias/biossíntese , Sulfetos/administração & dosagem , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Humanos , Quinase I-kappa B/genética , Proteínas I-kappa B/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/genética , Linfoma de Efusão Primária/genética , Linfoma de Efusão Primária/patologia , Camundongos , Inibidor de NF-kappaB alfa , NF-kappa B/metabolismo , Proteínas de Neoplasias/genética , Nitrilas/administração & dosagem , Fosforilação , Proteólise , Transdução de Sinais/efeitos dos fármacos , Sulfonas/administração & dosagem , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Curcumin has commonly been used for the treatment of various allergic diseases. However, its precise anti-allergic rhinitis effect and mechanism remain unknown. In the present study, the effect of curcumin on allergic responses in ovalbumin (OVA)-induced allergic rhinitis mouse was investigated. We explored the effect of curcumin on the release of allergic inflammatory mediators, such as histamine, OVA-specific IgE, and inflammatory cytokines. Also, we found that curcumin improved rhinitis symptoms, inhibited the histopathological changes of nasal mucosa, and decreased the serum levels of histamine, OVA-specific IgE and TNF-α in OVA-induced allergic rhinitis mice. In addition, curcumin suppressed the production of inflammatory cytokines, such as TNF-α, IL-1ß, IL-6 and IL-8. Moreover, curcumin significantly inhibited PMA-induced p-ERK, p-p38, p-JNK, p-Iκ-Bα and NF-κB. These findings suggest that curcumin has an anti-allergic effect through modulating mast cell-mediated allergic responses in allergic rhinitis, at least partly by inhibiting MAPK/NF-κB pathway.
Assuntos
Anti-Inflamatórios/farmacologia , Curcumina/farmacologia , Liberação de Histamina/efeitos dos fármacos , Mastócitos/imunologia , Rinite Alérgica/tratamento farmacológico , Animais , Linhagem Celular , Feminino , Histamina/sangue , Humanos , Proteínas I-kappa B/biossíntese , Imunoglobulina E/sangue , Interleucina-1beta/metabolismo , Interleucina-6/metabolismo , Interleucina-8/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Proteínas Quinases Ativadas por Mitógeno/antagonistas & inibidores , Proteínas Quinases Ativadas por Mitógeno/biossíntese , Miocárdio/imunologia , Inibidor de NF-kappaB alfa , Mucosa Nasal/imunologia , Ovalbumina , Estresse Oxidativo/efeitos dos fármacos , Rinite Alérgica/induzido quimicamente , Fator de Necrose Tumoral alfa/sangue , Fator de Necrose Tumoral alfa/metabolismoRESUMO
The inhibitor of nuclear factor kappa B zeta (IκBζ) is an atypical member of the IκB protein family. Its function in regulating the activity of the transcription factor nuclear factor kappa B (NFκB) as well as its involvement in cancer-associated processes is poorly understood. In glioma patients, enhanced expression of IκBζ in tumor specimen is associated with poor prognosis. Here we report that IκBζ is upregulated in a glioma cell line resistant towards NFκB-dependent non-apoptotic cell death. Upon γ-irradiation of glioma cells, IκBζ expression is enhanced, and subsequently serves as a transcriptional activator of the tumor promoting cytokines interleukin (IL-6), IL-8 and chemokine (C-X-C motif) ligand 1 (CXCL1) that are known to be involved in glioma associated inflammatory processes. In contrast, shRNA-mediated knockdown of IκBζ reduces the expression of the aforementioned cytokines. We propose a previously unappreciated role of IκBζ in the inflammatory micromilieu as well as progression in glioma.
Assuntos
Carcinogênese/genética , Glioma/genética , Proteínas I-kappa B/biossíntese , NF-kappa B/genética , Proteínas Nucleares/biossíntese , Proteínas Adaptadoras de Transdução de Sinal , Morte Celular/efeitos da radiação , Linhagem Celular Tumoral , Quimiocina CXCL1/biossíntese , Raios gama , Regulação Neoplásica da Expressão Gênica/efeitos da radiação , Humanos , Proteínas I-kappa B/antagonistas & inibidores , Proteínas I-kappa B/genética , Interleucina-6/biossíntese , Interleucina-8/biossíntese , NF-kappa B/biossíntese , Proteínas Nucleares/antagonistas & inibidores , Proteínas Nucleares/genética , Prognóstico , RNA Interferente PequenoRESUMO
Resveratrol, a natural polyphenolic compound found in foods and beverages, has attracted increasing attention in recent years because of its potent chemopreventive and anti-tumor effects. In this study, the effects of resveratrol on the expression of P-glycoprotein/multi-drug resistance protein 1 (P-gp/MDR1), and the underlying molecular mechanisms, were investigated in oxaliplatin (L-OHP)-resistant colorectal cancer cells (HCT116/L-OHP). Resveratrol downregulated MDR1 protein and mRNA expression levels and reduced MDR1 promoter activity. It also enhanced the intracellular accumulation of rhodamine 123, suggesting that resveratrol can reverse multi-drug resistance by downregulating MDR1 expression and reducing drug efflux. Resveratrol treatment also reduced nuclear factor-κB (NF-κB) activity, reduced phosphorylation levels of IκBα, and reduced nuclear translocation of the NF-κB subunit p65. Moreover, downregulation of MDR1 expression and promoter activity was mediated by resveratrol-induced AMP-activated protein kinase (AMPK) phosphorylation. The inhibitory effects of resveratrol on MDR1 expression and cAMP-responsive element-binding protein (CREB) phosphorylation were reversed by AMPKα siRNA transfection. We found that the transcriptional activity of cAMP-responsive element (CRE) was inhibited by resveratrol. These results demonstrated that the inhibitory effects of resveratrol on MDR1 expression in HCT116/L-OHP cells were closely associated with the inhibition of NF-κB signaling and CREB activation in an AMPK-dependent manner.
Assuntos
Proteínas Quinases Ativadas por AMP/biossíntese , Neoplasias Colorretais/genética , Estilbenos/administração & dosagem , eIF-2 Quinase/genética , Proteínas Quinases Ativadas por AMP/genética , Subfamília B de Transportador de Cassetes de Ligação de ATP/biossíntese , Subfamília B de Transportador de Cassetes de Ligação de ATP/genética , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/genética , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/patologia , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/genética , Doxorrubicina/administração & dosagem , Resistência a Múltiplos Medicamentos/genética , Resistencia a Medicamentos Antineoplásicos/genética , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Células HCT116 , Humanos , Proteínas I-kappa B/biossíntese , Proteínas I-kappa B/genética , Inibidor de NF-kappaB alfa , Compostos Organoplatínicos/administração & dosagem , Oxaliplatina , Resveratrol , Transdução de Sinais/efeitos dos fármacos , Ativação Transcricional/efeitos dos fármacos , eIF-2 Quinase/biossínteseRESUMO
Naringin exhibits antiinflammatory activity and is shown to induce bone formation. Yet the impact of naringin on inflammation-affected bone marrow-derived mesenchymal stem cell (BM-MSC), a promising tool for the regenerative treatment of bone injury, remained to be investigated. We first cultured and characterized the BM-MSCs in vitro and observe the effects of treatments of TNF-α, naringin, or the combination of both on osteogenic differentiation. TNF-α administered at the concentration of 20 ng/ml results in significant reductions in MSC's cell survival, alkaline phosphatase activity and expressions of two osteogenic genes, Runx2 and Osx. Simultaneous treatment of both TNF-α and naringin is able to rescue such reductions. Further mechanistic studies indicate that TNF-α treatment activates the NF-кB signaling pathway, evidenced by elevated p-IкBα level as well as the increased nuclear fraction of NF-кB subunit, p65. Finally, treatment with both TNF-α and naringin decreases expressions of p-IкBα and nuclear p65, and thus represses NF-кB pathway activated by sole TNF-α treatment. Our findings provide a molecular basis by which naringin restores the TNF-α-induced damage in MSCs and provide novel insights into the application of naringin in the MSC-based treatments for inflammation-induced bone injury.
Assuntos
Células da Medula Óssea/citologia , Flavanonas/farmacologia , Células-Tronco Mesenquimais/citologia , Osteogênese/efeitos dos fármacos , Fator de Necrose Tumoral alfa/farmacologia , Animais , Anti-Inflamatórios/farmacologia , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Subunidade alfa 1 de Fator de Ligação ao Core/biossíntese , Proteínas I-kappa B/biossíntese , Camundongos , Inibidor de NF-kappaB alfa , Osteogênese/fisiologia , Fator de Transcrição Sp7 , Fator de Transcrição RelA/biossíntese , Fatores de Transcrição/biossínteseRESUMO
Radiation-induced bystander effect, appearing as different biological changes in cells that are not directly exposed to ionizing radiation but are under the influence of molecular signals secreted by irradiated neighbors, have recently attracted considerable interest due to their possible implication for radiotherapy. However, various cells present diverse radiosensitivity and bystander responses that depend, inter alia, on genetic status including TP53, the gene controlling the cell cycle, DNA repair and apoptosis. Here we compared the ionizing radiation and bystander responses of human colorectal carcinoma HCT116 cells with wild type or knockout TP53 using a transwell co-culture system. The viability of exposed to X-rays (0-8 Gy) and bystander cells of both lines showed a roughly comparable decline with increasing dose. The frequency of micronuclei was also comparable at lower doses but at higher increased considerably, especially in bystander TP53-/- cells. Moreover, the TP53-/- cells showed a significantly elevated frequency of apoptosis, while TP53+/+ counterparts expressed high level of senescence. The cross-matched experiments where irradiated cells of one line were co-cultured with non-irradiated cells of opposite line show that both cell lines were also able to induce bystander effects in their counterparts, however different endpoints revealed with different strength. Potential mediators of bystander effects, IL-6 and IL-8, were also generated differently in both lines. The knockout cells secreted IL-6 at lower doses whereas wild type cells only at higher doses. Secretion of IL-8 by TP53-/- control cells was many times lower than that by TP53+/+ but increased significantly after irradiation. Transcription of the NFκBIA was induced in irradiated TP53+/+ mainly, but in bystanders a higher level was observed in TP53-/- cells, suggesting that TP53 is required for induction of NFκB pathway after irradiation but another mechanism of activation must operate in bystander cells.
Assuntos
Adenocarcinoma/genética , Apoptose/efeitos da radiação , Neoplasias Colorretais/genética , Genes p53 , Adenocarcinoma/patologia , Apoptose/genética , Efeito Espectador/efeitos da radiação , Linhagem Celular Tumoral/efeitos da radiação , Senescência Celular/efeitos da radiação , Neoplasias Colorretais/patologia , Regulação Neoplásica da Expressão Gênica/efeitos da radiação , Humanos , Proteínas I-kappa B/biossíntese , Proteínas I-kappa B/genética , Interleucina-6/biossíntese , Interleucina-6/genética , Interleucina-8/biossíntese , Interleucina-8/genética , Testes para Micronúcleos , Inibidor de NF-kappaB alfa , Proteínas de Neoplasias/biossíntese , Proteínas de Neoplasias/genética , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , RNA Neoplásico/biossíntese , RNA Neoplásico/genética , Proteína Supressora de Tumor p53/fisiologiaRESUMO
The objective of this study was to investigate the immunostimulatory effects of a polysaccharide fraction from the leaves of Diospyros kaki Thumb (PLE0) and the molecular mechanism responsible for its action in RAW 264.7 macrophages as well as in vivo effects on cyclophosphamide-induced immunosuppression in mice. PLE0 concentration-dependently activated the expressions of inducible nitric oxide synthase (iNOS) at the protein and mRNA levels and its promoter activity, and that these activations caused attendant increases the nitric oxide (NO) production. In addition, PLE0 increased the mRNA expressions of tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), and IL-6. Molecular data revealed that PLE0 increased the transcriptional activity and nuclear translocation of nuclear factor-κB (NF-κB) by inducing the degradation of inhibitory κBα (IκBα) and the phosphorylation of inhibitory κB kinase (IKK). Moreover, anti-toll-like receptor 2 (TLR2) antibody and myeloid differentiation primary response gene 88 (MyD88)-specific siRNA significantly reduce PLE0-induced NO production in RAW 264.7 macrophages. Pretreatment with PLE0 recovered cyclophosphamide-induced reductions in thymus and spleen indices as well as neutrophil counts. Taken together, our data suggest that PLE0 up-regulates the expressions of iNOS, TNF-α, IL-1ß, and IL-6 genes by activating TLR2-mediated NF-κB activations, and that these actions are responsible for its immunostimulatory effects.
Assuntos
Fatores Imunológicos/imunologia , Fatores Imunológicos/isolamento & purificação , Macrófagos/efeitos dos fármacos , Polissacarídeos/imunologia , Receptor 2 Toll-Like/genética , Animais , Diospyros/química , Regulação da Expressão Gênica/efeitos dos fármacos , Proteínas I-kappa B/biossíntese , Fatores Imunológicos/genética , Fatores Imunológicos/farmacologia , Macrófagos/imunologia , Camundongos , Inibidor de NF-kappaB alfa , Óxido Nítrico Sintase Tipo II/biossíntese , Folhas de Planta/química , Polissacarídeos/genética , Polissacarídeos/isolamento & purificação , Polissacarídeos/farmacologia , Transdução de Sinais/efeitos dos fármacos , Receptor 2 Toll-Like/biossíntese , Ativação Transcricional/efeitos dos fármacos , Fator de Necrose Tumoral alfa/biossínteseRESUMO
The therapeutic potential of pectic polysaccharides extracted from Rauvolfia verticillata (Lour.) Baill. var. hainanensis Tsiang in ulcerative colitis were investigated. This study showed that pectic polysaccharides extracted from Rauvolfia verticillata (Lour.) Baill. var. hainanensis Tsiang ameliorated ulcerative colitis and were proposed to exhibit anti-inflammatory effects via increased expression of IκB-α proteins and suppressing NF-αB translocation.
Assuntos
Colite Ulcerativa/tratamento farmacológico , Proteínas I-kappa B/biossíntese , Pectinas/farmacologia , Polissacarídeos/farmacologia , Rauwolfia/química , Animais , Colite Ulcerativa/patologia , Colo/patologia , Feminino , Camundongos , Camundongos Endogâmicos BALB C , Pectinas/química , Pectinas/uso terapêutico , Polissacarídeos/química , Polissacarídeos/uso terapêuticoRESUMO
Chronic inflammation is an important risk factor for lung cancer. Therefore, identification of chemopreventive agents that suppress inflammation-driven lung cancer is indispensable. We studied the efficacy of combinations of indole-3-carbinol (I3C) and silibinin (Sil), 20 µmol/g diet each, against mouse lung tumors induced by 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) and driven by lipopolysaccharide (LPS), a potent inflammatory agent and constituent of tobacco smoke. Mice treated with NNK + LPS developed 14.7±4.1 lung tumors/mouse, whereas mice treated with NNK + LPS and given combinations of I3C and Sil had 7.1±4.5 lung tumors/mouse, corresponding to a significant reduction of 52%. Moreover, the number of largest tumors (>1.0mm) was significantly reduced from 6.3±2.9 lung tumors/mouse in the control group to 1.0±1.3 and 1.6±1.8 lung tumors/mouse in mice given I3C + Sil and I3C alone, respectively. These results were paralleled by significant reductions in the level of proinflammatory and procarcinogenic proteins (pSTAT3, pIκBα and COX-2) and proteins that regulate cell proliferation (pAkt, cyclin D1, CDKs 2, 4, 6 and pRB). Further studies in premalignant bronchial cells showed that the antiproliferative effects of I3C + Sil were higher than the individual compounds and these effects were mediated by targeting cyclin D1, CDKs 2, 4 and 6 and pRB. I3C + Sil suppressed cyclin D1 by reducing its messenger RNA level and by enhancing its proteasomal degradation. Our results showed the potential lung cancer chemopreventive effects of I3C + Sil in smokers/former smokers with chronic pulmonary inflammatory conditions.
Assuntos
Transformação Celular Neoplásica/efeitos dos fármacos , Indóis/farmacologia , Inflamação/tratamento farmacológico , Neoplasias Pulmonares/tratamento farmacológico , Silimarina/farmacologia , Animais , Anticarcinógenos/farmacologia , Proteínas de Ciclo Celular/biossíntese , Linhagem Celular Tumoral , Proliferação de Células , Quimioprevenção , Ciclina D1/biossíntese , Ciclina D1/genética , Quinases Ciclina-Dependentes/biossíntese , Ciclo-Oxigenase 2/biossíntese , Combinação de Medicamentos , Feminino , Humanos , Proteínas I-kappa B/biossíntese , Interleucina-6/biossíntese , Lipopolissacarídeos , Pulmão/patologia , Camundongos , Camundongos Endogâmicos A , Inibidor de NF-kappaB alfa , Nitrosaminas/efeitos adversos , Proteínas Proto-Oncogênicas c-akt/biossíntese , Distribuição Aleatória , Proteína do Retinoblastoma/biossíntese , Fator de Transcrição STAT3/biossíntese , Silibina , Fumaça/efeitos adversos , Fator de Necrose Tumoral alfa/biossínteseRESUMO
BACKGROUND/AIMS: The Zucker diabetic fatty (ZDF, FA/FA) rat is a genetic model of type 2 diabetes, characterized by insulin resistance with progressive metabolic syndrome. We have previously demonstrated mitochondrial dysfunction and oxidative stress in the heart, kidneys and pancreas of ZDF rats. However, the precise molecular mechanism of disease progression is not clear. Our aim in the present study was to investigate oxidative stress and mitochondrial dysfunction in the liver and brain of ZDF rats. METHODS: In this study, we have measured mitochondrial oxidative stress, bioenergetics and redox homeostasis in the liver and brain of ZDF rats. RESULTS: Our results showed increased reactive oxygen species (ROS) production in the ZDF rat brain compared to the liver, while nitric oxide (NO) production was markedly increased both in the brain and liver. High levels of lipid and protein peroxidation were also observed in these tissues. Glutathione metabolism and mitochondrial respiratory functions were adversely affected in ZDF rats when compared to Zucker lean (ZL, +/FA) control rats. Reduced ATP synthesis was also observed in the liver and brain of ZDF rats. Western blot analysis confirmed altered expression of cytochrome P450 2E1, iNOS, p-JNK, and IκB-α confirming an increase in oxidative and metabolic stress in ZDF rat tissues. CONCLUSIONS: Our data shows that, like other tissues, ZDF rat liver and brain develop complications associated with redox homeostasis and mitochondrial dysfunction. These results, thus, might have implications in understanding the etiology and pathophysiology of diabesity which in turn, would help in managing the disease associated complications.
Assuntos
Encéfalo/metabolismo , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Fígado/metabolismo , Mitocôndrias/metabolismo , Animais , Encéfalo/patologia , Diabetes Mellitus Experimental/genética , Diabetes Mellitus Experimental/patologia , Diabetes Mellitus Tipo 2/genética , Diabetes Mellitus Tipo 2/patologia , Proteínas I-kappa B/biossíntese , Resistência à Insulina/genética , Fígado/patologia , Mitocôndrias/patologia , Inibidor de NF-kappaB alfa , Óxido Nítrico/biossíntese , Obesidade/genética , Obesidade/patologia , Oxirredução , Estresse Oxidativo/genética , Ratos , Ratos Zucker , Espécies Reativas de Oxigênio/metabolismoRESUMO
Evidence suggests that upregulation of soluble epoxide hydrolase (sEH) is associated with the development of myocardial infarction, dilated cardiomyopathy, cardiac hypertrophy, and heart failure. However, the upregulation mechanism is still unknown. In this study, we treated H9C2 cells with buthionine sulfoximine (BSO) to explore whether oxidative stress upregulates sEH gene expression and to identify the molecular and cellular mechanisms behind this upregulatory response. Real-time PCR and Western blot analyses were used to measure mRNA and protein expression, respectively. We demonstrated that BSO significantly upregulated sEH at mRNA levels in a concentration- and time-dependent manner, leading to a significant increase in the cellular hypertrophic markers, atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP). Furthermore, BSO significantly increased the cytosolic phosphorylated IκB-α and translocation of NF-κB p50 subunits, as measured by Western blot analysis. This level of translocation was paralleled by an increase in the DNA-binding activity of NF-κB P50 subunits. Moreover, our results demonstrated that pretreatment with the NF-κB inhibitor PDTC significantly inhibited BSO-mediated induction of sEH and cellular hypertrophic marker gene expression in a dose-dependent manner. Additionally, mitogen-activated protein kinases (MAPKs) were transiently phosphorylated by BSO treatment. To understand further the role of MAPKs pathway in BSO-mediated induction of sEH mRNA, we examined the role of extracellular signal-regulated kinase (ERK), c-JunN-terminal kinase (JNK), and p38 MAPK. Indeed, treatment with the MEK/ERK signal transduction inhibitor, PD98059, partially blocked the activation of IκB-α and translocation of NF-κB p50 subunits induced by BSO. Moreover, pretreatment with MEK/ERK signal transduction inhibitors, PD98059 and U0126, significantly inhibited BSO-mediated induction of sEH and cellular hypertrophic marker gene expression. These results clearly demonstrated that the NF-κB signaling pathway is involved in BSO-mediated induction of sEH gene expression, and appears to be associated with the activation of the MAPK pathway. Furthermore, our findings provide a strong link between sEH-induced cardiac dysfunction and involvement of NF-κB in the development of cellular hypertrophy.
Assuntos
Butionina Sulfoximina/farmacologia , Cardiomegalia/patologia , Epóxido Hidrolases/biossíntese , Sistema de Sinalização das MAP Quinases/fisiologia , Subunidade p50 de NF-kappa B/metabolismo , Animais , Antioxidantes/farmacologia , Fator Natriurético Atrial/biossíntese , Butadienos/farmacologia , Linhagem Celular , Sobrevivência Celular , Ativação Enzimática , Epóxido Hidrolases/genética , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Flavonoides/farmacologia , Regulação da Expressão Gênica/fisiologia , Glutationa/biossíntese , Insuficiência Cardíaca/patologia , Proteínas I-kappa B/biossíntese , Proteínas I-kappa B/metabolismo , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Mioblastos Cardíacos/metabolismo , Inibidor de NF-kappaB alfa , Subunidade p50 de NF-kappa B/antagonistas & inibidores , Subunidade p50 de NF-kappa B/biossíntese , Peptídeo Natriurético Encefálico/biossíntese , Nitrilas/farmacologia , Estresse Oxidativo , Fosforilação , Prolina/análogos & derivados , Prolina/farmacologia , RNA Mensageiro/biossíntese , Ratos , Tiocarbamatos/farmacologia , Fator de Transcrição RelA/metabolismo , Regulação para Cima , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
BACKGROUND: Activation of the nuclear factor-κB (NF-κB) pathway is central to the pathogenesis of lung injury and inflammation. We determined whether targeted overexpression of inhibitor-κBα (IκBα) in the lung could decrease the severity of ventilator-induced lung injury (VILI). METHODS: Anaesthetized adult male Sprague-Dawley rats were randomly allocated to undergo intratracheal instillation of: (i) vehicle alone (surfactant, n=10); (ii) 1×10(10) adeno-associated virus encoding IκBα (AAV-IκBα, n=10); (iii) 5×10(10) AAV-IκBα (n=10); and (iv) 1×10(10) AAV-Null (n=5). This was followed by 4 h of injurious mechanical ventilation. Subsequent experiments examined the effect of IκBα overexpression in animals undergoing 'protective' mechanical ventilation. RESULTS: IκBα overexpression increased survival duration at both the lower [3.8 h (0.4)] and higher [3.6 h (0.7)] doses compared with vehicle [2.7 h (1.0)] or the null transgene [2.2 h (0.8)]. IκBα overexpression reduced the alveolar-arterial oxygen gradient (kPa) at both the lower [53 (21)] and higher [52 (19)] doses compared with vehicle [75 (8.5)] or the null transgene [70 (15)], decreased alveolar neutrophil infiltration, and reduced alveolar concentrations of interleukin (IL)-1ß and IL-10. The lower IκBα dose was as effective as the higher dose. IκBα overexpression had no effect in the setting of protective lung ventilation. CONCLUSIONS: Inhibition of pulmonary NF-κB activity by IκBα overexpression reduced the severity of VILI in a rat model.
Assuntos
Terapia Genética/métodos , Proteínas I-kappa B/biossíntese , Pulmão/metabolismo , Lesão Pulmonar Induzida por Ventilação Mecânica/metabolismo , Lesão Pulmonar Induzida por Ventilação Mecânica/prevenção & controle , Animais , Dependovirus/genética , Modelos Animais de Doenças , Expressão Gênica , Vetores Genéticos , Proteínas I-kappa B/genética , Masculino , Inibidor de NF-kappaB alfa , NF-kappa B/metabolismo , Oxigênio/sangue , Pneumonia/metabolismo , Pneumonia/prevenção & controle , Ratos Sprague-Dawley , Respiração Artificial/métodos , Análise de Sobrevida , Transgenes , Lesão Pulmonar Induzida por Ventilação Mecânica/patologiaRESUMO
Resistance to antiestrogens is a major clinical problem in current breast cancer treatment and development of new treatment strategies for these tumors is highly prioritized. In this study, we have investigated the effects of 1α,25-dihydroxyvitamin D3 on the proliferation of tamoxifen-resistant cells. Further, we have investigated on a molecular level the effects of vitamin D on NFkB signaling in tamoxifen-resistant breast cancer cells. Parental human breast cancer MCF-7 cells and four tamoxifen-resistant sublines have been used to investigate the effects of 1α,25-dihydroxyvitamin D3 on cell proliferation using a colorimetric method, gene expression using quantitative PCR, protein phosphorylation using Western blot analysis and cellular localization of proteins using immunofluorescence microscopy. We found that 1α,25-dihydroxyvitamin D3 is able to strongly decrease the growth of both tamoxifen-sensitive and -resistant breast cancer cells and that this antiproliferative effect of 1α,25-dihydroxyvitamin D3 might be mediated via inhibition of the NFκB pathway. We found that 1α,25-dihydroxyvitamin D3 stimulates the gene expression of IkB, an NFκB-inhibiting protein, and that cells pretreated with 1α,25-dihydroxyvitamin D3 have a decreased sensitivity to TNFα stimulation. Further, we show that 1α,25-dihydroxyvitamin D3 treatment strongly decreases the TNFα-induced translocation of p65 into the nucleus. This manuscript reports novel findings regarding the effects of 1α,25-dihydroxyvitamin D3 on NFκB signaling in tamoxifen-resistant breast cancer cells and suggests that vitamin D might be interesting for further evaluation as a new strategy to treat antiestrogen-resistant breast cancers.