RESUMEN
Zearalenone (ZEN) is a non-steroidal estrogenic mycotoxin that induces severe health disturbances in humans and animals. This study aimed to determine the bioactive compounds in Costus speciosus extract (CSE) using GC-MS and evaluate its protective capability against ZEN-induced oxidative damage, genotoxicity, and cytotoxicity in rats. Six groups of male Sprague Dawley rats were treated orally for 15 days including the control group, CSE-treated groups at low (200 mg/kg b. w) or high (400 mg/kg b. w) dose, ZEN-treated group (40 µg/kg b. w), and the groups treated with ZEN plus the low or the high dose of CSE. Blood and tissue samples were collected for different assays and pathological analyses. The results of GC-MS indicated the identification of 6 compounds and Azulene was the major. Animals that received ZEN showed severe disturbances in serum biochemical, cytokines, oxidative stress indicators, mRNA expression of iNOS, Nrf2, and inflammatory-related genes. ZEN also increased micronucleated polychromatic erythrocytes (MNPCEs) and comet tail formation in bone marrow cells along with the disturbances in the histological architecture of the liver and kidney. Co-administration of CSE plus ZEN could normalize the majority of the tested parameters and the histological picture at a dose as low as 200 mg/kg b. w. Therefore, CSE protects against ZEN toxicity via its antioxidant activity, modulation of iNOS, inflammatory-related genes, and the Nrf2 pathway and it could be used in the endemic regions.
Asunto(s)
Costus , Citocinas , Estrés Oxidativo , Extractos Vegetales , Zearalenona , Animales , Costus/química , Citocinas/metabolismo , Expresión Génica/efectos de los fármacos , Masculino , Factor 2 Relacionado con NF-E2/genética , Factor 2 Relacionado con NF-E2/metabolismo , Óxido Nítrico Sintasa de Tipo II/biosíntesis , Óxido Nítrico Sintasa de Tipo II/genética , Óxido Nítrico Sintasa de Tipo II/metabolismo , Estrés Oxidativo/efectos de los fármacos , Extractos Vegetales/farmacología , Ratas , Ratas Sprague-Dawley , Zearalenona/toxicidadRESUMEN
Metabolic intermediates influence inflammation not only through signaling effects, but also by fueling the production of pro-inflammatory molecules. Microglial production of nitric oxide (NO) requires the consumption of NADPH. NADPH consumed in this process is regenerated from NADP+ primarily through the hexose monophosphate shunt, which can utilize only glucose as a substrate. These factors predict that glucose availability can be rate-limiting for glial NO production. To test this prediction, cultured astrocytes and microglia were incubated with lipopolysaccharide and interferon-γ to promote expression of inducible nitric oxide synthase, and the rate of NO production was assessed at defined glucose concentrations. Increased NO production was detected only in cultures containing microglia. The NO production was markedly slowed at glucose concentrations below 0.5 mM, and comparably reduced by inhibition of the hexose monophosphate shunt with 6-aminonicotinamide. Reduced NO production caused by glucose deprivation was partly reversed by malate, which fuels NADPH production by malate dehydrogenase, and by NADPH itself. These findings highlight the role of the hexose monophosphate shunt in fueling NO synthesis and suggest that microglial NO production in the brain may be limited at sites of low glucose availability, such as abscesses or other compartmentalized infections.
Asunto(s)
Glucosa/metabolismo , Microglía/metabolismo , Óxido Nítrico/biosíntesis , Animales , Animales Recién Nacidos , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/fisiología , Células Cultivadas , Femenino , Lipopolisacáridos/toxicidad , Masculino , Ratones , Ratones Endogámicos C57BL , Microglía/efectos de los fármacos , Óxido Nítrico Sintasa de Tipo II/biosíntesisRESUMEN
Fipronil (FIP) insecticide is extensively used in agriculture, public health and veterinary medicine. Although it is considered as a neurotoxin to insects (target organisms) and exhibits neurological signs upon vertebrates (non-target organisms) exposure, slight is known about its potential neurotoxic effects and its molecular mechanisms on vertebrates. The current study is designed to assess oxidative stress as a molecular mechanism of FIP neurotoxicity subordinated with apoptosis and neural tissue reactivity. Ten adult male albino rats received 10 mg/kg body weight fipronil technical grade by oral gavage daily for 45 days (subacute exposure). Brain neural tissue regions (hippocampus, cerebellum and caudate putamen) were processed to examine oxidative stress induced cellular macromolecular alterations as MDA, PCC and DNA fragmentation. Besides, TNF-α and Bcl-2 gene expression and immunoreactivity for caspase-3 (active form), iNOS and GFAP were evaluated. Also, histopathological assessment was conducted. We found that FIP significantly raised MDA, PCC and DNA fragmentation (p ≤ 0.05). Also, it significantly upregulated TNF-α and non-significantly down-regulated Bcl-2 gene expression (p ≤ 0.05). Further, significant increased immunoreactivity to GFAP, iNOS and caspase-3 (active form) in these brain neural tissue regions in FIP treated group was noticed (p ≤ 0.05). Histopathological findings, including alterations in the histological architecture and neuronal degeneration, were also observed in these brain regions of FIP treated group. In conclusion, we suggest the ability of FIP to induce oxidative stress mediated macromolecular alterations, leading to apoptosis and tissue reaction in these brain regions which showed variable susceptibility to FIP toxic effects.
Asunto(s)
Apoptosis/efectos de los fármacos , Fragmentación del ADN/efectos de los fármacos , Tejido Nervioso/metabolismo , Estrés Oxidativo/efectos de los fármacos , Pirazoles/efectos adversos , Animales , Caspasa 3/biosíntesis , Regulación de la Expresión Génica/efectos de los fármacos , Proteína Ácida Fibrilar de la Glía/biosíntesis , Masculino , Tejido Nervioso/patología , Óxido Nítrico Sintasa de Tipo II/biosíntesis , Proteínas Proto-Oncogénicas c-bcl-2/biosíntesis , Pirazoles/farmacología , Ratas , Ratas Wistar , Factor de Necrosis Tumoral alfa/biosíntesisRESUMEN
Carbon tetrachloride (CCl4) is an abundant environmental pollutant that can generate free radicals and induce oxidative stress in different human and animal organs like the kidney, lung, brain, and spleen, causing toxicity. The present study evaluated the alleviative mechanism of the isolated polyphenolic fraction from seedless (pulp and skin) black Vitis vinifera (VVPF) on systemic oxidative and necroinflammatory stress in CCl4-intoxicated rats. Here, we found that the administration of VVPF to CCl4-intoxicated rats for ten days was obviously ameliorated the CCl4-induced systemic elevation in ROS, NO and TBARS levels, as well as MPO activity. Also, it upregulated the cellular activities of the enzymatic (SOD, and GPx) and non-enzymatic (TAC and GSH) antioxidants. Furthermore, the gene expression of the ROS-related necroinflammatory mediators (NF-κB, iNOS, COX-2, and TNF-α) in the kidney, brain, and spleen, as well as IL-1ß, and IL-8 in the lung were greatly restored. The histopathological studies confirmed these biochemical results and showed a noticeable enhancing effect in the architecture of the studied organs after VVPF intake. Thus, this study indicated that VVPF had an alleviative effect on CCl4-induced necroinflammation and oxidative stress in rat kidney, lung, brain, and spleen via controlling the ROS/NF-κB pathway.
Asunto(s)
Antiinflamatorios/uso terapéutico , Antioxidantes/uso terapéutico , Intoxicación por Tetracloruro de Carbono/tratamiento farmacológico , FN-kappa B/antagonistas & inhibidores , Fitoterapia , Polifenoles/uso terapéutico , Especies Reactivas de Oxígeno/antagonistas & inhibidores , Vitis/química , Animales , Antiinflamatorios/aislamiento & purificación , Antioxidantes/aislamiento & purificación , Encéfalo/efectos de los fármacos , Encéfalo/metabolismo , Intoxicación por Tetracloruro de Carbono/metabolismo , Ciclooxigenasa 2/biosíntesis , Ciclooxigenasa 2/genética , Citocinas/biosíntesis , Citocinas/genética , Evaluación Preclínica de Medicamentos , Frutas/química , Concentración 50 Inhibidora , Riñón/efectos de los fármacos , Riñón/metabolismo , Pulmón/efectos de los fármacos , Pulmón/metabolismo , Óxido Nítrico Sintasa de Tipo II/biosíntesis , Óxido Nítrico Sintasa de Tipo II/genética , Estrés Oxidativo/efectos de los fármacos , Extractos Vegetales/aislamiento & purificación , Polifenoles/química , Polifenoles/aislamiento & purificación , Ratas , Transducción de Señal/efectos de los fármacos , Bazo/efectos de los fármacos , Bazo/metabolismo , Sustancias Reactivas al Ácido Tiobarbitúrico/análisisRESUMEN
Melittin (Mel), a natural detergent, is a major component of bee venom. Mel exhibits favorable clinical effects on the treatment of rheumatoid osteoarthritis, myositis, lumbar muscle strain, and peripheral neurological disorders. Interleukin-1ß (IL-1ß) contributes to the progression of osteoarthritis and is one of the key proinflammatory cytokines. However, the effect of Mel on IL-1ß-induced osteoarthritis has not been reported. We examined the effects of Mel on the expressions of inducible NO synthase (iNOS), nuclear transcription factor κB (NF-κB), and I kappa B (I-κB) in the knee joint cells of C518 rats induced by IL-1ß. Western blot and qPCR results showed that Mel at 0.1µg/mL or higher significantly inhibited iNOS expression. Similarly, 1µg/mL of Mel prevented IL-ß-induced I-κB degradation in the cytoplasm and NF-κB migration from cytoplasm to nucleus. Mel exerts an inhibitory effect on IL-ß-induced NF-κB activation by inhibiting both I-κB degradation and NF-κB migration and can potentially be developed as a new anti-osteoarthritis drug. Further research is needed to clarify the detailed mechanism.
Asunto(s)
Interleucina-1beta/toxicidad , Meliteno/farmacología , FN-kappa B/biosíntesis , Óxido Nítrico Sintasa de Tipo II/biosíntesis , Animales , Línea Celular , Relación Dosis-Respuesta a Droga , Expresión Génica , Masculino , FN-kappa B/genética , Óxido Nítrico Sintasa de Tipo II/genética , RatasRESUMEN
Macrophage accumulation and nitrosative stress are known mechanisms underlying age-related cardiovascular pathology and functional decline. The cardiac muscle microenvironment is known to change with age, yet the direct effects of these changes have yet to be studied in-depth. The present study sought to better elucidate the role that biochemical and biomechanical alterations in cardiac tissue have in the altered phenotype and functionality of cardiac resident macrophages observed with increasing age. To accomplish this, naïve bone marrow derived macrophages from young mice were seeded onto either functionalized poly-dimethyl-siloxane hydrogels ranging in stiffness from 2kPA to 64kPA or onto tissue culture plastic, both of which were coated with either young or aged solubilized mouse cardiac extracellular matrix (cECM). Both biomechanical and biochemical alterations were found to have a significant effect on macrophage polarization and function. Increased substrate stiffness was found to promote macrophage morphologies associated with pro-inflammatory macrophage activation, increased expression of pro-inflammatory inducible nitric oxide synthase protein with increased nitric oxide secretion, and attenuated arginase activity and protein expression. Additionally, exposure to aged cECM promoted attenuated responsivity to both canonical pro-inflammatory and anti-inflammatory cytokine signaling cues when compared to young cECM treated cells. These results suggest that both biomechanical and biochemical changes in the cardiovascular system play a role in promoting the age-related shift towards pro-inflammatory macrophage populations associated with cardiovascular disease development.
Asunto(s)
Microambiente Celular/fisiología , Corazón/fisiología , Macrófagos/fisiología , Macrófagos/ultraestructura , Envejecimiento/patología , Envejecimiento/fisiología , Animales , Arginasa/metabolismo , Fenómenos Biomecánicos , Células de la Médula Ósea , Citocinas/metabolismo , ADN/biosíntesis , Activación de Macrófagos , Ratones , Ratones Endogámicos C57BL , Miocardio/patología , Óxido Nítrico/metabolismo , Óxido Nítrico Sintasa de Tipo II/biosíntesis , Fenotipo , Transducción de Señal , Técnicas de Cultivo de TejidosRESUMEN
Lipopolysaccharide (LPS) - an endotoxin that is being extensively used in laboratory to mimic microbial infection that adversely affects male fertility. This study investigated the protective effects of melatonin on LPS-induced testicular nitro-oxidative stress, inflammation, and associated damages in the testes of male golden hamsters, Mesocricetus auratus. Hamsters were administered with melatonin and LPS for 7 days. Testes of LPS treated hamsters showed degenerative changes (appearance of vacuoles, exfoliation, and depletion of germ cells in the seminiferous tubules), adverse effects on spermatogenesis (sperm count and viability), and steroidogenesis (declined serum and testicular testosterone). Furthermore, LPS treatment decreased melatonin content, melatonin receptor (MT1), and antioxidant potential (catalase and SOD), and simultaneously increased nitro-oxidative stress (CRP, nitrate, TNFα). LPS upregulated NF-kB, COX-2, and iNOS expressions to increase testicular inflammatory load that resulted in the decrease of germ cell proliferation and survival, thus culminating into germ cell apoptosis as indicated by AO-EB staining and caspase-3 expression. Administration of melatonin with LPS showed improved testicular histoarchitecture, sperm parameters, and testosterone level. Melatonin increased testicular antioxidant status (SOD, catalase) to counteract the LPS-induced testicular ROS and thus reduced testicular nitro-oxidative stress. Furthermore, melatonin treatment upregulated testicular SIRT-1 expression to inhibit LPS-induced inflammatory proteins, i.e., NF-kB/COX-2/iNOS expression. The rescue effect of melatonin was further supported by increased germ cell survival (Bcl-2), proliferation (PCNA), and declined apoptosis (caspase-3). In conclusion, our result demonstrated that melatonin rescued testes from LPS-induced testicular nitro-oxidative stress, inflammation, and associated damages by upregulation of SIRT-1.
Asunto(s)
Ciclooxigenasa 2/biosíntesis , Melatonina/farmacología , FN-kappa B/biosíntesis , Óxido Nítrico Sintasa de Tipo II/biosíntesis , Sirtuina 1/biosíntesis , Testículo/metabolismo , Factor de Necrosis Tumoral alfa/biosíntesis , Animales , Cricetinae , Inhibidores de la Ciclooxigenasa 2/farmacología , Mediadores de Inflamación/metabolismo , Lipopolisacáridos/toxicidad , Masculino , Mesocricetus , FN-kappa B/antagonistas & inhibidores , Óxido Nítrico Sintasa de Tipo II/antagonistas & inhibidores , Estrés Nitrosativo/efectos de los fármacos , Estrés Nitrosativo/fisiología , Estrés Oxidativo/efectos de los fármacos , Estrés Oxidativo/fisiología , Testículo/efectos de los fármacos , Testículo/patologíaRESUMEN
Overexpression of the inducible isoform of the enzyme nitric oxide synthase (iNOS) has been associated to pathological processes in the kidney. Ethanol consumption induces the renal expression of iNOS; however, the contribution of this enzyme to the deleterious effects of ethanol in the kidney remains elusive. We examined whether iNOS plays a role in the renal dysfunction and oxidative stress induced by ethanol consumption. With this purpose, male C57BL/6 wild-type (WT) or iNOS-deficient (iNOS-/-) mice were treated with ethanol (20% v/v) for 10 weeks. Treatment with ethanol increased the expression of Nox4 as well as the concentration of thiobarbituric acid reactive substances and the levels of tumor necrosis factor α in the renal cortex of WT but not iNOS-/- mice. Augmented serum levels of creatinine and increased systolic blood pressure were found in WT and iNOS-/- mice treated with ethanol. WT mice treated with ethanol showed increased production of reactive oxygen species and myeloperoxidase activity, but these responses were attenuated in iNOS-/- mice. We concluded that iNOS played a role in ethanol-induced oxidative stress and pro-inflammatory cytokine production in the kidney. These are mechanisms that may contribute to the renal toxicity induced by ethanol.
Asunto(s)
Consumo de Bebidas Alcohólicas/metabolismo , Citocinas/metabolismo , Etanol/farmacología , Inflamación/patología , Enfermedades Renales/patología , Óxido Nítrico Sintasa de Tipo II/metabolismo , Consumo de Bebidas Alcohólicas/efectos adversos , Consumo de Bebidas Alcohólicas/patología , Animales , Antiinfecciosos Locales/toxicidad , Creatinina/metabolismo , Inflamación/enzimología , Inflamación/metabolismo , Mediadores de Inflamación/metabolismo , Enfermedades Renales/etiología , Enfermedades Renales/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Óxido Nítrico Sintasa de Tipo II/biosíntesis , Oxidación-Reducción , Estrés Oxidativo/efectos de los fármacos , Especies Reactivas de Oxígeno/metabolismoRESUMEN
Rheumatoid arthritis (RA) is a chronic autoimmune disorder and serious cause of disability. Despite considerable advances in RA management, challenges like extensive drug metabolism and rapid clearance causes poor bioavailability. Core-shell nanocarriers for co-delivery of glycyrrhizic acid (GA) and budesonide against RA were developed. GA-loaded gelatin nanoparticles (NPs) were synthesized and coated with budesonide encapsulated aminocellulose-grafted polycaprolactone (PCL-AC). GA- and budesonide-loaded PCL-AC-gel NPs had diameter of 200-225 nm. Dual drug-loaded (DDL) NPs reduced joint swelling and erythema in rats while markedly ameliorating bone erosion evidenced by radiological analysis, suppressed collagen destruction, restored synovial tissue, bone and cartilage histoarchitecture with reduced inflammatory cells infiltration. NPs also reduced various inflammatory biomarkers such as TNF-α, IL-1ß, COX-2, iNOS. Results of this study suggest that dual NPs exerted superior therapeutic effects in RA compared to free drugs which may be attributed to slow and sustained drug release and NPs' ability to inhibit inflammatory mediators.
Asunto(s)
Artritis Reumatoide/metabolismo , Artritis Reumatoide/terapia , Celulosa/química , Gelatina/química , Nanopartículas/química , Poliésteres/química , Animales , Biomarcadores/metabolismo , Huesos/efectos de los fármacos , Budesonida/farmacología , Cartílago/efectos de los fármacos , Colágeno/química , Ciclooxigenasa 2/biosíntesis , Sistemas de Liberación de Medicamentos , Femenino , Fibroblastos/metabolismo , Ácido Glicirrínico/farmacología , Humanos , Inflamación , Interleucina-1beta/biosíntesis , Cinética , Espectroscopía de Resonancia Magnética , Óxido Nítrico Sintasa de Tipo II/biosíntesis , Ratas , Ratas Wistar , Factor de Necrosis Tumoral alfa/biosíntesisRESUMEN
Tumor necrosis factor (TNF)/inducible nitric oxide synthase (iNOS)-producing dendritic cells (Tip-DCs) have profound impacts on host immune responses during infections. The mechanisms regulating Tip-DC development remain largely unknown. Here, using a mouse model of infection with African trypanosomes, we show that a deficiency in interleukin-27 receptor (IL-27R) signaling results in escalated intrahepatic accumulation of Ly6C-positive (Ly6C+) monocytes and their differentiation into Tip-DCs. Blocking Tip-DC development significantly ameliorates liver injury and increases the survival of infected IL-27R-/- mice. Mechanistically, Ly6C+ monocyte differentiation into pathogenic Tip-DCs in infected IL-27R-/- mice is driven by a CD4+ T cell-interferon gamma (IFN-γ) axis via cell-intrinsic IFN-γ signaling. In parallel, hyperactive IFN-γ signaling induces cell death of Ly6C-negative (Ly6C-) monocytes in a cell-intrinsic manner, which in turn aggravates the development of pathogenic Tip-DCs due to the loss of the negative regulation of Ly6C- monocytes on Ly6C+ monocyte differentiation into Tip-DCs. Thus, IL-27 inhibits the dual-track exacerbation of Tip-DC development induced by a CD4+ T cell-IFN-γ axis. We conclude that IL-27 negatively regulates Tip-DC development by preventing the cell-intrinsic effects of IFN-γ and that the regulation involves CD4+ T cells and Ly6C- monocytes. Targeting IL-27 signaling may manipulate Tip-DC development for therapeutic intervention.IMPORTANCE TNF/iNOS-producing dendritic cells (Tip-DCs) are at the front line as immune effector cells to fight off a broad range of invading microbes. Excessive development of Tip-DCs contributes to tissue destruction. Thus, identifying master regulators of Tip-DC development is fundamental for developing new therapeutic strategies. Here, we identify Tip-DCs as a terminal target of IL-27, which prevents Tip-DC-mediated early mortality during parasitic infections. We demonstrate that IL-27 inhibits Tip-DC development via a dual-track mechanism involving the complex interactions of effector CD4+ T cells, Ly6C- monocytes, and Ly6C+ monocytes. These findings delineate an in-depth view of mechanisms of Tip-DC differentiation that may have significant implications for the ongoing development of IL-27-based immunotherapy.
Asunto(s)
Diferenciación Celular/inmunología , Células Dendríticas/fisiología , Regulación de la Expresión Génica , Interleucinas/genética , Óxido Nítrico Sintasa de Tipo II/inmunología , Receptores de Interleucina/genética , Trypanosoma congolense/inmunología , Factor de Necrosis Tumoral alfa/inmunología , Animales , Linfocitos T CD4-Positivos/inmunología , Células Dendríticas/inmunología , Interferón gamma/genética , Interferón gamma/inmunología , Interferón gamma/metabolismo , Interleucinas/inmunología , Activación de Linfocitos , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Monocitos/inmunología , Monocitos/fisiología , Óxido Nítrico Sintasa de Tipo II/biosíntesis , Receptores de Interleucina/inmunología , Transducción de Señal/inmunología , Trypanosoma brucei brucei/inmunología , Factor de Necrosis Tumoral alfa/biosíntesisRESUMEN
Hip osteoarthritis (HOA) is characterized by degradation of the cartilage and synovitis. However, the pathohistological effects of synovial tissue inflammation on HOA are not clear. The aim of this study was to evaluate the expression of iNOS, BCL-2 and MMP-9 markers in different synovial cell populations. A total of 32 patients were evaluated retrospectively. Age, sex, height, weight, body mass index were recorded and lymphocyte, fibrocytes and macrophages were analysed in tissue sections. Osteoarthritis cartilage histopathology assessment system (OARSI), Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC), Krenn score, Harris Hip Score (HHS) and Kellgren-Lawrence (K-L) grading of the hip joints were performed. Total hip arthroplasty was performed on 32 patients and controls. Patients were divided into two groups according to their disease severity. The tissues were immunohistochemically analysed. K-L grade and Krenn score differ between all three groups, but also between moderate and severe OA. Synovial lining cell layer, resident cells in stroma and especially inflammatory infiltration were increasing with severity of OA. iNOS expression in both intima and subintima was positively correlated with Krenn score in moderate and severe osteoarthritis (OA) groups. Expression of BCL-2 in intima of severe OA patients was positively correlated with Krenn score. In conclusion, iNOS, BCL-2 and MMP-9 are involved in the regulation of HOA. Our study indicates a relationship between the pathohistological features, the synovial inflammation and the cartilage condition at the time of hip replacement due to OA or femoral neck fracture.
Asunto(s)
Regulación de la Expresión Génica , Metaloproteinasa 9 de la Matriz/biosíntesis , Óxido Nítrico Sintasa de Tipo II/biosíntesis , Osteoartritis de la Cadera/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/biosíntesis , Membrana Sinovial/metabolismo , Anciano , Estudios Transversales , Femenino , Genes bcl-2 , Humanos , Masculino , Metaloproteinasa 9 de la Matriz/genética , Persona de Mediana Edad , Óxido Nítrico Sintasa de Tipo II/genética , Receptor 1 de Factores de Crecimiento Endotelial Vascular/análisisRESUMEN
Artemisia lavandulaefolia, a traditional herbal medicine, has been utilized as anti-inflammatory and analgesia agent in clinic. Bioassay-guided fractionation resulted in a fraction (ALDF) with anti-inflammatory effect obtained from A. lavandulaefolia. Its main constituents were analyzed and identified by UPLC-ESI-Q-TOF-MS technology. ALDF showed the strong inhibitory activity on the nitrogen oxide (NO) production in LPS-induced RAW 264.7 macrophages with an IC50 value of 1.64±0.41â µg/mL. Further results displayed that ALDF also significantly suppressed the secretion of key pro-inflammatory mediators, including tumor necrosis factor-α (TNF-α), prostaglandin E2 (PGE2 ) and interleukin-1ß (IL-1ß), and the increase of the inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) protein expression induced by LPS stimulation. Mechanism study indicated that ALDF was able to block NF-κB signaling pathway through inhibiting IκB and p65 phosphorylation, as well as NF-κB p65 nuclear translocation. Furthermore, inâ vivo results in mice revealed that treatments with ALDF evoked significant inhibition on ear edema induced by xylene and on the writhing responses induced by acetic acid. These results suggest that ALDF holds great potential in the prevention and treatment of inflammatory disorders.
Asunto(s)
Antiinflamatorios/farmacología , Artemisia/química , Fitoquímicos/farmacología , Extractos Vegetales/farmacología , Ácido Acético , Animales , Antiinflamatorios/química , Antiinflamatorios/aislamiento & purificación , Supervivencia Celular/efectos de los fármacos , Células Cultivadas , Ciclooxigenasa 2/biosíntesis , Relación Dosis-Respuesta a Droga , Edema/inducido químicamente , Edema/tratamiento farmacológico , Femenino , Inflamación/tratamiento farmacológico , Inflamación/metabolismo , Interleucina-1beta/antagonistas & inhibidores , Interleucina-1beta/metabolismo , Lipopolisacáridos/antagonistas & inhibidores , Lipopolisacáridos/farmacología , Macrófagos/efectos de los fármacos , Ratones , Ratones Endogámicos BALB C , Estructura Molecular , Óxido Nítrico/antagonistas & inhibidores , Óxido Nítrico/biosíntesis , Óxido Nítrico Sintasa de Tipo II/biosíntesis , Dolor/inducido químicamente , Dolor/tratamiento farmacológico , Fitoquímicos/química , Fitoquímicos/aislamiento & purificación , Extractos Vegetales/química , Extractos Vegetales/aislamiento & purificación , Hojas de la Planta/química , Células RAW 264.7 , Estereoisomerismo , Relación Estructura-Actividad , Factor de Necrosis Tumoral alfa/antagonistas & inhibidores , Factor de Necrosis Tumoral alfa/metabolismo , XilenosRESUMEN
BACKGROUND: Microglia are resident immunocompetent and phagocytic cells in the CNS. Pro-inflammatory microglia, stimulated by microbial signals such as bacterial lipopolysaccharide (LPS), viral RNAs, or inflammatory cytokines, are neurotoxic and associated with pathogenesis of several neurodegenerative diseases. Long non-coding RNAs (lncRNA) are emerging as important tissue-specific regulatory molecules directing cell differentiation and functional states and may help direct proinflammatory responses of microglia. Characterization of lncRNAs upregulated in proinflammatory microglia, such as NR_126553 or 2500002B13Rik, now termed Nostrill (iNOS Transcriptional Regulatory Intergenic LncRNA Locus) increases our understanding of molecular mechanisms in CNS innate immunity. METHODS: Microglial gene expression array analyses and qRT-PCR were used to identify a novel long intergenic non-coding RNA, Nostrill, upregulated in LPS-stimulated microglial cell lines, LPS-stimulated primary microglia, and LPS-injected mouse cortical tissue. Silencing and overexpression studies, RNA immunoprecipitation, chromatin immunoprecipitation, chromatin isolation by RNA purification assays, and qRT-PCR were used to study the function of this long non-coding RNA in microglia. In vitro assays were used to examine the effects of silencing the novel long non-coding RNA in LPS-stimulated microglia on neurotoxicity. RESULTS: We report here characterization of intergenic lncRNA, NR_126553, or 2500002B13Rik now termed Nostrill (iNOS Transcriptional Regulatory Intergenic LncRNA Locus). Nostrill is induced by LPS stimulation in BV2 cells, primary murine microglia, and in cortical tissue of LPS-injected mice. Induction of Nostrill is NF-κB dependent and silencing of Nostrill decreased inducible nitric oxide synthase (iNOS) expression and nitric oxide (NO) production in BV2 and primary microglial cells. Overexpression of Nostrill increased iNOS expression and NO production. RNA immunoprecipitation assays demonstrated that Nostrill is physically associated with NF-κB subunit p65 following LPS stimulation. Silencing of Nostrill significantly reduced NF-κB p65 and RNA polymerase II recruitment to the iNOS promoter and decreased H3K4me3 activating histone modifications at iNOS gene loci. In vitro studies demonstrated that silencing of Nostrill in microglia reduced LPS-stimulated microglial neurotoxicity. CONCLUSIONS: Our data indicate a new regulatory role of the NF-κB-induced Nostrill and suggest that Nostrill acts as a co-activator of transcription of iNOS resulting in the production of nitric oxide by microglia through modulation of epigenetic chromatin remodeling. Nostrill may be a target for reducing the neurotoxicity associated with iNOS-mediated inflammatory processes in microglia during neurodegeneration.
Asunto(s)
Microglía/metabolismo , Óxido Nítrico Sintasa de Tipo II/biosíntesis , ARN Largo no Codificante/biosíntesis , Transcripción Genética/fisiología , Animales , Línea Celular , Células Cultivadas , Femenino , Lipopolisacáridos/toxicidad , Masculino , Ratones , Ratones Endogámicos C57BL , Microglía/efectos de los fármacos , Microglía/patología , Síndromes de Neurotoxicidad/genética , Síndromes de Neurotoxicidad/metabolismo , Síndromes de Neurotoxicidad/patología , Óxido Nítrico Sintasa de Tipo II/genética , ARN Largo no Codificante/genética , Transcripción Genética/efectos de los fármacosRESUMEN
Inflammation is one of the main mechanisms of pancreatic ß-cell damage and the development of type 1 diabetes (T1D). Carvedilol, a beta-adrenergic receptor blocker, has been demonstrated to have anti-inflammatory and antioxidant effects. The aim of this study was to investigate the protective effect of carvedilol against pancreatic ß-cell damage and the development of T1D in an experimental model. T1D was induced in mice by multiple low-dose streptozotocin (STZ) injections. Diabetic mice were treated with carvedilol (15 and 20 mg/kg/day, orally) for 14 days. Results showed that blood glucose levels, diabetes incidence, body weight loss and insulitis in the pancreatic tissue were significantly reduced in mice treated with carvedilol. Treatment of mice with carvedilol significantly increased the levels of antioxidants glutathione (GSH), superoxide dismutase (SOD), and catalase and decreased the levels of malondialdehyde (MDA), nitric oxide (NO) and myeloperoxidase (MPO) in the pancreatic tissue as compared with those in the STZ-induced diabetic mice. Carvedilol decreased the expression of nuclear factor kappa B (NF-κB), cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) as important modulators of inflammation and ß-cell damage in the pancreatic tissue. In addition, carvedilol significantly reduced the levels of proinflammatory cytokines tumor necrosis factor (TNF)-α, interleukin (IL)-1ß, IL-6 IL-12, IL-17, interferon (IFN)-γ and chemokine MCP-1, while increased the anti-inflammatory cytokine IL-10 in the pancreatic tissue. In conclusion, these findings suggest that carvedilol is able to prevent pancreatic ß-cell damage and the development of T1D in mice by the inhibition of inflammatory and oxidative mediators.
Asunto(s)
Carvedilol/farmacología , Ciclooxigenasa 2/biosíntesis , Diabetes Mellitus Tipo 1/prevención & control , Inflamación/metabolismo , Células Secretoras de Insulina/efectos de los fármacos , Subunidad p50 de NF-kappa B/biosíntesis , Óxido Nítrico Sintasa de Tipo II/biosíntesis , Estrés Oxidativo , Animales , Glucemia/metabolismo , Peso Corporal , Citocinas/biosíntesis , Citocinas/metabolismo , Glutatión/metabolismo , Insulina/sangre , Masculino , Malondialdehído/metabolismo , Ratones , Páncreas/metabolismoRESUMEN
Aortic dissection and aneurysm are associated with abnormal hemodynamic loads originating from hypertension. Our previous study demonstrated that cyclic mechanical stretch (CMS, mimicked hypertension) caused the death of rat aortic smooth muscle cells (RASMCs) in a mitogen activated-protein kinases (MAPKs)-dependent manner. The current study investigated the effects of inducible nitric oxide synthase (iNOS) on CMS-induced RASMC death. cDNA microarrays for CMS-treated RASMCs showed that iNOS expression levels were increased in response to CMS. Real-time polymerase chain reaction (PCR) analysis demonstrated that this increase was p38 MAPK (p38)-dependent. NO production was also increased. This increase could be inhibited by p38 and iNOS inhibitors. Thus, CMS-induced iNOS synthesized NO. CMS-induced cell death in RASMCs was increased by the iNOS inhibitor but abrogated by the long-acting NO donor DETA-NONOate. Increased iNOS expression was confirmed in the abdominal aortic constriction mouse model. Signal transducers and activators of transcription 1 (STAT1) was activated in stretched RASMCs, and iNOS expression and NO production were inhibited by the STAT1 inhibitor nifuroxazide. Our findings suggest that RASMCs were protected by iNOS from CMS-stimulated cell death through the STAT1 and p38 signal pathways independently.
Asunto(s)
Aorta/enzimología , Regulación Enzimológica de la Expresión Génica , Mecanotransducción Celular , Músculo Liso Vascular/enzimología , Miocitos del Músculo Liso/enzimología , Óxido Nítrico Sintasa de Tipo II/biosíntesis , Estrés Mecánico , Regulación hacia Arriba , Animales , Aorta/citología , Masculino , Músculo Liso Vascular/citología , Miocitos del Músculo Liso/citología , Ratas , Ratas Sprague-DawleyRESUMEN
Exposure to an electromagnetic field (EMF) can have adverse effects on many organs and tissues, including the reproductive system. This study aimed to investigate the effects of EMF exposure during prenatal and postnatal periods on ovarian development in rat offspring. In this study, rat pups born from eight pregnant rats were used. EMF exposure was initiated on the first day of pregnancy and continued until the 42nd postnatal day. The blood and ovarian tissue samples of female offspring in sham and EMF groups were collected when they reached the age of 42 days. Follicle-stimulating hormone levels were significantly higher in the EMF group than in the sham group. Estradiol levels were significantly lower in the EMF group than in the sham group. Tissue-inducible nitric oxide synthase (iNOS) levels and expression were significantly greater in the EMF group than in the sham group. In the EMF group, congestion, bleeding areas, and degeneration of follicle structures were observed in ovarian tissue. The findings suggest that exposure to 50-Hz, 3-mT EMF used in this study during prenatal and postnatal periods may lead to impaired ovarian structure and function in female offspring. EMF may affect ovarian physiology by increasing iNOS levels and may lead to fertility disorders.
Asunto(s)
Campos Electromagnéticos/efectos adversos , Ovario/efectos de la radiación , Efectos Tardíos de la Exposición Prenatal/veterinaria , Animales , Animales Recién Nacidos , Estradiol/biosíntesis , Femenino , Hormona Folículo Estimulante/biosíntesis , Óxido Nítrico Sintasa de Tipo II/biosíntesis , Embarazo , Ratas , Ratas Sprague-DawleyRESUMEN
Brain sexual differentiation is a developmental process leading to the establishment of stable neural sex differences. In birds and rodents, this process is largely driven by estrogens during a critical period. In rodents, estrogens drive the masculinization of the brain, a process that partly depends on microglia. In contrast, in birds, estrogens produced by females induce demasculinization, but whether microglia are involved in this process is unknown. This study assessed whether microglial number, morphology, and/or activity differ between the sexes in selected regions of the developing quail brain and whether they are influenced by estrogens. We found a robust female-biased sex difference in microglial numbers between embryonic day 9 and 12 in the medial preoptic nucleus (POM), a key region for the expression of male sexual behavior. This difference relies on estrogens produced during the sensitive period. Although most embryonic microglia express iNOS, the expression of iNOS in individual microglia does not differ between sexes. Finally, microglial number and the expression of iNOS were not affected by the microglia inhibitor minocycline. Together, these results revealed an estrogen-dependent sex difference in microglia during the critical period for the sexual differentiation of the quail brain. This difference mirrors the different role of estrogens in the development of birds and rodents and suggests a role for microglia in the sexual differentiation of the brain of birds, as in rodents, thus supporting the hypothesis of a conserved role of the neuroimmune system in the organization of the brain by estrogens.
Asunto(s)
Encéfalo/crecimiento & desarrollo , Encéfalo/metabolismo , Estrógenos/biosíntesis , Microglía/metabolismo , Caracteres Sexuales , Animales , Animales Recién Nacidos , Encéfalo/embriología , Coturnix , Femenino , Masculino , Óxido Nítrico Sintasa de Tipo II/biosíntesisRESUMEN
Macrophage activation is a key event that triggers inflammatory response. The activation is accompanied by metabolic shift such as upregulated glucose metabolism. There are accumulating evidences showing the anti-inflammatory activity of Momordica charantia. However, the effects of M. charantia on inflammatory response and glucose metabolism in activated macrophages have not been fully established. The present study aimed to examine the effect of M. charantia in modulating lipopolysaccharide (LPS)-induced inflammation and perturbed glucose metabolism in RAW264.7 murine macrophages. The results showed that LPS-induced NF-κB (p65) nuclear translocation was inhibited by M. charantia treatment. In addition, M. charantia was found to reduce the expression of inflammatory genes including IL6, TNF-α, IL1ß, COX2, iNOS, and IL10 in LPS-treated macrophages. Furthermore, the data showed that M. charantia reduced the expression of GLUT1 and HK2 genes and lactate production (-28%), resulting in suppression of glycolysis. Notably, its effect on GLUT1 gene expression was found to be independent of LPS-induced inflammation. A further experiment also indicated that the bioactivities of M. charantia may be attributed to its key bioactive compound, charantin. Taken together, the study provided supporting evidences showing the potential of M. charantia for the treatment of inflammatory disorders.
Asunto(s)
Antiinflamatorios , Regulación de la Expresión Génica/efectos de los fármacos , Glucólisis/efectos de los fármacos , Lipopolisacáridos/toxicidad , Macrófagos/metabolismo , Momordica charantia/química , Animales , Antiinflamatorios/química , Antiinflamatorios/farmacología , Ciclooxigenasa 2/biosíntesis , Citocinas/biosíntesis , Transportador de Glucosa de Tipo 1/biosíntesis , Hexoquinasa/biosíntesis , Inflamación/inducido químicamente , Inflamación/tratamiento farmacológico , Inflamación/metabolismo , Inflamación/patología , Ratones , Óxido Nítrico Sintasa de Tipo II/biosíntesis , Células RAW 264.7RESUMEN
Two new 12,23-epoxydammarane-type saponins, notoginsenosides NL-I (1) and NL-J (2), were isolated and identified from Panax notoginseng leaves through the combination of various chromatographies and extensive spectroscopic methods, as well as chemical reactions. Among them, notoginsenoside NL-J (2) had a new skeleton. Furthermore, the lipopolysaccharide (LPS)-induced RAW 264.7 macrophage model was used to identify the in vitro anti-inflammatory activity and mechanisms of compounds 1 and 2. Both of them exerted strong inhibition on nitric oxide (NO) productions in a concentration-dependent manner at 1, 10, and 25 µM. Moreover, these two compounds significantly decreased the secretion of tumor necrosis factor-alpha (TNF-α), interleukin 6 (IL-6), cyclooxygenase-2 (COX-2), nuclear factor kappa-B (NF-κB/p65), and nitric-oxide synthase (iNOS) in LPS-activated RAW 264.7 cells.
Asunto(s)
Antiinflamatorios , Panax notoginseng/química , Hojas de la Planta/química , Saponinas , Animales , Antiinflamatorios/química , Antiinflamatorios/aislamiento & purificación , Antiinflamatorios/farmacología , Ciclooxigenasa 2/biosíntesis , Citocinas/biosíntesis , Lipopolisacáridos/toxicidad , Ratones , Óxido Nítrico Sintasa de Tipo II/biosíntesis , Células RAW 264.7 , Saponinas/química , Saponinas/aislamiento & purificación , Saponinas/farmacología , Factor de Transcripción ReIA/metabolismoRESUMEN
In age-related macular degeneration, inflammatory events are presumed to contribute to disease development. A primary suspect of this contribution is the microglia, the innate immune cell of the retina. In addition, retinal pigment epithelium (RPE) cells can be inflammatorily activated. In this study, we investigate the effect of activated RPE cells on retinal microglia and on neuronal cells. RPE cells and microglia were harvested from porcine eyes. In addition, a neuronal cell line (SHSY-5Y) of human origin was used. For inflammatory activation, agonists of toll-like receptors in different concentrations were used: Pam2CSK4 (Pam; TLR-2), Polyinosinic:polycytidylic acid (Poly I:C; TLR-3) and lipopolysaccharid (LPS; TLR-4). Cell viability was investigated with an MTT assay. The secretion of cytokines was assessed in an ELISA and their expression in real-time PCR. There was no effect of the agonists on cell viability in RPE cells. All agonists induced the secretion of IL-6 and IL-8 in RPE cells with the strongest effect induced by LPS. In microglia, pro-inflammatory stimulation increased the metabolic activity. All agonists induced the secretion of IL-1ß, IL-8, and TNFα in microglia cells while in real-time PCR, LPS and Pam induced the expression of IL-6, IL-1ß and iNOS. Direct stimulation of SHSY-5Y with the agonists induced only minor alterations of viability. Stimulated RPE cell supernatant reduced the secretion of TNFα and IL-8 irrespective of the inducing agent in microglia cells. Additionally a slight induction of IL-1ß was found in microglia treated with supernatant of RPE cells treated with Pam. In real time PCR, the supernatant of RPE cells stimulated with LPS significantly reduced the expression of iNOS and IL-6, but not of IL-1ß. Of note, the expression of iNOS was also reduced by naive RPE cells. The treatment of the SHSY-5Y with supernatant of microglia previously treated with RPE conditioned medium significantly decreased SHSY-5Y viability with and without pro-inflammatory treatment. In conclusion, inflammatory activated RPE cells have a regulatory effect on the pro-inflammatory activation of microglia, stressing the importance of the interaction between these two retinal cell types. Microglia treated with RPE supernatant reduced viability of a neuronal cell line, indicating a neurotoxic effect.