RESUMO
MicroRNAs (miRs) act as important post-transcriptional regulators of gene expression in glial cells and have been shown to be involved in the pathogenesis of neurodegenerative diseases, including Alzheimer's disease (AD). Here, we investigated the effects of agathisflavone, a biflavonoid purified from the leaves of Cenostigma pyramidale (Tul.), on modulating the expression of miRs and inflammatory mediators in activated microglia. C20 human microglia were exposed to oligomers of the ß-amyloid peptide (Aß, 500 nM) for 4 h or to lipopolysaccharide (LPS, 1 µg/mL) for 24 h and then treated or not with agathisflavone (1 µM) for 24 h. We observed that ß-amyloid and LPS activated microglia to an inflammatory state, with increased expression of miR-146a, miR-155, IL1-ß, IL-6, and NOS2. Treatment with agathisflavone resulted in a significant reduction in miR146a and miR-155 induced by LPS or Aß, as well as inflammatory cytokines IL1-ß, IL-6, and NOS2. In cells stimulated with Aß, there was an increase in p-STAT3 expression that was reduced by agathisflavone treatment. These data identify a role for miRs in the anti-inflammatory effect of agathisflavone on microglia in models of neuroinflammation and AD.
Assuntos
Doença de Alzheimer , Biflavonoides , MicroRNAs , Humanos , Biflavonoides/farmacologia , Microglia/metabolismo , Interleucina-6/metabolismo , Lipopolissacarídeos/farmacologia , Doença de Alzheimer/metabolismo , Peptídeos beta-Amiloides/metabolismo , Citocinas/metabolismo , MicroRNAs/genética , Fator de Transcrição STAT3/metabolismoRESUMO
Quercetin (Q) is a bioflavonoid with biological potential; however, poor solubility in water, extensive enzymatic metabolism and a reduced bioavailability limit its biopharmacological use. The aim of this study was to perform structural modification in Q by acetylation, thus, obtaining the quercetin pentaacetate (Q5) analogue, in order to investigate the biological potentials (antioxidant, antileishmania, anti-inflammatory and cytotoxicity activities) in cell cultures. Q5 was characterized by FTIR, 1H and 13C NMR spectra. The antioxidant potential was evaluated against the radical ABTSâ¢+. The anti-inflammatory potential was evaluated by measuring the pro-inflammatory cytokine tumor necrosis factor (TNF) and the production of nitric oxide (NO) in peritoneal macrophages from BALB/c mice. Cytotoxicity tests were performed using the AlamarBlue method in cancer cells HepG2 (human hepatocarcinoma), HL-60 (promyelocytic leukemia) and MCR-5 (healthy human lung fibroblasts) as well as the MTT method for C6 cell cultures (rat glioma). Q and Q5 showed antioxidant activity of 29% and 18%, respectively, which is justified by the replacement of hydroxyls by acetyl groups. Q and Q5 showed concentration-dependent reductions in NO and TNF production (p < 0.05); Q and Q5 showed higher activity at concentrations > 40µM when compared to dexamethasone (20 µM). For the HL-60 lineage, Q5 demonstrated selectivity, inducing death in cancer cells, when compared to the healthy cell line MRC-5 (IC50 > 80 µM). Finally, the cytotoxic superiority of Q5 was verified (IC50 = 11 µM), which, at 50 µM for 24 h, induced changes in the morphology of C6 glioma cells characterized by a round body shape (not yet reported in the literature). The analogue Q5 had potential biological effects and may be promising for further investigations against other cell cultures, particularly neural ones.
Assuntos
Anti-Inflamatórios , Antineoplásicos , Antioxidantes , Antiprotozoários , Quercetina/análogos & derivados , Acetilação , Animais , Anti-Inflamatórios/síntese química , Anti-Inflamatórios/química , Anti-Inflamatórios/farmacologia , Antineoplásicos/síntese química , Antineoplásicos/química , Antineoplásicos/farmacologia , Antioxidantes/síntese química , Antioxidantes/química , Antioxidantes/farmacologia , Antiprotozoários/síntese química , Antiprotozoários/química , Antiprotozoários/farmacologia , Células HL-60 , Células Hep G2 , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Quercetina/síntese química , Quercetina/química , Quercetina/farmacologiaRESUMO
The new alkene lactone, (3E)-5,6-dihydro-5-(hydroxymethyl)-3-docdecylidenefuran-3(4H)-one (1), named majoranolide B, and three alkene lactones known as majorenolide (2), majoranolide (3) and majorynolide (4) were obtained from the aerial parts of Persea fulva (Lauraceae). The structures were elucidated in light of extensive spectroscopic analysis, including 1D, 2D NMR (1H, 13C, 1H-1H-COSY, HMBC and HSQC) and HR-ESI-MS. These compounds were screened for their in vitro antiproliferative activity in rat C6 glioma and astrocyte cells using MTT assay and in silico by molecular docking against targets that play a central role in controlling glioma cell cycle progression. Majoranolide (3) is the most active compound with IC50 6.69⯵M against C6 glioma cells, followed by the compounds 1 (IC50 9.06⯵M), 2 (IC50 12.04⯵M) and 4 (IC50 41.90⯵M). The alkene lactones 1-3 exhibited lower toxicity in non-tumor cells when compared to glioma cells. Molecular docking results showed that majoranolide establishes hydrogen bonds with all targets through its α,ß-unsaturated-γ-lactone moiety, whereas the long-chain alkyl group binds by means of several hydrophobic bonds. In the present study, it can be concluded from the anti-proliferative activity of isolates against C6 glioma cells that lactone constituents from P. fulva could have a great potential for the control of C6 glioma cells.
Assuntos
Antineoplásicos Fitogênicos/farmacologia , Lactonas/farmacologia , Lauraceae/química , Simulação de Acoplamento Molecular , Animais , Antineoplásicos Fitogênicos/química , Antineoplásicos Fitogênicos/isolamento & purificação , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Lactonas/química , Lactonas/isolamento & purificação , Estrutura Molecular , Folhas de Planta/química , Ratos , Relação Estrutura-Atividade , Células Tumorais CultivadasRESUMO
Agathisflavone, purified from Cenostigma pyramidale (Tul.) has been shown to be neuroprotective in in vitro models of glutamate-induced excitotoxicity and inflammatory damage. However, the potential role of microglial regulation by agathisflavone in these neuroprotective effects is unclear. Here we investigated the effects of agathisflavone in microglia submitted to inflammatory stimulus in view of elucidating mechanisms of neuroprotection. Microglia isolated from cortices of newborn Wistar rats were exposed to Escherichia coli lipopolysaccharide (LPS, 1 µg/mL) and treated or not with agathisflavone (1 µM). Neuronal PC12 cells were exposed to a conditioned medium from microglia (MCM) treated or not with agathisflavone. We observed that LPS induced microglia to assume an activated inflammatory state (increased CD68, more rounded/amoeboid phenotype). However, most microglia exposed to LPS and agathisflavone, presented an anti-inflammatory profile (increased CD206 and branched-phenotype), associated with the reduction in NO, GSH mRNA for NRLP3 inflammasome, IL1-ß, IL-6, IL-18, TNF, CCL5, and CCL2. Molecular docking also showed that agathisflavone bound at the NLRP3 NACTH inhibitory domain. Moreover, in PC12 cell cultures exposed to the MCM previously treated with the flavonoid most cells preserved neurites and increased expression of ß-tubulin III. Thus, these data reinforce the anti-inflammatory activity and the neuroprotective effect of agathisflavone, effects associated with the control of NLRP3 inflammasome, standing out it as a promising molecule for the treatment or prevention of neurodegenerative diseases.
RESUMO
Recent evidence shows that aminochrome induces glial activation related to neuroinflammation. This dopamine derived molecule induces formation and stabilization of alpha-synuclein oligomers, mitochondria dysfunction, oxidative stress, dysfunction of proteasomal and lysosomal systems, endoplasmic reticulum stress and disruption of the microtubule network, but until now there has been no evidence of effects on production of cytokines and neurotrophic factors, that are mechanisms involved in neuronal loss in Parkinson's disease (PD). This study examines the potential role of aminochrome on the regulation of NGF, GDNF, TNF-α and IL-1ß production and microglial activation in organotypic midbrain slice cultures from P8 - P9 Wistar rats. We demonstrated aminochrome (25⯵M, for 24â¯h) induced reduction of GFAP expression, reduction of NGF and GDNF mRNA levels, morphological changes in Iba1+ cells, and increase of both TNF-α, IL-1ß mRNA and protein levels. Moreover, aminochrome (25⯵M, for 48â¯h) induced morphological changes in the edge of slices and reduction of TH expression. These results demonstrate neuroinflammation, as well as negative regulation of neurotrophic factors (GDNF and NGF), may be involved in aminochrome-induced neurodegeneration, and they contribute to a better understanding of PD pathogenesis.
Assuntos
Encefalite/induzido quimicamente , Indolquinonas/toxicidade , Mesencéfalo/efeitos dos fármacos , Doença de Parkinson/metabolismo , Animais , Encefalite/metabolismo , Fator Neurotrófico Derivado de Linhagem de Célula Glial/metabolismo , Interleucina-1beta/metabolismo , Mesencéfalo/metabolismo , Microglia/efeitos dos fármacos , Microglia/metabolismo , Fator de Crescimento Neural/metabolismo , Ratos Wistar , Técnicas de Cultura de Tecidos , Fator de Necrose Tumoral alfa/metabolismo , Tirosina 3-Mono-Oxigenase/metabolismoRESUMO
Flavonoids are bioactive compounds that are known to be neuroprotective against glutamate-mediated excitotoxicity, one of the major causes of neurodegeneration. The mechanisms underlying these effects are unresolved, but recent evidence indicates flavonoids may modulate estrogen signaling, which can delay the onset and ameliorate the severity of neurodegenerative disorders. Furthermore, the roles played by glial cells in the neuroprotective effects of flavonoids are poorly understood. The aim of this study was to investigate the effects of the flavonoid agathisflavone (FAB) in primary neuron-glial co-cultures from postnatal rat cerebral cortex. Compared to controls, treatment with FAB significantly increased the number of neuronal progenitors and mature neurons, without increasing astrocytes or microglia. These pro-neuronal effects of FAB were suppressed by antagonists of estrogen receptors (ERα and ERß). In addition, treatment with FAB significantly reduced cell death induced by glutamate and this was associated with reduced expression levels of pro-inflammatory (M1) microglial cytokines, including TNFα, IL1ß and IL6, which are associated with neurotoxicity, and increased expression of IL10 and Arginase 1, which are associated with anti-inflammatory (M2) neuroprotective microglia. We also observed that FAB increased neuroprotective trophic factors, such as BDNF, NGF, NT4 and GDNF. The neuroprotective effects of FAB were also associated with increased expression of glutamate regulatory proteins in astrocytes, namely glutamine synthetase (GS) and Excitatory Amino Acid Transporter 1 (EAAT1). These findings indicate that FAB acting via estrogen signaling stimulates production of neurons in vitro and enhances the neuroprotective properties of microglia and astrocytes to significantly ameliorate glutamate-mediated neurotoxicity.