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1.
Pharm Res ; 36(10): 142, 2019 Aug 02.
Artigo em Inglês | MEDLINE | ID: mdl-31376020

RESUMO

BACKGROUND: With the recent approval of the first small interfering RNA (siRNA) therapeutic formulated as nanoparticles, there is increased incentive for establishing the factors of importance for the design of stable solid dosage forms of such complex nanomedicines. METHODS: The aims of this study were: (i) to identify factors of importance for the design of spray-dried siRNA-loaded lipidoid-poly(DL-lactic-co-glycolic acid) hybrid nanoparticles (LPNs), and (ii) to evaluate their influence on the resulting powders by using a quality-by-design approach. Critical formulation and process parameters were linked to critical quality attributes (CQAs) using design of experiments, and an optimal operating space (OOS) was identified. RESULTS: A series of CQAs were identified based on the quality target product profile. The loading (ratio of LPNs to the total solid content) and the feedstock concentration were determined as critical parameters, which were optimized systematically. Mannitol was chosen as stabilizing excipient due to the low water content of the resulting powders. The loading negatively affected the colloidal stability of the LPNs, whereas feedstock concentration correlated positively with the powder particle size. The optimal mannitol-based solid formulation, defined from the OOS, displayed a loading of 5% (w/w), mass median aerodynamic diameter of 3.3 ± 0.2 µm, yield of 60.6 ± 6.6%, and a size ratio of 1.15 ± 0.03. Dispersed micro-embedded LPNs had preserved physicochemical characteristics as well as in vitro siRNA release profile and gene silencing, as compared to non-spray-dried LPNs. CONCLUSION: The optimal solid dosage forms represent robust formulations suitable for higher scale-up manufacturing.


Assuntos
Dessecação/métodos , Lipídeos/química , Nanopartículas/química , Copolímero de Ácido Poliláctico e Ácido Poliglicólico/química , RNA Interferente Pequeno/química , Administração por Inalação , Animais , Composição de Medicamentos , Excipientes/química , Inativação Gênica , Técnicas de Transferência de Genes , Manitol/química , Camundongos , Nanomedicina , Tamanho da Partícula , Pós , Células RAW 264.7 , RNA Interferente Pequeno/administração & dosagem , Solubilidade , Solventes/química
2.
J Agric Food Chem ; 67(32): 9070-9078, 2019 Aug 14.
Artigo em Inglês | MEDLINE | ID: mdl-31343168

RESUMO

In this study, an immunologically active novel microparticulate mushroom ß-glucan (PRA-1p) was prepared using an alkali-soluble glucan PRA-1 by an emulsification and cross-linking method. PRA-1 was a hyperbranched (1→3),(1→6)-ß-d-glucan with a degree of branching of 0.89, isolated from the sclerotia of Polyporus rhinocerus. PRA-1 had a rod-like conformation, while PRA-1p exhibited a monodisperse and homogeneous spherical conformation with a diameter ranging from 0.3 to 2.0 µm in water. PRA-1p significantly induced nitric oxide and reactive oxygen species production as well as morphological changes of murine macrophages (RAW 264.7 cells) and upregulated their phagocytic activity. Furthermore, PRA-1p treatment markedly enhanced the secretion of cytokines, including cutaneous T cell-attracting chemokine 27, granulocyte-colony-stimulating factor, monocyte chemoattractant protein 1, macrophage inflammatory protein 1α, macrophage inflammatory protein 2, regulated on activation, normal T cell expressed and secreted, soluble tumor necrosis factor receptor 1, and tissue inhibitors of metalloproteinases. Activation of RAW 264.7 cells triggered by PRA-1p was associated with activation of inducible nitric oxide synthase, nuclear factor κB, extracellular signal-regulated kinase, and protein kinase B. This work suggests that novel PRA-1p derived from the mushroom sclerotia of P. rhinocerus has potential application as an immunostimulatory agent.


Assuntos
Fatores Imunológicos/química , Fatores Imunológicos/farmacologia , Extratos Vegetais/química , Extratos Vegetais/farmacologia , Polyporus/química , beta-Glucanas/química , beta-Glucanas/farmacologia , Animais , Quimiocina CCL27/genética , Quimiocina CCL27/imunologia , Fator Estimulador de Colônias de Granulócitos e Macrófagos/genética , Fator Estimulador de Colônias de Granulócitos e Macrófagos/imunologia , Fatores Imunológicos/isolamento & purificação , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Camundongos , NF-kappa B/genética , NF-kappa B/imunologia , Óxido Nítrico/imunologia , Fagocitose/efeitos dos fármacos , Extratos Vegetais/isolamento & purificação , Polyporus/imunologia , Células RAW 264.7 , Espécies Reativas de Oxigênio/imunologia , Linfócitos T/efeitos dos fármacos , Linfócitos T/imunologia , beta-Glucanas/isolamento & purificação
3.
J Agric Food Chem ; 67(32): 8810-8818, 2019 Aug 14.
Artigo em Inglês | MEDLINE | ID: mdl-31318199

RESUMO

Citrus grandis (L.) Osbeck is a popular fruit cultivated around the world, and its peels are sometimes used for the treatment of cough, abdominal pain, and indigestion in China. However, the peel is discarded after fruit consumption in most cases, and its chemical constituents and biological activities have not been validated before. The present study focused on evaluation of the chemical and pharmacological profile of coumarins from peels of C. grandis against inflammation. The extracts and phytochemicals from peels of C. grandis were prepared, and anti-inflammatory activities were carried out in vivo and in vitro, including inhibiting xylene-induced ear edema and carrageenan-induced paw edema in mice and the production of inflammatory cytokines (interleukin 1ß, prostaglandin 2, and tumor-necrosis factor α) in lipopolysaccharide (LPS)-induced RAW 264.7 cells. Results indicated that methanolic extract, ethyl acetate fraction, and four major coumarins (compounds 7, 8, 13, and 16) inhibited swelling induced by xylene and carrageenan, separately, in vivo. Furthermore, 18 coumarins inhibited inflammatory factor secretion in macrophages primed by LPS, in which compounds 4, 6, 7, 10, 17 showed the most pronounced change, which were comparable to dexamethasone. In summary, peel of C. grandis showed an anti-inflammatory effect and coumarin compounds were responsible for regulating inflammatory mediators and cytokines, which might provide a novel nutritional strategy for inflammatory diseases.


Assuntos
Anti-Inflamatórios/administração & dosagem , Citrus/química , Cumarínicos/administração & dosagem , Edema/tratamento farmacológico , Frutas/química , Extratos Vegetais/administração & dosagem , Animais , Anti-Inflamatórios/química , Anti-Inflamatórios/isolamento & purificação , Cumarínicos/química , Cumarínicos/isolamento & purificação , Dinoprostona/imunologia , Edema/genética , Edema/imunologia , Humanos , Interleucina-1beta/genética , Interleucina-1beta/imunologia , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Masculino , Camundongos , Camundongos Endogâmicos ICR , Extratos Vegetais/química , Extratos Vegetais/isolamento & purificação , Células RAW 264.7 , Fator de Necrose Tumoral alfa/genética , Fator de Necrose Tumoral alfa/imunologia , Resíduos/análise
4.
J Agric Food Chem ; 67(30): 8339-8347, 2019 Jul 31.
Artigo em Inglês | MEDLINE | ID: mdl-31291543

RESUMO

The dried seeds of Cuminum cyminum L. have been traditionally used as food and medicine. To explore its chemical composition and anti-inflammatory activity, four new compounds (1-4) along with five known compounds (5-9) were isolated from the seeds in the present study. The chemical structures of the new compounds were identified as follows: methyl 3-((7H-purin-2-yl) amino)-3-(4-isopropylphenyl) propanoate (1), 8-(amino(4-isopropylphenyl)methyl)-5-hydroxy-2-(4-hydroxyphenyl)-7-methoxy-4-oxo-4H-chromene-6-carboxylic acid (2), (3,4,5-trihydroxy-6-((4-isopropylbenzyl)oxy)tetrahydro-2H-pyran-2-yl)methyl (E)-3-(4-propoxyphenyl)acrylate (3), and (3,4,5-trihydroxy-6-((5-hydroxy-2-(4-hydroxyphenyl)-4-oxo-4H-chromen-7-yl)oxy)tetrahydro-2H-pyran-2-yl)methyl 3-(4-isopropylphenyl)-2-methoxypropanoate (4). Compound 2, an atypical nitrogen-containing flavonoid, exhibited the most active inhibitory effect on nitride oxide, with IC50 of 5.25 µM in the lipopolysaccharide-stimulated RAW264.7 cell assay. Compound 2 was found to suppress the expression levels of inducible nitric oxide synthase and cyclooxygenase-2. Furthermore, it was revealed that both nuclear factor κB and mitogen-activated protein kinase were involved in the anti-inflammatory process of compound 2.


Assuntos
Anti-Inflamatórios/química , Anti-Inflamatórios/farmacologia , Cuminum/química , Flavonoides/química , Flavonoides/farmacologia , Extratos Vegetais/química , Extratos Vegetais/farmacologia , Animais , Sobrevivência Celular/efeitos dos fármacos , Frutas/química , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Camundongos , Proteínas Quinases Ativadas por Mitógeno/genética , Proteínas Quinases Ativadas por Mitógeno/imunologia , Estrutura Molecular , NF-kappa B/genética , NF-kappa B/imunologia , Óxido Nítrico Sintase Tipo II/genética , Óxido Nítrico Sintase Tipo II/imunologia , Células RAW 264.7 , Sementes/química
5.
Chem Commun (Camb) ; 55(65): 9681-9684, 2019 Aug 21.
Artigo em Inglês | MEDLINE | ID: mdl-31347618

RESUMO

Here, we report a convenient, fast labeling strategy for the imaging of cell surface sialic acids (SAs, nine-carbon monosaccharides located at the terminals of cell surface sugar chains). This strategy is based on the synthesis of sticky, furry and fluorescent "wool-balls", which are wound into nanoclusters from p-benzoquinone/ethylenediamine polymer "wires". With abundant amino groups at the surface, the wool-balls can easily stick to the C-7 aldehyde group generated at the ends of periodate treated SAs in less than 30 min.


Assuntos
Benzoquinonas/química , Etilenodiaminas/química , Corantes Fluorescentes/química , Nanopartículas/química , Polímeros/química , Ácidos Siálicos/análise , Animais , Benzoquinonas/síntese química , Linhagem Celular Tumoral , Etilenodiaminas/síntese química , Fluorescência , Humanos , Camundongos , Microscopia de Fluorescência/métodos , Neuraminidase/química , Tamanho da Partícula , Polímeros/síntese química , Células RAW 264.7 , Bases de Schiff/síntese química , Bases de Schiff/química , Ácidos Siálicos/química
6.
J Agric Food Chem ; 67(28): 7855-7868, 2019 Jul 17.
Artigo em Inglês | MEDLINE | ID: mdl-31274310

RESUMO

Bee pollen (BP) collected from different floras possesses various potential bioactivities, but the mechanism-related research on anti-inflammatory effects is limited. Here, three types of BP originating from Camellia sinensis L. (BP-Cs), Nelumbo nucifera Gaertn. (BP-Nn), and Brassica campestris L. (BP-Bc) were assessed using molecular and metabolomics methods to determine their anti-inflammatory effects. The differences in polyphenolic abundance of three types of BP extracts were determined by HPLC-DAD/Q-TOF-MS. In vitro anti-inflammatory effects of three BP extracts were evaluated in a lipopolysaccharide (LPS)-induced RAW 264.7 cells model. BP-Cs extract with the most abundant polyphenols was found to be the most effective in reducing inflammation by downregulating inflammatory-related genes expression and blocking the activation of MAPK and NF-κB signaling pathways. Polyphenol-rich BP-Cs was further evaluated for their in vivo anti-inflammatory effect in a LPS-induced acute lung injury mouse model. An UPLC-Q-TOF/MS-based metabolomics approach was applied to analyze metabolite changes in mouse serum. Weshowed that the pretreated BP-Cs extract alleviated inflammation and regulated glycerophospholipid metabolism significantly. Our findings provide a foundation for developing and justifying BP as a potential anti-inflammatory ingredient in functional foods or nutraceutical formulations.


Assuntos
Lesão Pulmonar Aguda/tratamento farmacológico , Anti-Inflamatórios/administração & dosagem , Extratos Vegetais/administração & dosagem , Pólen/química , Lesão Pulmonar Aguda/genética , Lesão Pulmonar Aguda/imunologia , Animais , Anti-Inflamatórios/química , Abelhas , Brassica/química , Camellia sinensis/química , Cromatografia Líquida de Alta Pressão , Humanos , Lipopolissacarídeos/efeitos adversos , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Masculino , Espectrometria de Massas , Camundongos , Camundongos Endogâmicos ICR , Quinases de Proteína Quinase Ativadas por Mitógeno/genética , Quinases de Proteína Quinase Ativadas por Mitógeno/imunologia , NF-kappa B/genética , NF-kappa B/imunologia , Nelumbo/química , Extratos Vegetais/química , Polifenóis/administração & dosagem , Polifenóis/química , Células RAW 264.7
7.
Anticancer Res ; 39(6): 2871-2882, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31177125

RESUMO

BACKGROUND/AIM: Tumour-associated macrophages (TAMs) are highjacked M2-polarized macrophages that especially promote pancreatic cancer growth. The aim of this study was to identify an easy-to-use cell culture model suitable for studying this interaction and macrophage polarization. MATERIALS AND METHODS: Co-cultures of two cell lines, PDA6606 cells with RAW macrophages cells were used in vitro and in ovo. Macrophages were analyzed by microscopy, magnetic resonance imaging (MRI), and flow cytometry. RESULTS: By comparing chemically-induced M1 and M2 macrophages, a clear induction of the M2 phenotype of RAW macrophages by PDA6606 pancreatic cancer cells was observed in vitro. In ovo, PDA6606 cells and conditioned media polarized macrophages to the M2 phenotype, which in turn promoted tumour growth and angiogenesis via their surface marker profiles and VEGF production. CONCLUSION: PDA6606 pancreatic cancer cells expectantly and potently induced M2 polarization of RAW264.7 macrophages. This model may be used to study pancreatic cancer-macrophage plasticity in e.g. drug research in vitro and in ovo.


Assuntos
Técnicas de Cocultura/métodos , Neoplasias Pancreáticas/patologia , Células RAW 264.7/citologia , Fator A de Crescimento do Endotélio Vascular/metabolismo , Animais , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Polaridade Celular , Proliferação de Células/efeitos dos fármacos , Meios de Cultivo Condicionados/farmacologia , Camundongos , Neoplasias Pancreáticas/metabolismo , Células RAW 264.7/metabolismo , Transdução de Sinais/efeitos dos fármacos
8.
Vet Microbiol ; 233: 28-38, 2019 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-31176409

RESUMO

The anti-phagocytic abilities of bacteria often affect bacterial pathogenicity. Here, random mutant library of Streptococcus equi subsp. zooepidemicus (SEZ) was constructed using transposon mutagenesis. After careful screening, 30 transposon mutants with different transposon insertion sites were identified by conducting quantitative phagocytosis and insertion-site confirmation assays, whose anti-phagocytic abilities were significantly reduced relative to the wild-type strain. Insertion sites of 19 strains were monocistronic, including genes coding membrane proteins, transporters, and enzymes with unknown pathological function, such as sadM, adhP, purD, guaA, alpha-galactosidase coding gene, ABC transporter permease coding gene, metallo-beta-lactamase coding gene, and three secreted enzyme coding genes spuZ, slaB, and endoS, as well as known virulence factor coding genes, such as hasA and szM. The insertion sites of another 11 strains were polycistronic. We focused on four monocistronic-mutant strains: MhtpZ, MspuZ, MslaB, and MendoS. The anti-phagocytic abilities of not only the mutants that were precoincubated with the recombinant proteins, but also the complement strains were significantly more pronounced than those of all four corresponding mutants. The polyclonal antiserum against SlaB or EndoS also significantly decreased the anti-phagocytic capacity of wild-type SEZ. All four mutants exhibited significantly decreased viability in whole blood and reduced lethality in mice relative to the wild-type strain. Thus, we identified a variety of new anti-phagocytic factors, particularly multiple SEZ secreted enzymes. These factors are instrumental in the phagocytic resistance of SEZ in the absence of opsonin. Our results provide a framework for further studies of SEZ pathogenesis and relevant vaccine development for novel potential targets.


Assuntos
Genes Bacterianos , Óperon , Fagócitos/microbiologia , Fagocitose , Streptococcus equi/genética , Animais , Elementos de DNA Transponíveis , Feminino , Biblioteca Gênica , Camundongos , Camundongos Endogâmicos ICR , Mutagênese , Mutação , Células RAW 264.7 , Streptococcus equi/patogenicidade , Fatores de Virulência/genética
9.
Chem Commun (Camb) ; 55(54): 7729-7736, 2019 Jul 02.
Artigo em Inglês | MEDLINE | ID: mdl-31225574

RESUMO

In the last two decades, the paramount importance of Tumor Associated Carbohydrate Antigens (TACAs) as targets for anticancer vaccine development has been firmly assessed. The Tn antigen is an ideal target for immunotherapy, in that it is masked on normal cells, but exposed on cancer cells. However, it is difficult to elicit an effective and long-lasting response against Tn antigen and other TACAs. Here we report on the Tn antigen analogues developed to boost the latent Tn immune response. Hopefully, the results reported herein will be of help for the rational design of effective TACA-based immunostimulants.


Assuntos
Adjuvantes Imunológicos/química , Antígenos Glicosídicos Associados a Tumores/imunologia , Materiais Biomiméticos/química , Galactosídeos/imunologia , Adjuvantes Imunológicos/síntese química , Animais , Antígenos Glicosídicos Associados a Tumores/química , Materiais Biomiméticos/síntese química , Vacinas Anticâncer/imunologia , Galactosídeos/síntese química , Galactosídeos/química , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Neoplasias/imunologia , Neoplasias/terapia , Peptídeos Cíclicos/síntese química , Peptídeos Cíclicos/química , Peptídeos Cíclicos/imunologia , Células RAW 264.7
10.
Chem Commun (Camb) ; 55(56): 8130-8133, 2019 Jul 18.
Artigo em Inglês | MEDLINE | ID: mdl-31237579

RESUMO

A robust fluorescent probe, MCP1, was developed for triple-detection of H2S, H2Sn and biothiols for the first time. Introduction of H2S, H2Sn and biothiols to MCP1 lead to distinct emission peaks at 508, 576 and 469 nm, respectively, enabling simultaneous detection of H2S, H2Sn and biothiols from distinct emission channels.


Assuntos
Corantes Fluorescentes/química , Sulfeto de Hidrogênio/química , Compostos de Sulfidrila/química , Animais , Espectrometria de Massas/métodos , Camundongos , Células RAW 264.7 , Espectrometria de Fluorescência/métodos , Espectrofotometria Ultravioleta/métodos
11.
Photochem Photobiol Sci ; 18(7): 1851-1857, 2019 Jul 10.
Artigo em Inglês | MEDLINE | ID: mdl-31214682

RESUMO

Exploring techniques for monitoring the intracellular signaling molecule carbon monoxide (CO) in biosystems is important to help understand its various cellular functions. Therefore, a simple long-wavelength colorimetric fluorescent probe LW-CO was designed for selectively and sensitively detecting intracellular CO in living systems. Probe LW-CO is ultrasensitive and can track CO levels in the range of 0-1 µM, with a detection limit of about 3.2 nM. Additionally, the obvious color changes of probe LW-CO with CO (yellow to pink) provide a convenient way for on-site detection of CO with the naked eye. Probe LW-CO was applied to track the exogenous levels of CO in RAW264.7 cells. Probe LW-CO proved to be an efficient method for investigating various cellular functions of CO.


Assuntos
Monóxido de Carbono/análise , Colorimetria , Corantes Fluorescentes/química , Animais , Corantes Fluorescentes/síntese química , Camundongos , Microscopia de Fluorescência , Células RAW 264.7
12.
J Agric Food Chem ; 67(25): 7157-7166, 2019 Jun 26.
Artigo em Inglês | MEDLINE | ID: mdl-31146527

RESUMO

Lonicera caerulea berry polyphenols (LCBP) are known to reduce cholesterol accumulation. Currently, it is unknown whether LCBP can activate Sirtuin 1 (SIRT1) to regulate the formation of RAW264.7 macrophage foam cells. In this study, the effect of LCBP on lipid accumulation in macrophages was evaluated. Fluorescently labeled ox-LDL and 25-NBD cholesterol were used to detect the ox-LDL uptake and cholesterol outflow rate from macrophages. Gene silencing was performed using siRNA to detect changes in the expression of the ATP-binding cassette transporter A1 (ABCA1), sterol regulatory element-binding protein 2 (SREBP2), and SIRT1 proteins using Western blotting, and changes in the expression of miR-33 were detected by real-time polymerase chain reaction. The results showed that treatment with 80 µg/mL LCBP significantly inhibited the accumulation of lipids in RAW264.7 macrophages induced by ox-LDL and reduced intracellular cholesterol levels by activating SIRT1 to enhance the expression of ABCA1, a cholesterol efflux gene, but not independent effect. Of the three key LCBP components investigated, chlorogenic acid was found to activate SIRT1 and regulate the expression of the cholesterol-related factors ABCA1, SREBP2, and miR-33; cyanidin-3-glucoside and catechins were effective to a lesser extent. Our results suggest a novel hypolipidemic mechanism of LCBP.


Assuntos
Colesterol/metabolismo , Células Espumosas/efeitos dos fármacos , Lonicera/química , Macrófagos/efeitos dos fármacos , Polifenóis/farmacologia , Sirtuína 1/metabolismo , Transportador 1 de Cassete de Ligação de ATP/genética , Transportador 1 de Cassete de Ligação de ATP/metabolismo , Animais , Transporte Biológico/efeitos dos fármacos , Células Espumosas/metabolismo , Frutas/química , Lipoproteínas LDL/metabolismo , Macrófagos/metabolismo , Camundongos , Células RAW 264.7 , Sirtuína 1/genética , Proteína de Ligação a Elemento Regulador de Esterol 2/genética , Proteína de Ligação a Elemento Regulador de Esterol 2/metabolismo
13.
Nat Commun ; 10(1): 2377, 2019 05 30.
Artigo em Inglês | MEDLINE | ID: mdl-31147550

RESUMO

Glycans from microbial pathogens are well known pathogen-associated molecular patterns that are recognized by the host immunity; however, little is known about whether and how mammalian self-glycans activate the host immune response, especially in the context of autoimmune disease. Using biochemical fractionation and two-dimensional HPLC, we identify an abundant and bioactive free glycan, the Manß1-4GlcNAc disaccharide in TREX1-associated autoimmune diseases. We report that both monosaccharide residues and the ß1-4 linkage are critical for bioactivity of this disaccharide. We also show that Manß1-4GlcNAc is produced by oligosaccharyltransferase hydrolysis of lipid-linked oligosaccharides in the ER lumen, followed by ENGase and mannosidase processing in the cytosol and lysosomes. Furthermore, synthetic Manß1-4GlcNAc disaccharide stimulates a broad immune response in vitro, which is in part dependent on the STING-TBK1 pathway, and enhances antibody response in vivo. Together, our data identify Manß1-4GlcNAc as a novel innate immune modulator associated with chronic autoimmune diseases.


Assuntos
Doenças Autoimunes/imunologia , Autoimunidade/imunologia , Dissacarídeos/imunologia , Imunidade Inata/imunologia , Proteínas de Membrana/imunologia , Proteínas Serina-Treonina Quinases/imunologia , Animais , Doenças Autoimunes/genética , Modelos Animais de Doenças , Retículo Endoplasmático , Exodesoxirribonucleases/genética , Fibroblastos , Camundongos , Fosfoproteínas/genética , Células RAW 264.7
14.
Int J Nanomedicine ; 14: 4145-4155, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31239673

RESUMO

Background: There is emerging evidence which suggests that cellular ROS including nitric oxide (NO) are important mediators for inflammation and osteoarthritis (OA). Water-soluble polyhydroxylated fullerene C60 (fullerol) nanoparticle has been demonstrated to have an outstanding ability to scavenge ROS. Purpose: The objective of this study is to assess the effects of fullerol on inflammation and OA by in vitro and in vivo studies. Methods: For in vitro experiments, primary mouse peritoneal macrophages and a macrophage cell line RAW264.7 were stimulated to inflammatory phenotypes by lipopolysaccharide (LPS) in the presence of fullerol. For the animal study, OA model was created by intra-articular injection of monoiodoacetate into the knee joints of rats and fullerol was intravenously injected immediately after OA induction. Results: NO production and pro-inflammatory gene expression induced by LPS was significantly diminished by fullerol in both macrophage cell types. Meanwhile, fullerol could remarkably reduce phosphorylation of p38 mitogen-activated protein kinase, and protein level of transcription factors nuclear factor-kappaB and forkhead box transcription factor 1 within the nucleus. The animal study delineated that systematic administration of fullerol prevented OA, inhibiting inflammation of synovial membranes and the damage toward the cartilage chondrocytes in the OA joints. Conclusion: Antioxidative fullerol may have a potential therapeutic application for OA.


Assuntos
Antioxidantes/farmacologia , Fulerenos/farmacologia , Ativação de Macrófagos/efeitos dos fármacos , Nanopartículas/química , Osteoartrite/patologia , Animais , Células Cultivadas , Feminino , Interleucina-1beta/metabolismo , Interleucina-6/metabolismo , Articulações/efeitos dos fármacos , Articulações/patologia , Macrófagos Peritoneais/efeitos dos fármacos , Macrófagos Peritoneais/patologia , Masculino , Camundongos , Modelos Biológicos , NF-kappa B/metabolismo , Óxido Nítrico Sintase Tipo II/metabolismo , Nitritos/metabolismo , Osteoartrite/tratamento farmacológico , Células RAW 264.7 , Ratos Sprague-Dawley , Fator de Necrose Tumoral alfa/metabolismo
15.
Chem Biol Interact ; 309: 108718, 2019 Aug 25.
Artigo em Inglês | MEDLINE | ID: mdl-31211952

RESUMO

We have previously reported the isolation of four compounds, caffeoyloxy-5,6-dihydro-4-methyl-(2H)-pyran-2-one (CDMP), olinioside, caffeic acid and 3-hydroxylup-12-en-28-oic acid, from the leaves of Olinia usambarensis. Here, we evaluated the inhibitory effects of these compounds on lipopolysaccharide (LPS)-induced production of nitric oxide (NO) and prostaglandin E2 (PGE2) in RAW 264.7 macrophages, and found that CDMP is the most potent of these two pro-inflammatory mediators (IC50; 12.12 µM and 10.78 µM, respectively). Consistent with these results, CDMP also down-regulated inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), tumor necrosis factor α (TNF-α), interleukin 1ß (IL-1ß), and interleukin 6 (IL-6) at the protein and mRNA levels in LPS-treated RAW 264.7 macrophages. Furthermore, CDMP suppressed LPS-induced nuclear factor κB (NF-κB) activation by decreasing p65 nuclear translocation through the phosphorylation and degradation of the inhibitory κBα (IκBα). CDMP also attenuated LPS-induced transcriptional and DNA-binding activities of activator protein 1 (AP-1) by suppressing the phosphorylation and expression of c-Fos and c-Jun. Finally, CDMP considerably suppressed the LPS-induced phosphorylation of c-Jun N-terminal kinase (JNK), but did not affect the phosphorylation of p38 or extracellular signal-regulated kinase (ERK). Taken together, our data suggest that CDMP down-regulates genes encoding pro-inflammatory mediators and cytokines, such as iNOS, COX-2, TNF-α, IL-1ß, and IL-6 via NF-κB and JNK/AP-1 inactivation in LPS-induced RAW 264.7 macrophages.


Assuntos
Mediadores da Inflamação/metabolismo , Myrtales/química , NF-kappa B/metabolismo , Piranos/farmacologia , Transdução de Sinais/efeitos dos fármacos , Fator de Transcrição AP-1/metabolismo , Animais , Sobrevivência Celular/efeitos dos fármacos , Ciclo-Oxigenase 2/metabolismo , Citocinas/genética , Citocinas/metabolismo , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Lipopolissacarídeos/farmacologia , Camundongos , Myrtales/metabolismo , NF-kappa B/antagonistas & inibidores , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase Tipo II/metabolismo , Folhas de Planta/química , Folhas de Planta/metabolismo , Piranos/química , Células RAW 264.7 , Fator de Transcrição AP-1/antagonistas & inibidores
16.
Nat Commun ; 10(1): 2839, 2019 06 28.
Artigo em Inglês | MEDLINE | ID: mdl-31253798

RESUMO

Immunomodulatory therapies are becoming a paradigm-shifting treatment modality for cancer. Despite promising clinical results, cancer immunotherapy is accompanied with off-tumor toxicity and autoimmune adverse effects. Thus, the development of smarter systems to regulate immune responses with superior spatiotemporal precision and enhanced safety is urgently needed. Here we report an activatable engineered immunodevice that enables remote control over the antitumor immunity in vitro and in vivo with near-infrared (NIR) light. The immunodevice is composed of a rationally designed UV light-activatable immunostimulatory agent and upconversion nanoparticle, which acts as a transducer to shift the light sensitivity of the device to the NIR window. The controlled immune regulation allows the generation of effective immune response within tumor without disturbing immunity elsewhere in the body, thereby maintaining the antitumor efficacy while mitigating systemic toxicity. The present work illustrates the potential of the remote-controlled immunodevice for triggering of immunoactivity at the right time and site.


Assuntos
Raios Infravermelhos , Nanopartículas/efeitos da radiação , Fototerapia/métodos , Animais , Linhagem Celular Tumoral , Feminino , Imunomodulação , Macrófagos , Neoplasias Mamárias Experimentais/terapia , Camundongos , Camundongos Endogâmicos BALB C , Células RAW 264.7 , Raios Ultravioleta
17.
Eur J Med Chem ; 177: 457-466, 2019 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-31181405

RESUMO

Histone deacetylases (HDACs) play an important role in cancer, degenerative diseases and inflammation. The currently applied HDAC inhibitors in the clinic lack selectivity among HDAC isoforms, which limits their application for novel indications such as inflammatory diseases. Recent, literature indicates that HDAC 3 plays an important role among class I HDACs in gene expression in inflammation. In this perspective, the development and understanding of inhibitory selectivity among HDACs 1, 2 and 3 and their respective influence on gene expression need to be characterized to facilitate drug discovery. Towards this aim, we synthesized nine structural analogues of the class I HDAC inhibitor Entinostat and investigated their selectivity profile among HDACs 1, 2 and 3. We found that we can explain the observed structure activity relationships by small structural and conformational differences between HDAC 1 and HDAC 3 in the 'lid' interacting region. Cell-based studies indicated, however, that application of inhibitors with improved HDAC 3 selectivity did not provide an anti-inflammatory response in contrast to expectations from biochemical evidence in literature. Altogether, in this study, we identified structure activity relationships among class I HDACs and we connected isoform selectivity among class I HDACs with pro- and anti-inflammatory gene transcription in macrophages.


Assuntos
Anilidas/farmacologia , Benzamidas/farmacologia , Expressão Gênica/efeitos dos fármacos , Inibidores de Histona Desacetilases/farmacologia , Macrófagos/efeitos dos fármacos , Anilidas/síntese química , Anilidas/química , Anilidas/metabolismo , Animais , Benzamidas/síntese química , Benzamidas/química , Benzamidas/metabolismo , Domínio Catalítico , Histona Desacetilase 1/química , Histona Desacetilase 1/metabolismo , Histona Desacetilase 2/metabolismo , Inibidores de Histona Desacetilases/síntese química , Inibidores de Histona Desacetilases/química , Inibidores de Histona Desacetilases/metabolismo , Histona Desacetilases/química , Histona Desacetilases/metabolismo , Humanos , Inflamação/genética , Interleucina-10/genética , Interleucina-6/genética , Camundongos , Simulação de Acoplamento Molecular , Subunidade p50 de NF-kappa B/metabolismo , Óxido Nítrico Sintase Tipo II/genética , Ligação Proteica , Células RAW 264.7 , Estereoisomerismo , Relação Estrutura-Atividade , Fator de Necrose Tumoral alfa/genética
18.
J Microbiol Biotechnol ; 29(6): 989-998, 2019 Jun 28.
Artigo em Inglês | MEDLINE | ID: mdl-31154748

RESUMO

Autophagy is crucial for immune defense against Mycobacterium tuberculosis (Mtb) infection. Mtb can evade host immune attack and survival within macrophages by manipulating the autophagic process. MicroRNAs (miRNAs) are small, non-coding RNAs that are involved in regulating vital genes during Mtb infection. The precise role of miRNAs in autophagy with the exits of Mtb remains largely unknown. In this study, we found miR-1958, a new miRNA that could regulate autophagy by interacting with 3'UTR of autophagy-related gene 5 (Atg5). In addition, Mtb infection triggered miR-1958 expression in RAW264.7 cells. What's more, miR- 1958 overexpression blocked autophagic flux by impairing the fusion of autophagosomes and lysosomes. Overexpression of miR-1958 reduced Atg5 expression and LC3 puncta while inhibition of miR-1958 brought an increase of Atg5 and LC3 puncta; the opposite results were observed in detection of p62. The survival of Mtb in RAW264.7 cells transfected with mimic of miR-1958 was enhanced. Taken together, our research demonstrated that a novel miR-1958 could inhibit autophagy by interacting with Atg5 and favored intracellular Mtb survival in RAW264.7 cells.


Assuntos
Proteína 5 Relacionada à Autofagia/genética , Autofagia/fisiologia , MicroRNAs/metabolismo , Viabilidade Microbiana/genética , Mycobacterium tuberculosis/fisiologia , Regiões 3' não Traduzidas , Animais , Autofagossomos/metabolismo , Expressão Gênica , Evasão da Resposta Imune , Lisossomos/metabolismo , Macrófagos/metabolismo , Macrófagos/microbiologia , Macrófagos/patologia , Camundongos , MicroRNAs/genética , Mycobacterium tuberculosis/imunologia , Células RAW 264.7
19.
Sci Total Environ ; 684: 657-669, 2019 Sep 20.
Artigo em Inglês | MEDLINE | ID: mdl-31158627

RESUMO

Environmental pollution caused by plastic waste is a growing global problem. Discarded plastic products and debris (microplastic particles) in the oceans detrimentally affect marine ecosystems and may impact human. Humans are exposed to plastic debris via the consumption of seafood and drinking water, contact with food packaging, or inhalation of particles. The accumulation of microplastic particles in humans has potential health risks such as cytotoxicity, hypersensitivity, unwanted immune response, and acute response like hemolysis. We investigated the cellular responses of secondary polypropylene microplastics (PP particles) of approximately ~20 µm and 25-200 µm in different condition and size to normal cells, immune cells, blood cells, and murine immune cells by cytokine analysis, ROS assay, polarization assay and proliferation assay. We found that PP particles showed low cytotoxicity effect in size and concentration manner, however, a high concentration, small sized, DMSO method of PP particles stimulated the immune system and enhanced potential hypersensitivity to PP particles via an increase in the levels of cytokines and histamines in PBMCs, Raw 264.7 and HMC-1 cells.


Assuntos
Leucócitos/efeitos dos fármacos , Material Particulado/toxicidade , Polipropilenos/toxicidade , Poluentes Químicos da Água/toxicidade , Animais , Linhagem Celular , Humanos , Camundongos , Tamanho da Partícula , Células RAW 264.7
20.
Int J Food Microbiol ; 304: 89-96, 2019 Sep 02.
Artigo em Inglês | MEDLINE | ID: mdl-31176964

RESUMO

The environmental stability of enteric viruses and resistance to conventional treatments and common disinfectants, leads to their persistence in waters and food, causing serious implications on public health. Among non-thermal treatment methods, ionizing radiation is recognized as a useful and effective mean of disinfection. The objective of this study was to estimate the inactivation of enteric virus by gamma radiation in raw berry fruits, in order to evaluate the potential of this technology to be applied as a disinfection treatment. Fresh strawberries and raspberries were inoculated either individually with murine norovirus type 1 (MuNoV; as a human norovirus surrogate) and human adenovirus type 5 (HAdV) or with a viral pool of both viruses, and irradiated in a Co-60 equipment at doses of 1 kGy up to 11 kGy. The infectivity of viral particles of MuNoV and HAdV was assessed by plaque assay using Raw 264.7 and A549 cells, respectively. A 2 log PFU/g reduction on MuNoV and HAdV titers was obtained after treatment with a dose of 4 kGy for both fruits. However, non-linear inactivation survival curves were obtained for MuNoV and HAdV in fresh fruits, leading to the detection of infective viral particles at a dose of 11 kGy. The irradiation process indicated virucidal potential, although the estimated gamma radiation dose to attain food safety (> 7 kGy) would compromise the preservation of food quality. Nevertheless, the irradiation technology could be an effective virus mitigation tool to treat polluted waters, which are a major vehicle of contamination for fresh produce.


Assuntos
Adenovírus Humanos/efeitos da radiação , Desinfecção/métodos , Doenças Transmitidas por Alimentos/prevenção & controle , Fragaria/virologia , Raios gama , Norovirus/efeitos da radiação , Rubus/virologia , Células A549 , Animais , Linhagem Celular , Doenças Transmitidas por Alimentos/virologia , Frutas/virologia , Humanos , Camundongos , Células RAW 264.7
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