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ABSTRACT: Although it is caused by a single-nucleotide mutation in the ß-globin gene, sickle cell anemia (SCA) is a systemic disease with complex, incompletely elucidated pathologies. The mononuclear phagocyte system plays critical roles in SCA pathophysiology. However, how heterogeneous populations of hepatic macrophages contribute to SCA remains unclear. Using a combination of single-cell RNA sequencing and spatial transcriptomics via multiplexed error-robust fluorescence in situ hybridization, we identified distinct macrophage populations with diversified origins and biological functions in SCA mouse liver. We previously found that administering the von Willebrand factor (VWF)-cleaving protease ADAMTS13 alleviated vaso-occlusive episode in mice with SCA. Here, we discovered that the ADAMTS13-cleaved VWF was cleared from the circulation by a Clec4f+Marcohigh macrophage subset in a desialylation-dependent manner in the liver. In addition, sickle erythrocytes were phagocytized predominantly by Clec4f+Marcohigh macrophages. Depletion of macrophages not only abolished the protective effect of ADAMTS13 but exacerbated vaso-occlusive episode in mice with SCA. Furthermore, promoting macrophage-mediated VWF clearance reduced vaso-occlusion in SCA mice. Our study demonstrates that hepatic macrophages are important in the pathogenesis of SCA, and efficient clearance of VWF by hepatic macrophages is critical for the protective effect of ADAMTS13 in SCA mice.
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Anemia Falciforme , Doenças Vasculares , Camundongos , Animais , Fator de von Willebrand/genética , Hibridização in Situ Fluorescente , Anemia Falciforme/patologia , Macrófagos/patologia , Proteína ADAMTS13/genéticaRESUMO
BACKGROUND: Integrin-regulated monocyte recruitment and cellular responses of monocyte-derived macrophages are critical for the pathogenesis of atherosclerosis. In the canonical model, talin1 controls ligand binding to integrins, a prerequisite for integrins to mediate leukocyte recruitment and induce immune responses. However, the role of talin1 in the development of atherosclerosis has not been studied. Our study investigated how talin1 in myeloid cells regulates the progression of atherosclerosis. METHODS: On an Apoe-/- background, myeloid talin1-deficient mice and the control mice were fed with a high-fat diet for 8 or 12 weeks to induce atherosclerosis. The atherosclerosis development in the aorta and monocyte recruitment into atherosclerotic lesions were analyzed. RESULTS: Myeloid talin1 deletion facilitated the formation of atherosclerotic lesions and macrophage deposition in lesions. Talin1 deletion abolished integrin ß2-mediated adhesion of monocytes but did not impair integrin α4ß1-dependent cell adhesion in a flow adhesion assay. Strikingly, talin1 deletion did not prevent Mn2+- or chemokine-induced activation of integrin α4ß1 to the high-affinity state for ligands. In an in vivo competitive homing assay, monocyte infiltration into inflamed tissues was prohibited by antibodies to integrin α4ß1 but was not affected by talin1 deletion or antibodies to integrin ß2. Furthermore, quantitative polymerase chain reaction and ELISA (enzyme-linked immunosorbent assay) analysis showed that macrophages produced cytokines to promote inflammation and the proliferation of smooth muscle cells. Ligand binding to integrin ß3 inhibited cytokine generation in macrophages, although talin1 deletion abolished the negative effects of integrin ß3. CONCLUSIONS: Integrin α4ß1 controls monocyte recruitment during atherosclerosis. Talin1 is dispensable for integrin α4ß1 activation to the high-affinity state and integrin α4ß1-mediated monocyte recruitment. Yet, talin1 is required for integrin ß3 to inhibit the production of inflammatory cytokines in macrophages. Thus, intact monocyte recruitment and elevated inflammatory responses cause enhanced atherosclerosis in talin1-deficient mice. Our study provides novel insights into the roles of myeloid talin1 and integrins in the progression of atherosclerosis.
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Aterosclerose , Adesão Celular , Modelos Animais de Doenças , Macrófagos , Camundongos Endogâmicos C57BL , Camundongos Knockout para ApoE , Células Mieloides , Talina , Animais , Talina/metabolismo , Talina/genética , Aterosclerose/genética , Aterosclerose/patologia , Aterosclerose/metabolismo , Células Mieloides/metabolismo , Células Mieloides/patologia , Macrófagos/metabolismo , Doenças da Aorta/patologia , Doenças da Aorta/genética , Doenças da Aorta/metabolismo , Doenças da Aorta/imunologia , Doenças da Aorta/prevenção & controle , Masculino , Antígenos CD18/metabolismo , Antígenos CD18/genética , Integrina alfa4beta1/metabolismo , Integrina alfa4beta1/genética , Monócitos/metabolismo , Monócitos/imunologia , Placa Aterosclerótica , Camundongos , Células Cultivadas , Aorta/patologia , Aorta/metabolismo , Transdução de SinaisRESUMO
Microvascular thrombosis in patients with thrombotic thrombocytopenic purpura (TTP) is initiated by GPIbα-mediated platelet binding to von Willebrand factor (VWF). Binding of VWF to GPIbα causes activation of the platelet surface integrin αIIbß3. However, the mechanism of GPIbα-initiated activation of αIIbß3 and its clinical importance for microvascular thrombosis remain elusive. Deletion of platelet C-type lectin-like receptor 2 (CLEC-2) did not prevent VWF binding to platelets but specifically inhibited platelet aggregation induced by VWF binding in mice. Deletion of platelet CLEC-2 also inhibited αIIbß3 activation induced by the binding of VWF to GPIbα. Using a mouse model of TTP, which was created by infusion of anti-mouse ADAMTS13 monoclonal antibodies followed by infusion of VWF, we found that deletion of platelet CLEC-2 decreased pulmonary arterial thrombosis and the severity of thrombocytopenia. Importantly, prophylactic oral administration of aspirin, an inhibitor of platelet activation, and therapeutic treatment of the TTP mice with eptifibatide, an integrin αIIbß3 antagonist, reduced pulmonary arterial thrombosis in the TTP mouse model. Our observations demonstrate that GPIbα-mediated activation of integrin αIIbß3 plays an important role in the formation of thrombosis in TTP. These observations suggest that prevention of platelet activation with aspirin may reduce the risk for thrombosis in patients with TTP.
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Hipertensão Pulmonar , Púrpura Trombocitopênica Trombótica , Trombose , Aspirina , Plaquetas/metabolismo , Humanos , Lectinas Tipo C/genética , Lectinas Tipo C/metabolismo , Ativação Plaquetária , Complexo Glicoproteico GPIIb-IIIa de Plaquetas/metabolismo , Púrpura Trombocitopênica Trombótica/metabolismo , Trombose/etiologia , Fator de von Willebrand/metabolismoRESUMO
We previously demonstrated that fucoidan with a type II structure inhibited postprandial hyperglycemia by suppressing glucose uptake, but the mechanism remains elusive. Here, we aimed to assess whether the effect of glucose absorption inhibition was related to the basic structure of fucoidans and preliminarily clarified the underlying mechanism. Fucoidans with type II structure and type I structure were prepared from Ascophyllumnodosum (AnF) or Laminariajaponica (LjF) and Kjellmaniellacrassifolia (KcF), respectively. The effects of various fucoidans on suppressing postprandial hyperglycemia were investigated using in vitro (Caco-2 monolayer model), semi-in vivo (everted gut sac model), and in vivo (oral glucose tolerance test, OGTT) assays. The results showed that only AnF with a type II structure, but not LjF or KcF with type I structure, could inhibit the glucose transport in the Caco-2 monolayer and everted gut sac models. A similar result was seen in the OGTT of Kunming mice and leptin receptor-deficient (db/db) mice, where only AnF could effectively inhibit glucose transport into the bloodstream. Furthermore, AnF (400 mg/kg/d) treatment decreased the fasting blood glucose, HbA1c, and fasting insulin levels, while increasing the serum glucagon-like peptide-1 (GLP-1) level in obese leptin receptor-deficient (db/db) mice. Furthermore, surface plasmon resonance (SPR) analysis revealed the specific binding of AnF to Na+/glucose cotransporter 1 (SGLT1), which indicated the effect of AnF on postprandial hyperglycemia could be due to its suppression on SGLT1 activity. Taken together, this study suggests that AnF with a type II structure can be a promising candidate for hyperglycemia treatment.
Assuntos
Ascophyllum/química , Hiperglicemia/prevenção & controle , Polissacarídeos/farmacologia , Transportador 1 de Glucose-Sódio/antagonistas & inibidores , Animais , Glicemia/metabolismo , Células CACO-2 , Glucose/metabolismo , Teste de Tolerância a Glucose , Humanos , Laminaria/química , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Phaeophyceae/química , Polissacarídeos/isolamento & purificaçãoRESUMO
Recent studies have reported that dietary fiber improved metabolic syndrome (MetS). However, the effects of fucoidans on MetS were still not clear. In this study, we evaluated the activity of fucoidan from Fucus vesiculosus (FvF) on attenuating MetS and first elucidated the underlying mechanism. In vitro, FvF treatment remarkably lowered the level of reactive oxygen species (ROS) compared with the sodium palmitate (PA)-induced insulin resistance (IR) group. The phosphorylation level of c-Jun N-terminal kinase (JNK) was significantly decreased, while phosphorylation of protein kinase B (pAkt) level increased, compared with that of the HepG2 cells treated with PA. Thus, FvF increased glucose consumption and relieved IR via ROS-mediated JNK and Akt signaling pathways. In addition, these changes were accompanied by the activation of adenosine 5'-monophosphate-ativated protein kinase (AMPK) and its downstream targets (e.g., HMG-CoA reductase (HMGCR), acetyl-CoA carboxylase (ACC), and sterol-regulatory element-binding protein-1c (SREBP-1C)), which improved lipid metabolism in IR HepG2 cells. In vivo, FvF improved hyperglycemia and decreased serum insulin level in mice with MetS. Furthermore, we evaluated the inhibition of glucose transport by in vitro (Caco-2 monolayer model), semi-in vivo (everted gut sac model) and oral glucose tolerance test (OGTT), which indicated that FvF could significantly reduce the absorption of glucose into the blood stream, thus it could improve blood-glucose levels and IR in mice with MetS. Moreover, FvF decreased serum triglyceride (TG), total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C) levels and liver lipid accumulation, while increased the serum high density lipoprotein cholesterol (HDL-C) level in mice with MetS. Therefore, FvF could be considered as a potential candidate for the treatment of MetS by alleviating IR, inhibiting glucose transportation, and regulating lipid metabolism.
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Fucus/química , Síndrome Metabólica/tratamento farmacológico , Síndrome Metabólica/metabolismo , Polissacarídeos/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Proteínas Quinases Ativadas por AMP/metabolismo , Animais , Glicemia/metabolismo , Peso Corporal/efeitos dos fármacos , Modelos Animais de Doenças , Células Hep G2 , Humanos , Resistência à Insulina , Metabolismo dos Lipídeos/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Masculino , Camundongos , Estresse Oxidativo/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais/efeitos dos fármacosRESUMO
As an important glycosaminoglycan, keratan sulfate (KS) mainly exists in corneal and cartilage, possessing various biological activities. In this study, we purified KS from blue shark (Prionace glauca) cartilage and prepared KS oligosaccharides (KSO) through keratanase II-catalyzed hydrolysis. The structures of KS and KSO were characterized using multi-dimensional nuclear magnetic resonance (NMR) spectra and liquid chromatography-mass spectrometry (LC-MS). Shark cartilage KS was highly sulfated and modified with ~2.69% N-acetylneuraminic acid (NeuAc) through α(2,3)-linked to galactose. Additionally, KS exhibited binding affinity to Ricinus communis agglutinin I (RCA120) in a concentration-dependent manner, a highly toxic lectin from beans of the castor plant. Furthermore, KSO from dp2 to dp8 bound to RCA120 in the increasing trend while the binding affinity of dp8 was superior to polysaccharide. These results define novel structural features for KS from Prionace glauca cartilage and demonstrate the potential application on ricin-antidote exploitation.
Assuntos
Cartilagem/química , Sulfato de Queratano/química , Lectinas de Plantas/química , Tubarões/metabolismo , Acetilglucosaminidase/química , Animais , Cromatografia Líquida , Galactose/química , Hidrólise/efeitos dos fármacos , Espectroscopia de Ressonância Magnética/métodos , Oligossacarídeos/química , Espectrometria de Massas em Tandem/métodosRESUMO
An apigalacturonan (AGA)-rich polysaccharide, ZCMP, was isolated from the sea grass Zostera caespitosa Miki. The depolymerized fragments derived from ZCMP were obtained by either acidic degradation or pectinase degradation, and their structures were characterized by electrospray ionization collision-induced-dissociation mass spectrometry (ESI-CID-MS2) and nuclear magnetic resonance (NMR) spectroscopy. The average molecular weight of ZCMP was 77.2 kD and it consisted of galacturonic acid (GalA), apiosefuranose (Api), galactose (Gal), rhamnose (Rha), arabinose (Ara), xylose (Xyl), and mannose (Man), at a molar ratio of 51.4ê15.5ê6.0ê11.8ê4.2ê4.4ê4.2. There were two regions of AGA (70%) and rhamnogalacturonan-I (RG-Ι, 30%) in ZCMP. AGA was composed of an α-1,4-D-galactopyranosyluronan backbone mainly substituted at the O-3 position by single Api residues. RG-Ι possessed a backbone of repeating disaccharide units of â4GalAα1,2Rhaα1â, with a few α-L-arabinose and ß-D-galactose residues as side chains. The anti-angiogenesis assay showed that ZCMP inhibited the migratory activity of human umbilical vein endothelial cell (HUVECs), with no influence on endothelial cells growth. ZCMP also promoted macrophage phagocytosis. These findings of the present study demonstrated the potential anti-tumor activity of ZCMP through anti-angiogenic and immunoregulatory pathways.
Assuntos
Inibidores da Angiogênese/farmacologia , Polissacarídeos/farmacologia , Zosteraceae/química , Inibidores da Angiogênese/química , Inibidores da Angiogênese/isolamento & purificação , Animais , Antineoplásicos/química , Antineoplásicos/isolamento & purificação , Antineoplásicos/farmacologia , Linhagem Celular , Células Endoteliais da Veia Umbilical Humana , Humanos , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Espectroscopia de Ressonância Magnética , Camundongos , Fagocitose/efeitos dos fármacos , Polissacarídeos/química , Polissacarídeos/isolamento & purificação , Espectrometria de Massas por Ionização por ElectrosprayRESUMO
BACKGROUND & AIMS: Addition of sialic acids (sialylation) to glycoconjugates is a common capping step of glycosylation. Our study aims to determine the roles of the overall sialylation in intestinal mucosal homeostasis. METHODS: Mice with constitutive deletion of intestinal epithelial sialylation (IEC Slc35a1-/- mice) and mice with inducible deletion of sialylation in intestinal epithelium (TM-IEC Slc35a1-/- mice) were generated, which were used to determine the roles of overall sialylation in intestinal mucosal homeostasis by ex vivo and muti-omics studies. RESULTS: IEC Slc35a1-/- mice developed mild spontaneous microbiota-dependent colitis. Additionally, 30% of IEC Slc35a1-/- mice had spontaneous tumors in the rectum over the age of 12 months. TM-IEC Slc35a1-/- mice were highly susceptible to acute inflammation induced by 1% DSS vs controls. Loss of total sialylation was associated with reduced mucus thickness on fecal sections and within colon tissues. TM-IEC Slc35a1-/- mice showed altered microbiota with an increase in Clostridia disporicum, which is associated a global reduction in the abundance of at least 20 unique taxa; however, metabolomic analysis did not show any significant differences in short-chain fatty acid levels. Treatment with 5-fluorouracil (5-FU) led to more severe small intestine mucositis in the IEC Slc35a1-/- mice vs. WT littermates, which was associated with reduced Lgr5+ cell representation in small intestinal crypts in IEC Slc35a1-/-;Lgr5-GFP mice. CONCLUSIONS: Loss of overall sialylation impairs mucus stability and the stem cell niche leading to microbiota-dependent spontaneous colitis and tumorigenesis.
RESUMO
Although most cell membrane proteins are modified by glycosylation, our understanding of the role and actions of protein glycosylation is still very limited. ß1,3galactosyltransferase (C1GalT1) is a key glycosyltransferase that controls the biosynthesis of the Core 1 structure of O-linked mucin type glycans and is overexpressed by many common types of epithelial cancers. This study reports that suppression of C1GalT1 expression in human colon cancer cells caused substantial changes of protein glycosylation of cell membrane proteins, many of which were ligands of the galactoside-binding galectin-3 and the macrophage galactose-type lectin (MGL). This led to significant reduction of cancer cell proliferation, adhesion, migration and the ability of tumour cells to form colonies. Crucially, C1GalT1 suppression significantly reduced galectin-3-mediated tumour cell-cell interaction and galectin-3-promoted tumour cell activities. In the meantime, C1GalT1 suppression substantially increased MGL-mediated macrophage-tumour cell interaction and macrophage-tumour cell phagocytosis and cytokine secretion. C1GalT1-expressing cancer cells implanted in chick embryos resulted in the formation of significantly bigger tumours than C1GalT1-suppressed cells and the presence of galectin-3 increased tumour growth of C1GalT1-expressing but not C1GalT1-suppressed cells. More MGL-expressing macrophages and dendritic cells were seen to be attracted to the tumour microenvironment in ME C1galt1-/-/Erb mice than in C1galt1f/f /Erb mice. These results indicate that expression of C1GalT1 by tumour cells reciprocally controls tumour cell-cell and tumour-macrophage interactions mediated by galectin-3 and MGL with double impact on cancer development and progression. C1GalT1 overexpression in epithelial cancers therefore may represent a fundamental mechanism in cancer promotion and in reduction of immune response/surveillance in cancer progression.
Assuntos
Neoplasias do Colo , Galectina 3 , Embrião de Galinha , Humanos , Animais , Camundongos , Galectina 3/genética , Galactose , Neoplasias do Colo/genética , Glicosilação , Macrófagos , Microambiente TumoralRESUMO
Core 1-derived mucin-type O-glycans (O-glycans) are a major component of gastric mucus with an unclear role. To address this, we generated mice lacking gastric epithelial O-glycans (GEC C1galt1-/-). GEC C1galt1-/- mice exhibited spontaneous gastritis that progressed to adenocarcinoma with â¼80% penetrance by 1 yr. GEC C1galt1-/- gastric epithelium exhibited defective expression of a major mucus forming O-glycoprotein Muc5AC relative to WT controls, which was associated with impaired gastric acid homeostasis. Inflammation and tumorigenesis in GEC C1galt1-/- stomach were concurrent with activation of caspases 1 and 11 (Casp1/11)-dependent inflammasome. GEC C1galt1-/- mice genetically lacking Casp1/11 had reduced gastritis and gastric cancer progression. Notably, expression of Tn antigen, a truncated form of O-glycan, and CASP1 activation was associated with tumor progression in gastric cancer patients. These results reveal a critical role of O-glycosylation in gastric homeostasis and the protection of the gastric mucosa from Casp1-mediated gastric inflammation and cancer.
Assuntos
Gastrite/metabolismo , Mucinas/metabolismo , Polissacarídeos/metabolismo , Neoplasias Gástricas/metabolismo , Animais , Antígenos Glicosídicos Associados a Tumores/metabolismo , Carcinogênese/metabolismo , Caspase 1/metabolismo , Feminino , Mucosa Gástrica/metabolismo , Glicosilação , Homeostase/fisiologia , Humanos , Inflamação/metabolismo , Masculino , Camundongos , Muco/metabolismo , Neoplasias/metabolismoRESUMO
Colon mucus segregates the intestinal microbiota from host tissues, but how it organizes to function throughout the colon is unclear. In mice, we found that colon mucus consists of two distinct O-glycosylated entities of Muc2: a major form produced by the proximal colon, which encapsulates the fecal material including the microbiota, and a minor form derived from the distal colon, which adheres to the major form. The microbiota directs its own encapsulation by inducing Muc2 production from proximal colon goblet cells. In turn, O-glycans on proximal colon-derived Muc2 modulate the structure and function of the microbiota as well as transcription in the colon mucosa. Our work shows how proximal colon control of mucin production is an important element in the regulation of host-microbiota symbiosis.
Assuntos
Colo/metabolismo , Colo/microbiologia , Microbioma Gastrointestinal , Mucina-2/metabolismo , Muco/metabolismo , Animais , Fezes/microbiologia , Glicosilação , Camundongos , Camundongos Knockout , Mucina-2/genética , Transcrição GênicaRESUMO
Ulcerative colitis (UC) is a chronic inflammatory bowel disease characterized by defective intestinal barrier integrity toward the microbiota and epithelial damage. Double cortin-like kinase 1 (Dclk1), a marker of intestinal tuft cells, can regulate tissue regenerative responses, but its role in epithelial repair during bacterial-dependent chronic colitis is unclear. We addressed this question using our recently developed mouse model of spontaneous microbiota-dependent colitis induced by mucin-type O-glycan deficiency (DKO), which recapitulates most features of human UC. We generated DKO mice lacking intestinal epithelial Dclk1 (DKO;Dclk1ΔIEC) and analyzed colitis onset and severity using clinical and histologic indices, immune responses by qPCR and immunostaining, and epithelial responses using proliferation markers and organoid culture. We found 3-4-week-old DKO;Dclk1ΔIEC mice developed worsened spontaneous colitis characterized by reduced body weight, loose stool, severe colon thickening, epithelial lesions, and inflammatory cell infiltrates compared with DKO mice. The primary defect was an impaired epithelial proliferative response during inflammation. Dclk1 deficiency also reduced inflammation-induced proliferation and growth of colon organoids ex vivo. Mechanistically, Dclk1 expression was important for inflammation-induced Cox2 expression and prostaglandin E2 (PGE2) production in vivo, and PGE2 rescued proliferative defects in Dclk1-deficient colonic organoids. Although tuft cells were expanded in both DKO and DKO;Dclk1ΔIEC relative to WT mice, loss of Dclk1 was associated with reduced tuft cell activation (i.e., proliferation) during inflammation. Similar results were found in DKO vs. DKO;Dclk1ΔIEC mice at 3-6 months of age. Our results support that tuft cells, via Dclk1, are important responders to bacterial-induced colitis by enhancing epithelial repair responses, which in turn limits bacterial infiltration into the mucosa.
Assuntos
Apoptose/genética , Colite/genética , Inflamação/genética , Proteínas Serina-Treonina Quinases/genética , Animais , Proliferação de Células/genética , Doença Crônica/epidemiologia , Doença Crônica/prevenção & controle , Colite/metabolismo , Colite/patologia , Colo/metabolismo , Colo/patologia , Modelos Animais de Doenças , Quinases Semelhantes a Duplacortina , Células Epiteliais/metabolismo , Células Epiteliais/patologia , Humanos , Inflamação/patologia , Mucosa Intestinal/metabolismo , Mucosa Intestinal/patologia , Camundongos , Camundongos Knockout , Transdução de Sinais/genéticaRESUMO
Correction for 'Dietary fucoidan modulates the gut microbiota in mice by increasing the abundance of Lactobacillus and Ruminococcaceae' by Qingsen Shang et al., Food Funct., 2016, 7, 3224-3232.
RESUMO
The sulfated polysaccharide NP2 was isolated and purified from Nemacystus decipiens, the structure and antithrombotic activity of NP2 was further studied. NP2 was composed of fucose, glucuronic acid, galactose and xylose at molar ratios of 76.3:20.5:1.5:1.7. ES-CID-MS/MS results showed that NP2 had a backbone of α (1â¯ââ¯3)-linked fucose and a branch was composed of Fuc-(2â¯ââ¯1)-GlcA, which was agree with the results of NMR and methylation analysis. The results also show that the sulfate groups were substituted at the C2 or C4 positions of the fucose residues. In addition, analysis of the antithrombotic activity results indicated that NP2 can increase the percentage of t-PA/PAI-1, thereby suggesting that NP2 has high fibrinolytic activity and should be explored as a novel antithrombotic agent.
Assuntos
Fibrinolíticos/química , Fibrinolíticos/farmacologia , Phaeophyceae/química , Polissacarídeos/química , Polissacarídeos/farmacologia , Metilação , Inibidor 1 de Ativador de Plasminogênio/metabolismo , Ativador de Plasminogênio Tecidual/metabolismoRESUMO
We studied the mechanisms underlying the immunostimulatory effects of aß-1,3/1,6-glucan (BG136) from Durvillaea Antarctica. Our data showed that BG136 promoted the activation of MAPKs and NF-κB signaling pathways and cytokines production. BG136 did not increase MCP-1 or NO production or phosphorylation of NF-κB and MAPK in TLR4 siRNA knockdown cells, indicating that BG136 activates macrophages through TLR4. Flow cytometry analysis and confocal experiment showed that BG136 bound to TLR4 expressed on RAW264.7 macrophage cells surface. The affinity of BG136 for TLR4 was determined using Surface Plasmon Resonance (SPR) (KD: 4.51â¯×â¯10-6M). Altogether, our results showed that BG136 activates RAW264.7 cells by binding to TLR4 and then triggering TLR4-mediated signaling pathways to promote cytokines secretion.
Assuntos
Fatores Imunológicos/farmacologia , Macrófagos/efeitos dos fármacos , Phaeophyceae/química , Receptor 4 Toll-Like/metabolismo , beta-Glucanas/farmacologia , Animais , Sobrevivência Celular/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Técnicas de Silenciamento de Genes , Fatores Imunológicos/química , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Macrófagos/citologia , Macrófagos/imunologia , Macrófagos/metabolismo , Camundongos , NF-kappa B/metabolismo , Óxido Nítrico/biossíntese , Óxido Nítrico Sintase Tipo II/biossíntese , Células RAW 264.7 , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Receptor 4 Toll-Like/deficiência , Receptor 4 Toll-Like/genética , beta-Glucanas/químicaRESUMO
Carrageenan as a food additive has been used for years. However, controversy exists regarding to the safety of carrageenan and accumulating evidence indicates that it could induce colitis in experimental models. Here, to provide more information on this issue and solve the debate, we studied and compared in detail the toxic effects of different isomers of carrageenan (κ-, ι-, and λ-) on the colon of C57BL/6J mice. Interestingly, all isomers of carrageenan were found to induce colitis with a comparable activity. Given that carrageenan is unabsorbed after oral administration, and also in light of the fact that gut microbiota plays a pivotal role in the pathogenesis of colitis, we further investigated the effect of carrageenan on gut microbiota using high-throughput sequencing. Intriguingly, carrageenan-induced colitis was observed to be robustly correlated with changes in the composition of gut microbiota. Specifically, all carrageenans significantly decreased the abundance of a potent anti-inflammatory bacterium, Akkermansia muciniphila, in the gut, which is highly relevant for understanding the toxic effect of carrageenan. Altogether, our results corroborate previous studies demonstrating harmful gastrointestinal effect of carrageenan and, from a gut microbiota perspective, shed new light into the mechanism by which carrageenan induces colitis in experimental animals.
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Carragenina , Colite/microbiologia , Colo/microbiologia , Microbioma Gastrointestinal , Verrucomicrobia/crescimento & desenvolvimento , Animais , Carga Bacteriana , Colite/sangue , Colite/induzido quimicamente , Colo/metabolismo , Biologia Computacional , DNA Bacteriano/genética , Modelos Animais de Doenças , Sequenciamento de Nucleotídeos em Larga Escala , Interleucina-10/sangue , Interleucina-1beta/sangue , Interleucina-6/sangue , Camundongos Endogâmicos C57BL , Análise de Sequência de DNA/métodos , Fator de Necrose Tumoral alfa/sangue , Verrucomicrobia/classificação , Verrucomicrobia/genéticaRESUMO
Development of novel anti-influenza A virus (IAV) drugs with high efficiency and low toxicity is critical for preparedness against influenza outbreaks. Herein, we investigated the anti-IAV activities and mechanisms of fucoidan in vitro and in vivo. The results showed that a fucoidan KW derived from brown algae Kjellmaniella crassifolia effectively blocked IAV infection in vitro with low toxicity. KW possessed broad anti-IAV spectrum and low tendency of induction of viral resistance, superior to the anti-IAV drug amantadine. KW was capable of inactivating virus particles before infection and blocked some stages after adsorption. KW could bind to viral neuraminidase (NA) and inhibit the activity of NA to block the release of IAV. KW also interfered with the activation of EGFR, PKCα, NF-κB, and Akt, and inhibited both IAV endocytosis and EGFR internalization in IAV-infected cells, suggesting that KW may also inhibit cellular EGFR pathway. Moreover, intranasal administration of KW markedly improved survival and decreased viral titers in IAV-infected mice. Therefore, fucoidan KW has the potential to be developed into a novel nasal drop or spray for prevention and treatment of influenza in the future.
Assuntos
Receptores ErbB/metabolismo , Vírus da Influenza A/efeitos dos fármacos , Vírus da Influenza A/fisiologia , Neuraminidase/metabolismo , Polissacarídeos/farmacologia , Transdução de Sinais/efeitos dos fármacos , Replicação Viral/efeitos dos fármacos , Animais , Antivirais/farmacologia , Efeito Citopatogênico Viral , Cães , Farmacorresistência Viral , Endocitose/efeitos dos fármacos , Células Madin Darby de Rim Canino , Camundongos , Infecções por Orthomyxoviridae/metabolismo , Infecções por Orthomyxoviridae/virologia , Polissacarídeos/químicaRESUMO
The purpose of this study was to develop a promising wound dressing. Though chitosan cross-linked with genipin has been widely used as biomaterials, with the addition of partially oxidized Bletilla striata polysaccharide, the newly developed material in this study (coded as CSGB) showed less gelling time, more uniform aperture distribution, higher water retention, demanded mechanical strength and more L929 cell proliferation compared to the chitosan cross-linked only with genipin. Owning to partial blocking of free amino groups of chitosan, CSGB revealed almost no antibacterial activities, thus the bilayer composite of chitosan-silver nanoparticles (CS-AgG) on CSGB was designed to inhibit microbial invasion. The in vivo studies indicated that both CSGB and bilayer wound dressing significantly accelerated the healing rate of cutaneous wounds in mice, and the bilayer exhibited better mature epidermization with less inflammatory cells on Day 7. Therefore, this novel bilayer composite has great potential in wound dressing applications.
Assuntos
Bandagens , Quitosana/química , Nanopartículas Metálicas , Polissacarídeos/química , Cicatrização , Animais , Camundongos , PrataRESUMO
Recently, fucoidan has been proposed as a potential prebiotic agent for functional food and pharmaceutical development. However, while previous studies illustrated favorable modulations of gut microbiota by fucoidan, changes in the overall microbial structure remain elusive. In the present study, modulations of gut microbiota by different fucoidans were studied using high throughput sequencing and bioinformatics analysis. We found that at the expense of opportunistic pathogenic bacteria such as Peptococcus, the abundance of beneficial bacteria including Lactobacillus and Ruminococcaceae was significantly increased in response to fucoidan treatment. Besides, by maintaining a more balanced composition of gut microbiota, dietary fucoidan also significantly reduced the antigen load and the inflammatory response in the host as evidenced by the decreased serum lipopolysaccharide-binding protein levels. Collectively, our results indicate that fucoidan can be used as a gut microbiota modulator for health promotion and treatment of intestinal dysbiosis.
Assuntos
Clostridiales/crescimento & desenvolvimento , Microbioma Gastrointestinal , Lactobacillus/crescimento & desenvolvimento , Polissacarídeos/administração & dosagem , Prebióticos/administração & dosagem , Animais , Biologia Computacional , Alimento Funcional , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Peptococcus/patogenicidadeRESUMO
The inhibition of α-glucosidase is an effective therapeutic approach for type 2 diabetes mellitus that involves decreasing postprandial hyperglycemia. In the present study, the α-glucosidase and α-amylase inhibitory effects of 11 fucoidans extracted from different brown seaweeds were evaluated. Although no significant α-amylase inhibition was observed, fucoidan from Fucus vesiculosus (FvF) showed the highest α-glucosidase inhibitory activity, with an IC50 value of 67.9 µg/mL. In addition, FvF at a concentration of 200 µg/mL displayed very mild cytotoxicity to IEC-6 cells as indicated by the MTT assay. An in vivo study indicated that FvF decreased the fasting blood glucose and hemoglobin A1c (HbA1c) levels of db/db mice, with minimal effect on their weight. Therefore, our present in vitro and in vivo studies demonstrated that FvF could be a promising α-glucosidase inhibitor for the treatment of type 2 diabetes mellitus.