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1.
Proc Natl Acad Sci U S A ; 118(10)2021 03 09.
Artículo en Inglés | MEDLINE | ID: mdl-33658363

RESUMEN

Blood pH is tightly maintained between 7.35 and 7.45, and acidosis (pH <7.3) indicates poor prognosis in sepsis, wherein lactic acid from anoxic tissues overwhelms the buffering capacity of blood. Poor sepsis prognosis is also associated with low zinc levels and the release of High mobility group box 1 (HMGB1) from activated and/or necrotic cells. HMGB1 added to whole blood at physiological pH did not bind leukocyte receptors, but lowering pH with lactic acid to mimic sepsis conditions allowed binding, implying the presence of natural inhibitor(s) preventing binding at normal pH. Testing micromolar concentrations of divalent cations showed that zinc supported the robust binding of sialylated glycoproteins with HMGB1. Further characterizing HMGB1 as a sialic acid-binding lectin, we found that optimal binding takes place at normal blood pH and is markedly reduced when pH is adjusted with lactic acid to levels found in sepsis. Glycan array studies confirmed the binding of HMGB1 to sialylated glycan sequences typically found on plasma glycoproteins, with binding again being dependent on zinc and normal blood pH. Thus, HMGB1-mediated hyperactivation of innate immunity in sepsis requires acidosis, and micromolar zinc concentrations are protective. We suggest that the potent inflammatory effects of HMGB1 are kept in check via sequestration by plasma sialoglycoproteins at physiological pH and triggered when pH and zinc levels fall in late stages of sepsis. Current clinical trials independently studying zinc supplementation, HMGB1 inhibition, or pH normalization may be more successful if these approaches are combined and perhaps supplemented by infusions of heavily sialylated molecules.


Asunto(s)
Acidosis/sangre , Proteína HMGB1/sangre , Sepsis/sangre , Sialoglicoproteínas/sangre , Zinc/sangre , Acidosis/inmunología , Acidosis/metabolismo , Acidosis/patología , Proteínas Portadoras , Proteína HMGB1/farmacología , Humanos , Concentración de Iones de Hidrógeno , Inmunidad Innata , Lipopolisacáridos/farmacología , Polisacáridos/química , Sepsis/inmunología , Sepsis/patología , Ácidos Siálicos/química , Sialoglicoproteínas/química , Zinc/metabolismo
2.
Food Chem ; 349: 129164, 2021 Jul 01.
Artículo en Inglés | MEDLINE | ID: mdl-33550022

RESUMEN

Premna microphylla turcz leaf juice with polysaccharides (PMPs) as its main component, are raw material of jelly-like Chinese traditional food "Guanyin tofu", which were also experiencedly used to relieve inflammation-related symptoms. Here three kinds of PMPs were extracted in alkaline (APMP), water (WPMP) and acidic (HPMP) conditions, being characteristic of RG I, high- and low-methoxyl HG pectin, respectively, in amorphous form with diverse surface microstructures, among which APMP predominantly composed of Glucose instead of galacturonic acid, showing wider molecular weight distribution and more branched chains. PMPs showed remarkable radical scavenging capability, and especially APMP at concentrations above 50 µg/mL effectively inhibited the reactive oxygen species and malondialdehyde production in LPS-stimulated RAW 264.7 macrophages, by enhancing enzymatic activities of endogenous superoxide dismutase, glutathione peroxidase and catalase, and accordingly alleviated inflammatory cytokines. Thus, PMPs could be promising non-toxic natural dietary supplement to improve chronic inflammation-induced diseases.


Asunto(s)
Antiinflamatorios/farmacología , Antioxidantes/farmacología , Lamiaceae/química , Macrófagos/efectos de los fármacos , Pectinas/farmacología , Hojas de la Planta/química , Animales , Antiinflamatorios/química , Antioxidantes/química , Citocinas/metabolismo , Lipopolisacáridos/farmacología , Macrófagos/citología , Macrófagos/enzimología , Macrófagos/metabolismo , Ratones , Pectinas/química , Células RAW 264.7 , Especies Reactivas de Oxígeno/metabolismo
3.
Life Sci ; 270: 119138, 2021 Apr 01.
Artículo en Inglés | MEDLINE | ID: mdl-33524422

RESUMEN

AIMS: Sodium propionate (SP) has been reported to possess an anti-inflammatory and anti-apoptotic potential by inhibiting certain signaling pathways and helps in reducing the pathological damages of the mammary gland. However, the effects of sodium propionate on attenuating Lipopolysaccharide (LPS)-induced inflammatory condition and cell damage in bovine mammary epithelial cells (bMECs) are not comprehensively studied yet. Therefore, the aim of the current investigation was to evaluate the protective effects of sodium propionate on LPS-induced inflammatory conditions and to clarify the possible underlying molecular mechanism in bMECs. MAIN METHODS: The effects of increasing doses of SP on LPS-induced inflammation, oxidative stress and apoptosis was studied in vitro. Furthermore, the underlying protective mechanisms of SP on LPS-stimulated bMECs was investigated under different experimental conditions. KEY FINDINGS: The results reveled that increased inflammatory cytokines, chemokines and those of tight junction's mRNA expression was significantly attenuated dose-dependently by propionate. Biochemical analysis revealed that propionate pretreatment modulated the LPS-induced intercellular reactive oxygen species (ROS) accumulation, oxidative and antioxidant factors and apoptosis rate. Furthermore, we investigated that the LPS activated nuclear factor-kB (NF-kB), caspase/Bax apoptotic pathways and Histone deacetylases (HDAC) was significantly attenuated by propionate in bMECs. SIGNIFICANCE: Our results suggest that sodium propionate is a potent agent for ameliorating LPS-mediated cellular disruption and limiting detrimental inflammatory responses, partly via maintaining blood milk barrier integrity, inhibiting HDAC activity and NF-kB signaling pathway.


Asunto(s)
Leche/efectos de los fármacos , Propionatos/farmacología , Animales , Antiinflamatorios/farmacología , Apoptosis/efectos de los fármacos , Bovinos , Células Cultivadas , China , Citocinas/metabolismo , Células Epiteliales/metabolismo , Femenino , Inflamación/patología , Lipopolisacáridos/farmacología , Glándulas Mamarias Animales/efectos de los fármacos , Glándulas Mamarias Animales/metabolismo , Leche/metabolismo , FN-kappa B/metabolismo , Propionatos/metabolismo , Sustancias Protectoras/farmacología , Transducción de Señal/efectos de los fármacos
4.
Drug Deliv ; 28(1): 325-342, 2021 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-33517789

RESUMEN

The aim of the present study was to investigate the pharmacological mechanism of matrine in treatment of COVID-19 combined with liver injury. Potential targets related to matrine, COVID-19 and liver injury were identified from several databases. We constructed PPI network and screened the core targets according to the degree value. Then, GO and KEGG enrichment were carried out. Molecular docking technology was used to verify the affinity between matrine and the crystal structure of core target protein. Finally, real-time RT-PCR was used to detect the effects of matrine on hub gene expression in liver tissue of liver injury mice and lung tissue of lung injury mice to further confirm the results of network pharmacological analysis. The results show that six core targets including AKT1, TP53, TNF, IL6, BCL2L1 and ATM were identified. The potential therapeutic mechanism of matrine on COVID-19 combined with liver injury is closely related to regulate antiviral process, improve immune system and regulate the level of inflammatory factors. Molecular docking showed that matrine could spontaneously bind to the receptor protein and had strong binding force. Real-time RT-PCR demonstrated that matrine could significantly reduce the expression of AKT1, TP53, TNF, IL6 and ATM in mice with liver injury or lung injury (P < 0.05), and increase the expression of BCL2L1 to a certain extent (P > 0.05). Our results indicate that matrine can achieve simultaneous intervention of COVID-19 combined with liver injury by multi-dimensional pharmacological mechanism.


Asunto(s)
Alcaloides/farmacología , /epidemiología , Enfermedad Hepática Inducida por Sustancias y Drogas/epidemiología , Simulación del Acoplamiento Molecular/métodos , Quinolizinas/farmacología , Alcaloides/administración & dosificación , Animales , Antivirales/farmacología , Enfermedad Hepática Inducida por Sustancias y Drogas/etiología , Relación Dosis-Respuesta a Droga , Humanos , Lipopolisacáridos/farmacología , Hígado/efectos de los fármacos , Masculino , Ratones , Ratones Endogámicos C57BL , Quinolizinas/administración & dosificación , Reacción en Cadena en Tiempo Real de la Polimerasa , Transducción de Señal/efectos de los fármacos
5.
Phytochemistry ; 184: 112679, 2021 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-33550195

RESUMEN

A described monogalactosyldiacylglycerol (MGDG) and two undescribed digalactosyldiacylglycerols (DGDGs) were isolated from the leaves of Perilla frutescens (L.) Britton (Labiatae) by using silica gel column chromatography and semi-preparative high performance liquid chromatography. The elucidation of complete structure of these compounds were conducted by using MS and NMR techniques. The MGDG (7.5% of total lipids) was identified as 1,2-2-O-(9Z,12Z,15E-octadecatrienoyl)-3-O-(ß-D-galactopyranosyl)-sn-glycerol. The two DGDGs (2.8% and 1.0% of total lipids, respectively) were identified as 1-O-(9Z,12Z,15Z-octadecatrienoyl)-2-O-(6Z,9Z,12Z-octadecatrienoyl)-3-O-[ß-D-galactopyranosyl-(1″→6')-α-D-galactopyranosyl]-sn-glycerol and 1-O- hexadecanoyl -2-O-(9Z,12Z,15Z-octadecatrienoy -l)-3-O-[ß-D-galactopyranosyl-(1″→6')-α-D-galactopyranosyl]-sn-glycerol, respectively. All the isolated MGDG and DGDGs were evaluated for their anti-inflammatory activities in lipopolysaccharide (LPS)-stimulated murine macrophages RAW264.7 cells. All of them showed good inhibitory activities and significantly blocked the production of LPS-induced TNF-α, (IL)-1ß and IL-6. The above results shed some light on a better understanding of the traditional anti-inflammatory effect of Perilla frutescens and reveal the potential anti-inflammatory constituents.


Asunto(s)
Lamiaceae , Perilla frutescens , Animales , Antiinflamatorios/farmacología , Glucolípidos , Lipopolisacáridos/farmacología , Ratones , Hojas de la Planta
6.
Mol Cell ; 81(5): 953-968.e9, 2021 03 04.
Artículo en Inglés | MEDLINE | ID: mdl-33503407

RESUMEN

While the role of transcription factors and coactivators in controlling enhancer activity and chromatin structure linked to gene expression is well established, the involvement of corepressors is not. Using inflammatory macrophage activation as a model, we investigate here a corepressor complex containing GPS2 and SMRT both genome-wide and at the Ccl2 locus, encoding the chemokine CCL2 (MCP-1). We report that corepressors co-occupy candidate enhancers along with the coactivators CBP (H3K27 acetylase) and MED1 (mediator) but act antagonistically by repressing eRNA transcription-coupled H3K27 acetylation. Genome editing, transcriptional interference, and cistrome analysis reveals that apparently related enhancer and silencer elements control Ccl2 transcription in opposite ways. 4C-seq indicates that corepressor depletion or inflammatory signaling functions mechanistically similarly to trigger enhancer activation. In ob/ob mice, adipose tissue macrophage-selective depletion of the Ccl2 enhancer-transcribed eRNA reduces metaflammation. Thus, the identified corepressor-eRNA-chemokine pathway operates in vivo and suggests therapeutic opportunities by targeting eRNAs in immuno-metabolic diseases.


Asunto(s)
Quimiocina CCL2/genética , Proteínas Co-Represoras/genética , Elementos de Facilitación Genéticos , Péptidos y Proteínas de Señalización Intracelular/genética , Co-Represor 2 de Receptor Nuclear/genética , Obesidad/genética , Elementos Silenciadores Transcripcionales , Tejido Adiposo/inmunología , Tejido Adiposo/patología , Animales , Sistemas CRISPR-Cas , Quimiocina CCL2/inmunología , Proteínas Co-Represoras/inmunología , Edición Génica , Regulación de la Expresión Génica/efectos de los fármacos , Células HEK293 , Histona Acetiltransferasas/genética , Histona Acetiltransferasas/inmunología , Histonas/genética , Histonas/inmunología , Humanos , Péptidos y Proteínas de Señalización Intracelular/inmunología , Lipopolisacáridos/farmacología , Activación de Macrófagos/efectos de los fármacos , Masculino , Subunidad 1 del Complejo Mediador/genética , Subunidad 1 del Complejo Mediador/inmunología , Ratones , Ratones Obesos , Co-Represor 2 de Receptor Nuclear/inmunología , Obesidad/inmunología , Obesidad/patología , Células RAW 264.7 , ARN no Traducido/genética , ARN no Traducido/inmunología , Transducción de Señal
7.
Anal Chem ; 93(4): 2694-2705, 2021 02 02.
Artículo en Inglés | MEDLINE | ID: mdl-33397101

RESUMEN

Glycoproteins secreted by cells play essential roles in the regulation of extracellular activities. Secreted glycoproteins are often reflective of cellular status, and thus glycoproteins from easily accessible bodily fluids can serve as excellent biomarkers for disease detection. Cultured cells have been extensively employed as models in the research fields of biology and biomedicine, and global analysis of glycoproteins secreted from these cells provides insights into cellular activities and glycoprotein functions. However, comprehensive identification and quantification of secreted glycoproteins is a daunting task because of their low abundances compared with the high-abundance serum proteins required for cell growth and proliferation. Several studies employed serum-free media to analyze secreted proteins, but it has been shown that serum starvation, even for a short period of time, can alter protein secretion. To overcome these issues, we developed a method to globally characterize secreted glycoproteins and their N-glycosylation sites from cultured cells by combining selective enrichment of secreted glycoproteins with a boosting approach. The results demonstrated the importance of the boosting sample selection and the boosting-to-sample ratio for improving the coverage of secreted glycoproteins. The method was applied to globally quantify secreted glycoproteins from THP-1 monocytes and macrophages in response to lipopolysaccharides (LPS) and from Hep G2 cells treated with TGF-ß without serum starvation. We found differentially secreted glycoproteins in these model systems that showed the cellular response to the immune activation or the epithelial-to-mesenchymal transition. Benefiting from the selective enrichment and the signal enhancement of low-abundance secreted glycoproteins, this method can be extensively applied to study secreted glycoproteins without serum starvation, which will provide a better understanding of protein secretion and cellular activity.


Asunto(s)
Glicoproteínas/química , Técnicas de Cultivo de Célula , Química Clic , Glicoproteínas/metabolismo , Células Hep G2 , Humanos , Lipopolisacáridos/farmacología , Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Monocitos/efectos de los fármacos , Monocitos/metabolismo , Péptidos/química , Factor de Crecimiento Transformador beta/farmacología
8.
Life Sci ; 269: 119029, 2021 Mar 15.
Artículo en Inglés | MEDLINE | ID: mdl-33450256

RESUMEN

AIMS: The present study aimed to disclose a potent and selective GPR120 agonist LXT34 and its anti-diabetic effects. MAIN METHODS: Calcium mobilization assay was used to measure the agonistic potency and selectivity of LXT34 in GPR120 or GPR40-overexpression Chinese hamster ovary (CHO) cells. Glucagon-like peptide-1 (GLP-1) release and glucose-stimulated insulin secretion (GSIS) were evaluated in human colonic epithelial cell line NCI-H716 and mouse insulinoma cell line MIN6 by enzyme-linked immunosorbent assay (ELISA), respectively. The anti-inflammatory effect was determined in lipopolysaccharide (LPS)-induced murine macrophage cell line RAW264.7. Oral glucose tolerance test (OGTT) and insulin tolerance test (ITT) were performed to assess the anti-diabetic effects of LXT34 in db/db mice, and chronic inflammation in liver and adipose tissues were investigated using histomorphology, immunoblot and gene expression analysis. KEY FINDINGS: LXT34 was a potent GPR120 agonist with negligible activity toward human and mouse GPR40. LXT34 could potentiate GSIS and suppress LPS-induced inflammation in macrophages. LXT34 not only markedly improved glucose tolerance and insulin resistance, but also distinctly reduced macrophages infiltration, pro-inflammatory cytokines expression and JNK phosphorylation of both liver and adipose tissues in db/db mice. SIGNIFICANCE: LXT34, a novel and potent GPR120-selective agonist, showed beneficial effects on improving glucose homeostasis in obesity-related type 2 diabetes.


Asunto(s)
Inflamación/patología , Secreción de Insulina , Receptores Acoplados a Proteínas G/agonistas , Tejido Adiposo/patología , Animales , Enfermedad Crónica , Péptido 1 Similar al Glucagón/metabolismo , Glucosa/farmacología , Inflamación/sangre , Resistencia a la Insulina , Secreción de Insulina/efectos de los fármacos , Lipopolisacáridos/farmacología , Hígado/patología , Ratones , Ratones Endogámicos C57BL , Células RAW 264.7 , Receptores Acoplados a Proteínas G/metabolismo
9.
Bratisl Lek Listy ; 122(2): 138-144, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33502883

RESUMEN

BACKGROUND AND OBJECTIVES: Epithelial cells and macrophages play major roles in modulating the state of inlammatory response regulated by intracellular signaling pathways. The aim of this study was to characterize changes in cell proliferation, apoptosis and inflammation related intracellular signalling pathways MAPKs and NF-κB in Caco-2 epithelial cells and THP-1 macrophage-like monocytes contacted and filter-seperated co-cultures in the presence of LPS stimulation. METHODS: We assessed the apoptosis and inflammation by measuring caspase-3 activity, TNF-α and IL-10 cytokines, total and phosphorylated forms of intracellular signalling pathway molecules p53, JNK, Jun, ERK, p38, NF-κB p65 and IkB. RESULTS: The contacted co-culture of Caco-2 and THP-1 cells represented higher levels of JNK, jun and p38 MAPK pathway proteins associated with cell proliferation, whereas apoptosis related molecule caspase-3 and p53 increased in the filter-separated co-culture condition. Also, the contacted co-culture condition led to proinflammatory changes in NF-κB signalling pathways and cytokine responses with high phosphorylated NF-κB p65 and TNF-α, and low I-κB and IL-10 levels. CONCLUSION: Epithelia - monocytes co-culture stimulated with LPS regulated cell proliferation and inflammation in a contact dependent manner, whereas apoptosis was associated with non-contacted cell culture condition. Thus, co-culture models are important models for explaining the immunopathologies in the mucosal areas (Fig. 5, Ref. 30).


Asunto(s)
Macrófagos , Monocitos , Apoptosis , Células CACO-2 , Proliferación Celular , Técnicas de Cocultivo , Citocinas , Humanos , Inflamación , Lipopolisacáridos/farmacología , FN-kappa B
10.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi ; 37(2): 140-145, 2021 Feb.
Artículo en Chino | MEDLINE | ID: mdl-33504420

RESUMEN

Objective To investigate the effect of miR-21 on the expression of NF-κB and NLR family pyrin domain containing 3 (NLRP3) in RAW264.7 cells stimulated by lipopolysaccharide (LPS) and its possible mechanism. Methods Real-time fluorescent quantitative PCR was used to detect the expression of miR-21 after RAW264.7 cells were stimulated with 100 ng/mL LPS for 24 hours. miR-21 inhibitors or mimics were transfected to regulate the expression of miR-21 in RAW264.7 cells. After miR-21 inhibitors or mimics were stimulated by LPS, real-time quantitative PCR was used to detect its effect on the mRNA expression of interleukin 6 (IL-6), tumor necrosis factor α (TNF-α) and IL-1ß, and Western blotting was used to detect the effects on the expression of NF-κB, NLRP3 and A20 proteins. Results The expression of miR-21 significantly increased when RAW264.7 cells were stimulated by LPS. The expression of IL-6, TNF-α, IL-1ß, NF-κB and NLRP3 was raised when the expression of miR-21 was up-regulated. The expression of IL-6, TNF-α, IL-1ß, NF-κB and NLRP3 was reduced when the expression of miR-21 was down-regulated. miR-21 targeted the inhibition of A20 expression. Conclusion miR-21 can promote the expression of NF-κB and NLRP3 in RAW264.7 cells and its mechanism may be related to the targeted inhibition of A20.


Asunto(s)
MicroARNs , FN-kappa B , Animales , Interleucina-1beta/genética , Lipopolisacáridos/farmacología , Ratones , MicroARNs/genética , MicroARNs/fisiología , FN-kappa B/genética , Proteína con Dominio Pirina 3 de la Familia NLR/genética , Células RAW 264.7
11.
Methods Mol Biol ; 2270: 61-76, 2021.
Artículo en Inglés | MEDLINE | ID: mdl-33479893

RESUMEN

IL-10 is the best known and most studied anti-inflammatory cytokine and, in the last 20 years, it has acquired even greater fame as it has been associated with the regulatory phenotype of B cells. Indeed, although great efforts have been made to find a unique marker, to date IL-10 remains the main way to follow both murine and human regulatory B cells, hence the need of precise and reproducible methods to identify and purify IL-10-producing B cells for both functional and molecular downstream assays. In this chapter, we present our protocols to isolate these cells from the murine spleen and peritoneum and from human peripheral blood. Since the production of IL-10 by B cells is not only a weapon to counteract the adverse effect of pro-inflammatory cytokines but also a response to cellular activation, we focused on those B cells that are prone to IL-10 production and detectable following a short-term stimulation with phorbol-12-myristate-13-acetate, ionomycin, and lipopolysaccharide (murine system) or CpG (human system).


Asunto(s)
Subgrupos de Linfocitos B/citología , Linfocitos B Reguladores/citología , Separación Celular/métodos , Animales , Subgrupos de Linfocitos B/inmunología , Citocinas/inmunología , Expresión Génica/genética , Expresión Génica/inmunología , Humanos , Interleucina-10/metabolismo , Ionomicina/farmacología , Lipopolisacáridos/farmacología , Activación de Linfocitos/inmunología , Ratones , Ésteres del Forbol/farmacología , Bazo/citología , Acetato de Tetradecanoilforbol/farmacología
12.
Psychopharmacology (Berl) ; 238(3): 691-697, 2021 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-33410982

RESUMEN

BACKGROUND: Reduced motivation is one of the main symptomatic features of inflammation-induced depression. However, the exact nature of inflammation-induced alterations in motivation remains to be fully defined. As inflammation has been shown to increase sensitivity to negative stimuli, the present series of experiments was initiated to determine whether systemic inflammation induced by infra-septic doses of lipopolysaccharide (LPS) in mice influences consummatory and instrumental responding to successive negative contrast. METHODS: Successive negative contrast was operationally defined by a shift to a lower value reward than the one mice were trained with. Mice were trained to drink a high sucrose concentration solution and exposed to an acute shift to a lower concentration of sucrose. In another series of experiments, mice were trained to nose poke for chocolate pellets according to a fixed reinforcement schedule 10 (10 nose pokes for the food reinforcement) and exposed to a shift to a lower reward value (decreased number of chocolate pellets or replacement of chocolate pellets by less preferred grain pellets). Lipopolysaccharide (LPS) was administered at the dose of 0.33 1 mg/kg 24 h before the shift. RESULTS: Mice trained to drink a high sucrose concentration responded to the shift in reward value by a reduction in the volume of sucrose consumed and a decrease in lick numbers and bout durations. Mice trained to nose poke for chocolate pellets responded to the shift by alterations in their total number of nose pokes. In both conditions, LPS had no consistent effect on the response to the shift in reward value. CONCLUSIONS: These findings indicate a high variability in the effects of LPS on successive negative contrast and fail to provide evidence in favor of the hypothesis that LPS increases sensitivity to decreases in expected rewards.


Asunto(s)
Condicionamiento Operante/efectos de los fármacos , Conducta Consumatoria/efectos de los fármacos , Inflamación/psicología , Lipopolisacáridos/farmacología , Refuerzo en Psicología , Sacarosa/farmacología , Animales , Chocolate , Alimentos , Masculino , Ratones , Motivación/efectos de los fármacos , Esquema de Refuerzo , Recompensa , Sacarosa/administración & dosificación
13.
Mol Immunol ; 131: 97-111, 2021 03.
Artículo en Inglés | MEDLINE | ID: mdl-33461765

RESUMEN

Dehydroepiandrosterone (DHEA) is the major steroid hormone in humans and animals, which can regulate the body's inflammatory responses. However, the detail mechanism of this beneficial function is still poorly understood. The present study aimed to explore the anti-inflammation effect of DHEA and its underlying molecular mechanism in lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages. The findings showed that DHEA significantly inhibited the inflammation-related mediators production and pro-inflammatory cytokines expression level. Further research found that DHEA obviously blocked the LPS-stimulated PI3K/AKT, MAPK and NF-κB activation in RAW 264.7 cells. Meanwhile, DHEA enhanced the autophagy-dependent Keap1 protein degradation, subsequently activated the Nrf2 pathway to alleviate the redox imbalance and inflammatory responses. In conclusion, our data demonstrated that DHEA suppresses inflammatory responses through the activation of Nrf2 and inhibition of NF-κB in LPS-stimulated macrophages.


Asunto(s)
Antiinflamatorios/envenenamiento , Deshidroepiandrosterona/farmacología , Inflamación/tratamiento farmacológico , Macrófagos/efectos de los fármacos , Factor 2 Relacionado con NF-E2/metabolismo , Animales , Línea Celular , Citocinas/metabolismo , Inflamación/inducido químicamente , Inflamación/metabolismo , Mediadores de Inflamación/metabolismo , Lipopolisacáridos/farmacología , Macrófagos/metabolismo , Ratones , FN-kappa B/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Células RAW 264.7 , Transducción de Señal/efectos de los fármacos
14.
Am J Physiol Cell Physiol ; 320(3): C448-C461, 2021 03 01.
Artículo en Inglés | MEDLINE | ID: mdl-33471620

RESUMEN

Gram-negative bacterial lipopolysaccharide (LPS) increases the susceptibility of cells to pathogenic diseases, including inflammatory diseases and septic syndrome. In our experiments, we examined whether LPS induces epithelial barrier disruption in secretory epithelia and further investigated its underlying mechanism. The activities of Ca2+-activated Cl- channels (CACC) and epithelial Na+ channels (ENaC) were monitored with a short-circuit current using an Ussing chamber. Epithelial membrane integrity was estimated via transepithelial electrical resistance and paracellular permeability assays. We found that the apical application of LPS evoked short-circuit current (Isc) through the activation of CACC and ENaC. Although LPS disrupted epithelial barrier integrity, this was restored with the inhibition of CACC and ENaC, indicating the role of CACC and ENaC in the regulation of paracellular pathways. We confirmed that LPS, CACC, or ENaC activation evoked apical membrane depolarization. The exposure to a high-K+ buffer increased paracellular permeability. LPS induced the rapid redistribution of zonula occludens-1 (ZO-1) and reduced the expression levels of ZO-1 in tight junctions through apical membrane depolarization and tyrosine phosphorylation. However, the LPS-induced epithelial barrier disruption and degradation of ZO-1 were largely recovered by blocking CACC and ENaC. Furthermore, although LPS-impaired epithelial barrier became vulnerable to secondary bacterial infections, this vulnerability was prevented by inhibiting CACC and ENaC. We concluded that LPS induces the disruption of epithelial barrier integrity through the activation of CACC and ENaC, resulting in apical membrane depolarization and the subsequent tyrosine phosphorylation of ZO-1.


Asunto(s)
Canales de Cloruro/metabolismo , Células Epiteliales/efectos de los fármacos , Células Epiteliales/metabolismo , Epitelio/efectos de los fármacos , Epitelio/metabolismo , Lipopolisacáridos/farmacología , Canales de Sodio/metabolismo , Animales , Células Cultivadas , Masculino , Potenciales de la Membrana/efectos de los fármacos , Permeabilidad/efectos de los fármacos , Ratas , Ratas Sprague-Dawley , Uniones Estrechas/efectos de los fármacos , Uniones Estrechas/metabolismo , Proteína de la Zonula Occludens-1/metabolismo
15.
Nat Commun ; 12(1): 301, 2021 01 12.
Artículo en Inglés | MEDLINE | ID: mdl-33436596

RESUMEN

Macrophages are innate immune cells that contribute to fighting infections, tissue repair, and maintaining tissue homeostasis. To enable such functional diversity, macrophages resolve potentially conflicting cues in the microenvironment via mechanisms that are unclear. Here, we use single-cell RNA sequencing to explore how individual macrophages respond when co-stimulated with inflammatory stimuli LPS and IFN-γ and the resolving cytokine IL-4. These co-stimulated macrophages display a distinct global transcriptional program. However, variable negative cross-regulation between some LPS + IFN-γ-specific and IL-4-specific genes results in cell-to-cell heterogeneity in transcription. Interestingly, negative cross-regulation leads to mutually exclusive expression of the T-cell-polarizing cytokine genes Il6 and Il12b versus the IL-4-associated factors Arg1 and Chil3 in single co-stimulated macrophages, and single-cell secretion measurements show that these specialized functions are maintained for at least 48 h. This study suggests that increasing functional diversity in the population is one strategy macrophages use to respond to conflicting environmental cues.


Asunto(s)
Polaridad Celular , Macrófagos/citología , Animales , Arginasa/metabolismo , Polaridad Celular/efectos de los fármacos , Polaridad Celular/genética , Femenino , Regulación de la Expresión Génica/efectos de los fármacos , Interferón gamma/farmacología , Interleucina-12/metabolismo , Interleucina-6/metabolismo , Lipopolisacáridos/farmacología , Aprendizaje Automático , Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Masculino , Ratones Endogámicos C57BL , Redes Neurales de la Computación , Oportunidad Relativa , Análisis de la Célula Individual , Factores de Transcripción/metabolismo , Transcripción Genética/efectos de los fármacos
16.
Nat Commun ; 12(1): 86, 2021 01 04.
Artículo en Inglés | MEDLINE | ID: mdl-33397971

RESUMEN

Inflammation and cell death are closely linked arms of the host immune response to infection, which when carefully balanced ensure host survival. One example of this balance is the tightly regulated transition from TNFR1-associated pro-inflammatory complex I to pro-death complex II. By contrast, here we show that a TRIF-dependent complex containing FADD, RIPK1 and caspase-8 (that we have termed the TRIFosome) mediates cell death in response to Yersinia pseudotuberculosis and LPS. Furthermore, we show that constitutive binding between ZBP1 and RIPK1 is essential for the initiation of TRIFosome interactions, caspase-8-mediated cell death and inflammasome activation, thus positioning ZBP1 as an effector of cell death in the context of bacterial blockade of pro-inflammatory signaling. Additionally, our findings offer an alternative to the TNFR1-dependent model of complex II assembly, by demonstrating pro-death complex formation reliant on TRIF signaling.


Asunto(s)
Interleucina-1beta/metabolismo , Lipopolisacáridos/farmacología , Proteínas de Unión al ARN/metabolismo , Proteína Serina-Treonina Quinasas de Interacción con Receptores/metabolismo , Proteínas Adaptadoras del Transporte Vesicular/metabolismo , Animales , Caspasa 8/metabolismo , Muerte Celular/efectos de los fármacos , Ratones Endogámicos C57BL , Unión Proteica/efectos de los fármacos , Receptores Tipo I de Factores de Necrosis Tumoral/metabolismo , Yersinia
17.
Phytochemistry ; 183: 112642, 2021 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-33421888

RESUMEN

Fifteen eremophilane sesquiterpenoids, including nine undescribed congeners, septeremophilane A-H, and chaetopenoid G, together with four conjugated unsaturated polyketide fatty acids, including an undescribed derivative, were isolated from cultures of the fungus Septoria rudbeckiae, a plant pathogenic fungus isolated from the halophyte Karelinia caspia. Septeremophilane A represents an unprecedented tetranor-eremophilane sesquiterpenoid with an α,ß-unsaturated δ-lactone unit bearing a hemiacetal group, while septeremophilane B-H possesses a trinor-eremophilane skeleton. Their structures and absolute configurations were established based on spectroscopic data (NMR and HRESIMS), quantum chemical calculations and electronic circular dichroism (ECD) experiments. All metabolites were tested for nitric oxide (NO) production inhibition in lipopolysaccharide (LPS)-activated BV-2 microglial cells, while dendryphiellin D, septeremophilane D, and septeremophilane E were found to display significant inhibition, with IC50 values of 11.9 ± 1.0, 8.5 ± 0.1, and 6.0 ± 0.2 µM, respectively.


Asunto(s)
Ascomicetos , Sesquiterpenos , Lipopolisacáridos/farmacología , Sesquiterpenos Policíclicos , Sesquiterpenos/farmacología
18.
Gene ; 766: 145153, 2021 Jan 15.
Artículo en Inglés | MEDLINE | ID: mdl-32950633

RESUMEN

AIM: Acute lung injury (ALI) is the mild form of acute respiratory distress syndrome (ARDS) which is a common lung disease with a high incidence and mortality rate. Recent studies manifested that some circular RNAs were associated with ALI. In this study, we aimed to uncover the effect of circular RNA circ_0054633 on ALI initiation and progression and proposed a new mechanism related to ALI. METHODS: The lipopolysaccharides (LPS)-induced acute lung injury model were build both in vivo of rat and in vitro of primary murine pulmonary microvascular endothelial cells (MPVECs). Hematoxylin and eosin (H&E) was employed to observe the tissue morphology and estimate the degree of lung damage. We used real-time quantitative polymerase chain reaction (RT-qPCR) to measure the expression level of circ_0054633. The expression levels of inflammatory cytokines IL-17A and tumor necrosis factor-α (TNF-α) were detected by ELISA. The effects of circ_0054633 on MPVECs proliferation and apoptosis were detected with the help of CCK-8 and apoptosis assay, separately. The expression level of NF-κB p65 protein was measured by Western blot. RESULTS: circ_0054633, IL-17A, TNF-α and NF-κB p65 were all overexpressed in LPS-treated rat and MPVECs, and LPS enhanced the proliferation and apoptosis of MPVECs. While circ_0054633 silencing reversed the above promotion effects of LPS on IL-17A, TNF-α expression and MPVECs proliferation and apoptosis. CONCLUSIONS: Quietness of circ_0054633 alleviated LPS-induced ALI via NF-κB signaling pathway, implicating circ_0054633 may be a potential biomarker for diagnose and therapy of ALI.


Asunto(s)
Lesión Pulmonar Aguda/metabolismo , Proliferación Celular/fisiología , Células Endoteliales/metabolismo , Inflamación/metabolismo , FN-kappa B/metabolismo , ARN Circular/metabolismo , Lesión Pulmonar Aguda/inducido químicamente , Animales , Apoptosis/efectos de los fármacos , Apoptosis/fisiología , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Inflamación/inducido químicamente , Interleucina-17/metabolismo , Lipopolisacáridos/farmacología , Pulmón/efectos de los fármacos , Pulmón/metabolismo , Masculino , Ratones , Ratas , Ratas Sprague-Dawley , Factor de Transcripción ReIA/metabolismo , Factor de Necrosis Tumoral alfa/metabolismo
19.
Nat Neurosci ; 24(3): 368-378, 2021 03.
Artículo en Inglés | MEDLINE | ID: mdl-33328624

RESUMEN

It is unclear whether severe acute respiratory syndrome coronavirus 2, which causes coronavirus disease 2019, can enter the brain. Severe acute respiratory syndrome coronavirus 2 binds to cells via the S1 subunit of its spike protein. We show that intravenously injected radioiodinated S1 (I-S1) readily crossed the blood-brain barrier in male mice, was taken up by brain regions and entered the parenchymal brain space. I-S1 was also taken up by the lung, spleen, kidney and liver. Intranasally administered I-S1 also entered the brain, although at levels roughly ten times lower than after intravenous administration. APOE genotype and sex did not affect whole-brain I-S1 uptake but had variable effects on uptake by the olfactory bulb, liver, spleen and kidney. I-S1 uptake in the hippocampus and olfactory bulb was reduced by lipopolysaccharide-induced inflammation. Mechanistic studies indicated that I-S1 crosses the blood-brain barrier by adsorptive transcytosis and that murine angiotensin-converting enzyme 2 is involved in brain and lung uptake, but not in kidney, liver or spleen uptake.


Asunto(s)
Barrera Hematoencefálica/metabolismo , Glicoproteína de la Espiga del Coronavirus/farmacocinética , Administración Intranasal , Administración Intravenosa , Animales , Apolipoproteínas E/genética , Genotipo , Hipocampo/metabolismo , Humanos , Inflamación/inducido químicamente , Inflamación/metabolismo , Lipopolisacáridos/farmacología , Masculino , Ratones , Ratones Transgénicos , Bulbo Olfatorio/metabolismo , Caracteres Sexuales , Glicoproteína de la Espiga del Coronavirus/administración & dosificación , Distribución Tisular , Transcitosis
20.
J Dairy Sci ; 104(2): 1351-1363, 2021 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-33309364

RESUMEN

During the thermal processing of milk, Maillard reactions occur between proteins and lactose to generate glycated proteins. In this study, a lactose-glycated caseinate was hydrolyzed by trypsin. The obtained glycated caseinate (GCN) hydrolysate had a lactose content of 10.8 g/kg of protein. We identified its glycation sites and then assessed it for its protective effect against lipopolysaccharide-induced barrier injury using a rat intestinal epithelial cell line (IEC-6 cells) as a cell model and unglycated caseinate (CN) hydrolysate as a reference. Results from our liquid chromatography-mass spectrometry analysis of the GCN hydrolysate verified that lactose glycation occurred at the Lys residues in 3 casein components (αS1-casein, ß-casein, and κ-casein), and this resulted in the formation of 5 peptides with the following amino acid sequences: EMPFPKYPKYPVEPF, HIQKEDVPSE, GSENSEKTTMPL, NQDKTEIPT, and EGIHAQQKEPM. The results from cell experiments showed that the 2 hydrolysates could promote cell growth and decrease lactate dehydrogenase release in the lipopolysaccharide-injured cells; more importantly, they could partially protect the damaged barrier function of the cells by increasing trans-epithelial electrical resistance, decreasing epithelial permeability, and upregulating the expression of the 3 tight junction proteins zonula occludens-1, occludin, and claudin-1. However, compared with CN hydrolysate, GCN hydrolysate showed lower efficacy in protecting against cellular barrier dysfunction. We propose that the different chemical characteristics of the CN hydrolysate and the GCN hydrolysate (i.e., amino acid loss and lactose conjugation) contributed to the lower barrier-protective efficacy of the GCN hydrolysate. During dairy processing, protein glycation of the Maillard type might have a non-negligible, unfavorable effect on dairy proteins, in view of the resulting protein glycation we found and the critical function of proteins for maintaining the integrity of the intestinal barrier.


Asunto(s)
Caseínas/metabolismo , Células Epiteliales/efectos de los fármacos , Lactosa/metabolismo , Lipopolisacáridos/farmacología , Péptidos/farmacología , Animales , Sitios de Unión , Caseínas/química , Línea Celular , Claudina-1/metabolismo , Células Epiteliales/fisiología , Glicosilación , Hidrólisis , Intestinos/citología , Intestinos/efectos de los fármacos , Reacción de Maillard , Péptidos/química , Permeabilidad , Ratas , Tripsina/metabolismo
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