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1.
J Immunol ; 207(11): 2744-2753, 2021 12 01.
Artigo em Inglês | MEDLINE | ID: mdl-34725110

RESUMO

In response to infection or tissue damage, resident peritoneal macrophages (rpMACs) produce inflammatory lipid mediators from the polyunsaturated fatty acid (PUFA), arachidonic acid (AA). Long-chain acyl-CoA synthetase 4 (ACSL4) catalyzes the covalent addition of a CoA moiety to fatty acids, with a strong preference for AA and other PUFAs containing three or more double bonds. PUFA-CoA can be incorporated into phospholipids, which is the source of PUFA for lipid mediator synthesis. In this study, we demonstrated that deficiency of Acsl4 in mouse rpMACs resulted in a significant reduction of AA incorporated into all phospholipid classes and a reciprocal increase in incorporation of oleic acid and linoleic acid. After stimulation with opsonized zymosan (opZym), a diverse array of AA-derived lipid mediators, including leukotrienes, PGs, hydroxyeicosatetraenoic acids, and lipoxins, were produced and were significantly reduced in Acsl4-deficient rpMACs. The Acsl4-deficient rpMACs stimulated with opZym also demonstrated an acute reduction in mRNA expression of the inflammatory cytokines, Il6, Ccl2, Nos2, and Ccl5 When Acsl4-deficient rpMACs were incubated in vitro with the TLR4 agonist, LPS, the levels of leukotriene B4 and PGE2 were also significantly decreased. In LPS-induced peritonitis, mice with myeloid-specific Acsl4 deficiency had a significant reduction in leukotriene B4 and PGE2 levels in peritoneal exudates, which was coupled with reduced infiltration of neutrophils in the peritoneal cavity as compared with wild-type mice. Our data demonstrate that chronic deficiency of Acsl4 in rpMACs reduces the incorporation of AA into phospholipids, which reduces lipid mediator synthesis and inflammation.


Assuntos
Ácido Araquidônico/imunologia , Coenzima A Ligases/imunologia , Inflamação/imunologia , Fosfolipídeos/imunologia , Zimosan/biossíntese , Animais , Coenzima A Ligases/deficiência , Camundongos , Camundongos Transgênicos
4.
FASEB J ; 35(10): e21940, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-34551158

RESUMO

Macrophages are the principal innate immune cells that populate all major organs and provide the first line of cellular defense against infections and/or injuries. The immediate and early-responding macrophages must mount a robust pro-inflammatory response to protect the host by eliminating deleterious agents. The effective pro-inflammatory macrophage response requires the activation of complex transcriptional programs that modulate the dynamic regulation of inflammatory and metabolic gene expression. Therefore, transcription factors that govern pro-inflammatory and metabolic gene expression play an essential role in shaping the macrophage inflammatory response. Herein, we identify the basic helix-loop-helix family member e40 (BHLHE40), as a critical transcription factor that promotes broad pro-inflammatory and glycolytic gene expression by elevating HIF1α levels in macrophages. Our in vivo studies revealed that myeloid-BHLHE40 deficiency significantly attenuates macrophage and neutrophil recruitment to the site of inflammation. Our integrated transcriptomics and gene set enrichment analysis (GSEA) studies show that BHLHE40 deficiency broadly curtails inflammatory signaling pathways, hypoxia response, and glycolytic gene expression in macrophages. Utilizing complementary gain- and loss-of-function studies, our analyses uncovered that BHLHE40 promotes LPS-induced HIF1α mRNA and protein expression in macrophages. More importantly, forced overexpression of oxygen stable form of HIF1α completely reversed attenuated pro-inflammatory and glycolytic gene expression in BHLHE40-deficient macrophages. Collectively, these results demonstrate that BHLHE40 promotes macrophage pro-inflammatory gene expression and functions by elevating HIF1α expression in macrophages.


Assuntos
Fatores de Transcrição Hélice-Alça-Hélice Básicos/metabolismo , Regulação da Expressão Gênica , Proteínas de Homeodomínio/metabolismo , Inflamação/genética , Macrófagos/metabolismo , Animais , Fatores de Transcrição Hélice-Alça-Hélice Básicos/deficiência , Células Sanguíneas/metabolismo , Feminino , Glicólise/efeitos dos fármacos , Glicólise/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Inflamação/induzido quimicamente , Lipopolissacarídeos/farmacologia , Pulmão/efeitos dos fármacos , Pulmão/patologia , Masculino , Camundongos , Substâncias Protetoras , Zimosan/efeitos adversos , Zimosan/antagonistas & inibidores
5.
J Vis Exp ; (173)2021 07 16.
Artigo em Inglês | MEDLINE | ID: mdl-34338665

RESUMO

Mesenchymal stem cells (MSC) have traditionally been studied for their regenerative properties, but more recently, their immunoregulatory characteristics have been at the forefront. They interact with and regulate immune cell activity. The focus of this study is the MSC regulation of macrophage phagocytic activity. Macrophage (MΦ) phagocytosis is an important part of the innate immune system response to infection, and the mechanisms through which MSC modulate this response are under active investigation. Presented here is a method to study MΦ phagocytosis of non-opsonized zymosan particles conjugated to a pH-sensitive fluorescent molecule while in co-culture with MSC. As phagocytic activity increases and the labeled zymosan particles are enclosed within the acidic environment of the phagolysosome, the fluorescence intensity of the pH-sensitive molecule increases. With the appropriate excitation and emission wavelengths, phagocytic activity is measured using a fluorescent spectrophotometer and kinetic data is presented as changes in relative fluorescent units over a 70 min period. To support this quantitative data, the change in the phagocytic activity is visualized using dynamic imaging. Results using this method demonstrate that when in co-culture, MSC enhance MΦ phagocytosis of non-opsonized zymosan of both naive and IFN-γ treated MΦ. These data add to the current knowledge of MSC regulation of the innate immune system. This method can be applied in future investigations to fully delineate the underlying cellular and molecular mechanisms.


Assuntos
Células-Tronco Mesenquimais , Técnicas de Cocultura , Macrófagos , Fagocitose , Zimosan/farmacologia
6.
Sci Rep ; 11(1): 17312, 2021 08 27.
Artigo em Inglês | MEDLINE | ID: mdl-34453072

RESUMO

Resolvins, are specialized pro-resolving mediators (SPMs) derived from n-3 polyunsaturated fatty acids. They contribute actively to the resolution of inflammation, but little is known concerning their role in chronic inflammation, such as in rheumatoid arthritis (RA). Here, we performed lipid mediator (LM) profiling in tissues from the paws of SKG arthritic mice using lipid chromatography (LC)/mass spectrometry (MS)/MS-based LM metabololipidomics. We found elevated levels of SPMs including resolvin D5 (RvD5) in these tissues. Moreover, RvD5 levels were significantly correlated with arthritis disease activity. From experiments to assess the role of RvD5 in the pathology of RA, we concluded that RvD5 suppressed Th17 cell differentiation and facilitated regulatory T cell differentiation, as well as inhibiting CD4+ T cell proliferation. Furthermore, RvD5 attenuated osteoclast differentiation and interfered with osteoclastogenesis. Targeting the resolution of inflammation could be promising as a novel treatment for RA.


Assuntos
Artrite Reumatoide/fisiopatologia , Ácidos Docosa-Hexaenoicos/metabolismo , Osteogênese/fisiologia , Células Th17/metabolismo , Zimosan/farmacologia , Animais , Artrite Experimental , Diferenciação Celular , Proliferação de Células , Cromatografia Líquida de Alta Pressão , , Humanos , Inflamação , Camundongos , Espectrometria de Massas em Tandem
7.
J Immunol ; 207(6): 1652-1661, 2021 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-34426543

RESUMO

The IκB kinase (IKK) complex plays a vital role in regulating the NF-κB activation. Aberrant NF-κB activation is involved in various inflammatory diseases. Thus, targeting IKK activation is an ideal therapeutic strategy to cure and prevent inflammatory diseases related to NF-κB activation. In a previous study, we demonstrated that IKK-interacting protein (IKIP) inhibits the phosphorylation of IKKα/ß and the activation of NF-κB through disruption of the formation of IKK complex. In this study, we identified a 15-aa peptide derived from mouse IKIP (46-60 aa of IKIP), which specifically suppressed IKK activation and NF-κB targeted gene expression via disrupting the association of IKKß and NEMO. Importantly, administration of the peptide reduced LPS-induced acute inflammation and attenuated Zymosan-induced acute arthritis in mice. These findings suggest that this IKIP peptide may be a promising therapeutic reagent in the prevention and treatment of inflammatory diseases.


Assuntos
Quinase I-kappa B/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , NF-kappa B/metabolismo , Peptídeos/administração & dosagem , Transdução de Sinais/efeitos dos fármacos , Animais , Artrite Experimental/induzido quimicamente , Artrite Experimental/tratamento farmacológico , Linhagem Celular Tumoral , Ativação Enzimática/efeitos dos fármacos , Ativação Enzimática/genética , Regulação da Expressão Gênica/efeitos dos fármacos , Células HEK293 , Humanos , Inflamação/induzido quimicamente , Inflamação/tratamento farmacológico , Lipopolissacarídeos/efeitos adversos , Camundongos , Camundongos Knockout , Ligação Proteica , Transdução de Sinais/genética , Zimosan/efeitos adversos
8.
Proc Natl Acad Sci U S A ; 118(28)2021 07 13.
Artigo em Inglês | MEDLINE | ID: mdl-34244423

RESUMO

Optic neuropathies are leading causes of irreversible visual impairment and blindness, currently affecting more than 100 million people worldwide. Glaucoma is a group of optic neuropathies attributed to progressive degeneration of retinal ganglion cells (RGCs). We have previously demonstrated an increase in survival of RGCs by the activation of macrophages, whereas the inhibition of macrophages was involved in the alleviation on endotoxin-induced inflammation by antagonist of growth hormone-releasing hormone (GHRH). Herein, we hypothesized that GHRH receptor (GHRH-R) signaling could be involved in the survival of RGCs mediated by inflammation. We found the expression of GHRH-R in RGCs of adult rat retina. After optic nerve crush, subcutaneous application of GHRH agonist MR-409 or antagonist MIA-602 promoted the survival of RGCs. Both the GHRH agonist and antagonist increased the phosphorylation of Akt in the retina, but only agonist MR-409 promoted microglia activation in the retina. The antagonist MIA-602 reduced significantly the expression of inflammation-related genes Il1b, Il6, and Tnf Moreover, agonist MR-409 further enhanced the promotion of RGC survival by lens injury or zymosan-induced macrophage activation, whereas antagonist MIA-602 attenuated the enhancement in RGC survival. Our findings reveal the protective effect of agonistic analogs of GHRH on RGCs in rats after optic nerve injury and its additive effect to macrophage activation, indicating a therapeutic potential of GHRH agonists for the protection of RGCs against optic neuropathies especially in glaucoma.


Assuntos
Hormônio Liberador de Hormônio do Crescimento/agonistas , Macrófagos/patologia , Neuroproteção , Traumatismos do Nervo Óptico/patologia , Células Ganglionares da Retina/patologia , Animais , Sobrevivência Celular/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Hormônio do Crescimento/metabolismo , Hormônio Liberador de Hormônio do Crescimento/antagonistas & inibidores , Inflamação/genética , Inflamação/patologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Masculino , Microglia/efeitos dos fármacos , Microglia/metabolismo , Microglia/patologia , Neuroproteção/efeitos dos fármacos , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ratos Endogâmicos F344 , Receptores de Neuropeptídeos/metabolismo , Receptores de Hormônios Reguladores de Hormônio Hipofisário/metabolismo , Células Ganglionares da Retina/efeitos dos fármacos , Células Ganglionares da Retina/metabolismo , Fator de Transcrição STAT3/metabolismo , Sermorelina/análogos & derivados , Sermorelina/farmacologia , Transdução de Sinais/efeitos dos fármacos , Corpo Vítreo/efeitos dos fármacos , Corpo Vítreo/metabolismo , Zimosan/farmacologia
9.
Nat Commun ; 12(1): 3611, 2021 06 14.
Artigo em Inglês | MEDLINE | ID: mdl-34127673

RESUMO

Yeast is an integral part of mammalian microbiome, and like commensal bacteria, has the potential of being harnessed to influence immunity in clinical settings. However, functional specificities of yeast-derived immunoregulatory molecules remain elusive. Here we find that while under steady state, ß-1,3-glucan-containing polysaccharides potentiate pro-inflammatory properties, a relatively less abundant class of cell surface polysaccharides, dubbed mannan/ß-1,6-glucan-containing polysaccharides (MGCP), is capable of exerting potent anti-inflammatory effects to the immune system. MGCP, in contrast to previously identified microbial cell surface polysaccharides, through a Dectin1-Cox2 signaling axis in dendritic cells, facilitates regulatory T (Treg) cell induction from naïve T cells. Furthermore, through a TLR2-dependent mechanism, it restrains Th1 differentiation of effector T cells by suppressing IFN-γ expression. As a result, administration of MGCP display robust suppressive capacity towards experimental inflammatory disease models of colitis and experimental autoimmune encephalomyelitis (EAE) in mice, thereby highlighting its potential therapeutic utility against clinically relevant autoimmune diseases.


Assuntos
Imunomodulação/efeitos dos fármacos , Imunomodulação/imunologia , Polissacarídeos/imunologia , Saccharomyces cerevisiae/metabolismo , beta-Glucanas/imunologia , Animais , Linfócitos T CD4-Positivos , Diferenciação Celular/efeitos dos fármacos , Colite/imunologia , Colite/patologia , Ciclo-Oxigenase 2 , Células Dendríticas/imunologia , Encefalomielite Autoimune Experimental , Glucanos , Proteínas de Homeodomínio/genética , Imunidade , Lectinas Tipo C , Mananas , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Polissacarídeos/metabolismo , Polissacarídeos/farmacologia , Saccharomyces cerevisiae/genética , Linfócitos T Reguladores/efeitos dos fármacos , Linfócitos T Reguladores/imunologia , Células Th1 , Zimosan , beta-Glucanas/metabolismo , beta-Glucanas/farmacologia
10.
Sci Rep ; 11(1): 11923, 2021 06 07.
Artigo em Inglês | MEDLINE | ID: mdl-34099783

RESUMO

Ankylosing spondylitis is a male-predominant disease and previous study revealed that estrogens have an anti-inflammatory effect on the spondyloarthritis (SpA) manifestations in zymosan-induced SKG mice. This study aimed to evaluate the effect of selective estrogen receptor modulator (SERM) lasofoxifene (Laso) on disease activity of SpA. Mice were randomized into zymosan-treated, zymosan + 17ß-estradiol (E2)-treated, and zymosan + Laso-treated groups. Arthritis was assessed by 18F-fluorodeoxyglucose (18F-FDG) small-animal positron emission tomography/computed tomography and bone mineral density (BMD) was measured. Fecal samples were collected and 16S ribosomal RNA gene sequencing was used to determine gut microbiota differences. Both zymosan + E2-treated mice and zymosan + Laso-treated mice showed lower arthritis clinical scores and lower 18F-FDG uptake than zymosan-treated mice. BMD was significantly higher in zymosan + E2-treated mice and zymosan + Laso-treated mice than zymosan-treated mice, respectively. Fecal calprotectin levels were significantly elevated at 8 weeks after zymosan injection in zymosan-treated mice, but it was not significantly changed in zymosan + E2-treated mice and zymosan + Laso-treated mice. Gut microbiota diversity of zymosan-treated mice was significantly different from zymosan + E2-treated mice and zymosan + Laso-treated mice, respectively. There was no significant difference in gut microbiota diversity between zymosan + E2-treated mice and zymosan + Laso -treated mice. Laso inhibited joint inflammation and enhanced BMD in SKG mice, a model of SpA. Laso also affected the composition and biodiversity of gut microbiota. This study provides new knowledge regarding that selected SpA patients could benefit from SERM treatment.


Assuntos
Artrite Experimental/prevenção & controle , Microbioma Gastrointestinal/efeitos dos fármacos , Pirrolidinas/farmacologia , Moduladores Seletivos de Receptor Estrogênico/farmacologia , Espondilartrite/prevenção & controle , Tetra-Hidronaftalenos/farmacologia , Animais , Artrite Experimental/induzido quimicamente , Artrite Experimental/metabolismo , Bactérias/classificação , Bactérias/genética , Densidade Óssea/efeitos dos fármacos , Citocinas/genética , Citocinas/metabolismo , Estradiol/farmacologia , Estrogênios/farmacologia , Fezes/química , Fezes/microbiologia , Fluordesoxiglucose F18/metabolismo , Fluordesoxiglucose F18/farmacocinética , Microbioma Gastrointestinal/genética , Expressão Gênica/efeitos dos fármacos , Complexo Antígeno L1 Leucocitário/metabolismo , Camundongos , Tomografia por Emissão de Pósitrons combinada à Tomografia Computadorizada/métodos , RNA Ribossômico 16S/genética , Espondilartrite/induzido quimicamente , Espondilartrite/metabolismo , Zimosan
11.
Biologicals ; 71: 20-30, 2021 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-33994086

RESUMO

The rabbit pyrogen test (RPT) is a safety test conducted as a part of mandatory requirements of regulatory agencies. RPT is currently performed for routine quality control (QC) by manufacturers and for national lot release of biological products, such as plasma-derived products. However, RPT involves the use of many rabbits, counter to the international efforts to minimize the use of animals in research. Furthermore, pyrogen amount cannot be discerned from the test results and the results may be considerably affected by various factors. Therefore, a need exists for substituting RPT with in vitro assays. As a viable alternative to RPT, we here established a rabbit monocyte activation test (RMAT) based on the human MAT in the European Pharmacopoeia. RMAT uses rabbit peripheral blood mononuclear cells as the source of monocytes instead of live animals. The test detected endotoxin, lipoteichoic acid, peptidoglycan, and zymosan with high sensitivity, showing high correlation with the in vivo RPT results. The results of RMAT and RPT testing of non-pyrogenic plasma-derived products were also consistent. Furthermore, RMAT showed satisfactory recovery rates in an interference test with product samples and spiked-in pyrogens. We conclude that RMAT could replace the existing RPT for routine QC.


Assuntos
Alternativas aos Testes com Animais , Bioensaio , Monócitos , Pirogênios , Animais , Endotoxinas , Leucócitos Mononucleares , Lipopolissacarídeos , Peptidoglicano , Pirogênios/análise , Controle de Qualidade , Coelhos , Ácidos Teicoicos , Zimosan
12.
PLoS One ; 16(4): e0250352, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33878143

RESUMO

1,25-dihydroxyvitamin-D3 and its derivatives have shown anti-arthritic and chondroprotective effects in experimental animal models with prophylactic dosing. The purpose of this preliminary study was to test the efficacy and safety of calcipotriol, vitamin D analog, as a treatment for a fully-developed knee arthritis in Zymosan-induced arthritis (ZIA) model. Forty 5-month-old male Sprague-Dawley rats were randomized into three arthritis groups and a non-arthritic control group with no injections (10 rats/group). A day after Zymosan (0.1 mg) had been administrated into the right knee joints, the same knees were injected with calcipotriol (0.1 mg/kg), dexamethasone (0.1 mg/kg) or vehicle in a 100 µl volume. The left control knees were injected with saline (PBS) on two consecutive days. All injections, blood sampling and measurements were performed under general anesthesia on days 0, 1, 3 and 8. Internal organs and knees were harvested on day 8 and the histology of the whole knees was assessed blinded. Joints treated with calcipotriol showed a milder histological synovitis than those treated with vehicle (p = 0.041), but there was no statistically significant difference between the dexamethasone and vehicle groups. The clinical severity of arthritis did not differ between the arthritis groups measured by body temperature, swelling of the knee, thermal imaging, clinical scoring or cytokine levels on days 1, 3 and 8. Weight loss was bigger in rats treated with dexamethasone, propably due to loss of appetite,compared to other arthritis groups on days 2-3 (p<0.05). Study drugs did not influence serum calcium ion and glucose levels. Taken together, this preliminary study shows that a single intra-articular injection of calcipotriol reduces histological grade of synovitis a week after the local injection, but dexamethasone did not differ from the vehicle. Calcipotriol may have an early disease-modifying effect in the rat ZIA model without obvious side effects.


Assuntos
Anti-Inflamatórios/farmacologia , Artrite Experimental/tratamento farmacológico , Calcitriol/análogos & derivados , Sinovite/tratamento farmacológico , Animais , Artrite Experimental/sangue , Artrite Experimental/induzido quimicamente , Artrite Experimental/patologia , Glicemia/metabolismo , Calcitriol/farmacologia , Cálcio/sangue , Cartilagem Articular/efeitos dos fármacos , Cartilagem Articular/metabolismo , Cartilagem Articular/patologia , Dexametasona/farmacologia , Esquema de Medicação , Membro Posterior , Injeções Intra-Articulares , Masculino , Ratos , Ratos Sprague-Dawley , Sinovite/sangue , Sinovite/induzido quimicamente , Sinovite/patologia , Resultado do Tratamento , Zimosan/administração & dosagem
13.
Bull Exp Biol Med ; 170(6): 748-751, 2021 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-33893954

RESUMO

We analyzed functional status of blood leukocytes in diabetes mellitus and after addition of glucose in vitro. To this end, generation of ROS and reactive halogen species by monocytes and neutrophils from patients with diabetes mellitus and healthy donors was assayed using lucigenin- and luminol-dependent chemiluminescence after stimulation with phorbol 12-myristate 13-acetate or opsonized zymosan in vitro. Formation of neutrophil extracellular traps was evaluated in the blood after addition of glucose. In comparison with donors, leukocytes from patients with diabetes mellitus were primed and this effect can be modeled by addition of glucose to the blood in vitro. Addition of glucose to donor blood also triggered the formation of neutrophil extracellular traps.


Assuntos
Hiperglicemia/metabolismo , Leucócitos/metabolismo , Diabetes Mellitus/metabolismo , Armadilhas Extracelulares/metabolismo , Humanos , Medições Luminescentes , Neutrófilos/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Acetato de Tetradecanoilforbol/farmacologia , Zimosan/farmacologia
14.
Cells ; 10(4)2021 04 09.
Artigo em Inglês | MEDLINE | ID: mdl-33918872

RESUMO

Methyl-CpG-binding protein 2 (MECP2) is a critical transcriptional regulator for synaptic function. Dysfunction of synapses, as well as microglia-mediated neuroinflammation, represent the earliest pathological events in Alzheimer's disease (AD). Here, expression, protein levels, and activity-related phosphorylation changes of MECP2 were analyzed in post-mortem human temporal cortex. The effects of wild type and phosphorylation-deficient MECP2 variants at serine 423 (S423) or S80 on microglial and neuronal function were assessed utilizing BV2 microglial monocultures and co-cultures with mouse cortical neurons under inflammatory stress conditions. MECP2 phosphorylation at the functionally relevant S423 site nominally decreased in the early stages of AD-related neurofibrillary pathology in the human temporal cortex. Overexpression of wild type MECP2 enhanced the pro-inflammatory response in BV2 cells upon treatment with lipopolysaccharide (LPS) and interferon-γ (IFNγ) and decreased BV2 cell phagocytic activity. The expression of the phosphorylation-deficient MECP2-S423A variant, but not S80A, further increased the pro-inflammatory response of BV2 cells. In neurons co-cultured with BV2 cells, the MECP2-S423A variant increased the expression of several genes, which are important for the maintenance and protection of neurons and synapses upon inflammatory stress. Collectively, functional analyses in different cellular models suggest that MECP2 may influence the inflammatory response in microglia independently of S423 and S80 phosphorylation, while the S423 phosphorylation might play a role in the activation of neuronal gene expression, which conveys neuroprotection under neuroinflammation-related stress.


Assuntos
Regulação da Expressão Gênica , Inflamação/patologia , Proteína 2 de Ligação a Metil-CpG/metabolismo , Microglia/metabolismo , Microglia/patologia , Neurônios/metabolismo , Neurônios/patologia , Fosfosserina/metabolismo , Doença de Alzheimer/patologia , Animais , Encéfalo/patologia , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Moléculas de Adesão Celular Neuronais/metabolismo , Técnicas de Cocultura , Interferon gama , Lipopolissacarídeos , Camundongos Endogâmicos C57BL , Fagocitose , Fosforilação , Transcrição Genética , Zimosan
15.
Am J Physiol Renal Physiol ; 320(6): F1123-F1132, 2021 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-33818127

RESUMO

Peritonitis, due to a fungal or bacterial infection, leads to injury of the peritoneal lining and thereby forms a hazard for the long-term success of peritoneal dialysis (PD) and remains a lethal complication in patients with PD. This study investigated whether C1 inhibitor (C1-INH) could protect against the progression of peritoneal injuries with five daily administrations of zymosan after mechanical scraping of the rat peritoneum to mimic fungal peritonitis. Severe peritoneal injuries were seen in this model, accompanied by fibrinogen/fibrin exudation and peritoneal deposition of complement activation products such as activated C3 and C5b-9. However, intraperitoneal injection of C1-INH decreased peritoneal depositions of activated C3 and C5b-9, ameliorated peritoneal thickening, reduced the influx of inflammatory cells, and prevented the production of peritoneal fibrous layers with both one and two doses of C1-INH each day. Our results suggest that C1-INH might be useful to protect against peritoneal injuries after causes of peritonitis such as fungal infection. This clinically available agent may thus help extend the duration of PD.NEW & NOTEWORTHY Peritoneal injuries associated with peritonitis comprise an important issue to prevent long-term peritoneal dialysis (PD) therapy. Here, we showed that C1 inhibitor (C1-INH), as an anticomplement agent, protected against peritoneal injuries in a peritonitis animal model related to fungal infection. Therefore, C1-INH might be useful to protect against peritoneal injuries after peritonitis due to fungal infection. This clinically available agent may thus help extend the duration of PD.


Assuntos
Proteína Inibidora do Complemento C1/uso terapêutico , Peritônio/efeitos dos fármacos , Peritonite/induzido quimicamente , Zimosan/toxicidade , Animais , Células Epiteliais , Epitélio , Fibrina/metabolismo , Fibrinogênio/metabolismo , Masculino , Peritônio/citologia , Peritônio/patologia , Ratos , Ratos Sprague-Dawley
16.
Int J Mol Sci ; 22(5)2021 Feb 24.
Artigo em Inglês | MEDLINE | ID: mdl-33668084

RESUMO

The interaction of macrophages with apoptotic cells is required for efficient resolution of inflammation. While apoptotic cell removal prevents inflammation due to secondary necrosis, it also alters the macrophage phenotype to hinder further inflammatory reactions. The interaction between apoptotic cells and macrophages is often studied by chemical or biological induction of apoptosis, which may introduce artifacts by affecting the macrophages as well and/or triggering unrelated signaling pathways. Here, we set up a pure cell death system in which NIH 3T3 cells expressing dimerizable Caspase-8 were co-cultured with peritoneal macrophages in a transwell system. Phenotype changes in macrophages induced by apoptotic cells were evaluated by RNA sequencing, which revealed an unexpectedly dominant impact on macrophage proliferation. This was confirmed in functional assays with primary peritoneal macrophages and IC-21 macrophages. Moreover, inhibition of apoptosis during Zymosan-induced peritonitis in mice decreased mRNA levels of cell cycle mediators in peritoneal macrophages. Proliferation of macrophages in response to apoptotic cells may be important to increase macrophage numbers in order to allow efficient clearance and resolution of inflammation.


Assuntos
Apoptose , Proliferação de Células , Macrófagos Peritoneais/citologia , Peritonite/patologia , Animais , Células Cultivadas , Técnicas de Cocultura , Macrófagos Peritoneais/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Peritonite/induzido quimicamente , Peritonite/metabolismo , Fagocitose , Zimosan/toxicidade
17.
Theranostics ; 11(9): 4251-4261, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33754059

RESUMO

Rationale: The interaction between coagulation and inflammation resolution remains elusive. We recently highlighted a link between fibrinogen-like protein 2 (Fgl2) and a specialized pro-resolving mediator (SPM)-n-3 docosapentaenoic acid-derived resolvin D5 (RvD5n-3 DPA) in sepsis. This study aimed to investigate the functions of commonly used anticoagulants warfarin, dabigatran and heparin in regulating inflammation resolution. Methods: Peripheral blood was collected from clinical sepsis patients and healthy control for the determination of indicated indexes. Mouse sepsis models of zymosan-induced peritonitis and cecal ligation and puncture (CLP) were employed for the measurement of inflammation- and coagulation-related indexes. Western-blotting, ELISA and flow cytometry were applied to assess proteins. UPLC-MS/MS was used to evaluate lipid metabolites. Results: Here we report that the transmembrane Fgl2 (mFgl2) was positively associated with coagulation, while soluble Fgl2 (sFgl2) level correlated with the enhanced number of peripheral blood mononuclear cells in the sepsis patients. The anticoagulants dabigatran and warfarin attenuated zymosan-induced peritonitis, which was not shared by heparin, while only dabigatran significantly improved sepsis survival in the CLP sepsis mouse model. Although these anticoagulants consistently inhibited pro-inflammatory mediators including prostaglandin E2 and leukotriene B4, only dabigatran increased sFgl2 at both the initiation and resolution phases of inflammation. Mechanistically, dabigatran elicited the shedding of sFgl2 via prothrombin-related metalloproteases, thereby enhanced the subsequent biosynthesis of RvD5n-3 DPA via STAT6-ALOX15 axis. Blocking metalloproteases or ALOX15 significantly impaired dabigatran-enhanced macrophage efferocytosis in vitro, as well as delayed the dabigatran-accelerated inflammation resolution in vivo. Conclusions: Our findings identify the dual anti-inflammatory and pro-resolving actions of dabigatran, through promoting sFgl2-triggered RvD5n-3 DPA production, which has important implications for promoting tissue homeostasis of sepsis.


Assuntos
Dabigatrana/farmacologia , Ácidos Docosa-Hexaenoicos/metabolismo , Ácidos Graxos Insaturados/metabolismo , Fibrinogênio/metabolismo , Inflamação/metabolismo , Animais , Anticoagulantes/farmacologia , Cromatografia Líquida/métodos , Modelos Animais de Doenças , Humanos , Inflamação/tratamento farmacológico , Mediadores da Inflamação/metabolismo , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/metabolismo , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Masculino , Metaloproteases/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Protrombina/metabolismo , Sepse/tratamento farmacológico , Sepse/metabolismo , Espectrometria de Massas em Tandem/métodos , Zimosan/farmacologia
18.
Neurourol Urodyn ; 40(3): 763-770, 2021 03.
Artigo em Inglês | MEDLINE | ID: mdl-33604897

RESUMO

AIMS: Interstitial cystitis (IC) is a chronic pain syndrome that is characterized by suprapubic pain upon bladder filling. Bletilla striata, a well-known traditional Chinese herb with established efficacy in wound healing and anti-inflammation, was hypothesized to improve the symptoms of IC possibly though forming a physical barrier that could isolate the bladder tissue from irritants. This study was conducted to evaluate the beneficial effects of intravesical treatment with B. striata extract solution (BSES) on visceral pain and bladder function of rats with zymosan-induced IC. METHODS: Thirty female rats were randomly divided into control group, zymosan-induced cystitis rats treated with normal saline (Z + NS), and zymosan-induced cystitis rats treated with BSES (Z + BSES). All rats underwent evaluation for abdominal withdrawal reflex (AWR) scores to assess visceral hypersensitivity, cystometrography, and electromyogram (EMG) of both external urethral sphincter and bladder detrusor. Data were analyzed by one way analysis of variance. RESULTS: The Z + NS group had an increased visceral hypersensitivity as compared to control group. Rats treated with BSES (Z + BSES group) had decreased AWR scores and amplitude of bladder detrusor-EMG. Besides, BSES treatment improved overactive bladder with significant effects on the extend of micturition interval and increase of storage of urine. CONCLUSIONS: Intravesical instillation of BSES can significantly alleviate zymosan-induced visceral hypersensitivity and bladder overactivity associated with IC. This study suggested that intravesical instillation with BSES might be a promising treatment for IC.


Assuntos
Cistite Intersticial/induzido quimicamente , Cistite Intersticial/tratamento farmacológico , Extratos Vegetais/uso terapêutico , Polissacarídeos/uso terapêutico , Zimosan/efeitos adversos , Animais , Feminino , Polissacarídeos/farmacologia , Ratos , Ratos Sprague-Dawley
19.
Genes (Basel) ; 12(2)2021 01 29.
Artigo em Inglês | MEDLINE | ID: mdl-33572768

RESUMO

Immunobiotics are probiotics that promote intestinal health by modulating immune responses. Immunobiotics are recognized by Toll-like receptors (TLRs) and activate cytokine gene expression. This study aimed to characterize cytokine gene expression in the chicken peripheral blood mononuclear cells (PBMC) stimulated with purified TLR ligands and live probiotics. PBMC were isolated from the whole blood. PBMC were stimulated with: lipopolysaccharide (LPS), CpG ODN, Pam3CSK4, Zymosan, galactooligosaccharides (GOS), Lactococcuslactis subsp. cremoris (L. lactis), and Saccharomyces cerevisiae at 42.5 °C and 5% CO2 for 3 h, 6 h, and 9 h. After each time-point, PBMC were harvested for RNA isolation. Relative gene expression was analyzed with RT-qPCR for cytokine genes (IL-1ß, IL-2, IL-3, IL-4, IL-6, IL-8, IL-10, IL-12p40, and IFN-É£) and reference genes (ACTB and G6PDH). Genes were clustered into pro-inflammatory genes, Th1/Th2 genes, and Th1-regulators. The gene expression differed between treatments in IL1-ß, IL-6, IL-8, IL-10, and IL-12p40 (p < 0.001). The genes IL-1ß, IL-6, and IL-8 had the highest fold change of mRNA expression at 3 h in response to TLR ligands. L. lactis up-regulated the pro-inflammatory genes at the 6 h time-point. L. lactis did not activate the anti-inflammatory IL-10 gene, but activated IL-12p40 at 6 h. Hereby, L. lactis was proven to exert immunostimulatory properties in PBMC.


Assuntos
Galinhas/sangue , Citocinas/genética , Leucócitos Mononucleares/efeitos dos fármacos , Células Th1/efeitos dos fármacos , Animais , Galinhas/microbiologia , Citocinas/sangue , Citocinas/classificação , Regulação da Expressão Gênica , Leucócitos/efeitos dos fármacos , Leucócitos/imunologia , Leucócitos Mononucleares/imunologia , Leucócitos Mononucleares/microbiologia , Lipopeptídeos/farmacologia , Lipopolissacarídeos/farmacologia , Oligossacarídeos/farmacologia , Probióticos/farmacologia , Saccharomyces cerevisiae/genética , Células Th1/imunologia , Receptores Toll-Like/genética , Zimosan/farmacologia
20.
Eur J Immunol ; 51(4): 864-878, 2021 04.
Artigo em Inglês | MEDLINE | ID: mdl-33616974

RESUMO

Intestinal epithelial cells (IECs) are the first to encounter luminal antigens and play an active role in intestinal immune responses. We previously reported that ß-glucans, major fungal cell-wall glycans, induced chemokine secretion by IEC lines in a Dectin-1- and Syk-dependent manner. Here, we show that in contrast to ß-glucans, stimulation of IEC lines with Candida albicans and Saccharomyces cerevisiae did not induce secretion of any of the proinflammatory cytokines IL-8, CCL2, CXCL1, and GM-CSF. Commensal fungi and ß-glucans activated Syk and ERK in IEC lines. However, only ß-glucans activated p38, JNK, and the transcription factors NF-κB p65 and c-JUN, which were necessary for cytokine secretion. Furthermore, costimulation of IEC lines with ß-glucans and C. albicans yielded decreased cytokine secretion compared to stimulation with ß-glucans alone. Finally, ex vivo stimulation of human colonic mucosal explants with zymosan and C. albicans, leads to epithelial Syk and ERK phosphorylation, implying recognition of fungi and similar initial signaling pathways as in IEC lines. Lack of cytokine secretion in response to commensal fungi may reflect IECs' response to fungal glycans, other than ß-glucans, that contribute to mucosal tolerance. Skewed epithelial response to commensal fungi may impair homeostasis and contribute to intestinal inflammation.


Assuntos
Candida albicans/imunologia , Parede Celular/imunologia , Células Epiteliais/imunologia , Mucosa Intestinal/imunologia , beta-Glucanas/imunologia , Células CACO-2 , Candida albicans/metabolismo , Candida albicans/fisiologia , Linhagem Celular Tumoral , Parede Celular/metabolismo , Citocinas/imunologia , Citocinas/metabolismo , Células Epiteliais/metabolismo , Células Epiteliais/microbiologia , MAP Quinases Reguladas por Sinal Extracelular/imunologia , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Células HT29 , Interações Hospedeiro-Patógeno/imunologia , Humanos , Mucosa Intestinal/microbiologia , Fosforilação/imunologia , Quinase Syk/imunologia , Quinase Syk/metabolismo , Zimosan/imunologia , Zimosan/metabolismo , beta-Glucanas/metabolismo
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