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1.
J Infect Dis ; 221(9): 1542-1553, 2020 04 07.
Artigo em Inglês | MEDLINE | ID: mdl-31783409

RESUMO

BACKGROUND: Liver X receptors (LXRs) are nuclear receptors activated by oxidized lipids and were previously implicated in several metabolic development and inflammatory disorders. Although neutrophils express both LXR-α and LXR-ß, the consequences of their activation, particularly during sepsis, remain unknown. METHODS: We used the model of cecal ligation and puncture (CLP) to investigate the role of LXR activation during sepsis. RESULTS: In this study, we verified that LXR activation reduces neutrophil chemotactic and killing abilities in vitro. Mice treated with LXR agonists showed higher sepsis-induced mortality, which could be associated with reduced neutrophil infiltration at the infectious foci, increased bacteremia, systemic inflammatory response, and multiorgan failure. In contrast, septic mice treated with LXR antagonist showed increased number of neutrophils in the peritoneal cavity, reduced bacterial load, and multiorgan dysfunction. More important, neutrophils from septic patients showed increased ABCA1 messenger ribonucleic acid levels (a marker of LXR activation) and impaired chemotactic response toward CXCL8 compared with cells from healthy individuals. CONCLUSIONS: Therefore, our findings suggest that LXR activation impairs neutrophil functions, which might contribute to poor sepsis outcome.


Assuntos
Receptores X do Fígado/metabolismo , Neutrófilos/patologia , Sepse/imunologia , Sepse/metabolismo , Transportador 1 de Cassete de Ligação de ATP/metabolismo , Adulto , Animais , Ceco/microbiologia , Ceco/cirurgia , Modelos Animais de Doenças , Feminino , Humanos , Inflamação , Interleucina-8/metabolismo , Ligadura , Receptores X do Fígado/agonistas , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Pessoa de Meia-Idade , Insuficiência de Múltiplos Órgãos/imunologia , Insuficiência de Múltiplos Órgãos/microbiologia , Infiltração de Neutrófilos/imunologia , Neutrófilos/metabolismo , Punções , Sepse/microbiologia
2.
J Immunol ; 191(3): 1373-82, 2013 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-23817413

RESUMO

Type 1 diabetes enhances susceptibility to infection and favors the sepsis development. In addition, diabetic mice produced higher levels of histamine in several tissues and in the blood after LPS stimulation than nondiabetic mice. In this study, we aimed to explore the role of mast cells (MCs) and histamine in neutrophil migration and, consequently, infection control in diabetic mice with mild sepsis (MS) induced by cecum ligation and puncture. We used female BALB/c, MC-sufficient (WB/B6), MC-deficient (W/W(v)), and NOD mice. Diabetic mice given MS displayed 100% mortality within 24 h, whereas all nondiabetic mice survived for at least 5 d. The mortality rate of diabetic mice was reduced to 57% after the depletion of MC granules with compound 48/80. Moreover, this pretreatment increased neutrophil migration to the focus of infection, which reduced systemic inflammatory response and bacteremia. The downregulation of CXCR2 and upregulation of G protein-coupled receptor kinase 2 in neutrophils was prevented by pretreatment of diabetic mice given MS with compound 48/80. In addition, blocking the histamine H2 receptor restored neutrophil migration, enhanced CXCR2 expression, decreased bacteremia, and improved sepsis survival in alloxan-induced diabetic and spontaneous NOD mice. Finally, diabetic W/W(v) mice had neutrophil migration to the peritoneal cavity, increased CXCR2 expression, and reduced bacteremia compared with diabetic WB/B6 mice. These results demonstrate that histamine released by MCs reduces diabetic host resistance to septic peritonitis in mice.


Assuntos
Diabetes Mellitus Experimental/mortalidade , Quinase 2 de Receptor Acoplado a Proteína G/metabolismo , Mastócitos/imunologia , Neutrófilos/metabolismo , Receptores de Interleucina-8B/metabolismo , Aloxano , Animais , Bacteriemia/tratamento farmacológico , Movimento Celular , Diabetes Mellitus Experimental/complicações , Diabetes Mellitus Experimental/microbiologia , Regulação para Baixo/efeitos dos fármacos , Feminino , Histamina/metabolismo , Antagonistas dos Receptores H2 da Histamina , Inflamação/tratamento farmacológico , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos NOD , Receptores Histamínicos H2/metabolismo , Sepse/complicações , Sepse/microbiologia , Sepse/mortalidade , Regulação para Cima/efeitos dos fármacos , p-Metoxi-N-metilfenetilamina/farmacologia
3.
iScience ; 26(10): 107719, 2023 Oct 20.
Artigo em Inglês | MEDLINE | ID: mdl-37674984

RESUMO

Little is known about the effects of high-fat diet (HFD)-induced obesity on resident colonic lamina propria (LP) macrophages (LPMs) function and metabolism. Here, we report that obesity and diabetes resulted in increased macrophage infiltration in the colon. These macrophages exhibited the residency phenotype CX3CR1hiMHCIIhi and were CD4-TIM4-. During HFD, resident colonic LPM exhibited a lipid metabolism gene expression signature that overlapped that used to define lipid-associated macrophages (LAMs). Via single-cell RNA sequencing, we identified a sub-cluster of macrophages, increased in HFD, that were responsible for the LAM signature. Compared to other macrophages in the colon, these cells were characterized by elevated glycolysis, phagocytosis, and efferocytosis signatures. CX3CR1hiMHCIIhi colonic resident LPMs had fewer lipid droplets (LDs) and decreased triacylglycerol (TG) content compared to equivalent cells in lean mice and exhibited increased phagocytic capacity, suggesting that HFD induces adaptive responses in LPMs to limit bacterial translocation.

4.
Crit Care Med ; 40(9): 2631-7, 2012 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-22732279

RESUMO

OBJECTIVES: To investigate the role of toll-like receptor 9 on sepsis-induced failure of neutrophil recruitment to the site of infection. DESIGN: Prospective experimental study. SETTING: University research laboratory. INTERVENTIONS: Model of polymicrobial sepsis induced by cecal ligation and puncture in wild-type and toll-like receptor 9-deficient mice. MEASUREMENTS AND MAIN RESULTS: Toll-like receptor 9-deficient mice with cecal ligation and puncture-induced severe sepsis did not demonstrate failure of neutrophil migration and consequently had a low systemic inflammatory response and a high survival rate. Upon investigating the mechanism by which toll-like receptor 9 deficiency prevents the failure of neutrophil migration, it was found that neutrophils derived from toll-like receptor 9--deficient mice with cecal ligation and puncture-induced severe sepsis expressed high levels of chemokine C-X-C motif receptor 2 (CXCR2) and had reduced induction of G-protein-coupled receptor kinase 2. CONCLUSIONS: These findings suggest that the poor outcome of severe sepsis is associated with toll-like receptor 9 activation in neutrophils, which triggers G-protein-coupled receptor kinase 2 expression and CXCR2 downregulation. These events account for the reduction of neutrophil migration to the site of infection, with consequent spreading of the infection, onset of the systemic inflammatory response, and a decrease in survival.


Assuntos
Quimiotaxia/fisiologia , Quinase 2 de Receptor Acoplado a Proteína G/metabolismo , Infiltração de Neutrófilos/fisiologia , Sepse/metabolismo , Receptor Toll-Like 9/metabolismo , Animais , Sobrevivência Celular , Modelos Animais de Doenças , Feminino , Camundongos , Camundongos Endogâmicos C57BL , Distribuição Aleatória , Valores de Referência , Sensibilidade e Especificidade , Estatísticas não Paramétricas
5.
Am J Respir Crit Care Med ; 183(2): 234-42, 2011 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-20732989

RESUMO

RATIONALE: Sepsis is defined as a systemic inflammatory response to infection, which in its severe form is associated with multiple organ dysfunction syndrome (MODS). The precise mechanisms by which MODS develops remain unclear. Neutrophils have a pivotal role in the defense against infections; however, overwhelming activation of neutrophils is known to elicit tissue damage. OBJECTIVES: We investigated the role of the chemokine receptor CCR2 in driving neutrophil infiltration and eliciting tissue damage in remote organs during sepsis. METHODS: Sepsis was induced in wild-type mice treated with CCR2 antagonist (RS504393) or CCR2(-/-) mice by cecal ligation and puncture (CLP) model. Neutrophil infiltration into the organs was measured by myeloperoxidase activity and fluorescence-activated cell sorter. CCR2 expression and chemotaxis were determined in neutrophils stimulated with Toll-like receptor agonists or isolated from septic mice and patients. MEASUREMENTS AND MAIN RESULTS: CCR2 expression and responsiveness to its ligands was induced in circulating neutrophils during CLP-induced sepsis by a mechanism dependent on Toll-like receptor/nuclear factor-κB pathway. Genetic or pharmacologic inhibition of CCR2 protected mice from CLP-induced mortality. This protection was associated with lower infiltration of neutrophils into the lungs, heart, and kidneys and reduced serum biochemical indicators of organ injury and dysfunction. Importantly, neutrophils from septic patients express high levels of CCR2, and the severity of patient illness correlated positively with increasing neutrophil chemotaxis to CCR2 ligands. CONCLUSIONS: Collectively, these data identify CCR2 as a key receptor that drives the inappropriate infiltration of neutrophils into remote organs during sepsis. Therefore, CCR2 blockade is a novel potential therapeutic target for treatment of sepsis-induced MODS.


Assuntos
Insuficiência de Múltiplos Órgãos/sangue , Neutrófilos/metabolismo , Receptores CCR2/sangue , Choque Séptico/sangue , Animais , Biomarcadores/sangue , Quimiotaxia , Modelos Animais de Doenças , Ensaio de Imunoadsorção Enzimática/métodos , Citometria de Fluxo/métodos , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Insuficiência de Múltiplos Órgãos/etiologia , Peroxidase/sangue , Índice de Gravidade de Doença , Choque Séptico/complicações , Regulação para Cima
6.
Am J Respir Crit Care Med ; 183(7): 922-31, 2011 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-20971829

RESUMO

RATIONALE: The reduction of neutrophil migration to the bacterial focus is associated with poor outcome in sepsis. OBJECTIVES: The objective of this study was to identify soluble substances in the blood of septic mice that inhibit neutrophil migration. METHODS: A pool of serum obtained from mice 2 hours after the induction of severe sepsis by cecal ligation and puncture inhibited the neutrophil migration. The proteins with inhibitory activity on neutrophil migration were isolated by Blue-Sepharose chromatography, high-performance liquid chromatography, and electrophoresis, and identified by mass spectrometry. MEASUREMENTS AND MAIN RESULTS: Hemopexin was identified as the serum component responsible for the inhibition of neutrophil migration. In sepsis, the pretreatment of wild-type mice with hemopexin inhibited neutrophil migration to the focus of infection and decreased the survival rate from 87.5 to 50.0%. Hemopexin-null mice subjected to severe sepsis presented normal neutrophil migration, low bacteremia, and an improvement of 40% in survival rate. Moreover, hemopexin inhibited the neutrophil chemotaxis response evoked by C5a or macrophage inflammatory protein-2 and induced a reduction of CXCR2 and L-selectin as well as the up-regulation of CD11b expression in neutrophil membranes. The inhibitory effect of hemopexin on neutrophil chemotaxis was prevented by serine protease inhibitors or ATP. In addition, serum levels of ATP were decreased 2 hours after severe sepsis. CONCLUSIONS: These data demonstrate for the first time the inhibitory role of hemopexin in neutrophil migration during sepsis and suggest that the therapeutic inhibition of hemopexin or its protease activity could improve neutrophil migration to the focus of infection and survival in sepsis.


Assuntos
Movimento Celular/efeitos dos fármacos , Hemopexina/metabolismo , Neutrófilos/metabolismo , Sepse/metabolismo , Sepse/mortalidade , Análise de Variância , Animais , Antígeno CD11b/imunologia , Antígeno CD11b/metabolismo , Movimento Celular/imunologia , Quimiotaxia de Leucócito/imunologia , Modelos Animais de Doenças , Regulação para Baixo , Escherichia coli , Hemopexina/imunologia , Selectina L/metabolismo , Espectrometria de Massas , Camundongos , Camundongos Endogâmicos , Neutrófilos/efeitos dos fármacos , Neutrófilos/imunologia , Distribuição Aleatória , Receptores de Interleucina-8B/imunologia , Receptores de Interleucina-8B/metabolismo , Sepse/imunologia , Taxa de Sobrevida , Tioglicolatos/farmacologia , Regulação para Cima
7.
Proc Natl Acad Sci U S A ; 106(10): 4018-23, 2009 Mar 10.
Artigo em Inglês | MEDLINE | ID: mdl-19234125

RESUMO

Patients with sepsis have a marked defect in neutrophil migration. Here we identify a key role of Toll-like receptor 2 (TLR2) in the regulation of neutrophil migration and resistance during polymicrobial sepsis. We found that the expression of the chemokine receptor CXCR2 was dramatically down-regulated in circulating neutrophils from WT mice with severe sepsis, which correlates with reduced chemotaxis to CXCL2 in vitro and impaired migration into an infectious focus in vivo. TLR2 deficiency prevented the down-regulation of CXCR2 and failure of neutrophil migration. Moreover, TLR2(-/-) mice exhibited higher bacterial clearance, lower serum inflammatory cytokines, and improved survival rate during severe sepsis compared with WT mice. In vitro, the TLR2 agonist lipoteichoic acid (LTA) down-regulated CXCR2 expression and markedly inhibited the neutrophil chemotaxis and actin polymerization induced by CXCL2. Moreover, neutrophils activated ex vivo by LTA and adoptively transferred into naïve WT recipient mice displayed a significantly reduced competence to migrate toward thioglycolate-induced peritonitis. Finally, LTA enhanced the expression of G protein-coupled receptor kinases 2 (GRK2) in neutrophils; increased expression of GRK2 was seen in blood neutrophils from WT mice, but not TLR2(-/-) mice, with severe sepsis. Our findings identify an unexpected detrimental role of TLR2 in polymicrobial sepsis and suggest that inhibition of TLR2 signaling may improve survival from sepsis.


Assuntos
Movimento Celular , Neutrófilos/citologia , Receptores de Interleucina-8B/metabolismo , Sepse/imunologia , Sepse/microbiologia , Receptor 2 Toll-Like/metabolismo , Animais , Movimento Celular/efeitos dos fármacos , Quimiotaxia/efeitos dos fármacos , Regulação para Baixo/efeitos dos fármacos , Quinase 2 de Receptor Acoplado a Proteína G/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Imunidade Inata/efeitos dos fármacos , Lipopolissacarídeos/administração & dosagem , Lipopolissacarídeos/farmacologia , Camundongos , Neutrófilos/efeitos dos fármacos , Neutrófilos/enzimologia , Peritonite/complicações , Receptores de Interleucina-8B/genética , Sepse/complicações , Transdução de Sinais/efeitos dos fármacos , Análise de Sobrevida , Ácidos Teicoicos/administração & dosagem , Ácidos Teicoicos/farmacologia , Receptor 2 Toll-Like/agonistas , Receptor 2 Toll-Like/deficiência
8.
Proc Natl Acad Sci U S A ; 106(14): 5954-9, 2009 Apr 07.
Artigo em Inglês | MEDLINE | ID: mdl-19289819

RESUMO

IL-23/IL-17-induced neutrophil recruitment plays a pivotal role in rheumatoid arthritis (RA). However, the mechanism of the neutrophil recruitment is obscure. Here we report that prostaglandin enhances the IL-23/IL-17-induced neutrophil migration in a murine model of RA by inhibiting IL-12 and IFN gamma production. Methylated BSA (mBSA) and IL-23-induced neutrophil migration was inhibited by anti-IL-23 and anti-IL-17 antibodies, COX inhibitors, IL-12, or IFNgamma but was enhanced by prostaglandin E(2) (PGE(2)). IL-23-induced IL-17 production was increased by PGE(2) and suppressed by COX-inhibition or IL-12. Furthermore, COX inhibition failed to reduce IL-23-induced neutrophil migration in IL-12- or IFNgamma-deficient mice. IL-17-induced neutrophil migration was not affected by COX inhibitors, IL-12, or IFNgamma but was inhibited by MK886 (a leukotriene synthesis inhibitor), anti-TNFalpha, anti-CXCL1, and anti-CXCL5 antibodies and by repertaxin (a CXCR1/2 antagonist). These treatments all inhibited mBSA- or IL-23-induced neutrophil migration. IL-17 induced neutrophil chemotaxis through a CXC chemokines-dependent pathway. Our results suggest that prostaglandin plays an important role in IL-23-induced neutrophil migration in arthritis by enhancing IL-17 synthesis and by inhibiting IL-12 and IFNgamma production. We thus provide a mechanism for the pathogenic role of the IL-23/IL-17 axis in RA and also suggest an additional mechanism of action for nonsteroidal anti-inflammatory drugs.


Assuntos
Artrite Reumatoide/patologia , Inflamação/metabolismo , Interferon gama/antagonistas & inibidores , Interleucina-12/antagonistas & inibidores , Interleucina-17/imunologia , Interleucina-23/imunologia , Infiltração de Neutrófilos/imunologia , Prostaglandinas/fisiologia , Animais , Inibidores de Ciclo-Oxigenase/farmacologia , Dinoprostona/farmacologia , Interleucina-17/biossíntese , Camundongos
9.
J Nat Prod ; 74(2): 113-8, 2011 Feb 25.
Artigo em Inglês | MEDLINE | ID: mdl-21275387

RESUMO

Recent in vitro data have suggested that the flavonoid quercetin (1) does not affect the functioning of neutrophils. Therefore, we evaluated in vivo and in vitro whether or not 1 affects neutrophil function, focusing on recruitment. The in vivo treatment with 1 inhibited in a dose-dependent manner the recruitment of neutrophils to the peritoneal cavity of mice induced by known chemotatic factors such as CXCL1, CXCL5, LTB(4), and fMLP. Furthermore, 1 also inhibited in a concentration-dependent manner the chemoattraction of human neutrophils induced by CXCL8, LTB(4), and fMLP in a Boyden chamber. In vitro treatment with 1 did not affect human neutrophil surface expression of CXCR1, CXCR2, BLT1, or FLPR1, but rather reduced actin polymerization. These results suggest that 1 inhibits actin polymerization, hence, explaining the inhibition of neutrophil recruitment in vivo and in vitro and highlighting its possible usefulness to diminish excessive neutrophil migration during inflammation.


Assuntos
Actinas/metabolismo , Quimiocina CXCL5/imunologia , Interleucina-8/efeitos dos fármacos , Leucotrieno B4/imunologia , N-Formilmetionina Leucil-Fenilalanina/imunologia , Infiltração de Neutrófilos/efeitos dos fármacos , Neutrófilos/efeitos dos fármacos , Quercetina/farmacologia , Actinas/efeitos dos fármacos , Animais , Fatores Quimiotáticos/imunologia , Quimiotaxia/imunologia , Relação Dose-Resposta a Droga , Humanos , Inflamação/imunologia , Interleucina-8/imunologia , Masculino , Camundongos , Estrutura Molecular , Infiltração de Neutrófilos/imunologia , Neutrófilos/imunologia , Quercetina/química , Quercetina/imunologia
10.
Am J Respir Crit Care Med ; 182(3): 360-8, 2010 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-20339148

RESUMO

RATIONALE: Recovering the neutrophil migration to the infectious focus improves survival in severe sepsis. Recently, we demonstrated that the cystathionine gamma-lyase (CSE)/hydrogen sulfide (H(2)S) pathway increased neutrophil recruitment to inflammatory focus during sterile inflammation. OBJECTIVES: To evaluate if H(2)S administration increases neutrophil migration to infectious focus and survival of mice. METHODS: Sepsis was induced by cecal ligation and puncture (CLP). MEASUREMENTS AND MAIN RESULTS: The pretreatments of mice with H(2)S donors (NaHS or Lawesson's reagent) improved leukocyte rolling/adhesion in the mesenteric microcirculation as well as neutrophil migration. Consequently, bacteremia levels were reduced, hypotension and lung lesions were prevented, and the survival rate increased from approximately 13% to approximately 80%. Even when treatment was delayed (6 h after CLP), a highly significant reduction in mortality compared with untreated mice was observed. Moreover, H(2)S pretreatment prevented the down-regulation of CXCR2 and l-selectin and the up-regulation of CD11b and G protein-coupled receptor kinase 2 in neutrophils during sepsis. H(2)S also prevented the reduction of intercellular adhesion molecule-1 expression in the endothelium of the mesenteric microcirculation in severe sepsis. Confirming the critical role of H(2)S on sepsis outcome, pretreatment with dl-propargylglycine (a CSE inhibitor) inhibited neutrophil migration to the infectious focus, enhanced lung lesions, and induced high mortality in mice subjected to nonsevere sepsis (from 0 to approximately 80%). The beneficial effects of H(2)S were blocked by glibenclamide (a ATP-dependent K(+) channel blocker). CONCLUSIONS: These results showed that H(2)S restores neutrophil migration to the infectious focus and improves survival outcome in severe sepsis by an ATP-dependent K(+) channel-dependent mechanism.


Assuntos
Movimento Celular/efeitos dos fármacos , Sulfeto de Hidrogênio/farmacologia , Canais KATP/fisiologia , Neutrófilos/efeitos dos fármacos , Sepse/mortalidade , Sepse/patologia , Animais , Antígeno CD11b/fisiologia , Regulação para Baixo/efeitos dos fármacos , Endotélio Vascular , Molécula 1 de Adesão Intercelular/efeitos dos fármacos , Selectina L/fisiologia , Masculino , Mesentério/irrigação sanguínea , Camundongos , Neutrófilos/fisiologia , Receptores de Interleucina-8B/fisiologia , Regulação para Cima/efeitos dos fármacos
11.
Am J Respir Crit Care Med ; 182(6): 762-73, 2010 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-20508212

RESUMO

RATIONALE: Sepsis is a leading cause of death in the intensive care unit, characterized by a systemic inflammatory response (SIRS) and bacterial infection, which can often induce multiorgan damage and failure. Leukocyte recruitment, required to limit bacterial spread, depends on phosphoinositide-3 kinase γ (PI3Kγ) signaling in vitro; however, the role of this enzyme in polymicrobial sepsis has remained unclear. OBJECTIVES: This study aimed to determine the specific role of the kinase activity of PI3Kγ in the pathogenesis of sepsis and multiorgan damage. METHODS: PI3Kγ wild-type, knockout, and kinase-dead mice were exposed to cecal ligation and perforation-induced sepsis and assessed for survival; pulmonary, hepatic, and cardiovascular damage; coagulation derangements; systemic inflammation; bacterial spread; and neutrophil recruitment. Additionally, wild-type mice were treated either before or after the onset of sepsis with a PI3Kγ inhibitor and assessed for survival, neutrophil recruitment, and bacterial spread. MEASUREMENTS AND MAIN RESULTS: Both genetic and pharmaceutical PI3Kγ kinase inhibition significantly improved survival, reduced multiorgan damage, and limited bacterial decompartmentalization, while modestly affecting SIRS. Protection resulted from both neutrophil-independent mechanisms, involving improved cardiovascular function, and neutrophil-dependent mechanisms, through reduced susceptibility to neutrophil migration failure during severe sepsis by maintaining neutrophil surface expression of the chemokine receptor, CXCR2. Furthermore, PI3Kγ pharmacological inhibition significantly decreased mortality and improved neutrophil migration and bacterial control, even when administered during established septic shock. CONCLUSIONS: This study establishes PI3Kγ as a key molecule in the pathogenesis of septic infection and the transition from SIRS to organ damage and identifies it as a novel possible therapeutic target.


Assuntos
Insuficiência de Múltiplos Órgãos/enzimologia , Infiltração de Neutrófilos/fisiologia , Fosfatidilinositol 3-Quinases/fisiologia , Sepse/enzimologia , Animais , Quimiotaxia de Leucócito , Classe Ib de Fosfatidilinositol 3-Quinase , Modelos Animais de Doenças , Inibidores Enzimáticos/farmacologia , Isoenzimas/antagonistas & inibidores , Isoenzimas/fisiologia , Estimativa de Kaplan-Meier , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Insuficiência de Múltiplos Órgãos/prevenção & controle , Inibidores de Fosfoinositídeo-3 Quinase , Sepse/prevenção & controle , Transdução de Sinais/efeitos dos fármacos , Síndrome de Resposta Inflamatória Sistêmica/enzimologia , Síndrome de Resposta Inflamatória Sistêmica/prevenção & controle
12.
Ann Rheum Dis ; 69(9): 1697-703, 2010 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-20472598

RESUMO

OBJECTIVES: Interleukin 33 (IL-33) is a new member of the IL-1 family of cytokines which signals via its receptor, ST2 (IL-33R), and has an important role in Th2 and mast cell responses. This study shows that IL-33 orchestrates neutrophil migration in arthritis. METHODS AND RESULTS: Methylated bovine serum albumin (mBSA) challenge in the knee joint of mBSA-immunised mice induced local neutrophil migration accompanied by increased IL-33R and IL-33 mRNA expression. Cell migration was inhibited by systemic and local treatments with soluble (s)IL-33R, an IL-33 decoy receptor, and was not evident in IL-33R-deficient mice. IL-33 injection also induced IL-33R-dependent neutrophil migration. Antigen- and IL-33-induced neutrophil migration in the joint was dependent on CXCL1, CCL3, tumour necrosis factor alpha (TNFalpha) and IL-1beta synthesis. Synovial tissue, macrophages and activated neutrophils expressed IL-33R. IL-33 induces neutrophil migration by activating macrophages to produce chemokines and cytokines and by directly acting on neutrophils. Importantly, neutrophils from patients with rheumatoid arthritis successfully treated with anti-TNFalpha antibody (infliximab) expressed significantly lower levels of IL-33R than patients treated with methotrexate alone. Only neutrophils from patients treated with methotrexate alone or from normal donors stimulated with TNFalpha responded to IL-33 in chemotaxis. CONCLUSIONS: These results suggest that suppression of IL-33R expression in neutrophils, preventing IL-33-induced neutrophil migration, may be an important mechanism of anti-TNFalpha therapy of inflammation.


Assuntos
Antirreumáticos/farmacologia , Artrite Reumatoide/imunologia , Interleucinas/imunologia , Infiltração de Neutrófilos/imunologia , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Animais , Artrite Experimental/tratamento farmacológico , Artrite Experimental/imunologia , Artrite Reumatoide/tratamento farmacológico , Fatores Quimiotáticos/imunologia , Quimiotaxia de Leucócito/imunologia , Citocinas/imunologia , Regulação da Expressão Gênica/imunologia , Humanos , Proteína 1 Semelhante a Receptor de Interleucina-1 , Interleucina-33 , Interleucinas/biossíntese , Interleucinas/genética , Ativação de Macrófagos/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , RNA Mensageiro/genética , Receptores de Interleucina , Membrana Sinovial/imunologia
13.
J Immunol ; 181(6): 4287-98, 2008 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-18768887

RESUMO

In this study, we have addressed the role of H(2)S in modulating neutrophil migration in either innate (LPS-challenged naive mice) or adaptive (methylated BSA (mBSA)-challenged immunized mice) immune responses. Treatment of mice with H(2)S synthesis inhibitors, dl-propargylglycine (PAG) or beta-cyanoalanine, reduced neutrophil migration induced by LPS or methylated BSA (mBSA) into the peritoneal cavity and by mBSA into the femur/tibial joint of immunized mice. This effect was associated with decreased leukocyte rolling, adhesion, and P-selectin and ICAM-1 expression on endothelium. Predictably, treatment of animals with the H(2)S donors, NaHS or Lawesson's reagent, enhanced these parameters. Moreover, the NaHS enhancement of neutrophil migration was not observed in ICAM-1-deficient mice. Neither PAG nor NaHS treatment changed LPS-induced CD18 expression on neutrophils, nor did the LPS- and mBSA-induced release of neutrophil chemoattractant mediators TNF-alpha, keratinocyte-derived chemokine, and LTB(4). Furthermore, in vitro MIP-2-induced neutrophil chemotaxis was inhibited by PAG and enhanced by NaHS treatments. Accordingly, MIP-2-induced CXCR2 internalization was enhanced by PAG and inhibited by NaHS treatments. Moreover, NaHS prevented MIP-2-induced CXCR2 desensitization. The PAG and NaHS effects correlated, respectively, with the enhancement and inhibition of MIP-2-induced G protein-coupled receptor kinase 2 expression. The effects of NaHS on neutrophil migration both in vivo and in vitro, together with CXCR2 internalization and G protein-coupled receptor kinase 2 expression were prevented by the ATP-sensitive potassium (K(ATP)(+)) channel blocker, glybenclamide. Conversely, diazoxide, a K(ATP)(+) channel opener, increased neutrophil migration in vivo. Together, our data suggest that during the inflammatory response, H(2)S augments neutrophil adhesion and locomotion, by a mechanism dependent on K(ATP)(+) channels.


Assuntos
Adjuvantes Imunológicos/farmacologia , Moléculas de Adesão Celular/biossíntese , Endocitose/imunologia , Regulação da Expressão Gênica/imunologia , Sulfeto de Hidrogênio/farmacologia , Canais KATP/fisiologia , Infiltração de Neutrófilos/imunologia , Receptores de Interleucina-8B/antagonistas & inibidores , Adjuvantes Imunológicos/biossíntese , Animais , Bovinos , Moléculas de Adesão Celular/deficiência , Moléculas de Adesão Celular/genética , Endocitose/efeitos dos fármacos , Regulação da Expressão Gênica/efeitos dos fármacos , Sulfeto de Hidrogênio/metabolismo , Imunidade Inata/efeitos dos fármacos , Imunidade Inata/genética , Lipopolissacarídeos/farmacologia , Masculino , Metilação/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Infiltração de Neutrófilos/efeitos dos fármacos , Receptores de Interleucina-8B/metabolismo , Soroalbumina Bovina/administração & dosagem , Soroalbumina Bovina/imunologia , Soroalbumina Bovina/metabolismo
14.
J Photochem Photobiol B ; 209: 111918, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32531690

RESUMO

Folic acid (FA) regulates metabolic activities essential to the human body. FA receptor (FR) overexpression has been reported for many cancers, but there are still few or conflicting data about FRs in breast cancer cells. Quantum dots (QDs) have arisen as tools to elucidate aspects on FRs, due to their unique physicochemical properties. Herein, QDs conjugated to FA were explored to study the internalization and recycling of FRs in breast cancer cells, using HeLa as an out-group control. QDs were covalently conjugated to FA under different conditions. The best conjugate was applied to study FRs in HeLa, MCF7, MDA-MB231, and T47D cells applying confocal microscopy and flow cytometry analyses. The conjugation efficiency and specificity were evaluated, respectively, using fluorescence correlation spectroscopy (FCS) and saturation assays. FCS confirmed the effectiveness of the conjugation. HeLa and T47D had/internalized a higher amount of FRs (95% and 90% of labeling, respectively) than MDA-MB231 cells (68%). MCF7 cells seem to have very low functional FRs (3%). Saturation assays proved the specificity of QD-FA conjugates and suggested that FR recycling rate is low in the majority of cells studied, except for T47D. QD-FA conjugates were successfully developed. Therapies targeting FRs may be more effective for HeLa, T47D, and MDA-MB231.


Assuntos
Neoplasias da Mama/metabolismo , Endocitose , Ácido Fólico/metabolismo , Pontos Quânticos , Receptores de Superfície Celular/metabolismo , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Feminino , Citometria de Fluxo , Humanos , Microscopia Confocal , Espectrometria de Fluorescência
15.
Sci Rep ; 7(1): 5520, 2017 07 14.
Artigo em Inglês | MEDLINE | ID: mdl-28710436

RESUMO

Increased expression of Interleukin (IL)-33 has been detected in intestinal samples of patients with ulcerative colitis, a condition associated with increased risk for colon cancer, but its role in the development of colorectal cancer has yet to be fully examined. Here, we investigated the role of epithelial expressed IL-33 during development of intestinal tumors. IL-33 expression was detected in epithelial cells in colorectal cancer specimens and in the Apc Min/+ mice. To better understand the role of epithelial-derived IL-33 in the intestinal tumorigenesis, we generated transgenic mice expressing IL-33 in intestinal epithelial cells (V33 mice). V33 Apc Min/+ mice, resulting from the cross of V33 with Apc Min/+ mice, had increased intestinal tumor burden compared with littermate Apc Min/+ mice. Consistently, Apc Min/+ mice deficient for IL-33 receptor (ST2), had reduced polyp burden. Mechanistically, overexpression of IL-33 promoted expansion of ST2+ regulatory T cells, increased Th2 cytokine milieu, and induced alternatively activated macrophages in the gut. IL-33 promoted marked changes in the expression of antimicrobial peptides, and antibiotic treatment of V33 Apc Min/+ mice abrogated the tumor promoting-effects of IL-33 in the colon. In conclusion, elevated IL-33 signaling increases tumor development in the Apc Min/+ mice.


Assuntos
Proteína da Polipose Adenomatosa do Colo/genética , Transformação Celular Neoplásica/imunologia , Células Epiteliais/metabolismo , Interleucina-33/metabolismo , Neoplasias Intestinais/imunologia , Animais , Transformação Celular Neoplásica/genética , Células Cultivadas , Citocinas/metabolismo , Células Epiteliais/citologia , Regulação Neoplásica da Expressão Gênica , Humanos , Proteína 1 Semelhante a Receptor de Interleucina-1/metabolismo , Interleucina-33/genética , Neoplasias Intestinais/genética , Ativação de Macrófagos , Macrófagos/imunologia , Camundongos , Camundongos Transgênicos , Transdução de Sinais , Linfócitos T Reguladores/imunologia , Células Th2/imunologia , Regulação para Cima
16.
Inflammation ; 40(6): 2020-2032, 2017 12.
Artigo em Inglês | MEDLINE | ID: mdl-28780730

RESUMO

Rheumatoid arthritis (RA) is a chronic autoimmune disease characterized by debilitating pain, cartilage destruction, and loss of joint function. Management of RA includes drugs that target NF-κB and downstream cytokine production. Therefore, molecules that act by inhibiting this signaling pathway without the severe side effects of, for instance, corticoids would be suitable therapeutic strategies. Budlein A is a sesquiterpene lactone with antinociceptive and anti-inflammatory properties related to the inhibition of pro-inflammatory cytokines and neutrophil recruitment. In this study, the effect of budlein A was evaluated in antigen-induced arthritis (AIA) in mice. At the 26th day, leukocyte recruitment to the knee joint, knee contents of proteoglycans, blood levels of ALT and AST, stomach tissue myeloperoxidase activity, and RT-qPCR for pro-inflammatory gene mRNA expression in knee joint samples was performed. NF-κB luciferase activity was evaluated in RAW 264.7 macrophages. Budlein A treatment dose-dependently inhibited AIA-induced mechanical hyperalgesia, edema, total leukocytes and neutrophil recruitment, and proteoglycan degradation. Budlein A did not induce gastric or liver damage. Budlein also inhibited AIA-induced Il-33, Tnf, Il-1ß, preproET-1, and Cox-2 mRNA expression. In vitro, budlein reduced TNF- and IL-1ß-induced NF-κB activity in RAW 264.7 macrophages. Altogether, we demonstrate that budlein A ameliorates AIA-induced inflammation and pain by targeting NF-κB. Importantly, budlein A does not induce in vivo side effects, suggesting that it possesses a favorable pre-clinical profile as analgesic and it is a prosperous molecule to be further investigated for the treatment of RA.


Assuntos
Artrite Experimental/tratamento farmacológico , Lactonas/farmacologia , Sesquiterpenos/farmacologia , Animais , Antígenos/efeitos adversos , Artrite Experimental/induzido quimicamente , Citocinas/efeitos dos fármacos , Inflamação/prevenção & controle , Camundongos , NF-kappa B/antagonistas & inibidores , Dor/prevenção & controle , Células RAW 264.7
17.
Arthritis Rheumatol ; 67(7): 1751-9, 2015 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-25779331

RESUMO

OBJECTIVE: Infiltration of neutrophils into the joints plays an important role in bone erosion and articular destruction in rheumatoid arthritis (RA). Neutrophil trafficking during inflammation is a process that involves activation of chemotactic receptors. Recent findings suggest that changes in chemotactic receptor patterns could occur in neutrophils under certain inflammatory conditions. The aim of this study was to evaluate the gain of responsiveness of neutrophils to CCL2 in RA patients and to assess the role of CCL2 in driving neutrophil infiltration into the joints. METHODS: Neutrophils were purified from the peripheral blood of patients with RA or from mice with antigen-induced arthritis (AIA). Expression of CCR2 was evaluated using polymerase chain reaction, flow cytometry, and immunofluorescence analyses. In vitro chemotaxis to CCL2 was assayed to evaluate the functional significance of de novo CCR2 expression. The murine AIA model was used to evaluate the in vivo role of CCR2 in neutrophil infiltration into the joints. RESULTS: High CCR2 expression and responsiveness to CCL2 were observed in neutrophils from the blood of patients with early RA and in neutrophils from the blood and bone marrow of mice with AIA. Genetic deficiency or pharmacologic inhibition of CCR2 protected against the infiltration of neutrophils into the joints. This protection was not associated with an impairment of the neutrophil chemotactic ability or CXC chemokine production in the joints. Moreover, adoptive transfer of wild-type mouse neutrophils to CCR2-deficient mice restored neutrophil infiltration and the articular mechanical hyperalgesia associated with joint inflammation. CONCLUSION: These findings suggest that CCR2 is directly involved in the detrimental infiltration of neutrophils into the joints in patients with RA, showing a new inflammatory role of CCR2 during RA flares or active disease.


Assuntos
Artrite Reumatoide/patologia , Artrite Reumatoide/fisiopatologia , Movimento Celular/fisiologia , Articulações/patologia , Neutrófilos/patologia , Receptores CCR2/metabolismo , Animais , Artrite Reumatoide/metabolismo , Estudos de Casos e Controles , Movimento Celular/efeitos dos fármacos , Quimiocina CCL2/farmacologia , Quimiotaxia/fisiologia , Modelos Animais de Doenças , Feminino , Humanos , Técnicas In Vitro , Articulações/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Infiltração de Neutrófilos/efeitos dos fármacos , Infiltração de Neutrófilos/fisiologia , Neutrófilos/efeitos dos fármacos , Neutrófilos/metabolismo , Receptores CCR2/deficiência , Receptores CCR2/genética , Índice de Gravidade de Doença
18.
Cancer Chemother Pharmacol ; 73(2): 317-24, 2014 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-24258454

RESUMO

PURPOSE: To investigate the mechanisms affecting neutrophil migration capacity in breast cancer patients before and after chemotherapy. METHODS: Peripheral venous blood was collected at the time of diagnosis and immediately prior to the 4th cycle of an anthracycline-based chemotherapy regimen for patients diagnosed with different stages of breast cancer (n = 30), for experimental assays. Blood samples were also collected from a healthy control group (n = 17). RESULTS: IL-8 serum concentrations were higher in the patient group than in the control group (p = 0.02), and chemotherapy did not further affect this increase. Levels of TNF-α, IL-6, and IL-10 did not differ between controls and patients, or in relation to chemotherapy. Serum levels of nitric oxide (NO) metabolites were elevated following chemotherapy compared to levels detected prior to treatment (p = 0.01). When the supernatants of lipopolysaccharide-stimulated mononuclear cells and neutrophils obtained from the patients were assayed for levels of nitrite, these levels were significantly higher and unchanged, respectively, compared with controls. Expression levels of the chemokine receptors, CXCR1 and CXCR2, were significantly reduced in patients compared to controls, and chemotherapy did not further affect these differences. Furthermore, filamentous actin content for IL-8-activated neutrophils was reduced with chemotherapy (median 8.85; range 3.38-13.43) compared to the content detected prior to treatment (median 9.23; range 2.86-22.16) (p = 0.001). CONCLUSION: Elevated systemic levels of IL-8 and NO, desensitization to CXCR activation, and reduction in actin polymerization may affect neutrophil motility in patients before and after chemotherapy.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Neoplasias da Mama/sangue , Neoplasias da Mama/tratamento farmacológico , Movimento Celular/fisiologia , Doenças do Sistema Imunitário/patologia , Transtornos Leucocíticos/patologia , Neutrófilos/patologia , Actinas/sangue , Adulto , Idoso , Neoplasias da Mama/patologia , Estudos de Casos e Controles , Ciclofosfamida/administração & dosagem , Citocinas/sangue , Doxorrubicina/administração & dosagem , Epirubicina/administração & dosagem , Feminino , Fluoruracila/administração & dosagem , Humanos , Interleucina-8/sangue , Pessoa de Meia-Idade , Neutrófilos/metabolismo , Óxido Nítrico/sangue , Receptores CXCR/sangue
19.
PLoS One ; 9(8): e103734, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-25084278

RESUMO

Pathogen recognition and triggering of the inflammatory response following infection in mammals depend mainly on Toll-like and Nod-like receptors. Here, we evaluated the role of Nod1, Nod2 and MyD88-dependent signaling in the chemokine production and neutrophil recruitment to the infectious site during sepsis induced by cecal ligation and puncture (CLP) in C57Bl/6 mice. We demonstrate that Nod1 and Nod2 are not involved in the release of chemokines and recruitment of neutrophils to the infectious site during CLP-induced septic peritonitis because these events were similar in wild-type, Nod1-, Nod2-, Nod1/Nod2- and Rip2-deficient mice. Consequently, the local and systemic bacterial loads were not altered. Accordingly, neither Nod1 nor Nod2 was involved in the production of the circulating cytokines and in the accumulation of leukocytes in the lungs. By contrast, we showed that MyD88-dependent signaling is crucial for the establishment of the local inflammatory response during CLP-induced sepsis. MyD88-deficient mice were susceptible to sepsis because of an impaired local production of chemokines and defective neutrophil recruitment to the infection site. Altogether, these data show that Nod1, Nod2 and Rip2 are not required for local chemokine production and neutrophil recruitment during CLP-induced sepsis, and they reinforce the importance of MyD88-dependent signaling for initiation of a protective host response.


Assuntos
Fator 88 de Diferenciação Mieloide/metabolismo , Proteína Adaptadora de Sinalização NOD1/metabolismo , Proteína Adaptadora de Sinalização NOD2/metabolismo , Sepse/metabolismo , Animais , Modelos Animais de Doenças , Feminino , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fator 88 de Diferenciação Mieloide/genética , Proteína Adaptadora de Sinalização NOD1/genética , Proteína Adaptadora de Sinalização NOD2/genética , Proteína Serina-Treonina Quinase 2 de Interação com Receptor , Proteína Serina-Treonina Quinases de Interação com Receptores/genética , Proteína Serina-Treonina Quinases de Interação com Receptores/metabolismo , Sepse/genética
20.
Diabetes ; 61(6): 1584-91, 2012 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-22415874

RESUMO

The mechanisms underlying immune deficiency in diabetes are largely unknown. In the present study, we demonstrate that diabetic mice are highly susceptible to polymicrobial sepsis due to reduction in rolling, adhesion, and migration of leukocytes to the focus of infection. In addition, after sepsis induction, CXCR2 was strongly downregulated in neutrophils from diabetic mice compared with nondiabetic mice. Furthermore, CXCR2 downregulation was associated with increased G-protein-coupled receptor kinase 2 (GRK2) expression in these cells. Different from nondiabetic mice, diabetic animals submitted to mild sepsis displayed a significant augment in α1-acid glycoprotein (AGP) hepatic mRNA expression and serum protein levels. Administration of AGP in nondiabetic mice subjected to mild sepsis inhibited the neutrophil migration to the focus of infection, as well as induced l-selectin shedding and rise in CD11b of blood neutrophils. Insulin treatment of diabetic mice reduced mortality rate, prevented the failure of neutrophil migration, impaired GRK2-mediated CXCR2 downregulation, and decreased the generation of AGP. Finally, administration of AGP abolished the effect of insulin treatment in diabetic mice. Together, these data suggest that AGP may be involved in reduction of neutrophil migration and increased susceptibility to sepsis in diabetic mice.


Assuntos
Movimento Celular/imunologia , Diabetes Mellitus Experimental/metabolismo , Infiltração de Neutrófilos/imunologia , Neutrófilos/imunologia , Orosomucoide/metabolismo , Sepse/metabolismo , Animais , Antígeno CD11b/metabolismo , Movimento Celular/efeitos dos fármacos , Diabetes Mellitus Experimental/tratamento farmacológico , Diabetes Mellitus Experimental/imunologia , Suscetibilidade a Doenças/imunologia , Suscetibilidade a Doenças/metabolismo , Insulina/farmacologia , Insulina/uso terapêutico , Selectina L/metabolismo , Camundongos , Infiltração de Neutrófilos/efeitos dos fármacos , Neutrófilos/efeitos dos fármacos , Neutrófilos/metabolismo , Sepse/imunologia
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