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1.
Nat Immunol ; 24(12): 2021-2031, 2023 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-37903858

RESUMO

S100A8/S100A9 is a proinflammatory mediator released by myeloid cells during many acute and chronic inflammatory disorders. However, the precise mechanism of its release from the cytosolic compartment of neutrophils is unclear. Here, we show that E-selectin-induced rapid S100A8/S100A9 release during inflammation occurs in an NLRP3 inflammasome-dependent fashion. Mechanistically, E-selectin engagement triggers Bruton's tyrosine kinase-dependent tyrosine phosphorylation of NLRP3. Concomitant potassium efflux via the voltage-gated potassium channel KV1.3 mediates ASC oligomerization. This is followed by caspase 1 cleavage and downstream activation of pore-forming gasdermin D, enabling cytosolic release of S100A8/S100A9. Strikingly, E-selectin-mediated gasdermin D pore formation does not result in cell death but is a transient process involving activation of the ESCRT III membrane repair machinery. These data clarify molecular mechanisms of controlled S100A8/S100A9 release from neutrophils and identify the NLRP3/gasdermin D axis as a rapid and reversible activation system in neutrophils during inflammation.


Assuntos
Inflamassomos , Proteína 3 que Contém Domínio de Pirina da Família NLR , Humanos , Inflamassomos/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Gasderminas , Neutrófilos/metabolismo , Selectina E/metabolismo , Calgranulina A/metabolismo , Calgranulina B/metabolismo , Inflamação/metabolismo
2.
Immunity ; 54(3): 468-483.e5, 2021 03 09.
Artigo em Inglês | MEDLINE | ID: mdl-33484643

RESUMO

Tissue resident mast cells (MCs) rapidly initiate neutrophil infiltration upon inflammatory insult, yet the molecular mechanism is still unknown. Here, we demonstrated that MC-derived tumor necrosis factor (TNF) was crucial for neutrophil extravasation to sites of contact hypersensitivity-induced skin inflammation by promoting intraluminal crawling. MC-derived TNF directly primed circulating neutrophils via TNF receptor-1 (TNFR1) while being dispensable for endothelial cell activation. The MC-derived TNF was infused into the bloodstream by directional degranulation of perivascular MCs that were part of the vascular unit with access to the vessel lumen. Consistently, intravenous administration of MC granules boosted neutrophil extravasation. Pronounced and rapid intravascular MC degranulation was also observed upon IgE crosslinking or LPs challenge indicating a universal MC potential. Consequently, the directional MC degranulation of pro-inflammatory mediators into the bloodstream may represent an important target for therapeutic approaches aimed at dampening cytokine storm syndromes or shock symptoms, or intentionally pushing immune defense.


Assuntos
Vasos Sanguíneos/imunologia , Dermatite de Contato/imunologia , Inflamação/imunologia , Mastócitos/imunologia , Neutrófilos/imunologia , Pele/patologia , Fator de Necrose Tumoral alfa/metabolismo , Animais , Circulação Sanguínea , Degranulação Celular , Células Cultivadas , Doenças do Sistema Imunitário , Transtornos Leucocíticos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Ativação de Neutrófilo , Receptores Tipo I de Fatores de Necrose Tumoral/metabolismo , Vesículas Secretórias/metabolismo , Fator de Necrose Tumoral alfa/genética
3.
Proc Natl Acad Sci U S A ; 121(19): e2319057121, 2024 May 07.
Artigo em Inglês | MEDLINE | ID: mdl-38687790

RESUMO

Eosinophil recruitment is a pathological hallmark of many allergic and helminthic diseases. Here, we investigated chemokine receptor CCR3-induced eosinophil recruitment in sialyltransferase St3gal4-/- mice. We found a marked decrease in eosinophil extravasation into CCL11-stimulated cremaster muscles and into the inflamed peritoneal cavity of St3gal4-/- mice. Ex vivo flow chamber assays uncovered reduced adhesion of St3gal4-/- compared to wild type eosinophils. Using flow cytometry, we show reduced binding of CCL11 to St3gal4-/- eosinophils. Further, we noted reduced binding of CCL11 to its chemokine receptor CCR3 isolated from St3gal4-/- eosinophils. This was accompanied by almost absent CCR3 internalization of CCL11-stimulated St3gal4-/- eosinophils. Applying an ovalbumin-induced allergic airway disease model, we found a dramatic reduction in eosinophil numbers in bronchoalveolar lavage fluid following intratracheal challenge with ovalbumin in St3gal4-deficient mice. Finally, we also investigated tissue-resident eosinophils under homeostatic conditions and found reduced resident eosinophil numbers in the thymus and adipose tissue in the absence of ST3Gal-IV. Taken together, our results demonstrate an important role of ST3Gal-IV in CCR3-induced eosinophil recruitment in vivo rendering this enzyme an attractive target in reducing unwanted eosinophil infiltration in various disorders including allergic diseases.


Assuntos
Eosinófilos , Camundongos Knockout , Receptores CCR3 , Sialiltransferases , beta-Galactosídeo alfa-2,3-Sialiltransferase , Animais , Receptores CCR3/metabolismo , Receptores CCR3/genética , Sialiltransferases/metabolismo , Sialiltransferases/genética , Eosinófilos/metabolismo , Eosinófilos/imunologia , Camundongos , Quimiocina CCL11/metabolismo , Camundongos Endogâmicos C57BL , Ovalbumina/imunologia , Líquido da Lavagem Broncoalveolar
4.
Blood ; 139(23): 3402-3417, 2022 06 09.
Artigo em Inglês | MEDLINE | ID: mdl-35303071

RESUMO

Neutrophils are key players during host defense and sterile inflammation. Neutrophil dysfunction is a characteristic feature of the acquired immunodeficiency during kidney disease. We speculated that the impaired renal clearance of the intrinsic purine metabolite soluble uric acid (sUA) may account for neutrophil dysfunction. Indeed, hyperuricemia (HU, serum UA of 9-12 mg/dL) related or unrelated to kidney dysfunction significantly diminished neutrophil adhesion and extravasation in mice with crystal- and coronavirus-related sterile inflammation using intravital microscopy and an air pouch model. This impaired neutrophil recruitment was partially reversible by depleting UA with rasburicase. We validated these findings in vitro using either neutrophils or serum from patients with kidney dysfunction-related HU with or without UA depletion, which partially normalized the defective migration of neutrophils. Mechanistically, sUA impaired ß2 integrin activity and internalization/recycling by regulating intracellular pH and cytoskeletal dynamics, physiological processes that are known to alter the migratory and phagocytic capability of neutrophils. This effect was fully reversible by blocking intracellular uptake of sUA via urate transporters. In contrast, sUA had no effect on neutrophil extracellular trap formation in neutrophils from healthy subjects or patients with kidney dysfunction. Our results identify an unexpected immunoregulatory role of the intrinsic purine metabolite sUA, which contrasts the well-known immunostimulatory effects of crystalline UA. Specifically targeting UA may help to overcome certain forms of immunodeficiency, for example in kidney dysfunction, but may enhance sterile forms of inflammation.


Assuntos
Antígenos CD18 , Ácido Úrico , Animais , Antígenos CD18/metabolismo , Humanos , Imunidade Inata , Inflamação , Camundongos , Infiltração de Neutrófilos , Neutrófilos , Ácido Úrico/farmacologia , Ácido Úrico/urina
5.
Nephrol Dial Transplant ; 39(10): 1551-1564, 2024 Sep 27.
Artigo em Inglês | MEDLINE | ID: mdl-38115607

RESUMO

Neutrophils, the most abundant white blood cells in the human circulation, play crucial roles in various diseases, including kidney disease. Traditionally viewed as short-lived pro-inflammatory phagocytes that release reactive oxygen species, cytokines and neutrophil extracellular traps, recent studies have revealed their complexity and heterogeneity, thereby challenging this perception. Neutrophils are now recognized as transcriptionally active cells capable of proliferation and reverse migration, displaying phenotypic and functional heterogeneity. They respond to a wide range of signals and deploy various cargo to influence the activity of other cells in the circulation and in tissues. They can regulate the behavior of multiple immune cell types, exhibit innate immune memory, and contribute to both acute and chronic inflammatory responses while also promoting inflammation resolution in a context-dependent manner. Here, we explore the origin and heterogeneity of neutrophils, their functional diversity, and the cues that regulate their effector functions. We also examine their emerging role in infectious and non-infectious diseases with a particular emphasis on kidney disease. Understanding the complex behavior of neutrophils during tissue injury and inflammation may provide novel insights, thereby paving the way for potential therapeutic strategies to manage acute and chronic conditions. By deciphering their multifaceted role, targeted interventions can be developed to address the intricacies of neutrophil-mediated immune responses and improve disease outcomes.


Assuntos
Neutrófilos , Humanos , Neutrófilos/imunologia , Neutrófilos/metabolismo , Inflamação/imunologia , Nefropatias/imunologia , Nefropatias/etiologia , Animais , Imunidade Inata
6.
J Cell Sci ; 131(22)2018 11 21.
Artigo em Inglês | MEDLINE | ID: mdl-30333137

RESUMO

Integrins are α/ß heterodimers that interconvert between inactive and active states. In the active state the α/ß cytoplasmic domains recruit integrin-activating proteins and separate the transmembrane and cytoplasmic (TMcyto) domains (unclasped TMcyto). Conversely, in the inactive state the α/ß TMcyto domains bind integrin-inactivating proteins, resulting in the association of the TMcyto domains (clasped TMcyto). Here, we report the isolation of integrin cytoplasmic tail interactors using either lipid bicelle-incorporated integrin TMcyto domains (α5, αM, αIIb, ß1, ß2 and ß3 integrin TMcyto) or a clasped, lipid bicelle-incorporated αMß2 TMcyto. Among the proteins found to preferentially bind clasped rather than the isolated αM and ß2 subunits was L-plastin (LCP1, also known as plastin-2), which binds to and maintains the inactive state of αMß2 integrin in vivo and thereby regulates leukocyte adhesion to integrin ligands under flow. Our findings offer a global view on cytoplasmic proteins interacting with different integrins and provide evidence for the existence of conformation-specific integrin interactors.


Assuntos
Adesão Celular/fisiologia , Leucócitos/citologia , Leucócitos/metabolismo , Antígeno de Macrófago 1/metabolismo , Proteínas dos Microfilamentos/metabolismo , Animais , Membrana Celular/metabolismo , Citoplasma/metabolismo , Células HEK293 , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Ligação Proteica , Conformação Proteica , Células RAW 264.7
7.
Haematologica ; 105(7): 1845-1856, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-31699792

RESUMO

Leukocyte recruitment into inflamed tissue is highly dependent on the activation and binding of integrins to their respective ligands, followed by the induction of various signaling events within the cell referred to as outside-in signaling. Src family kinases (SFK) are the central players in the outside-in signaling process, assigning them a critical role for proper immune cell function. Our study investigated the role of SFK on neutrophil recruitment in vivo using Hck-/- Fgr-/- Lyn-/- mice, which lack SFK expressed in neutrophils. We show that loss of SFK strongly reduces neutrophil adhesion and post-arrest modifications in a shear force dependent manner. Additionally, we found that in the absence of SFK, neutrophils display impaired Rab27a-dependent surface mobilization of neutrophil elastase, VLA3 and VLA6 containing vesicles. This results in a defect in neutrophil vascular basement membrane penetration and thus strongly impaired extravasation. Taken together, we demonstrate that SFK play a role in neutrophil post-arrest modifications and extravasation during acute inflammation. These findings may support the current efforts to use SFK-inhibitors in inflammatory diseases with unwanted neutrophil recruitment.


Assuntos
Neutrófilos , Quinases da Família src , Animais , Membrana Basal , Camundongos , Camundongos Knockout , Proteínas Proto-Oncogênicas , Quinases da Família src/genética
8.
Eur J Clin Invest ; 48 Suppl 2: e12964, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-29873837

RESUMO

The recruitment of neutrophils to sites of inflammation, their battle against invading microorganisms through phagocytosis and the release of antimicrobial agents is a highly coordinated and tightly regulated process that involves the interplay of many different receptors, ion channels and signalling pathways. Changes in intracellular calcium levels, caused by cytosolic Ca2+ store depletion and the influx of extracellular Ca2+ via ion channels, play a critical role in synchronizing neutrophil activation and function. In this review, we provide an overview of how Ca2+ signalling is initiated in neutrophils and how changes in intracellular Ca2+ levels modulate neutrophil function.


Assuntos
Sinalização do Cálcio/fisiologia , Infiltração de Neutrófilos/fisiologia , Animais , Canais de Cálcio/fisiologia , Humanos , Canais Iônicos/fisiologia , Camundongos , Neutrófilos/fisiologia , Canais de Potássio/fisiologia , Receptores Purinérgicos P2X/fisiologia , Canais de Sódio/fisiologia , Canais de Potencial de Receptor Transitório/fisiologia
9.
Arterioscler Thromb Vasc Biol ; 37(6): 1076-1086, 2017 06.
Artigo em Inglês | MEDLINE | ID: mdl-28428216

RESUMO

OBJECTIVE: Platelet function has been intensively studied in the adult organism. However, little is known about the function and hemostatic capacity of platelets in the developing fetus as suitable in vivo models are lacking. APPROACH AND RESULTS: To examine fetal platelet function in vivo, we generated a fetal thrombosis model and investigated light/dye-induced thrombus formation by intravital microscopy throughout gestation. We observed that significantly less and unstable thrombi were formed at embryonic day (E) 13.5 compared with E17.5. Flow cytometry revealed significantly lower platelet counts in E13.5 versus E17.5 fetuses versus adult controls. In addition, fetal platelets demonstrated changed activation responses of surface adhesion molecules and reduced P-selectin content and mobilization. Interestingly, we also measured reduced levels of the integrin-activating proteins Kindlin-3, Talin-1, and Rap1 during fetal development. Consistently, fetal platelets demonstrated diminished spreading capacity compared with adults. Transfusion of adult platelets into the fetal circulation led to rapid platelet aggregate formation even in young fetuses. Yet, retrospective data analysis of a neonatal cohort demonstrated no correlation of platelet transfusion with closure of a persistent ductus arteriosus, a process reported to be platelet dependent. CONCLUSIONS: Taken together, we demonstrate an ontogenetic regulation of platelet function in vivo with physiologically low platelet numbers and hyporeactivity early during fetal development shedding new light on hemostatic function during fetal life.


Assuntos
Plaquetas/metabolismo , Hemostasia , Ativação Plaquetária , Trombose/sangue , Animais , Moléculas de Adesão Celular/sangue , Bases de Dados Factuais , Modelos Animais de Doenças , Permeabilidade do Canal Arterial/sangue , Feminino , Idade Gestacional , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Humanos , Recém-Nascido , Recém-Nascido Prematuro , Recém-Nascido de muito Baixo Peso , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Adesividade Plaquetária , Transfusão de Plaquetas , Nascimento Prematuro/sangue , Estudos Retrospectivos , Transdução de Sinais , Trombocitopenia/sangue
10.
J Leukoc Biol ; 2024 Sep 23.
Artigo em Inglês | MEDLINE | ID: mdl-39312228

RESUMO

Newborns are at high risk to develop sepsis. This is linked to innate immune responses at birth which are not completely adapted to postnatal life. Neutrophils are key players of innate immunity and exhibit a marked ontogenetic regulation of their functionality. Here, we studied the NLRP3 inflammasome in neonatal neutrophils and found lower baseline expression of NLRP3, pro-caspase-1 and the K+-channel KV1.3 compared to adult neutrophils. Following stimulation with LPS/Nigericin, ASC oligomerization, caspase-1 activation and IL-1ß release were significantly reduced in neonatal compared to adult neutrophils. Similarly, stimulation of neonatal neutrophils with E-selectin led to reduced NLRP3 inflammasome activation accompanied by diminished release of the alarmin S100A8/A9. Taken together, our results strongly indicate diminished NLRP3 inflammasome activation in neonatal neutrophils leading to a significant reduction of released IL-1ß and S100A8/A9. These findings identify reduced neutrophil NLRP3 inflammasome activation as critical component contributing to the inherent susceptibility to infections in neonates.

11.
Adv Sci (Weinh) ; : e2404661, 2024 Oct 04.
Artigo em Inglês | MEDLINE | ID: mdl-39364760

RESUMO

Exposure to nanoparticles (NPs) is frequently associated with adverse cardiovascular effects. In contrast, NPs in nanomedicine hold great promise for precise lung-specific drug delivery, especially considering the extensive pulmonary capillary network that facilitates interactions with bloodstream-suspended particles. Therefore, exact knowledge about effects of engineered NPs within the pulmonary microcirculation are instrumental for future application of this technology in patients. To unravel the real-time dynamics of intravenously delivered NPs and their effects in the pulmonary microvasculature, we employed intravital microscopy of the mouse lung. Only PEG-amine-QDs, but not carboxyl-QDs triggered rapid neutrophil recruitment in microvessels and their subsequent recruitment to the alveolar space and was linked to cellular degranulation, TNF-α, and DAMP release into the circulation, particularly eATP. Stimulation of the ATP-gated receptor P2X7R induced expression of E-selectin on microvascular endothelium thereby mediating the neutrophilic immune response. Leukocyte integrins LFA-1 and MAC-1 facilitated adhesion and decelerated neutrophil crawling on the vascular surface. In summary, this study unravels the complex cascade of neutrophil recruitment during NP-induced sterile inflammation. Thereby we demonstrate novel adverse effects for NPs in the pulmonary microcirculation and provide critical insights for optimizing NP-based drug delivery and therapeutic intervention strategies, to ensure their efficacy and safety in clinical applications.

12.
Cardiovasc Res ; 118(5): 1289-1302, 2022 03 25.
Artigo em Inglês | MEDLINE | ID: mdl-33881519

RESUMO

AIMS: Neutrophil trafficking within the vasculature strongly relies on intracellular calcium signalling. Sustained Ca2+ influx into the cell requires a compensatory efflux of potassium to maintain membrane potential. Here, we aimed to investigate whether the voltage-gated potassium channel KV1.3 regulates neutrophil function during the acute inflammatory process by affecting sustained Ca2+ signalling. METHODS AND RESULTS: Using in vitro assays and electrophysiological techniques, we show that KV1.3 is functionally expressed in human neutrophils regulating sustained store-operated Ca2+ entry through membrane potential stabilizing K+ efflux. Inhibition of KV1.3 on neutrophils by the specific inhibitor 5-(4-Phenoxybutoxy)psoralen (PAP-1) impaired intracellular Ca2+ signalling, thereby preventing cellular spreading, adhesion strengthening, and appropriate crawling under flow conditions in vitro. Using intravital microscopy, we show that pharmacological blockade or genetic deletion of KV1.3 in mice decreased neutrophil adhesion in a blood flow dependent fashion in inflamed cremaster muscle venules. Furthermore, we identified KV1.3 as a critical component for neutrophil extravasation into the inflamed peritoneal cavity. Finally, we also revealed impaired phagocytosis of Escherichia coli particles by neutrophils in the absence of KV1.3. CONCLUSION: We show that the voltage-gated potassium channel KV1.3 is critical for Ca2+ signalling and neutrophil trafficking during acute inflammatory processes. Our findings do not only provide evidence for a role of KV1.3 for sustained calcium signalling in neutrophils affecting key functions of these cells, they also open up new therapeutic approaches to treat inflammatory disorders characterized by overwhelming neutrophil infiltration.


Assuntos
Canais de Potássio de Abertura Dependente da Tensão da Membrana , Animais , Cálcio/metabolismo , Inflamação , Canal de Potássio Kv1.5 , Potenciais da Membrana/fisiologia , Camundongos , Infiltração de Neutrófilos
13.
Front Immunol ; 11: 606893, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33658993

RESUMO

During inflammation, neutrophils are one of the first responding cells of innate immunity, contributing to a fast clearance of infection and return to homeostasis. However, excessive neutrophil infiltration accelerates unsolicited disproportionate inflammation for instance in autoimmune diseases such as rheumatoid arthritis. The transient-receptor-potential channel-kinase TRPM7 is an essential regulator of immune system homeostasis. Naïve murine T cells with genetic inactivation of the TRPM7 enzyme, due to a point mutation at the active site, are unable to differentiate into pro-inflammatory T cells, whereas regulatory T cells develop normally. Moreover, TRPM7 is vital for lipopolysaccharides (LPS)-induced activation of murine macrophages. Within this study, we show that the channel-kinase TRPM7 is functionally expressed in neutrophils and has an important impact on neutrophil recruitment during inflammation. We find that human neutrophils cannot transmigrate along a CXCL8 chemokine gradient or produce reactive oxygen species in response to gram-negative bacterial lipopolysaccharide LPS, if TRPM7 channel or kinase activity are blocked. Using a recently identified TRPM7 kinase inhibitor, TG100-115, as well as murine neutrophils with genetic ablation of the kinase activity, we confirm the importance of both TRPM7 channel and kinase function in murine neutrophil transmigration and unravel that TRPM7 kinase affects Akt1/mTOR signaling thereby regulating neutrophil transmigration and effector function. Hence, TRPM7 represents an interesting potential target to treat unwanted excessive neutrophil invasion.


Assuntos
Infiltração de Neutrófilos , Neutrófilos/enzimologia , Peritonite/enzimologia , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Canais de Cátion TRPM/metabolismo , Animais , Células Cultivadas , Modelos Animais de Doenças , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Peritonite/induzido quimicamente , Peritonite/genética , Proteínas Serina-Treonina Quinases/genética , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais , Canais de Cátion TRPM/genética , Fator de Necrose Tumoral alfa
14.
Front Immunol ; 11: 588245, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33414784

RESUMO

Uromodulin (UMOD) is produced and secreted by tubular epithelial cells. Secreted UMOD polymerizes (pUMOD) in the tubular lumen, where it regulates salt transport and protects the kidney from bacteria and stone formation. Under various pathological conditions, pUMOD accumulates within the tubular lumen and reaches extratubular sites where it may interact with renal interstitial cells. Here, we investigated the potential of extratubular pUMOD to act as a damage associated molecular pattern (DAMP) molecule thereby creating local inflammation. We found that intrascrotal and intraperitoneal injection of pUMOD induced leukocyte recruitment in vivo and led to TNF-α secretion by F4/80 positive macrophages. Additionally, pUMOD directly affected vascular permeability and increased neutrophil extravasation independent of macrophage-released TNF-α. Interestingly, pUMOD displayed no chemotactic properties on neutrophils, did not directly activate ß2 integrins and did not upregulate adhesion molecules on endothelial cells. In obstructed neonatal murine kidneys, we observed extratubular UMOD accumulation in the renal interstitium with tubular atrophy and leukocyte infiltrates. Finally, we found extratubular UMOD deposits associated with peritubular leukocyte infiltration in kidneys from patients with inflammatory kidney diseases. Taken together, we identified extratubular pUMOD as a strong inducer of leukocyte recruitment, underlining its critical role in mounting an inflammatory response in various kidneys pathologies.


Assuntos
Inflamação/imunologia , Leucócitos/imunologia , Uromodulina/imunologia , Músculos Abdominais/imunologia , Animais , Moléculas de Adesão Celular/metabolismo , Células Cultivadas , Feminino , Células Endoteliais da Veia Umbilical Humana/metabolismo , Células Endoteliais da Veia Umbilical Humana/fisiologia , Humanos , Nefropatias/imunologia , Masculino , Camundongos Endogâmicos C57BL , Polimerização
15.
Endocr Connect ; 7(5): 749-761, 2018 May.
Artigo em Inglês | MEDLINE | ID: mdl-29700097

RESUMO

Although an inflammatory microenvironment is required for successful implantation, an inflammatory overreaction is one of the causes of unexplained recurrent pregnancy losses (uRPL). Prostaglandin E2 (PGE2) plays a pivotal role in regulating immune balance during early pregnancy, and it can stimulate inflammatory reactions via prostaglandin E2 receptor 3 (EP3). However, the role of PGE2 receptor signaling in the uRPL remains unknown. We aimed to investigate whether EP3 signaling is involved in the mechanism of uRPL. Via immunohistochemistry we could show that the expression of cyclooxygenase-2, EP3 and G protein alpha inhibitor 1 (Gi1) was enhanced in the decidua of the uRPL group in comparison to the control group in first-trimester placentas. In vitro, we demonstrated that sulprostone (an EP1/EP3 agonist) inhibited the secretion of beta-hCG and progesterone in JEG-3 cells and the secretion of beta-hCG in HTR-8/SVneo cells while it induced the expression of plasminogen activator inhibitor type 1 in JEG-3 cells. In addition, PGE2/sulprostone was able to stimulate the expression of Gi1, phosphorylated-extracellular signal-regulated kinases 1/2 (p-ERK1/2) and p53. L-798,106 (an EP3-specific antagonist) suppressed the expression of EP3 and p-ERK1/2 without affecting the secretion of beta-hCG. Elevated activation of EP3 signaling in first-trimester placentas plays an important role in regulating the inflammatory microenvironment, the hormone secretion of extravillous trophoblasts and the remodeling of extracellular matrix in the fetal-maternal interface. L-798,106 might be a 'potential therapeutic candidate' for the treatment of uRPL.

16.
Cell Metab ; 28(1): 175-182.e5, 2018 Jul 03.
Artigo em Inglês | MEDLINE | ID: mdl-29861387

RESUMO

Onset of cardiovascular complications as a consequence of atherosclerosis exhibits a circadian incidence with a peak in the morning hours. Although development of atherosclerosis extends for long periods of time through arterial leukocyte recruitment, we hypothesized that discrete diurnal invasion of the arterial wall could sustain atherogenic growth. Here, we show that myeloid cell recruitment to atherosclerotic lesions oscillates with a peak during the transition from the activity to the resting phase. This diurnal phenotype is regulated by rhythmic release of myeloid cell-derived CCL2, and blockade of its signaling abolished oscillatory leukocyte adhesion. In contrast, we show that myeloid cell adhesion to microvascular beds peaks during the early activity phase. Consequently, timed pharmacological CCR2 neutralization during the activity phase caused inhibition of atherosclerosis without disturbing microvascular recruitment. These findings demonstrate that chronic inflammation of large vessels feeds on rhythmic myeloid cell recruitment, and lay the foundation for chrono-pharmacology-based therapy.


Assuntos
Aterosclerose/terapia , Adesão Celular , Quimiocina CCL2/metabolismo , Células-Tronco Mesenquimais/metabolismo , Células Mieloides/metabolismo , Receptores CCR2/metabolismo , Animais , Inflamação/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Transdução de Sinais
17.
ACS Nano ; 11(2): 1498-1508, 2017 02 28.
Artigo em Inglês | MEDLINE | ID: mdl-28135073

RESUMO

Advances in the engineering of nanoparticles (NPs), which represent particles of less than 100 nm in one external dimension, led to an increasing utilization of nanomaterials for biomedical purposes. A prerequisite for their use in diagnostic and therapeutic applications, however, is the targeted delivery to the site of injury. Interactions between blood-borne NPs and the vascular endothelium represent a critical step for nanoparticle delivery into diseased tissue. Here, we show that the endothelial glycocalyx, which constitutes a glycoprotein-polysaccharide meshwork coating the luminal surface of vessels, effectively controls interactions of carboxyl-functionalized quantum dots with the microvascular endothelium. Glycosaminoglycans of the endothelial glycocalyx were found to physically cover endothelial adhesion and signaling molecules, thereby preventing endothelial attachment, uptake, and translocation of these nanoparticles through different layers of the vessel wall. Conversely, degradation of the endothelial glycocalyx promoted interactions of these nanoparticles with microvascular endothelial cells under the pathologic condition of ischemia-reperfusion, thus identifying the injured endothelial glycocalyx as an essential element of the blood-tissue border facilitating the targeted delivery of nanomaterials to diseased tissue.


Assuntos
Barreira Hematoencefálica/metabolismo , Endotélio Vascular/metabolismo , Glicocálix/metabolismo , Células Endoteliais da Veia Umbilical Humana/metabolismo , Nanopartículas/metabolismo , Pontos Quânticos/metabolismo , Animais , Barreira Hematoencefálica/química , Células Cultivadas , Endotélio Vascular/química , Glicocálix/química , Células Endoteliais da Veia Umbilical Humana/química , Humanos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Nanopartículas/química , Pontos Quânticos/química
18.
J Clin Invest ; 126(11): 4125-4139, 2016 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-27701149

RESUMO

Neutrophils need to penetrate the perivascular basement membrane for successful extravasation into inflamed tissue, but this process is incompletely understood. Recent findings have associated mammalian sterile 20-like kinase 1 (MST1) loss of function with a human primary immunodeficiency disorder, suggesting that MST1 may be involved in immune cell migration. Here, we have shown that MST1 is a critical regulator of neutrophil extravasation during inflammation. Mst1-deficient (Mst1-/-) neutrophils were unable to migrate into inflamed murine cremaster muscle venules, instead persisting between the endothelium and the basement membrane. Mst1-/- neutrophils also failed to extravasate from gastric submucosal vessels in a murine model of Helicobacter pylori infection. Mechanistically, we observed defective translocation of VLA-3, VLA-6, and neutrophil elastase from intracellular vesicles to the surface of Mst1-/- neutrophils, indicating that MST1 is required for this crucial step in neutrophil transmigration. Furthermore, we found that MST1 associates with the Rab27 effector protein synaptotagmin-like protein 1 (JFC1, encoded by Sytl1 in mice), but not Munc13-4, thereby regulating the trafficking of Rab27-positive vesicles to the cellular membrane. Together, these findings highlight a role for MST1 in vesicle trafficking and extravasation in neutrophils, providing an additional mechanistic explanation for the severe immune defect observed in patients with MST1 deficiency.


Assuntos
Fator de Crescimento de Hepatócito/imunologia , Neutrófilos/imunologia , Proteínas Proto-Oncogênicas/imunologia , Vesículas Secretórias/imunologia , Migração Transendotelial e Transepitelial/imunologia , Músculos Abdominais/irrigação sanguínea , Músculos Abdominais/imunologia , Animais , Membrana Basal/imunologia , Transporte Biológico Ativo/genética , Transporte Biológico Ativo/imunologia , Mucosa Gástrica/química , Mucosa Gástrica/imunologia , Fator de Crescimento de Hepatócito/genética , Humanos , Síndromes de Imunodeficiência/genética , Síndromes de Imunodeficiência/imunologia , Integrina alfa3beta1/genética , Integrina alfa3beta1/imunologia , Integrina alfa6beta1/genética , Integrina alfa6beta1/imunologia , Elastase de Leucócito/genética , Elastase de Leucócito/imunologia , Proteínas de Membrana/genética , Proteínas de Membrana/imunologia , Camundongos , Camundongos Knockout , Proteínas Proto-Oncogênicas/genética , Vesículas Secretórias/genética , Migração Transendotelial e Transepitelial/genética , Vênulas/imunologia , Proteínas de Transporte Vesicular
19.
Nat Commun ; 6: 6915, 2015 Apr 20.
Artigo em Inglês | MEDLINE | ID: mdl-25892652

RESUMO

Myeloid-related proteins (MRPs) 8 and 14 are cytosolic proteins secreted from myeloid cells as proinflammatory mediators. Currently, the functional role of circulating extracellular MRP8/14 is unclear. Our present study identifies extracellular MRP8/14 as an autocrine player in the leukocyte adhesion cascade. We show that E-selectin-PSGL-1 interaction during neutrophil rolling triggers Mrp8/14 secretion. Released MRP8/14 in turn activates a TLR4-mediated, Rap1-GTPase-dependent pathway of rapid ß2 integrin activation in neutrophils. This extracellular activation loop reduces leukocyte rolling velocity and stimulates adhesion. Thus, we identify Mrp8/14 and TLR4 as important modulators of the leukocyte recruitment cascade during inflammation in vivo.


Assuntos
Antígenos CD18/metabolismo , Calgranulina A/metabolismo , Calgranulina B/metabolismo , Adesão Celular/fisiologia , Migração e Rolagem de Leucócitos/fisiologia , Neutrófilos/fisiologia , Animais , Antígenos CD18/genética , Calgranulina A/genética , Calgranulina B/genética , Regulação da Expressão Gênica , Receptores de Hialuronatos/genética , Receptores de Hialuronatos/metabolismo , Inflamação/induzido quimicamente , Inflamação/metabolismo , Macrófagos/fisiologia , Masculino , Camundongos , Camundongos Knockout , Ligação Proteica
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