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1.
Front Immunol ; 14: 1240597, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37753073

RESUMO

Venous thromboembolism (VTE) is a leading cause of preventable deaths in hospitals, and its incidence is not decreasing despite extensive efforts in clinical and laboratory research. Venous thrombi are primarily formed in the valve pockets of deep veins, where activated monocytes play a crucial role in bridging innate immune activation and hemostatic pathways through the production of inflammatory cytokines, chemokines, and tissue factor (TF) - a principal initiator of coagulation. In the valve pocket inflammation and hypoxia (sustained/intermittent) coexist, however their combined effects on immunothrombotic processes are poorly understood. Inflammation is strongly associated with VTE, while the additional contribution of hypoxia remains largely unexplored. To investigate this, we modelled the intricate conditions of the venous valve pocket using a state-of-the-art hypoxia chamber with software-controlled oxygen cycling. We comprehensively studied the effects of sustained and intermittent hypoxia alone, and in combination with VTE-associated inflammatory stimuli on primary monocytes. TF expression and activity was measured in monocytes subjected to sustained and intermittent hypoxia alone, or in combination with IL-1ß. Monocyte responses were further analyzed in detailed by RNA sequencing and validated by ELISA. Stimulation with IL-1ß alone promoted both transcription and activity of TF. Interestingly, the stimulatory effect of IL-1ß on TF was attenuated by sustained hypoxia, but not by intermittent hypoxia. Our transcriptome analysis further confirmed that sustained hypoxia limited the pro-inflammatory response induced by IL-1ß, and triggered a metabolic shift in monocytes. Intermittent hypoxia alone had a modest effect on monocyte transcript. However, in combination with IL-1ß intermittent hypoxia significantly altered the expression of 2207 genes and enhanced the IL-1ß-stimulatory effects on several chemokine and interleukin genes (e.g., IL-19, IL-24, IL-32, MIF), as well as genes involved in coagulation (thrombomodulin) and fibrinolysis (VEGFA, MMP9, MMP14 and PAI-1). Increased production of CCL2, IL-6 and TNF following stimulation with intermittent hypoxia and IL-1ß was confirmed by ELISA. Our findings provide valuable insights into how the different hypoxic profiles shape the immunothrombotic response of monocytes and shed new light on the early events in the pathogenesis of venous thrombosis.


Assuntos
Monócitos , Tromboembolia Venosa , Humanos , Tromboembolia Venosa/metabolismo , Citocinas/metabolismo , Hipóxia/metabolismo , Inflamação/metabolismo , Tromboplastina/metabolismo
2.
J Thromb Haemost ; 20(4): 899-908, 2022 04.
Artigo em Inglês | MEDLINE | ID: mdl-35000275

RESUMO

BACKGROUND: Microvesicles (MVs) are small double-membrane encapsulated particles shed from cells. Case-control studies have reported elevated plasma levels of platelet-derived MVs (PDMVs) in patients with venous thromboembolism (VTE). However, it is not known whether high PDMV levels is a risk factor or a consequence of the acute VTE event. OBJECTIVES: To investigate the association between PDMVs in plasma and risk of future incident VTE. METHODS: We performed a population-based nested case-control study with 314 VTE cases and 705 age- and sex-matched controls (from The Tromsø Study) to investigate the association between the proportion of PDMVs (PDMVs%) in plasma and risk of future incident VTE. MVs isolated from plasma sampled at baseline (i.e., before VTE) were stained for platelet markers and analyzed by flow cytometry. PDMVs% were defined as the number of PDMVs divided by the total number of MVs. Odds ratios (ORs) with 95% confidence intervals (CI) for VTE risk were estimated across quartiles of PDMVs%. RESULTS: Subjects with PDMVs% in the highest quartile had an OR for VTE of 1.78 (95% CI: 1.21-2.64) and 1.99 (95% CI: 1.24-3.26) for provoked VTE, compared to those in the lowest quartile. The association was moderately affected by multivariable adjustment for age, sex, body mass index, C-reactive protein, platelet count, and cancer. The OR for VTE was higher when the time between blood sampling and event was shorter. CONCLUSIONS: Our results show that high proportions of PDMVs are associated with future risk of incident VTE and imply a role of platelet activation in the pathogenesis of VTE.


Assuntos
Tromboembolia Venosa , Plaquetas/patologia , Estudos de Casos e Controles , Humanos , Razão de Chances , Fatores de Risco , Tromboembolia Venosa/diagnóstico , Tromboembolia Venosa/epidemiologia
3.
Sci Rep ; 10(1): 15328, 2020 09 18.
Artigo em Inglês | MEDLINE | ID: mdl-32948789

RESUMO

Pulmonary sarcoidosis has unknown etiology, a difficult diagnostic procedure and no curative treatment. Extracellular vesicles including exosomes are nano-sized entities released from all cell types. Previous studies of exosomes from bronchoalveolar lavage fluid (BALF) of sarcoidosis patients have revealed pro-inflammatory components and abilities, but cell sources and mechanisms have not been identified. In the current study, we found that BALF exosomes from sarcoidosis patients, but not from healthy individuals, induced a dose-dependent elevation of intracellular IL-1ß in monocytes. Analyses of supernatants showed that patient exosomes also induced release of IL-1ß, IL-6 and TNF from both PBMCs and enriched monocytes, suggesting that the observed effect is direct on monocytes. The potently chemotactic chemokine CCL2 was induced by exosomes from a subgroup of patients, and in a blocking assay the exosome-induced CCL2 was reduced for 13 out of 19 patients by the asthma drug Montelukast, a cysteinyl leukotriene receptor antagonist. Further, reactive oxygen species generation by PBMCs was induced to a higher degree by patient exosomes compared to healthy exosomes. These findings add to an emerging picture of exosomes as mediators and disseminators of inflammation, and open for further investigations of the link between CCL2 and exosomal leukotrienes in sarcoidosis.


Assuntos
Quimiocina CCL2/metabolismo , Citocinas/metabolismo , Exossomos/metabolismo , Monócitos/metabolismo , Sarcoidose Pulmonar/patologia , Acetatos/farmacologia , Adulto , Líquido da Lavagem Broncoalveolar/citologia , Estudos de Casos e Controles , Ciclopropanos/farmacologia , Exossomos/efeitos dos fármacos , Exossomos/patologia , Feminino , Humanos , Interleucina-1beta/metabolismo , Masculino , Pessoa de Meia-Idade , Quinolinas/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Sulfetos/farmacologia
4.
Cancer Lett ; 444: 1-8, 2019 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-30508568

RESUMO

Tumor-derived exosomes can modulate the cancer microenvironment and induce metastatic spread. Exosomes may carry enzymes for leukotriene (LT) biosynthesis, but the role of exosomal LTs has not been studied in cancer. We isolated exosomes and malignant cells from pleura exudates from 14 patients with non-small cell lung cancer. Lipidomic profiles, migration and apoptosis were determined. Both exosomes and primary cancer cells contained γ-glutamyl transpeptidase 1 (GGT-1) and avidly transformed exogenous LTC4 to pro-tumorigenic LTD4, for the cells to levels 100-fold above their endogenous CysLT production. This suggests that cancer cells promote their own survival via LTD4 if supplied with LTC4, which in the exudates was produced by monocytic cells. Furthermore, exosomes promoted migration of cancer cells, which was counteracted by the CysLT1 antagonist montelukast. Montelukast also induced apoptosis of cancer cells, and this was partially inhibited by exosomes. Our results demonstrate how cancer cells and exosomes, together with monocytic cells in lung cancer tissue, can produce high amounts of LTD4, to stimulate cancer cell migration and survival. This suggests that part of the pro-metastatic effect of exosomes is mediated by the leukotriene machinery, further supporting the use of CysLT1 antagonists for lung cancer therapy.


Assuntos
Movimento Celular , Exossomos/metabolismo , Leucotrieno C4/metabolismo , Leucotrieno D4/metabolismo , Neoplasias Pulmonares/patologia , Neoplasias Pleurais/patologia , Receptores de Leucotrienos/metabolismo , Acetatos/farmacologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Apoptose , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Proliferação de Células , Ciclopropanos , Exossomos/efeitos dos fármacos , Feminino , Seguimentos , Humanos , Antagonistas de Leucotrienos/farmacologia , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Masculino , Pessoa de Meia-Idade , Neoplasias Pleurais/genética , Neoplasias Pleurais/metabolismo , Prognóstico , Quinolinas/farmacologia , Receptores de Leucotrienos/genética , Sulfetos , Taxa de Sobrevida , Células Tumorais Cultivadas
5.
Sci Rep ; 7(1): 17095, 2017 12 06.
Artigo em Inglês | MEDLINE | ID: mdl-29213052

RESUMO

Extracellular vesicles (EV), including exosomes and microvesicles (MV), represent a rapidly expanding field of research with diagnostic and therapeutic applications. Although many aspects of EV function remain to be revealed and broad investigations are warranted, most published findings focus on only one vesicle category or a non-separated mix of EVs. In this paper, we investigated both MVs and exosomes from Ovalbumin (OVA)-pulsed dendritic cells for their immunostimulatory potential side-by-side in vivo. Only exosomes induced antigen-specific CD8+ T-cells, and were more efficient than MVs in eliciting antigen-specific IgG production. Further, mainly exosome-primed mouse splenocytes showed significant ex vivo interferon gamma production in response to antigen restimulation. Exosomes carried high levels of OVA, while OVA in MVs was barely detectable, which could explain the more potent antigen-specific response induced by exosomes. Moreover, exosomes induced increased germinal center B cell proportions, whereas MVs had no such effect. Immunisation with both vesicle types combined showed neither inhibitory nor synergistic effects. We conclude that DC-derived MVs and exosomes differ in their capacity to incorporate antigen and induce immune responses. The results are of importance for understanding the role of EVs in vivo, and for future design of vesicle-based immunotherapies and vaccines.


Assuntos
Antígenos/imunologia , Micropartículas Derivadas de Células/imunologia , Exossomos/imunologia , Animais , Linfócitos B/citologia , Linfócitos B/imunologia , Linfócitos B/metabolismo , Células da Medula Óssea/citologia , Linfócitos T CD8-Positivos/citologia , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Micropartículas Derivadas de Células/química , Células Dendríticas/citologia , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Exossomos/química , Imunoglobulina G/sangue , Interferon gama/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Ovalbumina/imunologia , Baço/citologia , Baço/imunologia , Baço/metabolismo
6.
Front Cell Dev Biol ; 5: 39, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28491866

RESUMO

Cells of the airways are constantly exposed to environmental hazards including cigarette smoke, irritants, pathogens, and mechanical insults. Maintaining barrier integrity is vital, and mounting responses to threats depends on intercellular communication. Extracellular vesicles (EVs), including exosomes and microvesicles, are major signal mediators between cells, shuttling cargo in health and disease. Depending on the state of the originating cells, EVs are capable of inducing proinflammatory effects including antigen presentation, cellular migration, apoptosis induction, and inflammatory cytokine release. Cells of the airways release EVs, which can be found in bronchoalveolar lavage fluid. EVs of the airways can support inflammation in the lung, but may also exit into the circulation and carry a cocktail of pro-inflammatory molecules to recipient cells in distant organs. In this review, we discuss the possibility that EVs originating from the airways contribute to dissemination of inflammation in both lung disorders and systemic inflammatory conditions.

7.
J Allergy Clin Immunol ; 139(4): 1186-1194, 2017 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-27566455

RESUMO

BACKGROUND: Sarcoidosis is an inflammatory granulomatous disorder characterized by accumulation of TH1-type CD4+ T cells and immune effector cells within affected organs, most frequently the lungs. Exosomes are extracellular vesicles conveying intercellular communication with possible diagnostic and therapeutic applications. OBJECTIVES: We aimed to provide an understanding of the proinflammatory role of bronchoalveolar lavage fluid (BALF) exosomes in patients with sarcoidosis and to find candidates for disease biomarkers. METHODS: We performed a mass spectrometric proteomics characterization of BALF exosomes from 15 patients with sarcoidosis and 5 healthy control subjects and verified the most interesting results with flow cytometry, ELISA, and Western blot analyses in an additional 39 patients and 22 control subjects. RESULTS: More than 690 proteins were identified in the BALF exosomes, several of which displayed significant upregulation in patients, including inflammation-associated proteins, such as leukotriene A4 hydrolase. Most of the complement-activating factors were upregulated, whereas the complement regulator CD55 was seen less in patients compared with healthy control subjects. In addition, for the first time, we detected vitamin D-binding protein in BALF exosomes, which was more abundant in patients. To evaluate exosome-associated vitamin D-binding protein as a biomarker for sarcoidosis, we investigated plasma exosomes from 23 patients and 11 healthy control subjects and found significantly higher expression in patients. CONCLUSION: Together, these data contribute to understanding the role of exosomes in lung disease and provide suggestions for highly warranted sarcoidosis biomarkers. Furthermore, the validation of an exosome-associated biomarker in the blood of patients provides novel, and less invasive, opportunities for disease diagnosis.


Assuntos
Biomarcadores/análise , Exossomos/metabolismo , Sarcoidose Pulmonar/metabolismo , Proteína de Ligação a Vitamina D/metabolismo , Adulto , Biomarcadores/metabolismo , Western Blotting , Líquido da Lavagem Broncoalveolar/química , Líquido da Lavagem Broncoalveolar/imunologia , Cromatografia Líquida , Ensaio de Imunoadsorção Enzimática , Exossomos/patologia , Feminino , Citometria de Fluxo , Humanos , Inflamação/metabolismo , Inflamação/patologia , Masculino , Pessoa de Meia-Idade , Proteômica , Sarcoidose Pulmonar/patologia , Espectrometria de Massas em Tandem
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