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1.
J Immunol ; 212(10): 1531-1539, 2024 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-38506555

RESUMO

Immune thrombocytopenia (ITP) is an autoimmune disease characterized by low platelet counts primarily due to antiplatelet autoantibodies. Anti-D is a donor-derived polyclonal Ab against the rhesus D Ag on erythrocytes used to treat ITP. Unfortunately, adverse inflammatory/hypersensitivity reactions and a Food and Drug Administration-issued black box warning have limited its clinical use. This underscores the imperative to understand the inflammatory pathway associated with anti-erythrocyte Ab-based therapies. TER119 is an erythrocyte-specific Ab with anti-D-like therapeutic activity in murine ITP, while also exhibiting a distinct inflammatory signature involving production of CCL2, CCL5, and CXCL9 but not IFN-γ. Therefore, TER119 has been used to elucidate the potential mechanism underlying the adverse inflammatory activity associated with anti-erythrocyte Ab therapy in murine ITP. Prior work has demonstrated that TER119 administration is associated with a dramatic decrease in body temperature and inflammatory cytokine/chemokine production. The work presented in the current study demonstrates that inhibiting the highly inflammatory platelet-activating factor (PAF) pathway with PAF receptor antagonists prevents TER119-driven changes in body temperature and inhibits the production of the CCL2, CCL5, and CXCL9 inflammatory cytokines in CD-1 mice. Phagocytic cells and a functional TER119 Fc region were found to be necessary for TER119-induced body temperature changes and increases in CXCL9 and CCL2. Taken together, this work reveals the novel requirement of the PAF pathway in causing adverse inflammatory activity associated with anti-erythrocyte Ab therapy in a murine model and provides a strategy of mitigating these potential reactions without altering therapeutic activity.


Assuntos
Quimiocina CCL2 , Eritrócitos , Inflamação , Fator de Ativação de Plaquetas , Glicoproteínas da Membrana de Plaquetas , Púrpura Trombocitopênica Idiopática , Animais , Camundongos , Fator de Ativação de Plaquetas/imunologia , Púrpura Trombocitopênica Idiopática/imunologia , Púrpura Trombocitopênica Idiopática/tratamento farmacológico , Eritrócitos/imunologia , Inflamação/imunologia , Glicoproteínas da Membrana de Plaquetas/antagonistas & inibidores , Glicoproteínas da Membrana de Plaquetas/imunologia , Quimiocina CCL2/imunologia , Quimiocina CCL5/imunologia , Quimiocina CXCL9/imunologia , Receptores Acoplados a Proteínas G/imunologia , Transdução de Sinais/imunologia , Camundongos Endogâmicos C57BL , Autoanticorpos/imunologia , Modelos Animais de Doenças
2.
Int J Hematol ; 115(3): 322-328, 2022 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-35037230

RESUMO

Autoimmune hematological disorders are rare complications after allogeneic hematopoietic stem cell transplantation (allo-HSCT). Diagnosis of immune thrombocytopenia (ITP) is challenging, especially after allo-HSCT, because various complications such as graft-versus-host disease, disease relapse, viral infection, thrombotic microangiopathy, and drug side effects can also cause thrombocytopenia. Assessment of reticulated platelets (RP) and plasma thrombopoietin (TPO) levels may be useful to distinguish between ITP and hypoplastic thrombocytopenia. ITP is generally characterized by an increased percentage of RP, and a normal or slightly increased plasma TPO level. We now report three cases of thrombocytopenia after allo-HSCT. RP% was elevated in these patients, as it is in primary ITP. However, in contrast to primary ITP, plasma TPO levels were high in two of three patients. Anti-αIIbß3 and anti-GPIb/IX-specific direct IgG antibodies were detected as well, suggesting occurrence of immune-mediated platelet destruction in addition to bone marrow suppression in two patients. All three patients were successfully treated with corticosteroids and/or thrombopoietin receptor agonists (TPO-RAs). These results suggest that increased RP% and detection of glycoprotein-specific platelet autoantibodies are useful for the diagnosis of ITP after HSCT.


Assuntos
Autoanticorpos/sangue , Plaquetas , Transplante de Células-Tronco Hematopoéticas/efeitos adversos , Contagem de Plaquetas , Glicoproteínas da Membrana de Plaquetas/imunologia , Complicações Pós-Operatórias/diagnóstico , Complicações Pós-Operatórias/etiologia , Púrpura Trombocitopênica Idiopática/diagnóstico , Púrpura Trombocitopênica Idiopática/etiologia , Transplante Homólogo/efeitos adversos , Adolescente , Corticosteroides/uso terapêutico , Adulto , Biomarcadores/sangue , Plaquetas/imunologia , Feminino , Humanos , Imunoglobulina G/sangue , Masculino , Pessoa de Meia-Idade , Púrpura Trombocitopênica Idiopática/tratamento farmacológico
4.
J Clin Invest ; 131(10)2021 05 17.
Artigo em Inglês | MEDLINE | ID: mdl-33830943

RESUMO

A complete carcinogen, ultraviolet B (UVB) radiation (290-320 nm), is the major cause of skin cancer. UVB-induced systemic immunosuppression that contributes to photocarcinogenesis is due to the glycerophosphocholine-derived lipid mediator platelet-activating factor (PAF). A major question in photobiology is how UVB radiation, which only absorbs appreciably in the epidermal layers of skin, can generate systemic effects. UVB exposure and PAF receptor (PAFR) activation in keratinocytes induce the release of large numbers of microvesicle particles (MVPs; extracellular vesicles ranging from 100 to 1000 nm in size). MVPs released from skin keratinocytes in vitro in response to UVB (UVB-MVPs) are dependent on the keratinocyte PAFR. Here, we used both pharmacologic and genetic approaches in cells and mice to show that both the PAFR and enzyme acid sphingomyelinase (aSMase) were necessary for UVB-MVP generation. Our discovery that the calcium-sensing receptor is a keratinocyte-selective MVP marker allowed us to determine that UVB-MVPs leaving the keratinocyte can be found systemically in mice and humans following UVB exposure. Moreover, we found that UVB-MVPs contained bioactive contents including PAFR agonists that allowed them to serve as effectors for UVB downstream effects, in particular UVB-mediated systemic immunosuppression.


Assuntos
Micropartículas Derivadas de Células/imunologia , Tolerância Imunológica/efeitos da radiação , Queratinócitos/imunologia , Raios Ultravioleta , Animais , Linhagem Celular , Micropartículas Derivadas de Células/genética , Feminino , Humanos , Camundongos , Camundongos Knockout , Fator de Ativação de Plaquetas/genética , Fator de Ativação de Plaquetas/imunologia , Glicoproteínas da Membrana de Plaquetas/genética , Glicoproteínas da Membrana de Plaquetas/imunologia , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/imunologia , Esfingomielina Fosfodiesterase/genética , Esfingomielina Fosfodiesterase/imunologia
5.
Pharmacol Ther ; 217: 107630, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-32681846

RESUMO

Subendothelial collagen exposed to platelets after rupture of atherosclerotic plaques is the main trigger of platelet activation und acute arterial thrombotic occlusion leading to myocardial infarction or ischemic stroke. An efficacious antiplatelet therapy is essential to prevent atherothrombotic events. However, increasing potency of antiplatelet treatment is associated with an increased risk of bleeding limiting the clinical benefit for the patient since current antiplatelet strategies concomitantly affect hemostasis. Therefore, an unmet clinical need remains to develop antiplatelet strategies that selectively inhibit atherothrombosis without interfering with control of bleeding. Platelet glycoprotein VI (GPVI) plays a crucial role in collagen-induced activation and aggregation of platelets. Since GPVI is platelet-specific and strongly involved in the pathogenesis of arterial thrombosis without great impact on normal hemostasis, GPVI moved into the focus of novel approaches of antithrombotic therapy strategies. This review summarizes ligands, properties, function and downstream signaling of GPVI and discusses the potential of GPVI as target for novel therapeutic strategies in thrombotic and inflammatory diseases on the basis of recent scientific findings and currently ongoing clinical phase I and phase II trials.


Assuntos
Aterosclerose/prevenção & controle , Ativação Plaquetária/fisiologia , Inibidores da Agregação Plaquetária/farmacologia , Glicoproteínas da Membrana de Plaquetas/metabolismo , Trombose/prevenção & controle , Animais , Biomarcadores , Plaquetas/fisiologia , Colágeno/metabolismo , Hemorragia/induzido quimicamente , Humanos , Ligantes , Neoplasias/fisiopatologia , Agregação Plaquetária/fisiologia , Inibidores da Agregação Plaquetária/efeitos adversos , Glicoproteínas da Membrana de Plaquetas/antagonistas & inibidores , Glicoproteínas da Membrana de Plaquetas/imunologia , Transdução de Sinais
6.
Platelets ; 32(3): 314-324, 2021 Apr 03.
Artigo em Inglês | MEDLINE | ID: mdl-32896192

RESUMO

Platelets are increasingly being recognized for playing roles beyond thrombosis and hemostasis. Today we know that they mediate inflammation by direct interactions with innate immune cells or secretion of cytokines/chemokines. Here we review their interactions with neutrophils and monocytes/macrophages in infection and sepsis, stroke, myocardial infarction and venous thromboembolism. We discuss new roles for platelet surface receptors like GPVI or GPIb and also look at platelet contributions to the formation of neutrophil extracellular traps (NETs) as well as to deep vein thrombosis during infection, e.g. in COVID-19 patients.


Assuntos
Plaquetas/imunologia , COVID-19/imunologia , Infarto do Miocárdio/imunologia , Neutrófilos/imunologia , Sepse/imunologia , Acidente Vascular Cerebral/imunologia , Tromboembolia Venosa/imunologia , Plaquetas/patologia , COVID-19/genética , COVID-19/patologia , COVID-19/virologia , Comunicação Celular/genética , Comunicação Celular/imunologia , Citocinas/genética , Citocinas/imunologia , Armadilhas Extracelulares/genética , Armadilhas Extracelulares/imunologia , Regulação da Expressão Gênica , Humanos , Inflamação , Macrófagos/imunologia , Macrófagos/patologia , Monócitos/imunologia , Monócitos/patologia , Infarto do Miocárdio/genética , Infarto do Miocárdio/patologia , Neutrófilos/patologia , Complexo Glicoproteico GPIb-IX de Plaquetas/genética , Complexo Glicoproteico GPIb-IX de Plaquetas/imunologia , Glicoproteínas da Membrana de Plaquetas/genética , Glicoproteínas da Membrana de Plaquetas/imunologia , Sepse/genética , Sepse/patologia , Acidente Vascular Cerebral/genética , Acidente Vascular Cerebral/patologia , Tromboembolia Venosa/genética , Tromboembolia Venosa/patologia
7.
J Immunol Methods ; 487: 112868, 2020 12.
Artigo em Inglês | MEDLINE | ID: mdl-32941887

RESUMO

Antibodies against human platelets cause a variety of thrombocytopenic disorders, which lead to potentially fatal haemorrhage. Therefore, their prompt detection is mandatory for successful patient treatment. Solid phase red cell adherence (SPRCA) assay allows for platelet antibody detection widely. However, preparation of fresh platelets with HLA-I and human platelet antigens (HPA)1-5,15 genotyped as target cells is inconvenient and fresh platelets have a short shelf life. In this study, the lyophilised human platelets for antibody detection in SPRCA were prepared. Firstly, platelets were resuspended in lyophilisation buffer and freeze-dried. Then the characteristics of lyophilised platelet were analysed. Rehydrated platelets were recovered with a mean rate of 80.91% ± 2.87%, and still retained spherical morphology. Indirect flow cytometry showed that glycoproteins IIb/IIIa, Ia/IIa, Ib/IX, IV, CD109, and HLA class I were present on the surface of the lyophilised platelets at a comparable level to that of fresh platelets. The consistent results obtained with WHO reference reagents containing anti-HPA-1a, anti-HPA-3a, and anti-HPA-5b, as well as clinical samples from the same donors containing anti-HLA antibodies when reacting with lyophilised versus fresh platelets confirmed good antigenicity preservation of platelets after freeze-drying. Further investigation showed that the lyophilised platelets could be stored at 2-8 °C for up to 14 months and the reconstituted suspension was stable for 48 h. Therefore, lyophilised platelets can be a convenient alternative to fresh platelets to use for anti-platelet antibody detection in SPRCA tests.


Assuntos
Antígenos de Plaquetas Humanas/imunologia , Plaquetas/imunologia , Reação de Imunoaderência , Isoanticorpos/sangue , Glicoproteínas da Membrana de Plaquetas/imunologia , Trombocitopenia/diagnóstico , Biomarcadores/sangue , Coleta de Amostras Sanguíneas , Estudos de Casos e Controles , Liofilização , Histocompatibilidade , Humanos , Integrina beta3 , Valor Preditivo dos Testes , Trombocitopenia/sangue , Trombocitopenia/imunologia
8.
Thromb Haemost ; 120(6): 977-993, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32492725

RESUMO

Glycoprotein VI (GPVI), a platelet collagen receptor, is crucial in mediating atherothrombosis. Besides collagen, injured plaques expose tissue factor (TF) that triggers fibrin formation. Previous studies reported that GPVI also is a platelet receptor for fibrinogen and fibrin. We studied the effect of anti-GPVI antibodies and inhibitors of GPVI signaling kinases (Syk and Btk) on platelet adhesion and aggregate formation onto immobilized fibrinogen and different types of fibrin under arterial flow conditions. Fibrin was prepared from isolated fibrinogen ("pure fibrin"), recombinant fibrinogen ("recombinant fibrin"), or generated more physiologically from endogenous fibrinogen in plasma ("plasma fibrin") or by exposing TF-coated surfaces to flowing blood ("blood fibrin"). Inhibition of GPVI and Syk did not inhibit platelet adhesion and aggregate formation onto fibrinogen. In contrast anti-GPVI antibodies, inhibitors of Syk and Btk and the anti-GPIb antibody 6B4 inhibited platelet aggregate formation onto pure and recombinant fibrin. However, inhibition of GPVI and GPVI signaling did not significantly reduce platelet coverage of plasma fibrin and blood fibrin. Plasma fibrin contained many proteins incorporated during clot formation. Advanced optical imaging revealed plasma fibrin as a spongiform cushion with thicker, knotty, and long fibers and little activation of adhering platelets. Albumin intercalated in plasma fibrin fibers left only little space for platelet attachment. Pure fibrin was different showing a dense mesh of thin fibers with strongly activated platelets. We conclude that fibrin formed in plasma and blood contains plasma proteins shielding GPVI-activating epitopes. Our findings do not support a role of GPVI for platelet activation by physiologic fibrin.


Assuntos
Plaquetas/metabolismo , Fibrina/metabolismo , Glicoproteínas da Membrana de Plaquetas/fisiologia , Receptores de Peptídeos/metabolismo , Tirosina Quinase da Agamaglobulinemia/sangue , Tirosina Quinase da Agamaglobulinemia/fisiologia , Ativação Enzimática , Fibrinogênio/metabolismo , Hemorreologia , Humanos , Microscopia Confocal/métodos , Plasma , Adesividade Plaquetária , Agregação Plaquetária , Complexo Glicoproteico GPIb-IX de Plaquetas/metabolismo , Glicoproteínas da Membrana de Plaquetas/antagonistas & inibidores , Glicoproteínas da Membrana de Plaquetas/imunologia , Ligação Proteica , Proteínas Recombinantes/metabolismo , Quinase Syk/antagonistas & inibidores , Quinase Syk/sangue , Quinase Syk/fisiologia , Tromboplastina/metabolismo
9.
Int J Lab Hematol ; 42(4): 371-379, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32202052

RESUMO

INTRODUCTION: Hemorrhage and infection are two main causes of death in patients with myelodysplastic syndromes (MDS), and it is becoming increasingly clear that platelet dysfunction can also affect the process of hemostasis and anti-infection. The aim of this study was to evaluate activation function and immune-related function of platelets in MDS. METHODS: We included 29 MDS patients and divided them into different subgroups (low-risk group and high-risk group; untreated group and treated group; pretransfusion group and post-transfusion group) according to IPSS-R score, hypomethylating agents (HMAs) therapy, and platelet transfusion history. Platelet light scatter properties, expression of CD41a, activation-associated phenotypes (CD62p and CD63), and immune-associated phenotypes (CD154 and TLR4) were detected by multiparameter flow cytometry. RESULTS: Expression of CD41a was decreased (P < .05), and no difference was found in platelet light scatter properties between MDS patients and healthy subjects (P > .05). Significantly decreased expression frequency and intensity of activation phenotype CD63 were found in patients with MDS (P < .05). Low-risk MDS showed lower expression frequency while high-risk MDS showed reduced mean fluorescence intensity (MFI) of CD63. Decreased expression of CD154 and TLR4 was found in MDS patients (P < .05) which was significantly elevated after HMAs therapy (P < .05). Particularly, MFI of CD154 and TLR4 reduced in high-risk MDS patients (P < .05). CONCLUSION: Myelodysplastic syndromes patients displayed defective expression of both activation- and immune-associated platelet phenotypes, with differential mechanisms between low-risk and high-risk groups regarding phenotype alterations. The findings confirmed impaired platelet phenotypes in MDS which may assist in the diagnosis and identification of MDS patients.


Assuntos
Plaquetas/metabolismo , Regulação da Expressão Gênica , Síndromes Mielodisplásicas/sangue , Ativação Plaquetária , Glicoproteínas da Membrana de Plaquetas/biossíntese , Adulto , Idoso , Idoso de 80 Anos ou mais , Plaquetas/imunologia , Plaquetas/patologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Síndromes Mielodisplásicas/imunologia , Síndromes Mielodisplásicas/patologia , Síndromes Mielodisplásicas/terapia , Glicoproteínas da Membrana de Plaquetas/imunologia
10.
Front Immunol ; 10: 1260, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31244834

RESUMO

Obstruction of a coronary artery causes ischemia of heart tissue leading to myocardial infarction. Prolonged oxygen deficiency provokes tissue necrosis, which can result in heart failure and death of the patient. Therefore, restoration of coronary blood flow (reperfusion of the ischemic area) by re-canalizing the affected vessel is essential for a better patient outcome. Paradoxically, sudden reperfusion also causes tissue injury, thereby increasing the initial ischemic damage despite restoration of blood flow (=ischemia/reperfusion injury, IRI). Myocardial IRI is a complex event that involves various harmful mechanisms (e.g., production of reactive oxygen species and local increase in calcium ions) as well as inflammatory cells and signals like chemokines and cytokines. An involvement of platelets in the inflammatory reaction associated with IRI was discovered several years ago, but the underlying mechanisms are not yet fully understood. This mini review focusses on platelet contributions to the intricate picture of myocardial IRI. We summarize how upregulation of platelet surface receptors and release of immunomodulatory mediators lead to aggravation of myocardial IRI and subsequent cardiac damage by different mechanisms such as recruitment and activation of immune cells or modification of the cardiac vascular endothelium. In addition, evidence for cardioprotective roles of distinct platelet factors during IRI will be discussed.


Assuntos
Plaquetas/metabolismo , Suscetibilidade a Doenças , Traumatismo por Reperfusão Miocárdica/etiologia , Traumatismo por Reperfusão Miocárdica/metabolismo , Animais , Plaquetas/imunologia , Comunicação Celular/imunologia , Citocinas/metabolismo , Modelos Animais de Doenças , Células Endoteliais/imunologia , Células Endoteliais/metabolismo , Humanos , Imunomodulação , Mediadores da Inflamação/metabolismo , Leucócitos/imunologia , Leucócitos/metabolismo , Traumatismo por Reperfusão Miocárdica/patologia , Estresse Oxidativo , Glicoproteínas da Membrana de Plaquetas/imunologia , Glicoproteínas da Membrana de Plaquetas/metabolismo , Espécies Reativas de Oxigênio/metabolismo
11.
Haematologica ; 104(6): 1237-1243, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-30923095

RESUMO

Platelet autoantibody-induced platelet clearance represents a major pathomechanism in immune thrombocytopenia (ITP). There is growing evidence for clinical differences between anti-glycoprotein IIb/IIIa and anti-glycoprotein Ib/IX mediated ITP. Glycoprotein V is a well characterized target antigen in Varicella-associated and drug-induced thrombocytopenia. We conducted a systematic study assessing the prevalence and functional capacity of autoantibodies against glycoprotein V. A total of 1140 patients were included. In one-third of patients, platelet-bound autoantibodies against glycoproteins Ib/IX, IIb/IIIa, or V were detected in a monoclonal antibody immobilization of platelet antigen assay; platelet-bound autoantiglycoprotein V was present in the majority of samples (222 out of 343, 64.7%). Investigation of patient sera revealed the presence of free autoantibodies against glycoprotein V in 13.5% of these patients by an indirect monoclonal antibody immobilization of platelet antigen assay, but in 39.6% by surface plasmon resonance technology. These antibodies showed significantly lower avidity (association/dissociation ratio 0.32±0.13 vs 0.73±0.14; P<0.001). High- and low-avidity antibodies induced comparable amounts of platelet uptake in a phagocytosis assay using CD14+ positively-selected human macrophages [mean phagocytic index, 6.81 (range, 4.75-9.86) vs 6.01 (range, 5.00-6.98); P=0.954]. In a NOD/SCID mouse model, IgG prepared from both types of anti-glycoprotein V autoantibodies eliminated human platelets with no detectable difference between the groups from the murine circulation [mean platelet survival at 300 minutes, 40% (range, 27-55) vs 35% (16-46); P=0.025]. Our data establish glycoprotein V as a relevant immune target in immune thrombocytopenia. We would suggest that further studies including glycoprotein V will be required before ITP treatment can be tailored according to platelet autoantibody specificity.


Assuntos
Autoanticorpos/imunologia , Autoantígenos/imunologia , Suscetibilidade a Doenças/imunologia , Glicoproteínas da Membrana de Plaquetas/imunologia , Púrpura Trombocitopênica Idiopática/imunologia , Animais , Plaquetas/imunologia , Plaquetas/metabolismo , Modelos Animais de Doenças , Feminino , Humanos , Masculino , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Fagocitose , Prevalência , Ligação Proteica/imunologia , Púrpura Trombocitopênica Idiopática/epidemiologia
12.
Thromb Res ; 176: 108-114, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-30802692

RESUMO

BACKGROUND: The main role of platelets is to control haemostasis when there is a blood vessel injury in order to minimise blood loss at the injury site. Under normal circumstances, platelets flow freely within blood vessels as the endothelial cells provide a non-adhesion surface. Naturally, bioactive mediators are released from endothelial cells to prevent and control platelet activation. However, when the vascular endothelium is ruptured, the local concentration of nitric oxide and prostaglandin is diminished and receptors containing a sequence of amino acids known as, immunoreceptor tyrosine-based inhibition motifs (ITIMs), serve as natural inhibitors within platelets. The main role of ITIMs is to decrease immunoreceptor tyrosine-based activation motif (ITAM) signalling in platelets; however, some studies have revealed their novel role in integrin αIIbß3 activation. This review highlights the main structural and functional features of immunoreceptors in platelets.


Assuntos
Plaquetas/imunologia , Hemostasia , Motivo de Inibição do Imunorreceptor Baseado em Tirosina , Animais , Antígenos CD/imunologia , Moléculas de Adesão Celular/imunologia , Humanos , Motivo de Ativação do Imunorreceptor Baseado em Tirosina , Ativação Plaquetária , Molécula-1 de Adesão Celular Endotelial a Plaquetas/imunologia , Complexo Glicoproteico GPIIb-IIIa de Plaquetas/imunologia , Glicoproteínas da Membrana de Plaquetas/imunologia , Receptores de IgG/imunologia
13.
J Clin Invest ; 129(1): 12-23, 2019 01 02.
Artigo em Inglês | MEDLINE | ID: mdl-30601137

RESUMO

Although platelets are best known for their role in hemostasis, they are also crucial in development, host defense, inflammation, and tissue repair. Many of these roles are regulated by the immune-like receptors glycoprotein VI (GPVI) and C-type lectin receptor 2 (CLEC-2), which signal through an immunoreceptor tyrosine-based activation motif (ITAM). GPVI is activated by collagen in the subendothelial matrix, by fibrin and fibrinogen in the thrombus, and by a remarkable number of other ligands. CLEC-2 is activated by the transmembrane protein podoplanin, which is found outside of the vasculature and is upregulated in development, inflammation, and cancer, but there is also evidence for additional ligands. In this Review, we discuss the physiological and pathological roles of CLEC-2 and GPVI and their potential as targets in thrombosis and thrombo-inflammatory disorders (i.e., disorders in which inflammation plays a critical role in the ensuing thrombosis) relative to current antiplatelet drugs.


Assuntos
Matriz Extracelular/imunologia , Lectinas Tipo C/imunologia , Glicoproteínas de Membrana/imunologia , Proteínas de Neoplasias/imunologia , Neoplasias/imunologia , Glicoproteínas da Membrana de Plaquetas/imunologia , Trombose/imunologia , Motivos de Aminoácidos , Animais , Colágeno/imunologia , Matriz Extracelular/patologia , Fibrina/imunologia , Fibrinogênio/imunologia , Humanos , Inflamação/imunologia , Inflamação/patologia , Neoplasias/tratamento farmacológico , Neoplasias/patologia , Inibidores da Agregação Plaquetária/uso terapêutico , Trombose/tratamento farmacológico , Trombose/patologia
14.
Biomaterials ; 167: 107-120, 2018 06.
Artigo em Inglês | MEDLINE | ID: mdl-29567387

RESUMO

Collagen, a strong platelet activator, is recognized by integrin α2ß1 and GPVI. It induces aggregation, if added to suspended platelets, or platelet adhesion if immobilized to a surface. The recombinant non-prolylhydroxylated mini-collagen FC3 triple helix containing one α2ß1 integrin binding site is a tool to specifically study how α2ß1 integrin activates platelet. Whereas soluble FC3 monomers antagonistically block collagen-induced platelet activation, immobilization of several FC3 molecules to an interface or to colloidal nanobeads determines the agonistic action of FC3. Nanopatterning of FC3 reveals that intermolecular distances below 64 nm between α2ß1 integrin binding sites trigger signaling through dot-like clusters of α2ß1 integrin, which are visible in high resolution microscopy with dSTORM. Upon signaling, these integrin clusters increase in numbers per platelet, but retain their individual size. Immobilization of several FC3 to 100 nm-sized nanobeads identifies α2ß1 integrin-triggered signaling in platelets to occur at a twentyfold slower rate than collagen, which activates platelet in a fast integrative signaling via different platelet receptors. As compared to collagen stimulation, FC3-nanobead-triggered signaling cause a significant stronger activation of the protein kinase BTK, a weak and dispensable activation of PDK1, as well as a distinct phosphorylation pattern of PDB/Akt.


Assuntos
Tirosina Quinase da Agamaglobulinemia/imunologia , Plaquetas/citologia , Colágeno/imunologia , Integrina alfa2beta1/imunologia , Ativação Plaquetária , Sítios de Ligação , Plaquetas/imunologia , Colágeno/química , Humanos , Proteínas Imobilizadas/química , Proteínas Imobilizadas/imunologia , Glicoproteínas da Membrana de Plaquetas/imunologia , Transdução de Sinais
15.
Biochem Biophys Res Commun ; 495(4): 2475-2481, 2018 01 22.
Artigo em Inglês | MEDLINE | ID: mdl-29278700

RESUMO

Myocardial ischemia/reperfusion (I/R) still have high morbidity and mortality worldwide. Platelet activating factor (PAF) is a potent phospholipid regulator of inflammation. PAF acts on a single receptor (PAFR), which is expressed on cellular and nuclear membranes of various cell types. The study is aimed to explore if PAFR could modulate myocardial I/R injury in mice. PAFR expressions began to up-regulate at 1 h, and reached peak at 24 h. PAFR deletion markedly attenuated myocardial I/R injury, evidenced by the reduced infarct size and the improved cardiac function. Furthermore, PAFR-knockout inhibited inflammatory response, as demonstrated by down-regulated pro-inflammatory cytokines and chemokine, as well as the inactivation of nuclear factor κB (NF-κB). Additionally, PAFR-absence ameliorated oxidative stress induced by myocardial I/R, associated with the up-regulation of superoxide dismutase (SOD) and nuclear respiratory factor 2 (Nrf-2) activity. Finally, PAFR-deficiency impeded apoptosis, which was proved by the decreasing in terminal deoxynucleotidyl transferase (TdT)-mediated dNTP nick end labeling (TUNEL)-positive myocytes, and Caspase-3 cleavage. And the activation of Janus kinase 1-signal transducer and activator of transcription 1 (JAK1/STAT1) pathway was also suppressed by PAFR-knockout. The findings above were confirmed in lipopolysaccharide (LPS)-incubated cardiomyocytes with or without PAFR expressions in vitro. In summary, we supposed that inhibiting PAFR reduced inflammation, oxidative stress and apoptosis, and thus might be a promising therapeutic strategy to alleviate myocardial I/R injury.


Assuntos
Apoptose/imunologia , Traumatismo por Reperfusão Miocárdica/imunologia , Miocardite/imunologia , Estresse Oxidativo/imunologia , Animais , Citocinas/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Traumatismo por Reperfusão Miocárdica/patologia , Miocardite/patologia , Glicoproteínas da Membrana de Plaquetas/genética , Glicoproteínas da Membrana de Plaquetas/imunologia , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/imunologia
16.
Nat Commun ; 8(1): 2239, 2017 12 21.
Artigo em Inglês | MEDLINE | ID: mdl-29269852

RESUMO

Platelets play a critical role in vascular inflammation through the podoplanin and collagen/fibrin receptors, C-type-lectin-like-2 (CLEC-2) and glycoprotein VI (GPVI), respectively. Both receptors regulate endothelial permeability and prevent peri-vascular bleeding in inflammation. Here we show that platelet-specific deletion of CLEC-2 but not GPVI leads to enhanced systemic inflammation and accelerated organ injury in two mouse models of sepsis-intra-peritoneal lipopolysaccharide and cecal ligation and puncture. CLEC-2 deficiency is associated with reduced numbers of podoplanin-expressing macrophages despite increased cytokine and chemokine levels in the infected peritoneum. Pharmacological inhibition of the interaction between CLEC-2 and podoplanin regulates immune cell infiltration and the inflammatory reaction during sepsis, suggesting that activation of podoplanin underlies the anti-inflammatory action of platelet CLEC-2. We suggest podoplanin-CLEC-2 as a novel anti-inflammatory axis regulating immune cell recruitment and activation in sepsis.


Assuntos
Plaquetas/imunologia , Inflamação/imunologia , Lectinas Tipo C/imunologia , Macrófagos/imunologia , Glicoproteínas de Membrana/imunologia , Insuficiência de Múltiplos Órgãos/imunologia , Sepse/imunologia , Animais , Ceco/cirurgia , Quimiocinas/imunologia , Citocinas/imunologia , Injeções Intraperitoneais , Rim/imunologia , Rim/patologia , Lectinas Tipo C/genética , Ligadura , Lipopolissacarídeos/toxicidade , Glicoproteínas de Membrana/genética , Camundongos , Fagocitose/imunologia , Glicoproteínas da Membrana de Plaquetas/genética , Glicoproteínas da Membrana de Plaquetas/imunologia , Punções , Sepse/induzido quimicamente
17.
J Leukoc Biol ; 102(3): 871-880, 2017 09.
Artigo em Inglês | MEDLINE | ID: mdl-28760822

RESUMO

Formyl peptide receptor (FPR)-desensitized neutrophils display increased production/release of superoxide (O2-) when activated by platelet-activating factor (PAF), a priming of the response achieved through a unique receptor crosstalk mechanism. The aim of this study was to determine the effect of an inhibitor selective for small, heterotrimeric G proteins belonging to the Gαq subclass on that receptor crosstalk. We show that signals generated by FPRs and the PAF receptor (PAFR) induce activation of the neutrophil O2-, producing NADPH-oxidase, and that response was sensitive to Gαq inhibition in cells activated by PAF, but no inhibition was obtained in cells activated by FPR agonists. Signaling in naive neutrophils is terminated fairly rapidly, and the receptors become homologously desensitized. The downstream sensitivity to Gαq inhibition in desensitized cells displaying increased production/release of O2- through the PAFR receptor crosstalk mechanism also comprised the reactivation of the FPRs, and the activation signals were redirected from the PAFR to the desensitized/reactivated FPRs. The Gαq-dependent activation signals generated by the PAFRs activate the Gαi-coupled FPRs, a receptor crosstalk that represents a novel pathway by which G protein-coupled receptors can be regulated and signaling can be turned on and off.


Assuntos
Subunidades alfa Gq-G11 de Proteínas de Ligação ao GTP/imunologia , Neutrófilos/imunologia , Glicoproteínas da Membrana de Plaquetas/imunologia , Receptores de Formil Peptídeo/imunologia , Receptores Acoplados a Proteínas G/imunologia , Transdução de Sinais/imunologia , Humanos , NADPH Oxidases/imunologia , Fator de Ativação de Plaquetas/imunologia , Superóxidos/imunologia
18.
J Immunol ; 199(3): 1184-1195, 2017 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-28667161

RESUMO

Over 89% of asthmatic children in underdeveloped countries demonstrate sensitivity to house dust mites (HDMs). The allergic response to HDMs is partially mediated by epithelial cell-derived cytokines that activate group 2 innate lymphoid cells, induce migration and activation of dendritic cells, and promote effector differentiation of HDM-specific TH2 cells. However, the contribution of innate receptor engagement on epithelial or dendritic cells by HDMs that ultimately mediates said innate and adaptive allergic responses is poorly understood. We and other investigators have demonstrated that HDMs express phosphorylcholine (PC) moieties. The major PC receptors involved in immune responses include CD36 and platelet-activating factor receptor (PAFR). Because CD36 and PAFR are expressed by epithelial cells and dendritic cells, and expression of these receptors is higher in human asthmatics, we determined whether engagement of CD36 or PAFR on epithelial or dendritic cells contributes to HDM allergy development. Testing bone marrow chimeric mice revealed that CD36 engagement on radioresistant cells and PAFR engagement on radioresistant and radiosensitive cells in the lung promote allergic responses to HDMs. Additionally, passive anti-PC IgM Abs administered intratracheally with HDMs decreased allergen uptake by epithelial cells and APCs in the lungs of C57BL/6 mice but not CD36-/- or PAFR-/- mice. These results show that CD36 and PAFR are important mediators of HDM allergy development and that inhibiting HDM engagement with PC receptors in the lung protects against allergic airway disease.


Assuntos
Antígenos CD36/imunologia , Antígenos CD36/metabolismo , Glicoproteínas da Membrana de Plaquetas/imunologia , Glicoproteínas da Membrana de Plaquetas/metabolismo , Pyroglyphidae/imunologia , Receptores Acoplados a Proteínas G/imunologia , Receptores Acoplados a Proteínas G/metabolismo , Alérgenos/imunologia , Animais , Antígenos de Dermatophagoides/imunologia , Asma/imunologia , Asma/prevenção & controle , Antígenos CD36/deficiência , Antígenos CD36/genética , Células Dendríticas/imunologia , Células Epiteliais/imunologia , Hipersensibilidade/imunologia , Hipersensibilidade/prevenção & controle , Imunidade Inata , Imunoglobulina M/administração & dosagem , Pulmão/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Fosforilcolina/química , Fosforilcolina/imunologia , Pyroglyphidae/química , Hipersensibilidade Respiratória/imunologia , Hipersensibilidade Respiratória/prevenção & controle
19.
Blood Coagul Fibrinolysis ; 28(8): 617-622, 2017 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-28654425

RESUMO

: Immune thrombocytopenic purpura (ITP) is an autoimmune disease in which increased platelet destruction and thrombocytopenia are diagnostic features. In fact, the exact pathogenesis of this disease is still unknown, but genetic changes can be a potential factor in the development of ITP. In this study, the relationship between polymorphisms with platelet destruction has been studied, which leads to decreased platelet count. Relevant literature was identified by a PubMed search (2000-2016) of English language papers using the terms 'ITP', 'polymorphism,' and 'immune system'. The majority of genetic changes (polymorphisms) occur in immune system genes, including interferon (IFN)-γ gene. These changes lead to the dysfunction of immune system and production of pathogenic antibodies against platelet surface glycoproteins such as glycoprotein IIb/IIIa, which eventually result in the destruction of platelets and increasing disease severity. In addition, IFN-γ as well as factors and cytokines involved in megakaryopoiesis, including stem cell factor and interleukin-3 (IL-3), leads to the differentiation of megakaryocytes and platelet release. Considering the fact that IFN-γ is a factor of inflammation and thrombocytopenia, coexistence of this cytokine with thrombopoietin, stem cell factor, and IL-3 results in megakaryocytes differentiation and platelet production, which can be effective to reduce disease severity and increase the platelet counts.


Assuntos
Polimorfismo Genético , Púrpura Trombocitopênica Idiopática/genética , Citocinas , Humanos , Sistema Imunitário , Interferon gama/genética , Fenótipo , Glicoproteínas da Membrana de Plaquetas/imunologia
20.
Eur J Clin Pharmacol ; 73(8): 949-956, 2017 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-28523478

RESUMO

INTRODUCTION: Glycoprotein VI (GPVI) is the major platelet receptor for collagen-mediated platelet adhesion and activation. SAR264565 is an anti-GPVI-Fab, binds to GPVI with high affinity, and blocks GPVI function in human platelets in vitro. METHODS: The effect of SAR26456 on platelet responsiveness in the blood of 21 healthy male subjects was investigated using Sakariassen's ex vivo thrombogenesis perfusion chamber model on a collagen-coated surface under conditions mimicking arterial flow. Ex vivo effects of SAR264565 (10 and 100 µg/mL) were investigated before administration of aspirin or clopidogrel to study subjects (baseline), after aspirin (2× 300 mg) administration alone, and after combined aspirin (2× 300 mg)/clopidogrel (600 mg) administration. Additional ex vivo and in vitro platelet tests were also performed. RESULTS: Addition of SAR264565 to the perfusion chamber dose-dependently reduced platelet and fibrin deposition, reaching statistical significance at 100 µg/mL (415 ± 67 compared to 137 ± 36 platelets/cm2, [p < 0.01] and fibrin 0.095 ± 0.014 compared to 0.032 ± 0.008 µg/cm2, [p < 0.001]). Aspirin administration caused an additive and dose-dependent reduction of SAR264565-induced platelet and fibrin deposition. Combined aspirin/clopidogrel administration did not lead to additional SAR264565-induced inhibition of platelet or fibrin deposition. CONCLUSION: GPVI antagonism by the anti-GPVI-Fab fragment SAR264565 dose-dependently inhibits platelet adhesion and fibrin formation on a collagen surface under arterial shear. Additive inhibition is observed after prior aspirin administration with no further amplification on top of a combination of aspirin with clopidogrel. Ex vivo antiplatelet tests confirmed a selective inhibiting effect of SAR264565 on collagen-induced platelet activation.


Assuntos
Plaquetas/efeitos dos fármacos , Fragmentos Fab das Imunoglobulinas/farmacologia , Glicoproteínas da Membrana de Plaquetas/antagonistas & inibidores , Adulto , Plaquetas/fisiologia , Fibrina/metabolismo , Humanos , Masculino , Agregação Plaquetária/efeitos dos fármacos , Glicoproteínas da Membrana de Plaquetas/imunologia , Adulto Jovem
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