RESUMEN
Antiphospholipid syndrome (APS) is an autoimmune disease characterized by recurrent arterial, venous, and microvascular thrombosis where activated neutrophils play a determinant role. However, neutrophils are challenging to target given their short lifespan in circulation and spontaneous activation. Screening of a small library of gold nanoparticles (AuNPs) led to the discovery of a formulation capable of targeting activated neutrophil attachment and has demonstrated that star-shaped, anti-PSGL-1-antibody-coated AuNPs (aPSGL-1-AuNPs) were more efficacious compared with other shapes of AuNPs. Our findings further revealed an exciting and safe targeting mode toward activated neutrophils in the APS mouse model induced by human-patient-derived antiphospholipid IgGs. Our studies demonstrate that targeting is dependent on the specific topographical features of the highly segregated PSGL-1 on the activated neutrophil surface for which a high affinity shape-driven nanomedicine can be designed and implemented. As such, star-shaped aPSGL-1-AuNPs serve as a promising nanoimmunotherapy for immunothrombosis associated with neutrophil adhesion in APS.
Asunto(s)
Síndrome Antifosfolípido , Nanopartículas del Metal , Trombosis , Animales , Ratones , Humanos , Síndrome Antifosfolípido/tratamiento farmacológico , Síndrome Antifosfolípido/complicaciones , Síndrome Antifosfolípido/diagnóstico , Neutrófilos , Oro/uso terapéutico , Nanopartículas del Metal/uso terapéutico , Trombosis/tratamiento farmacológico , Inmunoglobulina GRESUMEN
Background Proper function of endothelial cells is critical for vascular integrity and organismal survival. Studies over the past 2 decades have identified 2 members of the KLF (Krüppel-like factor) family of proteins, KLF2 and KLF4, as nodal regulators of endothelial function. Strikingly, inducible postnatal deletion of both KLF2 and KLF4 resulted in widespread vascular leak, coagulopathy, and rapid death. Importantly, while transcriptomic studies revealed profound alterations in gene expression, the molecular mechanisms underlying these changes remain poorly understood. Here, we seek to determine mechanisms of KLF2 and KLF4 transcriptional control in multiple vascular beds to further understand their roles as critical endothelial regulators. Methods and Results We integrate chromatin occupancy and transcription studies from multiple transgenic mouse models to demonstrate that KLF2 and KLF4 have overlapping yet distinct binding patterns and transcriptional targets in heart and lung endothelium. Mechanistically, KLFs use open chromatin regions in promoters and enhancers and bind in context-specific patterns that govern transcription in microvasculature. Importantly, this occurs during homeostasis in vivo without additional exogenous stimuli. Conclusions Together, this work provides mechanistic insight behind the well-described transcriptional and functional heterogeneity seen in vascular populations, while also establishing tools into exploring microvascular endothelial dynamics in vivo.
Asunto(s)
Endotelio , Factor 4 Similar a Kruppel , Factores de Transcripción de Tipo Kruppel , Animales , Ratones , Cromatina/metabolismo , Células Endoteliales/metabolismo , Endotelio/metabolismo , Expresión Génica , Factor 4 Similar a Kruppel/genética , Factor 4 Similar a Kruppel/metabolismo , Factores de Transcripción de Tipo Kruppel/genética , Factores de Transcripción de Tipo Kruppel/metabolismoRESUMEN
Endothelial activation and sickle red blood cell (RBC) adhesion are central to the pathogenesis of sickle cell disease (SCD). Quantitatively, RBC-derived extracellular vesicles (REVs) are more abundant from SS RBCs compared with healthy RBCs (AA RBCs). Sickle RBC-derived REVs (SS REVs) are known to promote endothelial cell (EC) activation through cell signalling and transcriptional regulation at longer terms. However, the SS REV-mediated short-term non-transcriptional response of EC is unclear. Here, we examined the impact of SS REVs on acute microvascular EC activation and RBC adhesion at 2 h. Compared with AA REVs, SS REVs promoted human pulmonary microvascular ECs (HPMEC) activation indicated by increased von Willebrand factor (VWF) expression. Under microfluidic conditions, we found abnormal SS RBC adhesion to HPMECs exposed to SS REVs. This enhanced SS RBC adhesion was reduced by haeme binding protein haemopexin or VWF cleaving protease ADAMTS13 to a level similar to HPMECs treated with AA REVs. Consistent with these observations, haemin- or SS REV-induced microvascular stasis in SS mice with implanted dorsal skin-fold chambers that was inhibited by ADAMTS13. The adhesion induced by SS REVs was variable and was higher with SS RBCs from patients with increased markers of haemolysis (lactate dehydrogenase and reticulocyte count) or a concomitant clinical diagnosis of deep vein thrombosis. Our results emphasise the critical contribution made by REVs to the pathophysiology of SCD by triggering acute microvascular EC activation and abnormal RBC adhesion. These findings may help to better understand acute pathophysiological mechanism of SCD and thereby the development of new treatment strategies using VWF as a potential target.
Asunto(s)
Anemia de Células Falciformes , Células Endoteliales , Humanos , Animales , Ratones , Células Endoteliales/patología , Factor de von Willebrand/metabolismo , Adhesión Celular , Eritrocitos/metabolismoRESUMEN
Arterial and venous thrombosis constitutes a major source of morbidity and mortality worldwide. Long considered as distinct entities, accumulating evidence indicates that arterial and venous thrombosis can occur in the same populations, suggesting that common mechanisms are likely operative. Although hyperactivation of the immune system is a common forerunner to the genesis of thrombotic events in both vascular systems, the key molecular control points remain poorly understood. Consequently, antithrombotic therapies targeting the immune system for therapeutics gain are lacking. Here, we show that neutrophils are key effectors of both arterial and venous thrombosis and can be targeted through immunoregulatory nanoparticles. Using antiphospholipid antibody syndrome (APS) as a model for arterial and venous thrombosis, we identified the transcription factor Krüppel-like factor 2 (KLF2) as a key regulator of neutrophil activation. Upon activation through genetic loss of KLF2 or administration of antiphospholipid antibodies, neutrophils clustered P-selectin glycoprotein ligand 1 (PSGL-1) by cortical actin remodeling, thereby increasing adhesion potential at sites of thrombosis. Targeting clustered PSGL-1 using nanoparticles attenuated neutrophil-mediated thrombosis in APS and KLF2 knockout models, illustrating the importance and feasibility of targeting activated neutrophils to prevent pathological thrombosis. Together, our results demonstrate a role for activated neutrophils in both arterial and venous thrombosis and identify key molecular events that serve as potential targets for therapeutics against diverse causes of immunothrombosis.
Asunto(s)
Síndrome Antifosfolípido , Trombosis , Trombosis de la Vena , Anticuerpos Antifosfolípidos , Humanos , Neutrófilos/metabolismo , Trombosis/etiologíaRESUMEN
The thrombotic microangiopathies (TMAs) are a heterogenous group of disorders with distinct pathophysiologies that cause occlusive microvascular or macrovascular thrombosis, and are characterized by microangiopathic hemolytic anemia, thrombocytopenia, and/or end-organ ischemia. TMAs are associated with significant morbidity and mortality, and data on the management of certain TMAs are often lacking. The nomenclature, classification, and management of various TMAs is constantly evolving as we learn more about these rare syndromes. Thorough clinical and laboratory evaluation is essential to distinguish various TMAs and arrive at an accurate diagnosis, which is key for appropriate management. In this illustrated review, we focus on thrombotic thrombocytopenic purpura (TTP), Shiga toxin-associated hemolytic uremic syndrome, complement-mediated hemolytic uremic syndrome, hematopoietic cell transplant-associated TMA, and drug-induced TMA, and describe their incidence, pathophysiology, diagnosis, and management. We also highlight emerging complement-directed therapies under investigation for the management of complement-mediated TMAs.
RESUMEN
It is widely recognized that inflammation plays a critical role in cardiac hypertrophy and heart failure. However, clinical trials targeting cytokines have shown equivocal effects, indicating the need for a deeper understanding of the precise role of inflammation and inflammatory cells in heart failure. Leukocytes from human subjects and a rodent model of heart failure were characterized by a marked reduction in expression of Klf2 mRNA. Using a mouse model of angiotensin II-induced nonischemic cardiac dysfunction, we showed that neutrophils played an essential role in the pathogenesis and progression of heart failure. Mechanistically, chronic angiotensin II infusion activated a neutrophil KLF2/NETosis pathway that triggered sporadic thrombosis in small myocardial vessels, leading to myocardial hypoxia, cell death, and hypertrophy. Conversely, targeting neutrophils, neutrophil extracellular traps (NETs), or thrombosis ameliorated these pathological changes and preserved cardiac dysfunction. KLF2 regulated neutrophil activation in response to angiotensin II at the molecular level, partly through crosstalk with HIF1 signaling. Taken together, our data implicate neutrophil-mediated immunothrombotic dysregulation as a critical pathogenic mechanism leading to cardiac hypertrophy and heart failure. This neutrophil KLF2-NETosis-thrombosis mechanism underlying chronic heart failure can be exploited for therapeutic gain by therapies targeting neutrophils, NETosis, or thrombosis.
Asunto(s)
Cardiomegalia/metabolismo , Insuficiencia Cardíaca/metabolismo , Factores de Transcripción de Tipo Kruppel/metabolismo , Activación Neutrófila , Neutrófilos/metabolismo , Trombosis/metabolismo , Animales , Modelos Animales de Enfermedad , Humanos , RatonesRESUMEN
Venous thromboembolism (VTE) in individuals with sickle cell disease is common and portends a poor prognosis. The role of leukocyte count and its subsets on risk of VTE in sickle cell disease are not known. We conducted a retrospective case-control study and analyzed for leukocyte count at the time of VTE and 3 months prior. Leukocyte and neutrophil counts were elevated at the time of VTE (p = 0.003 and p = 0.0006, respectively) and 3 months prior (p = 0.001 and p = 0.0096, respectively) when compared to controls. Baseline leukocytosis and neutrophilia may be associated with subsequent risk for thrombosis in sickle cell disease.
Asunto(s)
Anemia de Células Falciformes , Tromboembolia Venosa , Anemia de Células Falciformes/complicaciones , Estudios de Casos y Controles , Humanos , Leucocitosis , Estudios Retrospectivos , Factores de Riesgo , Tromboembolia Venosa/etiologíaRESUMEN
OBJECTIVE: To determine whether mild or moderate thrombocytopenia is associated with postoperative complications after benign hysterectomy. METHODS: A retrospective study of data from women who underwent benign hysterectomy included in the American College of Surgeons National Surgical Quality Improvement Project Database. The data were stratified by normal platelet count, mild thrombocytopenia (100-149 × 103 platelets/µl), and moderate thrombocytopenia (50-99 × 103 platelets/µl). Multivariable logistic regression was used to determine the relationship between mild or moderate thrombocytopenia and the main outcome measures. RESULTS: Moderate thrombocytopenia was associated with an increased risk of perioperative transfusion (adjusted odds ratio [aOR], 2.87; 95% confidence interval [CI], 1.96-4.21) and reoperation (aOR, 4.03; 95% CI, 1.94-17.33), but mild thrombocytopenia was not. There was an increased risk of infection among women with both mild (aOR, 1.38; 95% CI, 1.12-1.69) and moderate (aOR, 2.00; 95% CI,1.23-3.22) thrombocytopenia. There was no association between either mild or moderate thrombocytopenia and readmission, prolonged hospital stay, or longer surgical time. CONCLUSION: Thrombocytopenia was found to be associated with increased infectious morbidity after hysterectomy, and moderate thrombocytopenia was associated with an increased risk of perioperative transfusion and reoperation.
Asunto(s)
Histerectomía/efectos adversos , Complicaciones Posoperatorias/epidemiología , Trombocitopenia/epidemiología , Adulto , Transfusión Sanguínea , Estudios de Cohortes , Femenino , Humanos , Tiempo de Internación , Persona de Mediana Edad , Tempo Operativo , Complicaciones Posoperatorias/etiología , Reoperación , Estudios Retrospectivos , Trombocitopenia/etiologíaRESUMEN
Substantial evidence implicates crosstalk between metabolic tissues and the immune system in the inception and progression of obesity. However, molecular regulators that orchestrate metaflammation both centrally and peripherally remains incompletely understood. Here, we identify myeloid Krüppel-like factor 2 (KLF2) as an essential regulator of obesity and its sequelae. In mice and humans, consumption of a fatty diet downregulates myeloid KLF2 levels. Under basal conditions, myeloid-specific KLF2 knockout mice (K2KO) exhibit increased feeding and weight gain. High-fat diet (HFD) feeding further exacerbates the K2KO metabolic disease phenotype. Mechanistically, loss of myeloid KLF2 increases metaflammation in peripheral and central tissues. A combination of pair-feeding, bone marrow-transplant, and microglial ablation implicate central and peripheral contributions to K2KO-induced metabolic dysfunction observed. Finally, overexpression of myeloid KLF2 protects mice from HFD-induced obesity and insulin resistance. Together, these data establish myeloid KLF2 as a nodal regulator of central and peripheral metabolic inflammation in homeostasis and disease.
Asunto(s)
Factores de Transcripción de Tipo Kruppel/inmunología , Enfermedades Metabólicas/inmunología , Células Mieloides/inmunología , Obesidad/inmunología , Animales , Sistema Nervioso Central/inmunología , Dieta Alta en Grasa/efectos adversos , Ingestión de Alimentos , Humanos , Inflamación , Resistencia a la Insulina , Factores de Transcripción de Tipo Kruppel/genética , Masculino , Enfermedades Metabólicas/genética , Enfermedades Metabólicas/metabolismo , Enfermedades Metabólicas/fisiopatología , Ratones , Ratones Noqueados , Obesidad/etiología , Obesidad/genética , Obesidad/fisiopatología , Sistema Nervioso Periférico/inmunologíaRESUMEN
Inflammation is a vital physiological response of the immune system meant to protect against the invasion of pathogens. However, accumulating evidence describes an intimate link between inflammation and thrombosis and cellular elements of the immune system of the immune system such as neutrophils and monocytes/macrophages are emerging as key players in the generation of a prothrombotic milieu suggesting that anti-inflammatory therapy may have a role in the management of thrombosis that is driven by inflammation. Tongji 2 (TJ2) is a traditional Chinese medication manufactured as granules by Tongji hospital of Tongji University (Shanghai, China) with known anti-inflammatory properties. In this study, we examine the effects of TJ2 on inflammation and thrombosis. Our study shows that TJ2 modulates NF-κB activation and thus generates a prominent anti-inflammatory effect. Further, we use mouse models of thrombosis to demonstrate that TJ2 has a beneficial effect in both arterial and venous thrombosis that occurs in the absence of alterations in platelet activation or coagulation.
Asunto(s)
Antiinflamatorios/farmacología , Medicamentos Herbarios Chinos/farmacología , Células Endoteliales/efectos de los fármacos , Fibrinolíticos/farmacología , FN-kappa B/metabolismo , Trombosis de la Vena/tratamiento farmacológico , Animales , Antiinflamatorios/uso terapéutico , Células Cultivadas , Medicamentos Herbarios Chinos/uso terapéutico , Células Endoteliales/metabolismo , Endotelio Vascular/citología , Fibrinolíticos/uso terapéutico , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Células RAW 264.7RESUMEN
Background: Immune thrombotic thrombocytopenic purpura (iTTP) is a rare, life-threatening disorder managed with plasma exchange (PLEX) and steroids. Addition of rituximab (RTX) to initial disease treatment has been shown to lower future relapse rates. Information as to whether upfront cyclophosphamide (CTX) treatment is helpful in reducing relapse is not known. Methods: In a retrospective cohort study, we identified all patients at our institution diagnosed with iTTP between 2010 and 2019. We analyzed outcomes of cumulative incidence of relapse (CIR) and duration of remission. Results: Thirty Nine patients were studied. Group A (n = 10) included patients who received upfront PLEX and steroids alone, and Group B (n = 28) included those who received either upfront RTX (n = 23) or CTX (n = 5) in addition to PLEX and steroids. The 2-year CIR was 50% in Group A and 27.7% in Group B, with a median duration of remission of 43.6 months vs. 108.3 months, respectively (p = 0.04). Group A was associated with a HR=8.7 (95% CI: 1.27, 59.45), p = 0.027 for duration of remission. There was no significant difference between CTX and RTX in both outcomes of CIR and duration of remission. We observed a potential impact on remission duration based on the presenting absolute neutrophil count (HR = 0.74, 95% CI: 0.58, 0.96) and serum creatinine (HR = 1.42, 95% CI: 1.03, 1.94). Conclusion: There was no significant difference in iTTP relapse outcomes between upfront RTX and CTX. Absolute neutrophil count and serum creatinine may have a role in predicting relapse. Larger, prospective studies are needed to evaluate these findings.
RESUMEN
The 2019 coronavirus disease (COVID-19) presents with a large variety of clinical manifestations ranging from asymptomatic carrier state to severe respiratory distress, multiple organ dysfunction and death. While it was initially considered primarily a respiratory illness, rapidly accumulating data suggests that COVID-19 results in a unique, profoundly prothrombotic milieu leading to both arterial and venous thrombosis. Consistently, elevated D-dimer level has emerged as an independent risk factor for poor outcomes, including death. Several other laboratory markers and blood counts have also been associated with poor prognosis, possibly due to their connection to thrombosis. At present, the pathophysiology underlying the hypercoagulable state is poorly understood. However, a growing body of data suggests that the initial events occur in the lung. A severe inflammatory response, originating in the alveoli, triggers a dysfunctional cascade of inflammatory thrombosis in the pulmonary vasculature, leading to a state of local coagulopathy. This is followed, in patients with more severe disease, by a generalized hypercoagulable state that results in macro- and microvascular thrombosis. Of concern, is the observation that anticoagulation may be inadequate in many circumstances, highlighting the need for alternative or additional therapies. Numerous ongoing studies investigating the pathophysiology of the COVID-19 associated coagulopathy may provide mechanistic insights that can direct appropriate interventional strategies.
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Coagulación Sanguínea , Tratamiento Farmacológico de COVID-19 , COVID-19 , Inflamación/tratamiento farmacológico , Trombofilia , Trombosis , Tromboembolia Venosa , Animales , Antiinflamatorios/uso terapéutico , Anticoagulantes/uso terapéutico , Coagulación Sanguínea/efectos de los fármacos , COVID-19/sangre , COVID-19/epidemiología , COVID-19/fisiopatología , Humanos , Incidencia , Inflamación/sangre , Inflamación/epidemiología , Inflamación/fisiopatología , Trombofilia/sangre , Trombofilia/epidemiología , Trombofilia/fisiopatología , Trombofilia/prevención & control , Trombosis/sangre , Trombosis/epidemiología , Trombosis/fisiopatología , Trombosis/terapia , Tromboembolia Venosa/sangre , Tromboembolia Venosa/epidemiología , Tromboembolia Venosa/fisiopatología , Tromboembolia Venosa/terapiaRESUMEN
Estrogen, in the clinical setting is used primarily for contraception and hormone replacement therapy. It has been well established that estrogen increases the risk of both arterial and venous thrombosis. While estrogen is known to induce a prothrombotic milieu through various effects on the hemostatic pathways, the exact molecular mechanism leading to those effects is not known. The most common clinical presentation of estrogen-related thrombosis is venous thromboembolism (VTE) of the deep veins of the legs or pulmonary vessels, usually within the first few months of use. Estrogen has also been associated with increased risk of "unusual site" thromboses, as well as arterial thrombosis. Women at high-risk of thrombosis need careful evaluation and counseling for contraception, pregnancy, menopausal hormonal therapy and other estrogen-related conditions or treatments in order to lower the risk of thromboses. We review the most recent evidence on management of high-estrogen states in women at high-risk of thrombosis, as well as emerging data on unique populations such as transgender women. More studies are needed to better understand the pathophysiology of hormone-related thrombosis, as well as more comprehensive techniques to stratify risks for thrombosis so as to enable tailoring of recommendations for each individual.
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Trombosis , Tromboembolia Venosa , Trombosis de la Vena , Anticoncepción , Terapia de Reemplazo de Estrógeno/efectos adversos , Estrógenos/efectos adversos , Femenino , Humanos , Factores de Riesgo , Tromboembolia Venosa/inducido químicamenteRESUMEN
BACKGROUND: Antibodies against factor VIII (FVIII), seen in acquired (AHA) and congenital haemophilia A, lead to severe bleeding diatheses. Current first-line treatment includes bypass agents. Recombinant porcine sequence FVIII (rpFVIII) was developed as an alternative therapy. AIM: To describe our institutional experience with the use of rpFVIII. METHODS: A retrospective chart review of five patients treated with rpVIII between 2016 and 2019. RESULTS: Five patients (four AHA, one congenital haemophilia with inhibitors) were treated with rpFVIII. No patient had an adverse event during infusion. All patients initially exhibited a response evidenced by increased FVIII levels from baseline <1% to 81%-170%, normalization of the activated partial thromboplastin time (aPTT) and resolution of bleeding. However, all five patients were subsequently noted to have decreasing peak FVIII levels and aPTT prolongation, either within the initial treatment course or upon later re-administration. Resistance to rpFVIII was recognized after an average of 12.4 exposure days. Porcine FVIII inhibitor levels measured afterwards were present (detectable-170 Bethesda units) in all patients. Three out of four AHA subjects also developed an increase in the anti-human FVIII inhibitor titres after receiving rpFVIII. CONCLUSION: rpFVIII was safe and initially effective in all patients. However, its use is associated with development of an inhibitor to rpFVIII, decreasing its efficacy and duration of effect. Further, rpFVIII use may lead to an increase in patient anti-human FVIII inhibitor titres. A larger study is necessary to appropriately assess the incidence of these outcomes.
Asunto(s)
Factor VIII/antagonistas & inhibidores , Hemofilia A/terapia , Proteínas Recombinantes/farmacología , Adulto , Anciano , Animales , Autoanticuerpos/inmunología , Pruebas de Coagulación Sanguínea/métodos , Factor VIII/química , Factor VIII/inmunología , Femenino , Hemofilia A/genética , Hemofilia A/inmunología , Hemorragia/prevención & control , Humanos , Infusiones Intravenosas , Masculino , Proteínas Recombinantes/administración & dosificación , Proteínas Recombinantes/uso terapéutico , Estudios Retrospectivos , Seguridad , Porcinos , Resultado del TratamientoRESUMEN
Upregulated expression of P-selectin on activated endothelium and platelets significantly contributes to the initiation and progression of vaso-occlusive crises (VOC), a major cause of morbidity in sickle cell disease (SCD). Crizanlizumab (ADAKVEO®), a humanized monoclonal antibody against P-selectin, primarily inhibits the interaction between leukocytes and P-selectin, and has been shown to decrease the frequency of VOCs in clinical trials. However, the lack of reliable in vitro assays that objectively measure leukocyte adhesion to P-selectin remains a critical barrier to evaluating and improving the therapeutic treatment in SCD. Here, we present a standardized microfluidic BioChip whole blood adhesion assay to assess leukocyte adhesion to P-selectin under physiologic flow conditions. Our results demonstrated heterogeneous adhesion by leukocytes to immobilized P-selectin, and dose-dependent inhibition of this adhesion following pre-exposure to Crizanlizumab. Importantly, treatment with Crizanlizumab following adhesion to P-selectin promoted detachment of rolling, but not of firmly adherent leukocytes. Taken together, our results suggest that the microfluidic BioChip system is a promising in vitro assay with which to screen patients, monitor treatment response, and guide current and emerging anti-adhesive therapies in SCD.