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1.
Transfusion ; 2024 Jul 09.
Artículo en Inglés | MEDLINE | ID: mdl-38979976

RESUMEN

BACKGROUND: Neutrophils in sickle cell disease (SCD) are activated, contributing to disease. Red cell exchange (RCE), with the goal of lowering hemoglobin S (HbS), is an important part of therapy for many SCD patients. Whether RCE impacts neutrophil reactivity is unknown. STUDY DESIGN AND METHODS: To determine the effect of RCE on neutrophil activation, SCD patients undergoing RCE in steady-state were enrolled. Neutrophil degranulation responses were examined before/after RCE. Kinetic studies were completed to determine the duration of the effect of RCE on neutrophil function. Degranulation results were examined in relation to white blood cell count, neutrophil count, and HbS levels. The effect of RCE on RBC phosphatidylserine (PS) exposure was examined as a possible contributor to modulation of neutrophil function by RCE. RESULTS: Twenty-two patients with SCD, genotype SS, who underwent RCE (average pre-RCE HbS 33 ± 14%) were included for the study. RCE significantly decreased neutrophil degranulation responses. The effect of RCE on neutrophil activation was unrelated to cell count and instead directly correlated with HbS. The effect of RCE on neutrophil activation was sustained over several days post-apheresis. Furthermore, while increased RBC PS exposure results in increased neutrophil degranulation, RCE decreases RBC PS exposure. DISCUSSION: To our knowledge, this is the first study demonstrating that RCE significantly decreases neutrophil activation in a sustained HbS-dependent manner. Modulation of PS exposure by RCE may be a contributing mechanism by which RCE modulates neutrophil activation. These studies raise the possibility that modulation of neutrophil activation contributes significantly to the therapeutic effect of RCE.

3.
Blood Adv ; 3(17): 2586-2597, 2019 09 10.
Artículo en Inglés | MEDLINE | ID: mdl-31484636

RESUMEN

Sickle red blood cells (SSRBCs) are adherent to the endothelium, activate leukocyte adhesion, and are deficient in bioactive nitric oxide (NO) adducts such as S-nitrosothiols (SNOs), with reduced ability to induce vasodilation in response to hypoxia. All these pathophysiologic characteristics promote vascular occlusion, the hallmark of sickle cell disease (SCD). Loading hypoxic SSRBCs in vitro with NO followed by reoxygenation significantly decreased epinephrine-activated SSRBC adhesion to the endothelium, the ability of activated SSRBCs to mediate leukocyte adhesion in vitro, and vessel obstruction in vivo. Because transfusion is frequently used in SCD, we also determined the effects of banked (SNO-depleted) red blood cells (RBCs) on vaso-occlusion in vivo. Fresh or 14-day-old normal RBCs (AARBCs) reduced epinephrine-activated SSRBC adhesion to the vascular endothelium and prevented vaso-occlusion. In contrast, AARBCs stored for 30 days failed to decrease activated SSRBC adhesivity or vaso-occlusion, unless these RBCs were loaded with NO. Furthermore, NO loading of SSRBCs increased S-nitrosohemoglobin and modulated epinephrine's effect by upregulating phosphorylation of membrane proteins, including pyruvate kinase, E3 ubiquitin ligase, and the cytoskeletal protein 4.1. Thus, abnormal SSRBC NO/SNO content both contributes to the vaso-occlusive pathophysiology of SCD, potentially by affecting at least protein phosphorylation, and is potentially amenable to correction by (S)NO repletion or by RBC transfusion.


Asunto(s)
Anemia de Células Falciformes/complicaciones , Adhesión Celular/efectos de los fármacos , Eritrocitos/patología , Óxido Nítrico/farmacología , Enfermedades Vasculares/prevención & control , Anemia de Células Falciformes/etiología , Anemia de Células Falciformes/fisiopatología , Endotelio Vascular/metabolismo , Transfusión de Eritrocitos , Eritrocitos/metabolismo , Hemoglobinas/metabolismo , Humanos , Proteínas de la Membrana/metabolismo , Óxido Nítrico/uso terapéutico , Oxígeno/farmacología , Fosforilación/efectos de los fármacos , Enfermedades Vasculares/etiología
4.
Br J Haematol ; 175(5): 935-948, 2016 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-27549988

RESUMEN

Sevuparin is a novel drug candidate in phase II development as a treatment for vaso-occlusive crises (VOC) in patients with sickle cell disease (SCD). As a heparin-derived polysaccharide, sevuparin has been designed to retain anti-adhesive properties, while the antithrombin-binding domains have been eliminated, substantially diminishing its anticoagulant activity. Here, we demonstrate that sevuparin inhibits the adhesion of human sickle red blood cells (SS-RBCs) to stimulated cultured endothelial cells in vitro. Importantly, sevuparin prevents vaso-occlusion and normalizes blood flow in an in vivo mouse model of SCD vaso-occlusion. Analyses by surface plasmon resonance (SPR) and fluorescence correlation spectroscopy (FCS) demonstrate that sevuparin binds to P- and L-selectins, thrombospondin, fibronectin and von Willebrand factor, all of which are thought to contribute to vaso-occlusion in SCD. Despite low anticoagulation activity, sevuparin has anti-adhesive efficacy similar to the low molecular weight heparin tinzaparin both in vitro and in vivo. These results suggest that the anti-adhesive properties rather than the anticoagulant effects of heparinoids are critical for the treatment of vaso-occlusion in SCD. Therefore, sevuparin is now being evaluated in SCD patients hospitalized for treatment of VOC.


Asunto(s)
Anemia de Células Falciformes/tratamiento farmacológico , Heparina de Bajo-Peso-Molecular/farmacología , Heparina/análogos & derivados , Anemia de Células Falciformes/complicaciones , Anemia de Células Falciformes/patología , Animales , Arteriopatías Oclusivas/tratamiento farmacológico , Adhesión Celular/efectos de los fármacos , Células Cultivadas , Células Endoteliales/metabolismo , Eritrocitos/metabolismo , Heparina/farmacología , Heparina/uso terapéutico , Heparina de Bajo-Peso-Molecular/uso terapéutico , Humanos , Ratones , Unión Proteica , Tinzaparina
5.
Clin Transl Sci ; 5(6): 437-44, 2012 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-23253664

RESUMEN

Sickle red blood cells (SSRBCs) adhere to both endothelial cells (ECs) and the extracellular matrix. Epinephrine elevates cyclic adenosine monophosphate in SSRBCs and increases adhesion of SSRBCs to ECs in a ß-adrenergic receptor and protein kinase A-dependent manner. Studies in vitro as well as in vivo have suggested that adrenergic stimuli like epinephrine may contribute to vaso-occlusion associated with physiologic stress. We conducted both animal studies and a Phase I dose-escalation study in sickle cell disease (SCD) patients to investigate whether systemically administered propranolol inhibits SSRBC adhesion and to document the safety of propranolol in SCD. Systemically administered propranolol prevented SSRBC adhesion and associated vaso-occlusion in a mouse model. In patients receiving a single oral dose of 10, 20, or 40 mg propranolol, SSRBC adhesion to ECs was studied before and after propranolol, with and without stimulation with epinephrine. Propranolol administration significantly reduced epinephrine-stimulated SSRBC adhesion in a dose dependent manner (p = 0.03), with maximum inhibition achieved at 40 mg. Adverse events were not severe, did not show dose dependence, and were likely unrelated to drug. No significant heart rate changes occurred. These results imply that ß-blockers may have a role as antiadhesive therapy for SCD.


Asunto(s)
Anemia de Células Falciformes/tratamiento farmacológico , Anemia de Células Falciformes/patología , Antidrepanocíticos/uso terapéutico , Propranolol/uso terapéutico , Administración Oral , Adulto , Anemia de Células Falciformes/fisiopatología , Animales , Antidrepanocíticos/farmacología , Presión Sanguínea/efectos de los fármacos , Vasos Sanguíneos/efectos de los fármacos , Vasos Sanguíneos/patología , Adhesión Celular/efectos de los fármacos , Células Endoteliales/efectos de los fármacos , Células Endoteliales/patología , Epinefrina/farmacología , Epinefrina/uso terapéutico , Eritrocitos/efectos de los fármacos , Eritrocitos/patología , Femenino , Frecuencia Cardíaca/efectos de los fármacos , Humanos , Inyecciones Intravenosas , Ratones , Ratones Desnudos , Propranolol/efectos adversos , Propranolol/farmacología
6.
Nucleic Acid Ther ; 21(4): 275-83, 2011 Aug.
Artículo en Inglés | MEDLINE | ID: mdl-21793788

RESUMEN

Patients with sickle cell disease (SCD) often suffer painful vaso-occlusive episodes caused in part by the adhesion of sickle erythrocytes (SS-RBC) to the vascular endothelium. To investigate inhibition of SS-RBC adhesion as a possible treatment for vaso-occlusion, 2 adhesion molecules, α(v)ß(3) and P-selectin, were targeted by high-affinity RNA aptamers. An in vitro flow chamber assay was used to test the antiadhesion activity of α(v)ß(3) aptamer clone 17.16. Human SS-RBC were passed across a confluent monolayer of thrombin-stimulated human umbilical vein endothelial cells (HUVEC) at a constant rate. α(v)ß(3) aptamer reduced SS-RBC adhesion to activated endothelial cells to the level seen with untreated HUVEC. An aptamer reactive with complement component 8 was used as a negative control and exerted no inhibition, confirming the specificity of α(v)ß(3) aptamer (P=0.04). At 2 dyn/cm(2) shear stress, 30 nM α(v)ß(3) aptamer showed maximal effect in decreasing SS-RBC adhesion to HUVEC. The antiadhesive activity of the P-selectin aptamer clone PF377 was also tested using HUVEC pretreated with IL-13 to upregulate expression of P-selectin as seen in activated endothelial cells. At 1 dyn/cm(2) shear stress, 60 nM of P-selectin aptamer had antiadhesion activity similar to heparin, a known inhibitor of SS-RBC adhesion to P-selectin. A negative control did not prevent adhesion (P=0.05). These data show the potential utility of aptamers to block endothelial adhesion molecules to prevent or treat vaso-occlusion in SCD.


Asunto(s)
Anemia de Células Falciformes/tratamiento farmacológico , Antidrepanocíticos/farmacología , Aptámeros de Nucleótidos/farmacología , Anemia de Células Falciformes/patología , Antidrepanocíticos/uso terapéutico , Aptámeros de Nucleótidos/uso terapéutico , Adhesión Celular/efectos de los fármacos , Células Cultivadas , Eritrocitos/efectos de los fármacos , Eritrocitos/patología , Células Endoteliales de la Vena Umbilical Humana/efectos de los fármacos , Células Endoteliales de la Vena Umbilical Humana/metabolismo , Humanos , Integrina alfaVbeta3/antagonistas & inhibidores , Integrina alfaVbeta3/metabolismo , Selectina-P/metabolismo
7.
Blood ; 112(8): 3474-83, 2008 Oct 15.
Artículo en Inglés | MEDLINE | ID: mdl-18664622

RESUMEN

Infusion of epinephrine-activated human sickle erythrocytes (SS RBCs) into nude mice promotes both SS RBC and murine leukocyte adhesion to vascular endothelium in vivo. We hypothesized that interaction of epinephrine-stimulated SS RBCs with leukocytes leads to activation of leukocytes, which then adhere to endothelial cells (ECs). In exploring the underlying molecular mechanisms, we have found that coincubation in vitro of epinephrine-treated SS RBCs with human peripheral blood mononuclear cells (PBMCs) results in robust adhesion of PBMCs to ECs. Sham-treated SS RBCs had a similar but less pronounced effect, whereas neither sham- nor epinephrine-treated normal RBCs activated PBMC adhesion. PBMC activation was induced via at least 2 RBC adhesion receptors, LW and CD44. In response to SS RBCs, leukocyte CD44 and beta2 integrins mediated PBMC adhesion to ECs, a process that involved endothelial E-selectin and fibronectin. SS RBCs activated adhesion of both PBMC populations, lymphocytes and monocytes. Thus, our findings reveal a novel mechanism that may contribute to the pathogenesis of vaso-occlusion in sickle cell disease, in which SS RBCs act via LW and CD44 to stimulate leukocyte adhesion to endothelium, and suggest that RBC LW and CD44 may serve as potential targets for antiadhesive therapy designed to prevent vaso-occlusion.


Asunto(s)
Anemia de Células Falciformes/sangre , Endotelio Vascular/metabolismo , Eritrocitos Anormales/citología , Linfocitos/citología , Monocitos/citología , Anemia de Células Falciformes/metabolismo , Animales , Antígenos CD18/metabolismo , Adhesión Celular , Epinefrina/farmacología , Humanos , Receptores de Hialuranos/biosíntesis , Leucocitos Mononucleares/citología , Ratones , Ratones Desnudos , Venas Umbilicales/citología
8.
Blood ; 110(7): 2708-17, 2007 Oct 01.
Artículo en Inglés | MEDLINE | ID: mdl-17609430

RESUMEN

Sickle red cell (SS RBC) adhesion is believed to contribute to the process of vaso-occlusion in sickle cell disease (SCD). We previously found that the LW RBC adhesion receptor can be activated by epinephrine to mediate SS RBC adhesion to endothelial alphavbeta3 integrin. To determine the contribution of LW activation to vaso-occlusive events in vivo, we investigated whether in vitro treatment of SS RBCs by epinephrine resulted in vaso-occlusion in intact microvasculature after RBC infusion into nude mice. Epinephrine enhanced human SS but not normal RBC adhesion to murine endothelial cells in vitro and to endothelium in vivo, promoting vaso-occlusion and RBC organ sequestration. Murine sickle RBCs also responded to epinephrine with increased adhesion to postcapillary endothelium in nude mice. Epinephrine-induced SS RBC adhesion, vaso-occlusion, and RBC organ trapping could be prevented by the beta-adrenergic receptor (beta-AR) antagonist, propranolol. Infusion of soluble recombinant LW also significantly reduced adhesion and vaso-occlusion. In addition, epinephrine-treated SS RBCs induced activation of murine leukocyte adhesion to endothelium as well. We conclude that LW activation by epinephrine via beta-AR stimulation can promote both SS RBC and leukocyte adhesion as well as vaso-occlusion, suggesting that both epinephrine and LW play potentially pathophysiological roles in SCD.


Asunto(s)
Anemia de Células Falciformes/patología , Moléculas de Adhesión Celular/metabolismo , Epinefrina/farmacología , Anemia de Células Falciformes/metabolismo , Animales , Adhesión Celular/efectos de los fármacos , Supervivencia Celular , Células Cultivadas , Endotelio/irrigación sanguínea , Endotelio/efectos de los fármacos , Endotelio/patología , Eritrocitos/citología , Eritrocitos/efectos de los fármacos , Humanos , Leucocitos/citología , Leucocitos/efectos de los fármacos , Leucocitos/inmunología , Ratones , Ratones Desnudos , Receptores Adrenérgicos beta 2/metabolismo
9.
Am J Hematol ; 75(2): 63-72, 2004 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-14755370

RESUMEN

Red blood cells from patients with sickle cell disease (SS RBC) adhere to laminin and over-express the high-affinity laminin receptor basal cell adhesion molecule/Lutheran protein (B-CAM/LU). This receptor has recently been shown to undergo activation in vitro through a protein kinase A-dependent mechanism. Low-density SS RBC express two-thirds more B-CAM/LU than high-density SS RBC. However, high-density SS RBC have been identified as most adherent to laminin under flow conditions. We investigated the ability of low- and high-density SS RBC to interact with laminin under various conditions and explored factors that might be responsible for the differences in B-CAM/LU-laminin interaction between high- and low-density SS RBC. We confirmed that high-density SS RBC adhere to laminin more strongly than low-density SS RBC under flow conditions. However, low-density SS RBC bind soluble laminin most strongly and are the most adherent to laminin under static conditions. Soluble recombinant Lutheran extracellular domain protein completely blocked SS RBC adhesion to laminin under both static and flow conditions. The protein kinase A inhibitor 14-22 amide inhibited adhesion to laminin during flow by high-density SS RBC from patients with strongly adherent cells but had no effect on adhesion observed after a static phase. Deletion of the cytoplasmic domain of B-CAM as well as mutation of the juxtamembranous tyrosine residue failed to reduce B-CAM-mediated adhesion to laminin by transfected MEL cells. These studies confirm that B-CAM/LU is the most critical receptor mediating adhesion to laminin under both static and flow conditions. Dense SS RBC are most adherent to laminin despite bearing fewer laminin receptors, apparently due to a reversible protein kinase A-dependent process that is unlikely to involve direct phosphorylation of B-CAM/LU. Our results also suggest that the nature of the interaction of B-CAM/LU with laminin may be different under static and flow conditions.


Asunto(s)
Anemia de Células Falciformes/sangre , Moléculas de Adhesión Celular/sangre , Moléculas de Adhesión Celular/genética , Eritrocitos/metabolismo , Laminina/sangre , Proteínas de Neoplasias/sangre , Proteínas de Neoplasias/genética , Adulto , Secuencia de Bases , Adhesión Celular , Cartilla de ADN , Citometría de Flujo , Humanos , Sistema del Grupo Sanguíneo Lutheran , Reticulocitos/metabolismo , Reticulocitos/patología
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