Minimally modified human blood coagulation factor X to bypass direct factor Xa inhibitors.

BACKGROUND: Direct oral factor (F)Xa inhibitors are widely used as alternatives to conventional vitamin K antagonists in managing venous thromboembolism and nonvalvular atrial fibrillation. Unfortunately, bleeding-related adverse events remain a major concern in clinical practice. In case of bleeding or emergency surgery, rapid-onset reversal agents may be required to counteract the anticoagulant activity. OBJECTIVES: The ability of FXa variants to bypass the direct oral FXa inhibitors was assessed. METHODS: Human FXa variants were generated through substitution of phenylalanine 174 (F174) for either alanine, isoleucine, or serine. FXa variants were stably expressed in HEK293 cells and purified to homogeneity using ion-exchange chromatography. RESULTS: F174-substituted human FX variants demonstrated efficacy in restoring thrombin generation in plasma containing direct FXa inhibitors (apixaban, rivaroxaban, edoxaban). Their ability to bypass the anticoagulant effects stems from a significantly reduced sensitivity for the direct FXa inhibitors due to a decrease in binding affinity determined using molecular dynamics simulations and free energy computation. Furthermore, F174 modification resulted in a partial loss of inhibition by tissue factor pathway inhibitor, enhancing the procoagulant effect of F174-substituted FX. Consequently, the F174A- and F174S-substituted FX variants effectively counteracted the effects of 2 widely used anticoagulants, apixaban and rivaroxaban, in plasma of atrial fibrillation and venous thromboembolism patients. CONCLUSION: These human FX variants have the potential to serve as a rescue reversal strategy to overcome the effect of direct FXa inhibitors in case of life-threatening bleeding events or emergency surgical interventions.

Understanding the Angiogenic Characteristics of Clinical-Grade Mesenchymal Stromal Cells Isolated from Human Umbilical Cord.

Addressing the challenges in managing ischemic tissue repair and remodelling remains a prominent clinical concern. Current research is heavily concentrated on identifying innovative cell-based therapies with the potential to enhance revascularization in patients affected by these diseases. We have previously developed and validated a manufacturing process for human umbilical cord mesenchymal stromal cells (UC-MSCs)-based cell therapy medicinal product, according to Good Manufacturing Practices. In this study, we demonstrate that these UC-MSCs enhance the proliferation and migration of endothelial cells and the formation of capillary structures. Moreover, UC-MSCs and endothelial cells interact, allowing UC-MSCs to acquire a perivascular cell phenotype and consequently provide direct support to the newly formed vascular network. This characterization of the proangiogenic properties of this UC-MSCs based-cell therapy medicinal product is an essential step for its therapeutic assessment in the clinical context of vascular regeneration.

VEGF-A plasma levels are associated with impaired DLCO and radiological sequelae in long COVID patients.

BACKGROUND: Long COVID, also known as post-acute sequelae of COVID-19 (PASC), is characterized by persistent clinical symptoms following COVID-19. OBJECTIVE: To correlate biomarkers of endothelial dysfunction with persistent clinical symptoms and pulmonary function defects at distance from COVID-19. METHODS: Consecutive patients with long COVID-19 suspicion were enrolled. A panel of endothelial biomarkers was measured in each patient during clinical evaluation and pulmonary function test (PFT). RESULTS: The study included 137 PASC patients, mostly male (68%), with a median age of 55 years. A total of 194 PFTs were performed between months 3 and 24 after an episode of SARS-CoV-2 infection. We compared biomarkers evaluated in PASC patients with 20 healthy volunteers (HVs) and acute hospitalized COVID-19 patients (n = 88). The study found that angiogenesis-related biomarkers and von Willebrand factor (VWF) levels were increased in PASC patients compared to HVs without increased inflammatory or platelet activation markers. Moreover, VEGF-A and VWF were associated with persistent lung CT scan lesions and impaired diffusing capacity of the lungs for carbon monoxide (DLCO) measurement. By employing a Cox proportional hazards model adjusted for age, sex, and body mass index, we further confirmed the accuracy of VEGF-A and VWF. Following adjustment, VEGF-A emerged as the most significant predictive factor associated with persistent lung CT scan lesions and impaired DLCO measurement. CONCLUSION: VEGF-A is a relevant predictive factor for DLCO impairment and radiological sequelae in PASC. Beyond being a biomarker, we hypothesize that the persistence of angiogenic disorders may contribute to long COVID symptoms.

Combined Platelet and Red Blood Cell Recovery during On-pump Cardiac Surgery Using same™ by i-SEP Autotransfusion Device: A First-in-human Noncomparative Study (i-TRANSEP Study).

BACKGROUND: Centrifugation-based autotransfusion devices only salvage red blood cells while platelets are removed. The same™ device (Smart Autotransfusion for ME; i-SEP, France) is an innovative filtration-based autotransfusion device able to salvage both red blood cells and platelets. The authors tested the hypothesis that this new device could allow a red blood cell recovery exceeding 80% with a posttreatment hematocrit exceeding 40%, and would remove more than 90% of heparin and 75% of free hemoglobin. METHODS: Adults undergoing on-pump elective cardiac surgery were included in a noncomparative multicenter trial. The device was used intraoperatively to treat shed and residual cardiopulmonary bypass blood. The primary outcome was a composite of cell recovery performance, assessed in the device by red blood cell recovery and posttreatment hematocrit, and of biologic safety assessed in the device by the washout of heparin and free hemoglobin expressed as removal ratios. Secondary outcomes included platelet recovery and function and adverse events (clinical and device-related adverse events) up to 30 days after surgery. RESULTS: The study included 50 patients, of whom 18 (35%) underwent isolated coronary artery bypass graft, 26 (52%) valve surgery, and 6 (12%) aortic root surgery. The median red blood cell recovery per cycle was 86.1% (25th percentile to 75th percentile interquartile range, 80.8 to 91.6) with posttreatment hematocrit of 41.8% (39.7 to 44.2). Removal ratios for heparin and free hemoglobin were 98.9% (98.2 to 99.7) and 94.6% (92.7 to 96.6), respectively. No adverse device effect was reported. Median platelet recovery was 52.4% (44.2 to 60.1), with a posttreatment concentration of 116 (93 to 146) · 109/l. Platelet activation state and function, evaluated by flow cytometry, were found to be unaltered by the device. CONCLUSIONS: In this first-in-human study, the same™ device was able to simultaneously recover and wash both platelets and red blood cells. Compared with preclinical evaluations, the device achieved a higher platelet recovery of 52% with minimal platelet activation while maintaining platelet ability to be activated in vitro.

Impact of age on in vitro metabolism of clopidogrel: a potential explanation for high on-treatment platelet reactivity in the elderly?

Background: High on-treatment platelet reactivity has been reported in 30% of patients on clopidogrel and 50% in elderly patients; however, little is known about the mechanisms of this biological resistance. One hypothesis is an age-related impaired hepatic metabolism of the prodrug clopidogrel, leading to a lower formation of its active metabolite (clopidogrel-AM). Objectives: To compare the levels of clopidogrel-AM formed in vitro using "old" and "young" human liver microsomes (HLMs) and their consequences on platelet functions. Methods: We developed an in vitro model using "old" (73.6 ± 2.3 years) and "young" (51.2 ± 8.5 years) HLMs, added to platelet-rich plasma from 21 healthy donors with or without clopidogrel (50 µM) and incubated at 37 °C for 30 (T30) and 45 minutes (T45). Clopidogrel-AM was quantified by liquid chromatography-mass spectrometry/mass spectrometry method. Platelet aggregation was performed by light transmission aggregometry. Results: The generation of clopidogrel-AM increased over time and reached concentrations comparable with those reported in treated patients. At T30, mean clopidogrel-AM concentrations were significantly higher with "young" (8.56 µg/L; 95% CI, 5.87-11.24) than with "old" HLMs (7.64 µg/L; 95% CI, 5.14-10.14; P = .002); and at T45, 11.40 µg/L; 95% CI (7.57-15.22) vs 10.63 µg/L, 95% CI (7.10-14.15), P = .02 (n = 21). Despite a significant inhibition of platelet aggregation, no significant difference was found in light transmission aggregometry (adenosine diphosphate, 10 µM) after clopidogrel metabolism by "old" or "young" HLMs, probably because of low sensitivity of the method to small variations of clopidogrel-AM. Conclusion: In this original model combining metabolic and functional approaches, less clopidogrel-AM was produced with HLMs from older patients. This provides support for a decreased CYP450 activity that may contribute to high on-treatment platelet reactivity in elderly patients.

Assessment of a Quantra-Guided Hemostatic Algorithm in High-Bleeding-Risk Cardiac Surgery.

OBJECTIVES: To assess whether a Quantra-guided hemostatic algorithm would reduce transfusion requirement and major bleeding compared with laboratory-guided testing in patients facing high-bleeding-risk cardiac surgery. DESIGN: Single-center before-and-after study. SETTING: University hospital. PARTICIPANTS: Patients facing high-bleeding-risk cardiac surgery with cardiopulmonary bypass. INTERVENTIONS: Hemostatic algorithm was based on standard laboratory testing during the control period, then on the Quantra during the Quantra period. The primary endpoint was the number of red blood cell (RBC) units transfused on day 1 after surgery. MEASUREMENTS AND MAIN RESULTS: After propensity-score matching, 66 patients were included in the Quantra group and 117 in the control group. The Quantra group received fewer RBC units on day 1 than the control group (2 [0-5] v 4 [2-6], p = 0.016, respectively). Intraoperatively, the Quantra group received fewer RBC (2 [0-3] v 3 [1-5], p = 0.005), less fresh frozen plasma (0 [0-3] v 3[2-5], p < 0.0001), and fewer platelet units (7.5 [0-10] v 8.2 [6.3-11.7], p = 0.014). The intraoperative rates of RBC, plasma, and platelet transfusion were reduced (64% v 78%, p = 0.05; 41% v 85%, p < 0.001; 55% v 82%, p = 0.001, respectively). The RBC and plasma transfusions were reduced on days 1, 2, and 7. The incidence of major bleeding on day 1 also was reduced (36% v 56%, p = 0.014). In multivariate analysis, implementation of the Quantra-guided hemostatic algorithm was associated independently with reductions in major bleeding. CONCLUSION: Implementation of a Quantra-based hemostatic algorithm was associated with a decrease in transfusion requirement and major bleeding after high-bleeding-risk cardiac surgery. Randomized trials are needed to confirm these results.

Assessment of DOAC in GEriatrics (Adage Study): Rivaroxaban/Apixaban Concentrations and Thrombin Generation Profiles in NVAF Very Elderly Patients.

BACKGROUND: Although a growing number of very elderly patients with atrial fibrillation (AF), multiple conditions, and polypharmacy receive direct oral anticoagulants (DOACs), few studies specifically investigated both apixaban/rivaroxaban pharmacokinetics and pharmacodynamics in such patients. AIMS: To investigate: (1) DOAC concentration-time profiles; (2) thrombin generation (TG); and (3) clinical outcomes 6 months after inclusion in very elderly AF in-patients receiving rivaroxaban or apixaban. METHODS: Adage-NCT02464488 was an academic prospective exploratory multicenter study, enrolling AF in-patients aged ≥80 years, receiving DOAC for at least 4 days. Each patient had one to five blood samples at different time points over 20 days. DOAC concentrations were determined using chromogenic assays. TG was investigated using ST-Genesia (STG-ThromboScreen, STG-DrugScreen). RESULTS: We included 215 patients (women 71.1%, mean age: 87 ± 4 years), 104 rivaroxaban and 111 apixaban, and 79.5% receiving reduced-dose regimen. We observed important inter-individual variabilities (coefficient of variation) whatever the regimen, at C max [49-46%] and C min [75-61%] in 15 mg rivaroxaban and 2.5 mg apixaban patients, respectively. The dose regimen was associated with C max and C min plasma concentrations in apixaban (p = 0.0058 and p = 0.0222, respectively), but not in rivaroxaban samples (multivariate analysis). Moreover, substantial variability of thrombin peak height (STG-ThromboScreen) was noticed at a given plasma concentration for both xabans, suggesting an impact of the underlying coagulation status on TG in elderly in-patients. After 6-month follow-up, major bleeding/thromboembolic event/death rates were 6.7%/1.0%/17.3% in rivaroxaban and 5.4%/3.6%/18.9% in apixaban patients, respectively. CONCLUSION: Our study provides original data in very elderly patients receiving DOAC in a real-life setting, showing great inter-individual variability in plasma concentrations and TG parameters. Further research is needed to understand the potential clinical impact of these findings.

Antiplatelet Therapy for Atherothrombotic Disease in 2022-From Population to Patient-Centered Approaches.

Antiplatelet agents, with aspirin and P2Y12 receptor antagonists as major key molecules, are currently the cornerstone of pharmacological treatment of atherothrombotic events including a variety of cardio- and cerebro-vascular as well as peripheral artery diseases. Over the last decades, significant changes have been made to antiplatelet therapeutic and prophylactic strategies. The shift from a population-based approach to patient-centered precision medicine requires greater awareness of individual risks and benefits associated with the different antiplatelet strategies, so that the right patient gets the right therapy at the right time. In this review, we present the currently available antiplatelet agents, outline different management strategies, particularly in case of bleeding or in perioperative setting, and develop the concept of high on-treatment platelet reactivity and the steps toward person-centered precision medicine aiming to optimize patient care.

Induced forms of α2-macroglobulin neutralize heparin and direct oral anticoagulant effects.

Alpha2-macroglobulin (α2M) is a physiological macromolecule that facilitates the clearance of many proteinases, cytokines and growth factors in human. Here, we explored the effect of induced forms of α2M on anticoagulant drugs. Gla-domainless factor Xa (GDFXa) and methylamine (MA)-induced α2M were prepared and characterized by electrophoresis, immunonephelometry, chromogenic, clot waveform and rotational thromboelastometry assays. Samples from healthy volunteers and anticoagulated patients were included. In vivo neutralization of anticoagulants was evaluated in C57Bl/6JRj mouse bleeding-model. Anticoagulant binding sites on induced α2M were depicted by computer-aided energy minimization modeling. GDFXa-induced α2M neutralized dabigatran and heparins in plasma and whole blood. In mice, a single IV dose of GDFXa-induced α2M following anticoagulant administration significantly reduced blood loss and bleeding time. Being far easier to prepare, we investigated the efficacy of MA-induced α2M. It neutralized rivaroxaban, apixaban, dabigatran and heparins in spiked samples in a concentration-dependent manner and in samples from treated patients. Molecular docking analysis evidenced the ability of MA-induced α2M to bind non-covalently these compounds via some deeply buried binding sites. Induced forms of α2M have the potential to neutralize direct oral anticoagulants and heparins, and might be developed as a universal antidote in case of major bleeding or urgent surgery.

Role of Membrane Lipid Rafts in MRP4 (ABCC4) Dependent Regulation of the cAMP Pathway in Blood Platelets.

BACKGROUND: Platelet cytosolic cyclic adenosine monophosphate (cAMP) levels are balanced by synthesis, degradation, and efflux. Efflux can occur via multidrug resistant protein-4 (MRP4; ABCC4) present on dense granule and/or plasma membranes. As lipid rafts have been shown to interfere on cAMP homeostasis, we evaluated the relationships between the distribution and activity of MRP4 in lipid rafts and cAMP efflux. METHODS: Platelet activation and cAMP homeostasis were analyzed in human and wild-type or MRP4-deleted mouse platelets in the presence of methyl-ß-cyclodextrin (MßCD) to disrupt lipid rafts, and of activators of the cAMP signalling pathways. Human platelet MRP4 and effector proteins of the cAMP pathway were analyzed by immunoblots in lipid rafts isolated by differential centrifugation. RESULTS: MßCD dose dependently inhibited human and mouse platelet aggregation without affecting per se cAMP levels. An additive inhibitory effect existed between the adenylate cyclase (AC) activator forskolin and MßCD that was accompanied by an overincrease of cAMP, and which was significantly enhanced upon MRP4 deletion. Finally, an efflux of cAMP out of resting platelets incubated with prostaglandin E1 (PGE1) was observed that was partly dependent on MRP4. Lipid rafts contained a small fraction (≈15%) of MRP4 and most of the inhibitory G-protein Gi, whereas Gs protein, AC3, and phosphodiesterases PDE2 and PDE3A were all present as only trace amounts. CONCLUSION: Our results are in favour of part of MRP4 present at the platelet surface, including in lipid rafts. Lipid raft integrity is necessary for cAMP signalling regulation, although MRP4 and most players of cAMP homeostasis are essentially located outside rafts.

Placental growth factor level in plasma predicts COVID-19 severity and in-hospital mortality.

BACKGROUND: Coronavirus disease 2019 (COVID-19) is a respiratory disease associated with vascular inflammation and endothelial injury. OBJECTIVES: To correlate circulating angiogenic markers vascular endothelial growth factor A (VEGF-A), placental growth factor (PlGF), and fibroblast growth factor 2 (FGF-2) to in-hospital mortality in COVID-19 adult patients. METHODS: Consecutive ambulatory and hospitalized patients with COVID-19 infection were enrolled. VEGF-A, PlGF, and FGF-2 were measured in each patient ≤48 h following admission. RESULTS: The study enrolled 237 patients with suspected COVID-19: 208 patients had a positive diagnostic for COVID-19, of whom 23 were mild outpatients and 185 patients hospitalized after admission. Levels of VEGF-A, PlGF, and FGF-2 significantly increase with the severity of the disease (P < .001). Using a logistic regression model, we found a significant association between the increase of FGF-2 or PlGF and mortality (odds ratio [OR] 1.11, 95% confidence interval [CI; 1.07-1.16], P < .001 for FGF-2 and OR 1.07 95% CI [1.04-1.10], P < .001 for PlGF) while no association were found for VEGF-A levels. Receiver operating characteristic curve analysis was performed and we identified PlGF above 30 pg/ml as the best predictor of in-hospital mortality in COVID-19 patients. Survival analysis for PlGF confirmed its interest for in-hospital mortality prediction, by using a Kaplan-Meier survival curve (P = .001) and a Cox proportional hazard model adjusted to age, body mass index, D-dimer, and C-reactive protein (3.23 95% CI [1.29-8.11], P = .001). CONCLUSION: Angiogenic factor PlGF is a relevant predictive factor for in-hospital mortality in COVID-19 patients. More than a biomarker, we hypothesize that PlGF blocking strategies could be a new interesting therapeutic approach in COVID-19.

Role of oculocerebrorenal syndrome of Lowe (OCRL) protein in megakaryocyte maturation, platelet production and functions: a study in patients with Lowe syndrome.

Lowe syndrome (LS) is an oculocerebrorenal syndrome of Lowe (OCRL1) genetic disorder resulting in a defect of the OCRL protein, a phosphatidylinositol-4,5-bisphosphate 5-phosphatase containing various domains including a Rho GTPase-activating protein (RhoGAP) homology domain catalytically inactive. We previously reported surgery-associated bleeding in patients with LS, suggestive of platelet dysfunction, accompanied with a mild thrombocytopenia in several patients. To decipher the role of OCRL in platelet functions and in megakaryocyte (MK) maturation, we conducted a case-control study on 15 patients with LS (NCT01314560). While all had a drastically reduced expression of OCRL, this deficiency did not affect platelet aggregability, but resulted in delayed thrombus formation on collagen under flow conditions, defective platelet spreading on fibrinogen and impaired clot retraction. We evidenced alterations of the myosin light chain phosphorylation (P-MLC), with defective Rac1 activity and, inversely, elevated active RhoA. Altered cytoskeleton dynamics was also observed in cultured patient MKs showing deficient proplatelet extension with increased P-MLC that was confirmed using control MKs transfected with OCRL-specific small interfering(si)RNA (siOCRL). Patients with LS also had an increased proportion of circulating barbell-shaped proplatelets. Our present study establishes that a deficiency of the OCRL protein results in a defective actomyosin cytoskeleton reorganisation in both MKs and platelets, altering both thrombopoiesis and some platelet responses to activation necessary to ensure haemostasis.

Circulating Von Willebrand factor and high molecular weight multimers as markers of endothelial injury predict COVID-19 in-hospital mortality.

BACKGROUND: Coronavirus disease 2019 (COVID-19) is a respiratory disease associated with endotheliitis and microthrombosis. OBJECTIVES: To correlate endothelial dysfunction to in-hospital mortality in a bi-centric cohort of COVID-19 adult patients. METHODS: Consecutive ambulatory and hospitalized patients with laboratory-confirmed COVID-19 were enrolled. A panel of endothelial biomarkers and von Willebrand factor (VWF) multimers were measured in each patient ≤ 48 h following admission. RESULTS: Study enrolled 208 COVID-19 patients of whom 23 were mild outpatients and 189 patients hospitalized after admission. Most of endothelial biomarkers tested were found increased in the 89 critical patients transferred to intensive care unit. However, only von Willebrand factor antigen (VWF:Ag) scaled according to clinical severity, with levels significantly higher in critical patients (median 507%, IQR 428-596) compared to non-critical patients (288%, 230-350, p < 0.0001) or COVID-19 outpatients (144%, 133-198, p = 0.007). Moreover, VWF high molecular weight multimers (HMWM) were significantly higher in critical patients (median ratio 1.18, IQR 0.86-1.09) compared to non-critical patients (0.96, 1.04-1.39, p < 0.001). Among all endothelial biomarkers measured, ROC curve analysis identified a VWF:Ag cut-off of 423% as the best predictor for in-hospital mortality. The accuracy of VWF:Ag was further confirmed in a Kaplan-Meier estimator analysis and a Cox proportional Hazard model adjusted on age, BMI, C-reactive protein and D-dimer levels. CONCLUSION: VWF:Ag is a relevant predictive factor for in-hospital mortality in COVID-19 patients. More than a biomarker, we hypothesize that VWF, including excess of HMWM forms, drives microthrombosis in COVID-19.

Human Aortic Valve Interstitial Cells Display Proangiogenic Properties During Calcific Aortic Valve Disease.

OBJECTIVE: The study's aim was to analyze the capacity of human valve interstitial cells (VICs) to participate in aortic valve angiogenesis. Approach and Results: VICs were isolated from human aortic valves obtained after surgery for calcific aortic valve disease and from normal aortic valves unsuitable for grafting (control VICs). We examined VIC in vitro and in vivo potential to differentiate in endothelial and perivascular lineages. VIC paracrine effect was also examined on human endothelial colony-forming cells. A pathological VIC (VICp) mesenchymal-like phenotype was confirmed by CD90+/CD73+/CD44+ expression and multipotent-like differentiation ability. When VICp were cocultured with endothelial colony-forming cells, they formed microvessels by differentiating into perivascular cells both in vivo and in vitro. VICp and control VIC conditioned media were compared using serial ELISA regarding quantification of endothelial and angiogenic factors. Higher expression of VEGF (vascular endothelial growth factor)-A was observed at the protein level in VICp-conditioned media and confirmed at the mRNA level in VICp compared with control VIC. Conditioned media from VICp induced in vitro a significant increase in endothelial colony-forming cell proliferation, migration, and sprouting compared with conditioned media from control VIC. These effects were inhibited by blocking VEGF-A with blocking antibody or siRNA approach, confirming VICp involvement in angiogenesis by a VEGF-A dependent mechanism. CONCLUSIONS: We provide here the first proof of an angiogenic potential of human VICs isolated from patients with calcific aortic valve disease. These results point to a novel function of VICp in valve vascularization during calcific aortic valve disease, with a perivascular differentiation ability and a VEGF-A paracrine effect. Targeting perivascular differentiation and VEGF-A to slow calcific aortic valve disease progression warrants further investigation.

Interleukin-8 Receptors CXCR1 and CXCR2 Are Not Expressed by Endothelial Colony-forming Cells.

Endothelial colony-forming cells (ECFCs) are human vasculogenic cells described as potential cell therapy product and good candidates for being a vascular liquid biopsy. Since interleukin-8 (IL-8) is a main actor in senescence, its ability to interact with ECFCs has been explored. However, expression of CXCR1 and CXCR2, the two cellular receptors for IL-8, by ECFCs remain controversial as several teams published contradictory reports. Using complementary technical approaches, we have investigated the presence of these receptors on ECFCs isolated from cord blood. First, CXCR1 and CXCR2 were not detected on several clones of cord blood- endothelial colony-forming cell using different antibodies available, in contrast to well-known positive cells. We then compared the RT-PCR primers used in different papers to search for the presence of CXCR1 and CXCR2 mRNA and found that several primer pairs used could lead to non-specific DNA amplification. Last, we confirmed those results by RNA sequencing. CXCR1 and CXCR2 were not detected in ECFCs in contrary to human-induced pluripotent stem cell-derived endothelial cells (h-iECs). In conclusion, using three different approaches, we confirmed that CXCR1 and CXCR2 were not expressed at mRNA or protein level by ECFCs. Thus, IL-8 secretion by ECFCs, its effects in angiogenesis and their involvement in senescent process need to be reanalyzed according to this absence of CXCR-1 and - 2 in ECFCs.Graphical Abstract.

No impact of cancer and plague-relevant FPR1 polymorphisms on COVID-19.

Formyl peptide receptor 1 (FPR1) is a pattern-recognition receptor that detects bacterial as well as endogenous danger-associated molecular patterns to trigger innate immune responses by myeloid cells. A single nucleotide polymorphism, rs867228 (allelic frequency 19-20%), in the gene coding for FPR1 accelerates the manifestation of multiple carcinomas, likely due to reduced anticancer immunosurveillance secondary to a defect in antigen presentation by dendritic cells. Another polymorphism in FPR1, rs5030880 (allelic frequency 12-13%), has been involved in the resistance to plague, correlating with the fact that FPR1 is the receptor for Yersinia pestis. Driven by the reported preclinical effects of FPR1 on lung inflammation and fibrosis, we investigated whether rs867228 or rs5030880 would affect the severity of coronavirus disease-19 (COVID-19). Data obtained on patients from two different hospitals in Paris refute the hypothesis that rs867228 or rs5030880 would affect the severity of COVID-19.

[Usefulness of immature platelet fraction measurement for diagnosis and monitoring of iron deficiency associated thrombocytopenia: about two cases]. / Apport de la mesure de la fraction de plaquettes immatures dans le diagnostic et le suivi des thrombopénies ferriprives : à propos de deux cas.

Iron deficiency anemia is frequently associated with thrombocytosis. However, in some rare cases of very severe iron deficiency, a thrombocytopenia may occur. This condition may lead to a misdiagnosis of immune thrombocytopenic purpura and thus to unnecessary tests in this context. Here we report two patients who presented with iron deficiency associated thrombocytopenia rapidly corrected after martial supplementation. We then discuss the value of measuring immature platelet fraction (IPF), which represents the population of newly formed platelets containing a greater amount of residual RNA. For both cases, low IPF values at admission indicated a central origin of thrombocytopenia with decreased platelet production, which is the pathophysiological mechanism of iron deficiency associated thrombocytopenia.

Comparative In Vitro Study of Various α2-Adrenoreceptor Agonist Drugs for Ticagrelor Reversal.

Ticagrelor, an antiplatelet adenosine diphosphate (ADP)-P2Y12 receptor antagonist, increases the risk of bleeding. Its management is challenging because platelet transfusion is ineffective and no specific antidote is currently available. Epinephrine, a vasopressor catecholamine prescribed during shock, restores platelet functions inhibited by ticagrelor through stimulation of α2A-adrenoreceptors. It subsequently inhibits cyclic adenosine monophosphate (cAMP) pathway and PI3K signaling. However, since epinephrine may expose a patient to deleterious hemodynamic effects, we hypothesized that other α2-adrenoreceptor agonist drugs used in clinical practice with fewer side effects could reverse the antiplatelet effects of ticagrelor. We compared in vitro the efficacy of clonidine, dexmedetomidine, brimonidine, and norepinephrine with epinephrine to restore ADP- and PAR-1-AP-induced washed platelet aggregation inhibited by ticagrelor, as well as resulting platelet cAMP levels. In ticagrelor-free samples, none of the α2-adrenoreceptor agonists induced aggregation by itself but all of them potentiated ADP-induced aggregation. Compared with epinephrine, norepinephrine, and brimonidine partially restored ADP- and fully restored PAR-1-AP-induced aggregation inhibited by ticagrelor while clonidine and dexmedetomidine were ineffective. Indeed, this lack of effect resulted from a lower decrease in cAMP concentration elicited by these partial α2-adrenoreceptor agonists, clonidine, and dexmedetomidine, compared with full α2-agonists. Our results support the development of specific full and systemic α2-adrenoreceptor agonists for ticagrelor reversal.

P2Y12 Inhibition beyond Thrombosis: Effects on Inflammation.

The P2Y12 receptor is a key player in platelet activation and a major target for antithrombotic drugs. The beneficial effects of P2Y12 receptor antagonists might, however, not be restricted to the primary and secondary prevention of arterial thrombosis. Indeed, it has been established that platelet activation also has an essential role in inflammation. Additionally, nonplatelet P2Y12 receptors present in immune cells and vascular smooth muscle cells might be effective players in the inflammatory response. This review will investigate the biological and clinical impact of P2Y12 receptor inhibition beyond its platelet-driven antithrombotic effects, focusing on its anti-inflammatory role. We will discuss the potential molecular and cellular mechanisms of P2Y12-mediated inflammation, including cytokine release, platelet-leukocyte interactions and neutrophil extracellular trap formation. Then we will summarize the current evidence on the beneficial effects of P2Y12 antagonists during various clinical inflammatory diseases, especially during sepsis, acute lung injury, asthma, atherosclerosis, and cancer.

Epinephrine restores platelet functions inhibited by ticagrelor: A mechanistic approach.

Ticagrelor, an antagonist of the platelet adenosine diphosphate (ADP)-P2Y12 receptor is recommended for patients with acute coronary syndromes. However, ticagrelor exposes to a risk of bleeding, the management of which is challenging because platelet transfusion is ineffective, and no antidote is yet available. We hypothesized that the vasopressor drug epinephrine could counter the antiplatelet effects of ticagrelor and restore platelet functions. We assessed in vitro the efficiency of epinephrine in restoring platelet aggregation inhibited by ticagrelor and investigated the underlying mechanisms. Washed platelet aggregation and secretion were measured upon stimulation by epinephrine alone or in combination with ADP, in the presence or absence of ticagrelor. Mechanistic investigations used P2Y1 and phosphoinositide 3-kinase (PI3K) inhibitors and included vasodilator-stimulated phosphoprotein (VASP) and Akt phosphorylation assays as well as measurement of Ca2+ mobilisation. We found that epinephrine restored ADP-induced platelet aggregation, but not dense granule release. Epinephrine alone failed to induce aggregation whereas it fully induced VASP dephosphorylation and Akt phosphorylation regardless of the presence of ticagrelor. In the presence of ticagrelor, blockage of the P2Y1 receptor prevented restoration of platelet aggregation by the combination of epinephrine and ADP, as well as intracellular Ca2+ mobilisation. In combination with ADP, epinephrine induced platelet aggregation of ticagrelor-treated platelets through inhibition of the cAMP pathway and activation of the PI3K pathway, thus enabling the P2Y1 receptor signalling and subsequent Ca2+ mobilisation. This proof-of-concept study needs to be challenged in vivo for the management of bleeding in ticagrelor-treated patients.