Age-induced alterations of granulopoiesis generate atypical neutrophils that aggravate stroke pathology.

Aging accounts for increased risk and dismal outcome of ischemic stroke. Here, we investigated the impact of age-related changes in the immune system on stroke. Upon experimental stroke, compared with young mice, aged mice had increased neutrophil clogging of the ischemic brain microcirculation, leading to worse no-reflow and outcomes. Aged mice showed an enhanced granulopoietic response to stroke that led to the accumulation of CD101+CD62Llo mature and CD177hiCD101loCD62Llo and CD177loCD101loCD62Lhi immature atypical neutrophils in the blood, endowed with increased oxidative stress, phagocytosis and procoagulant features. Production of CXCL3 by CD62Llo neutrophils of the aged had a key role in the development and pathogenicity of aging-associated neutrophils. Hematopoietic stem cell rejuvenation reverted aging-associated neutropoiesis and improved stroke outcome. In elderly patients with ischemic stroke, single-cell proteome profile of blood leukocytes identified CD62Llo neutrophil subsets associated with worse reperfusion and outcome. Our results unveil how stroke in aging leads to a dysregulated emergency granulopoiesis impacting neurological outcome.

Whole-Exome Sequencing in a Family with an Unexplained Tendency for Venous Thromboembolism: Multicomponent Prediction of Low-Frequency Variant Deleteriousness and of Individual Protein Interaction.

Whole-exome sequencing (WES) in families with an unexplained tendency for venous thromboembolism (VTE) may favor detection of low-frequency variants in genes with known contribution to hemostasis or associated with VTE-related phenotypes. WES analysis in six family members, three of whom affected by documented VTE, filtered for MAF < 0.04 in 192 candidate genes, revealed 22 heterozygous (16 missense and six synonymous) variants in patients. Functional prediction by multi-component bioinformatics tools, implemented by a database/literature search, including ClinVar annotation and QTL analysis, prioritized 12 missense variants, three of which (CRP Leu61Pro, F2 Asn514Lys and NQO1 Arg139Trp) were present in all patients, and the frequent functional variants FGB Arg478Lys and IL1A Ala114Ser. Combinations of prioritized variants in each patient were used to infer functional protein interactions. Different interaction patterns, supported by high-quality evidence, included eight proteins intertwined in the "acute phase" (CRP, F2, SERPINA1 and IL1A) and/or in the "fibrinogen complex" (CRP, F2, PLAT, THBS1, VWF and FGB) significantly enriched terms. In a wide group of candidate genes, this approach highlighted six low-frequency variants (CRP Leu61Pro, F2 Asn514Lys, SERPINA1 Arg63Cys, THBS1 Asp901Glu, VWF Arg1399His and PLAT Arg164Trp), five of which were top ranked for predicted deleteriousness, which in different combinations may contribute to disease susceptibility in members of this family.

Pulmonary Vascular Thrombosis in COVID-19 Pneumonia.

OBJECTIVES: During severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, dramatic endothelial cell damage with pulmonary microvascular thrombosis have been was hypothesized to occur. The aim was to assess whether pulmonary vascular thrombosis (PVT) is due to recurrent thromboembolism from peripheral deep vein thrombosis or to local inflammatory endothelial damage, with a superimposed thrombotic late complication. DESIGN: Observational study. SETTING: Medical and intensive care unit wards of a teaching hospital. PARTICIPANTS: The authors report a subset of patients included in a prospective institutional study (CovidBiob study) with clinical suspicion of pulmonary vascular thromboembolism. INTERVENTIONS: Computed tomography pulmonary angiography and evaluation of laboratory markers and coagulation profile. MEASUREMENTS AND MAIN RESULTS: Twenty-eight of 55 (50.9%) patients showed PVT, with a median time interval from symptom onset of 17.5 days. Simultaneous multiple PVTs were identified in 22 patients, with bilateral involvement in 16, mostly affecting segmental/subsegmental pulmonary artery branches (67.8% and 96.4%). Patients with PVT had significantly higher ground glass opacity areas (31.7% [22.9-41] v 17.8% [10.8-22.1], p < 0.001) compared with those without PVT. Remarkably, in all 28 patients, ground glass opacities areas and PVT had an almost perfect spatial overlap. D-dimer level at hospital admission was predictive of PVT. CONCLUSIONS: The findings identified a specific radiologic pattern of coronavirus disease 2019 (COVID-19) pneumonia with a unique spatial distribution of PVT overlapping areas of ground-glass opacities. These findings supported the hypothesis of a pathogenetic relationship between COVID-19 lung inflammation and PVT and challenged the previous definition of pulmonary embolism associated with COVID-19 pneumonia.

A Case of Covid-19 Patient with Acute Limb Ischemia and Heparin Resistance.

Heparin resistance is an uncommon phenomenon defined as the need for high-dose unfractionated heparin (UFH) of more than 35,000 IU/day to achieve the target activated partial-thromboplastin time ratio or the failure to achieve the desired activated clotting time after a full UFH dose. This rare phenomenon is being more commonly observed in Covid-19 patients in a hypercoagulable state. We describe a Covid-19 patient confirmed by reverse-transcriptase polymerase chain reaction assay, with acute limb ischemia, who developed heparin resistance. The patient was managed by the departments of vascular surgery, anesthesia and intensive care, and the Coagulation Service and Thrombosis Research from San Raffaele Scientific Institute, Milan, Italy.

Lentiviral gene therapy corrects platelet phenotype and function in patients with Wiskott-Aldrich syndrome.

BACKGROUND: Thrombocytopenia is a serious issue for all patients with classical Wiskott-Aldrich syndrome (WAS) and X-linked thrombocytopenia (XLT) because it causes severe and life-threatening bleeding. Lentiviral gene therapy (GT) for WAS has shown promising results in terms of immune reconstitution. However, despite the reduced severity and frequency of bleeding events, platelet counts remain low in GT-treated patients. OBJECTIVE: We carefully investigated platelet defects in terms of phenotype and function in untreated patients with WAS and assessed the effect of GT treatment on platelet dysfunction. METHODS: We analyzed a cohort of 20 patients with WAS/XLT, 15 of them receiving GT. Platelet phenotype and function were analyzed by using electron microscopy, flow cytometry, and an aggregation assay. Platelet protein composition was assessed before and after GT by means of proteomic profile analysis. RESULTS: We show that platelets from untreated patients with WAS have reduced size, abnormal ultrastructure, and a hyperactivated phenotype at steady state, whereas activation and aggregation responses to agonists are decreased. GT restores platelet size and function early after treatment and reduces the hyperactivated phenotype proportionally to WAS protein expression and length of follow-up. CONCLUSIONS: Our study highlights the coexistence of morphologic and multiple functional defects in platelets lacking WAS protein and demonstrates that GT normalizes the platelet proteomic profile with consequent restoration of platelet ultrastructure and phenotype, which might explain the observed reduction of bleeding episodes after GT. These results are instrumental also from the perspective of a future clinical trial in patients with XLT only presenting with microthrombocytopenia.

Missense changes in the catalytic domain of coagulation factor X account for minimal function preventing a perinatal lethal condition.

INTRODUCTION: Inherited deficiencies in the coagulation pathway provide diversified models to investigate the molecular bases of perinatal lethality associated with null-like variants. Differently from X-linked haemophilias, homozygous/doubly heterozygous null variants in the rare autosomally inherited deficiency of factor X (FX) might be incompatible with perinatal survival. AIM: To provide experimental evidence about the null/close-to-null FX function. METHODS: The residual secreted (ELISA) and functional (thrombin generation assays) protein levels associated with the novel nonsense (c.1382G>A; p.Trp461Ter) and missense (c.752T>C; p.Leu251Pro) variants, found in the proposita with life-threatening symptoms at birth, were characterized through recombinant (r)FX expression. RESULTS: The rFX-461Ter showed very low secretion and undetectable function. Expression and function of the predicted readthrough-deriving missense variants (rFX-461Tyr, rFX-461Gln) were also severely impaired. These unfavourable features, due to nucleotide and protein sequence constraints, precluded functional readthrough over the 461 stop codon. Differently, the poorly secreted rFX-251Pro variant displayed residual function that was characterized by anti-TFPI aptamer-based amplification or selective inhibition of activated FX function by fondaparinux in plasma and found to be reduced by approximately three orders of magnitude. Similarly to the rFX-251Pro, a group of catalytic domain missense variants cause poorly secreted molecules with modest function in FX-deficient patients with life-threatening symptoms. CONCLUSIONS: Our data, contributing to the knowledge of the very severe FX deficiency forms, support life-saving requirement of trace FX function, clearly exemplified by the dysfunctional but not completely inactive rFX-251Pro variant that, albeit with severely reduced function, is compatible with a residual activity ensuring minimal haemostasis and permitting perinatal survival.

Low molecular weight heparins prevent the induction of autophagy of activated neutrophils and the formation of neutrophil extracellular traps.

The protection exerted by neutrophils against invading microbes is partially mediated via the generation of neutrophil extracellular traps (NETs). In sterile conditions NETs are damaging species, enriched in autoantigens and endowed with the ability to damage the vessel wall and bystander tissues, to promote thrombogenesis, and to impair wound healing. To identify and reposition agents that can be used to modulate the formation of NETs is a priority in the research agenda. Low molecular weight heparins (LMWH) are currently used, mostly on an empirical basis, in conditions in which NETs play a critical role, such as pregnancy complications associated to autoimmune disease. Here we report that LMWHs induce a profound change in the ability of human neutrophils to generate NETs and to mobilize the content of the primary granules in response to unrelated inflammatory stimuli, such as IL-8, PMA and HMGB1. Autophagy consistently accompanies NET generation in our system and autophagy inhibitors, 3-MA and wortmannin, prevent NET generation. Pretreatment with LMWH in vitro critically jeopardizes neutrophil ability to activate autophagy, a mechanism that might contribute to neutrophil unresponsiveness. Finally, we verified that treatment of healthy volunteers with a single prophylactic dose of parnaparin abrogated the ability of neutrophils to activate autophagy and to generate NETs. Together, these results support the contention that neutrophils, and NET generation in particular, might represent a preferential target of the anti-inflammatory action of LMWH.

Low-grade endotoxemia and clotting activation in the early phase of pneumonia.

BACKGROUND AND OBJECTIVE: Community-acquired pneumonia (CAP) is associated with an increased risk of arterial and venous thrombosis but the underlying pathophysiological mechanisms are still unclear. We investigated if, in patients with CAP, a pro-thrombotic state does exist and its relationship with serum levels of endotoxins. METHODS: A total of 104 consecutive patients with CAP were prospectively recruited and followed up until discharge. At admission and at discharge, serum endotoxins, systemic markers of clotting activation and zonulin, a marker of gut permeability, were analysed. Hospitalized patients matched for gender, age and comorbidities but without infections were used as control. RESULTS: At admission, CAP patients showed higher plasma levels of F1+2 , a marker of thrombin generation (P = 0.023), and lower levels of protein C (PC; P < 0.001) and activated PC (aPC) (P < 0.001) compared with controls. At discharge, plasma levels of both PC and aPC significantly increased while F1+2 significantly decreased (P < 0.001). Baseline serum endotoxins and zonulin were higher in CAP patients than controls (P < 0.001) and significantly decreased at discharge; a significant correlation between serum endotoxins and zonulin was detected (R = 0.575; P < 0.001) CONCLUSION: This study provides the first evidence that CAP patients disclose an ongoing pro-thrombotic state and suggests a role for endotoxemia in determining enhanced thrombin generation.

Hyperfunctional coagulation factor IX improves the efficacy of gene therapy in hemophilic mice.

Gene therapy may provide a cure for hemophilia and overcome the limitations of protein replacement therapy. Increasing the potency of gene transfer vectors may allow improvement of their therapeutic index, as lower doses can be administered to achieve therapeutic benefit, reducing toxicity of in vivo administration. Here we generated codon-usage optimized and hyperfunctional factor IX (FIX) transgenes carrying an R338L amino acid substitution (FIX Padua), previously associated with clotting hyperactivity and thrombophilia. We delivered these transgenes to hemophilia B mice by hepatocyte-targeted integration-competent and -defective lentiviral vectors. The hyperfunctional FIX transgenes increased FIX activity reconstituted in the plasma without detectable adverse effects, allowing correction of the disease phenotype at lower vector doses and resulting in improved hemostasis in vivo. The combined effect of codon optimization with the hyperactivating FIX-R338L mutation resulted in a robust 15-fold gain in potency and therefore provides a promising strategy to improve the efficacy, feasibility, and safety of hemophilia gene therapy.

Head-to-head comparison of four COVID-19 vaccines on platelet activation, coagulation and inflammation. The TREASURE study.

INTRODUCTION: Studies exploring alterations in blood coagulation and platelet activation induced by COVID-19 vaccines are not concordant. We aimed to assess the impact of four COVID-19 vaccines on platelet activation, coagulation, and inflammation considering also the immunization dose and the history of SARS-CoV-2 infection. METHODS: TREASURE study enrolled 368 consecutive subjects (161 receiving viral vector vaccines -ChAdOx1-S/Vaxzevria or Janssen- and 207 receiving mRNA vaccines -Comirnaty/Pfizer-BioNTech or Spikevax/Moderna). Blood was collected the day before and 8 ± 2 days after the vaccination. Platelet activation markers (P-selectin, aGPIIbIIIa and Tissue Factor expression; number of platelet-monocyte and -granulocyte aggregates) and microvesicle release were analyzed by flow cytometry. Platelet thrombin generation (TG) capacity was measured using the Calibrated Automated Thrombogram. Plasma coagulation and inflammation markers and immune response were evaluated by ELISA. RESULTS: Vaccination did not induce platelet activation and microvesicle release. IL-6 and CRP levels (+30%), D-dimer, fibrinogen and F1+2 (+13%, +3.7%, +4.3%, respectively) but not TAT levels significantly increased upon immunization with all four vaccines, with no difference among them and between first and second dose. An overall minor post-vaccination reduction of aPC, TM and TFPI, all possibly related to endothelial function, was observed. No anti-PF4 seroconversion was observed. CONCLUSION: This study showed that the four COVID-19 vaccines administered to a large population sample induce a transient inflammatory response, with no onset of platelet activation. The minor changes in clotting activation and endothelial function might be potentially involved at a population level in explaining the very rare venous thromboembolic complications of COVID-19 vaccination.

Platelet activation state in early stages of COVID-19.

BACKGROUND: Platelet activation at the early stage of COVID-19 is poorly described. The need for antiplatelet therapy in patients with COVID-19 remains controversial. We characterized the platelet activation profile in hospitalized patients at the early stage of COVID-19 using the modified prothrombinase Platelet Activation State (PAS) Assay. METHODS: Sixteen patients admitted to the emergency department of the IRCCS San Raffaele Hospital (Milan, Italy) between February 8 and April 2021 were enrolled. All patients presented with respiratory symptoms and tested positive for severe acute respiratory syndrome Coronavirus 2 (SARS-CoV-2). Platelet activation was measured via the PAS Assay within 24 hours from patients' hospital admission. Data were compared with those measured in N.=24 healthy subjects (controls). RESULTS: Platelet activation was significantly higher in COVID-19 patients with respect to controls (PAS=0.63 [0.58-0.98%] vs. 0.46 [0.40-0.65%], respectively; P=0.03). Of note, highest PAS values were measured in the two patients with the worst clinical outcome, i.e., death because of respiratory failure (PAS=2.09% and 1.20%, respectively). No differences in standard coagulation parameters were noted between these two patients and those who were later discharged home. CONCLUSIONS: This study provides evidence of significant platelet activation state at the early stage of COVID-19 and suggests that the patient-specific platelet activation profile is a reliable clinical marker to stratify COVID-19 patients at high risk of poor clinical outcome who might potentially benefit from antiplatelet therapy.

Bleeding in patients with continuous-flow left ventricular assist devices: acquired von Willebrand disease or antithrombotics?

OBJECTIVES: To evaluate the competing pro-haemorrhagic contribution of acquired von Willebrand (vW) disease and antithrombotic therapy in patients implanted with continuous-flow left ventricular assist devices (LVADs). METHODS: We compared the extent of vW factor (vWf) degradation [vWf antigen (vWf:Ag)] and a decrease of functional activity of large vWf multimers [vWf collagen binding (vWf:CB)] in LVAD patients who did and did not suffer from bleeding. Data were measured pre-implant, at short-term (t1: <3 months) and long-term (t2: >12 months) follow-up. The occurrence of primary bleeding events, as well as bleeding recurrence, was correlated with patient-specific vWf profile and antithrombotic regimen. Indeed, patients were discharged on warfarin (international normalized ratio: 2-2.5) and aspirin, with the latter withhold after a first bleeding episode. RESULTS: Fifty-three patients were enrolled. The median follow-up was 324 (226-468) days. We recorded 25 primary bleeding events (47% of patients). All primary events occurred in patients on warfarin and aspirin. Both vWf:Ag and vWf:CB decreased significantly post-implant (P = 0.0003 and P < 0.0001), and patients showing pathological vWf:CB/vWf:Ag ratio (<0.7) increased progressively over the time of support (pre-implant = 26%, t1 = 58%, t2 = 74%; P < 0.0001). Of note, activity of large vWf multimers of bleeders was significantly lower at t2 with respect to non-bleeders (vWf:CB: 61 (36-115) vs 100 (68-121), P = 0.04; vWf:CB/vWf:Ag ratio: 0.36 (0.26-0.61) vs 0.58 (0.33-0.96), P = 0.04). Despite these marked differences in the vWf profile, following aspirin discontinuation only 3 patients had bleeding recurrence. CONCLUSIONS: Aspirin contributes significantly to haemorrhagic events in the background of acquired vW disease; its discontinuation significantly reduces bleeding recurrence. CLINICAL TRIAL REGISTRATION: https://clinicaltrials.gov/ct2/show/NCT03255928; ClinicalTrials.gov Identifier: NCT03255928.

A novel evidence-based algorithm to predict thromboembolism in patients with COVID-19: preliminary data from a single-center cohort.

BACKGROUND: Severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2)-related disease (COVID-19) is an infectious disease characterized by systemic inflammation, which might enhance baseline thrombotic risk, especially in hospitalized patients. Little is, however, known about predictors of thrombotic complications in patients with COVID-19. METHODS: We prospectively followed up 180 hospitalized COVID-19 patients. Demographics, clinical and laboratory features at presentation and past medical history were tested as predictors of the first thrombotic complication through multivariate Cox regression analysis and a categorical score generated based on the results. RESULTS: Sixty-four thromboses were recorded in 54 patients, of whom seven with thrombosis on admission and 47 with thrombosis during hospitalization. Patients with thrombosis were mainly Caucasian and diabetic, had marked baseline signs of inflammation and organ damage, lower PaO2/FiO2 ratio, higher D-dimer levels and history of major hemorrhages. The latter three variables were independently associated to thrombotic complications and concurred to a 0-5 score, which accounted for 80% of the total sample variability. Patients with three or more points of the newly generated score were at higher risk for thrombotic complications (HR=4.9, P<0.001). Patients with thrombotic complications were more likely to be admitted to intensive care and/or to die (HR=1.9, P=0.036). Five of 180 patients were diagnosed with disseminated intravascular coagulation and three of them died. Eleven minor and no major bleeding events were observed. CONCLUSIONS: Patients with COVID-19 are at increased risk for thrombosis and might be stratified on admission based on lower Pao2/FiO2 ratio, higher D-dimer levels and history of major hemorrhages.

High Risk of Secondary Infections Following Thrombotic Complications in Patients With COVID-19.

Background: This study's primary aim was to evaluate the impact of thrombotic complications on the development of secondary infections. The secondary aim was to compare the etiology of secondary infections in patients with and without thrombotic complications. Methods: This was a cohort study (NCT04318366) of coronavirus disease 2019 (COVID-19) patients hospitalized at IRCCS San Raffaele Hospital between February 25 and June 30, 2020. Incidence rates (IRs) were calculated by univariable Poisson regression as the number of cases per 1000 person-days of follow-up (PDFU) with 95% confidence intervals. The cumulative incidence functions of secondary infections according to thrombotic complications were compared with Gray's method accounting for competing risk of death. A multivariable Fine-Gray model was applied to assess factors associated with risk of secondary infections. Results: Overall, 109/904 patients had 176 secondary infections (IR, 10.0; 95% CI, 8.8-11.5; per 1000-PDFU). The IRs of secondary infections among patients with or without thrombotic complications were 15.0 (95% CI, 10.7-21.0) and 9.3 (95% CI, 7.9-11.0) per 1000-PDFU, respectively (P = .017). At multivariable analysis, thrombotic complications were associated with the development of secondary infections (subdistribution hazard ratio, 1.788; 95% CI, 1.018-3.140; P = .043). The etiology of secondary infections was similar in patients with and without thrombotic complications. Conclusions: In patients with COVID-19, thrombotic complications were associated with a high risk of secondary infections.

Thromboembolism risk among patients with diabetes/stress hyperglycemia and COVID-19.

PURPOSE: Individuals with diabetes/stress hyperglycemia carry an increased risk for adverse clinical outcome in case of SARS-CoV-2 infection. The purpose of this study was to evaluate whether this risk is, at least in part, modulated by an increase of thromboembolic complications. METHODS: We prospectively followed 180 hospitalized patients with confirmed COVID-19 pneumonia admitted to the Internal Medicine Units of San Raffaele Hospital. Data from 11 out of 180 patients were considered incomplete and excluded from the analysis. We analysed inflammation, tissue damage biomarkers, hemostatic parameters, thrombotic events (TEs) and clinical outcome according to the presence of diabetes/stress hyperglycemia. RESULTS: Among 169 patients, 51 (30.2%) had diabetes/stress hyperglycemia. Diabetes/stress hyperglycemia and fasting blood glucose (FBG) were associated with increased inflammation and tissue damage circulating markers, higher D-dimer levels, increased prothrombin time and lower antithrombin III activity. Forty-eight venous and 10 arterial TEs were identified in 49 (29%) patients. Diabetes/stress hyperglycemia (HR 2.71, p = 0.001), fasting blood glucose (HR 4.32, p < 0.001) and glucose variability (HR 1.6, p < 0.009) were all associated with an increased risk of thromboembolic complication. TEs significantly increased the risk for an adverse clinical outcome only in the presence of diabetes/stress hyperglycemia (HR 3.05, p = 0.010) or fasting blood glucose ≥7 mmol/L (HR 3.07, p = 0.015). CONCLUSIONS: Thromboembolism risk is higher among patients with diabetes/stress hyperglycemia and COVID-19 pneumonia and is associated to poor clinical outcome. In case of SARS-Cov-2 infection patients with diabetes/stress hyperglycemia could be considered for a more intensive prophylactic anticoagulation regimen.

Incidence of deep venous thrombosis in COVID-19 hospitalized patients during the first peak of the Italian outbreak.

OBJECTIVES: A high rate of thrombotic events has been reported in COVID-19 population. The study aims to assess the incidence of deep vein thrombosis (DVT) in COVID-19 patients admitted to a single tertiary hospital. METHODS: From April 2nd to April 18th, 2020, hospitalized patients with SARS-CoV-2 infection were screened by lower limb duplex ultrasound (DUS). Patients were on (low molecular weight heparin) LMWH prophylaxis in medical wards, and on therapeutic anticoagulation in intensive care unit (ICU). DVT risk factors, reported by the Padua prediction score and blood tests, were retrieved from institutional electronic charts. The study primary endpoint was the incidence of DVT in the in-hospital COVID-19 population and its association with clinical and laboratory risk factors. The secondary endpoint was the association of DVT with mortality. RESULTS: Two hundred patients (median age 62 years, 72% male, 40 in ICU) received DUS screening. DVT was observed in 29 patients (14.5%), with proximal extension in 16 patients, and in association with symptoms in four patients. The DVT rate was similar in ICU (12.5%) and non-ICU patients (15%). Eighty-seven patients underwent a computed tomography angiography (CTA) that showed pulmonary embolism in 35 patients (40.2%) not associated with DVT in 25/35 cases (71.4%). DVT in the ten patients with pulmonary embolism were symptomatic in four and with a proximal localization in eight cases. A D-dimer level ≥5 mg/l at admission was predictive of DVT (OR 1.02; IC95% 1.03-1.16; p = .003). At the multivariate analysis in-hospital mortality was predicted by age (OR 1.06; 95% CI 0.02-1.15; p = .004) and by being an ICU patient (OR 1.23; 95% CI 0.30-2.25; p = .01). CONCLUSIONS: Despite LMWH prophylaxis or full anticoagulant therapy, the incidence of DVT, mainly asymptomatic, in hospitalized COVID-19 patients was 14.5%. Further research should focus on the appropriate antithrombotic therapy for COVID-19 patients.

Influence of Different Antithrombotic Regimens on Platelet-Mediated Thrombin Generation in Patients with Left Ventricular Assist Devices.

We characterized the biologic background of prothrombotic platelet function in the setting of durable left ventricular assist devices (LVADs) evaluating the role of different antithrombotic regimens. Platelet-mediated thrombin generation was quantified using the Platelet Activity State (PAS) Assay and the Thrombin Generation Test (TGT) in 78 patients implanted with the HeartMate II (n = 10, 13%), the HeartMate 3 (HM3) (n = 30, 38%), or the HVAD (n = 38, 49%) and managed with oral anticoagulation plus aspirin (n = 46, 59%) or anticoagulation alone (n = 32, 41%). Coagulation parameters (platelet count, International Normalized Ratio (INR), activated Partial Thromboplastin Time, Fibrinogen and D-Dimer levels) and hemolysis (lactate dehydrogenase levels [LDH]) were also recorded to comprehensively characterize the hemostatic profile in the two groups. In patients without aspirin, the PAS assay revealed low-intensity increase in platelet prothrombinase activity (1.11-fold, p = 0.03). Similarly the TGT revealed moderate higher platelet reactivity when compared with patients receiving aspirin, consistent with reduction in lag time (0.87-fold, p < 0.001), increase in peak of thrombin generation (1.5-fold, p = 0.002) and thrombin generation rate (2-fold, p = 0.02), but comparable endogenous thrombin potential (p = 0.50). Coagulation parameters and LDH were comparable in the two groups (p > 0.05). Moreover, no differences were noted in platelet prothrombinase activity of patients implanted with the HM3 or HVAD. Our results suggest that, in the setting of durable LVADs, aspirin minimally modulates the biochemical pathway of platelet-mediated thrombin generation. Accordingly, re-evaluation of current antithrombotic management criteria in patients stratified according to bleeding/thromboembolic risk might be safe and beneficial to prevent adverse events.

Microvascular COVID-19 lung vessels obstructive thromboinflammatory syndrome (MicroCLOTS): an atypical acute respiratory distress syndrome working hypothesis.

We suggest the use of MicroCLOTS (microvascular COVID-19 lung vessels obstructive thromboinflammatory syndrome) as a new name for severe pulmonary coronavirus disease 2019 (COVID-19). We hypothesise that, in predisposed individuals, alveolar viral damage is followed by an inflammatory reaction and by microvascular pulmonary thrombosis. This progressive endothelial thromboinflammatory syndrome may also involve the microvascular bed of the brain and other vital organs, leading to multiple organ failure and death. Future steps in the understanding of the disease and in the identification of treatments may benefit from this definition and hypothesised sequence of events.

Educational Impact on Apixaban Adherence in Atrial Fibrillation (the AEGEAN STUDY): A Randomized Clinical Trial.

INTRODUCTION: Adherence to non-vitamin-K oral anticoagulants (NOACs) may be lower than to vitamin K antagonists because NOACs do not require routine monitoring. OBJECTIVE: We assessed the impact of an educational program on adherence and persistence with apixaban in patients with non-valvular atrial fibrillation (NVAF). METHODS: Patients with NVAF eligible for NOACs with one or more stroke risk factor (prior stroke/transient ischemic attack, age ≥ 75 years, hypertension, diabetes, or symptomatic heart failure) were randomized (1:1) to standard of care (SOC) or SOC with additional educational (information booklet, reminder tools, virtual clinic access). The primary outcome was adherence to apixaban (2.5 or 5 mg twice daily) at 24 weeks. Patients receiving the educational program were re-randomized (1:1) to continue the program for 24 further weeks or to switch to secondary SOC. Implementation adherence and persistence were reassessed at 48 weeks. RESULTS: In total, 1162 patients were randomized (SOC, 583; educational program, 579). Mean implementation adherence ± standard deviation (SD) at 24 weeks was 91.6% ± 17.1 for SOC and 91.9% ± 16.1 for the educational program arm; results did not differ significantly between groups at any time-point. At 48 weeks, implementation adherence was 90.4% ± 18.0, 90.1% ± 18.6, and 89.3% ± 18.1 for continued educational program, SOC, and secondary SOC, respectively; and corresponding persistence was 86.1% (95% confidence interval [CI] 81.3-89.7), 85.2% (95% CI 81.5-88.2), and 87.8% (95% CI 83.4-91.1). Serious adverse events were similar across groups. CONCLUSION: High implementation adherence and persistence with apixaban were observed in patients with NVAF receiving apixaban. The educational program did not show additional benefits. CLINICAL TRIAL REGISTRATION: This study is registered at ClinicalTrials.gov [NCT01884350].

Telomere length in peripheral blood mononuclear cells is associated with folate status in men.

Human chromosomes are capped by telomeres, which consist of tandem repeats of DNA and associated proteins. The length of the telomeres is reduced with increasing cell divisions except when the enzyme telomerase is active, as in stem cells and germ cells. Telomere dysfunction has been associated with development of age-related pathologies, including cancer, cardiovascular disease, Alzheimer's disease, and Parkinson's disease. DNA damage in the telomeric region causes attrition of telomeres. Because folate provides precursors for nucleotide synthesis and thus affects the integrity of DNA, including that of the telomeric region, folate status has the potential to influence telomere length. Telomere length is epigenetically regulated by DNA methylation, which in turn could be modulated by folate status. In this study, we determined whether folate status and the 677C > T polymorphism of the methylene tetrahydrofolate reductase (MTHFR) gene are associated with the telomere length of peripheral blood mononuclear cells in healthy men. The results of our study showed that plasma concentration of folate was associated with telomere length of peripheral blood mononuclear cells in a nonlinear manner. When plasma folate concentration was above the median, there was a positive relationship between folate and telomere length. In contrast, there was an inverse relationship between folate and telomere length when plasma folate concentration was below the median. The MTHFR 677C > T polymorphism was weakly associated (P = 0.065) with increased telomere length at below-median folate status. We propose that folate status influences telomere length by affecting DNA integrity and the epigenetic regulation of telomere length through DNA methylation.