Evaluation of the Effect of Favipiravir Use on Inr, Pt, Aptt Tests in Covid-19 Patients

Objective: Studies have shown that high mortality rates associated with abnormal coagulation response, bleeding and coagulation disorders in COVID-19 patients. In our study, it was aimed to investigate the effect of the use of favipiravir on coagulation tests such as INR, PTT and Aptt. Materials-Methods: 50 patients who had a positive RT-PCR in nasal and throat swabs result and were diagnosed with COVID-19 using favipiravir and 50 non-users favipiravir COVID-19 patients were included. INR, PT, Aptt data were evaluated for all patients. Result(s): Results of patients using favipiravir;INR 1.3+/-0.2, PT(s) 16.4+/-3.4, Aptt(s) 40.7+/-10.1, while the results of patients who did not use favipiravir were INR 1,2+/-0.2, PT(s) 14.6+/-2.5, Aptt(s) was found 38.4+/-7.8. While PT and INR were found to be significantly higher in patients using favipiravir (p<0.05), the elevation in Aptt values was not significant. Conclusion(s): As a result, it was observed that favipiravir prolongs the clotting time. In the light of these RESULTS, it is recommended to consider this in anticoagulant therapy used for treatment.

Protein S Deficiency with Favourable Foetomaternal Outcome: A Case Report

Protein S is a multifunctional plasma protein, whose deficiency, results in a rare congenital thrombophilia, inherited in an autosomal dominant pattern. It can aggravate the hypercoagulable state of pregnancy, when it presents in parallel with the condition, leading to adverse maternal outcomes and foetal loss. A 35-year-old female third gravida having previous 2 deliveries by Lower Segment Caesarean Section (LSCS) presented to emergency at 10 weeks pregnancy with chief complaints of pain and swelling in left thigh since 4-5 days. After thorough investigations and work-up, the patient was diagnosed with Protein S deficiency. She was managed conservatively and was delivered by elective LSCS with bilateral tubal ligation at 38 weeks of gestation with good foetal and maternal outcomes.The rarity of Protein S deficiency along with the successful outcome of the pregnancy makes this a unique case.Copyright © 2023 Journal of Clinical and Diagnostic Research. All rights reserved.

Impact of endothelial function on the prognosis of COVID19 patients

Introduction: The pathophysiological mechanisms related to the severity of the clinical picture of Sars-Cov-2 infection remain questions that the medical community seeks to resolve and whose knowledge will allow the design of therapeutic strategies. Purpose(s): To evaluate the association between endothelial function and mortality in patients with COVID-19. Material(s) and Method(s): Prospective cohort study. Patients with confirmed diagnosis of COVID-19, who required hospitalization, oxygen saturation <60% and laboratory data of endothelial function markers were included;all those with incomplete data were excluded. Result(s): The age of the population was 57.06 +/- 13.405, 69% were intubated, those who died had poorer ventilatory parameters;peak pressure (30.06 +/- 6.13 vs 24.82 +/- 6.06, p<0.001), Fi02 (57.5 [40 - 80] vs 45 [40 -80], p <0.001) and PaO2/FiO2 calculation (127.84 +/- 50.08 vs 163.36 +/- 45.47, p<0.001). In addition, they had greater laboratory alterations: procalcitonin (0.49 [0.16 - 4] vs. 18 [0.07 - 0.52], p=0.03) and CRP (13.34[7.67 - 19.03] vs 6.69 [2.4 - 15 89], p=0.05), higher levels of clotting times, PT(14.8 [14 - 16.6] vs 14.4 [13.7 -15.6], p=0.5) and INR (1.04 [0.98 - 1.17] vs 1.01 [.95 - 1.1], p=0.05). In blood biometry, higher numbers of leukocytes of (11.3 [8.65 - 14.4] vs. 9.9 [6.7 - 13], p=0.041), neutrophils (8.8 [7.4 - 12.5] vs. 8.1 [4.74 - 11.1], p=0.01) and lower numbers of lymphocytes (. 7 [.40 - 1.1] vs 1.1 [.7 - 1.5], p<0.001) and with significant statistical trend, hemoglobin(11.26 +/- 2.55 vs 12.42 +/- 2.4, p=0.07) compared to surviving subjects. Conclusion(s): Hematological and liver alterations are markers of higher mortality in patients with COVID-19 as an expression of multiorgan disease.

Monitoring of argatroban in COVID-19 ICU patients: A prospective study comparing aPTT, point-of-care viscoelastic testing with ECA-test and dTT to LC/MS/MS

Introduction Argatroban is indicated for treatment of heparin-induced thrombocytopenia, but is also used in critical ill COVID-19 patients presenting with extensive thrombin overload. Direct drug monitoring is not available and argatroban dosing is mainly based on activated partial plasmin time (aPTT), which has limitations in hypercoagulable patients with increased FVIII [1, 2]. The aim of this study was to compare correlation of routine clotting tests (aPTT, ecarin clotting time [ECA-CT] and diluted thrombin time [dTT]) [3] to argatroban plasma levels measured by gold standard mass spectrometry (LC/MS/MS). Method From 06/2021 to 03/2022, 205 samples from 22 COVID-19 ICU patients were analyzed: aPTT and dTT on STA R Max3-Analyzer (Stago Deutschland GmbH, Germany) using the BIOPHEN DTI Kit with Argatroban-calibration (CoaChrom Diagnostica GmbH, Austria);ECA-CT was measured using ClotPro ecarin assay. LC/MS/MS was performed using an RP column, a solvent gradient and an API4000 mass spectrometer with electrospray. Correlation was analyzed using Pearson correlation coefficient r in R version 3.2.4. This study was approved by the Ethics Committee of the Technical University of Dresden, Germany (BO-EK-64022022) and registered with German Clinical Trials Register DRKS00028689. Results From 205 samples with LC/MS/MS analysis, 195 were compared to aPTT, 153 to ECA-CT and 105 to dTT. In 40 samples, dTT was not measureable due high bilirubin values. Compared to LC/MS/MS, correlation of dTT was highest (r = 0.924), followed by ECA-CT (r 0.609) and aPTT (r 0.367;p < 0.001;Fig. 1). When recommended cut-offs for argatroban plasma levels (500-1000 ng/ml according to SmPC) were applied, dTT (when measurable) and ECA-CT better identified critical values of argatroban plasma values > 1000ng/ml than aPTT (Fig. 2). Conclusion Argatroban in critical ill COVID-19 patients should be monitored using dTT. If dTT is not possible or measurements are highly time-sensitive, point-of-care ClotPro ECA-test should be preferably used instead of aPTT. (Table Presented).

Monitoring of unfractionated heparin in critical ill ICU patients: A prospective study comparing aPTT and point-of-care viscoelastic testing with IN /HI ratio to anti-Xa

Introduction Anticoagulation is indicated for the prevention or therapy of thromboembolic events, but remains highly challenging considering the high risk of bleeding events in critically ill patients.Unfractionated heparin (UFH) is widely used as preferred anticoagulation for patients on intensive care units (ICU) due to its beneficial short half time and fast elimination. For monitoring of UFH, activated partial thromboplastin time (aPTT) is mainly used, but aPTT can be misleading in both directions [1]. While high factor VIII plasma values may decrease aPTT, a reduced factor XII under extracorporal circulation may prolong aPTT that no longer correlates to anticoagulation intensity [2]. Anticoagulation monitoring using specific UFH calibrated anti-Xa levels is an established alternative to overcome aPTT limitations but is rarely available 24/7 [3, 4]. Using point-of-care (POC) viscoelastic testing (VET) [5] with a specific ratio between clotting time (CT) in intrinsic test (IN-test) compared to heparinase test (HI-test) - which includes the inactivation of heparin in the probe - might help to determine the UFH effect in critically ill patients [6]. Method From 09/2020 to 07/2022, 467 samples from 120 adult ICU patients receiving UFH therapy were prospectively collected. Samples for aPTT, anti-Xa measurement and POC VET using ClotPro (Haemonetics, Boston, Massachusetts, USA) were simultaneously collected. Measurement for aPTT (C.K. Prest) and anti-Xa (Liquid AntiXa) were performed using STA R Max 3 device (Stago Deutschland GmbH, Dusseldorf). Correlation was analyzed using Kruskal-Wallis test in SPSS version 27 and R version 3.2.4. This study was approved by the local Ethics Committee at the Technische Universitat Dresden, Germany (BOEK- 374072021) and registered with the German Clinical Trials Register DRKS00028689. Results 467 samples under UFH treatment were included in this analysis, the majority of these patients were treated for COVID-19 associated acute distress syndrome. According to our institutional guidelines, anti-Xa target levels for UFH were set at 0.3-0.5 IE/ml for standard high-risk prophylaxis and 0.5-0.7 IE/ ml for therapeutic anticoagulation therapy with values < 0.3 and > 0.7 being defined as under- or over-treatment. Table 1 presents the median aPTT and CT IN/HI ratio values for patients within these anti-Xa categories. CT IN/HI ratio correlation to anti-Xa levels was considerably better than aPTT correlation (Tab. 1). Notably, aPTT could not exactly discriminate between patients receiving UFH dosages correlating to high-risk prophylaxis or therapeutic anticoagulation. Conclusion Whole blood POC VET using a specific heparinase-approach (IN/ HI ratio) is superior to aPTT in detecting patients in or out of targeted anti-Xa levels. POC VET should be made available for ICUs as bedside test and might help to guide anticoagulation management in critical ill patients, being faster and potentially more widely available than lab-based anti-Xa testing (Fig. 1). (Table Presented).

Recovery after COVID-19 and persisting endothelial cell activation and hypercoagulability-The prospective observational ROADMAP-post COVID-19 study

Background: Hypercoagulable state and endothelial cell activation are common alterations in patients with COVID-19. Nevertheless, the hypothesis of persistent hypercoagulability and endothelial cell activation following recovery from COVID-19 remains an unresolved issue. Aim(s): To investigate the persistence of endothelial cell activation and hypercoagulability after recovery from COVID-19. Patients/ Methods. COVID-19 survivors (n = 208) and 30 healthy individuals were enrolled in this study. Method(s): The following biomarkers were measured: Procoagulant phospholipid-dependent clotting time (PPL-ct), D-Dimer, fibrin monomers (FM), free Tissue factor pathway inhibitor (free-TFP) I, heparinase, and soluble thrombomodulin (sTM). Antibodies against SARS-CoV- 2 (IgG and IgA) were also measured. Result(s): The median interval between symptom onset and screening for SARS-CoV- 2 antibodies was 62 days (IQR = 22 days). Survivors showed significantly higher levels of D-Dimers, FM, TFPI, and heparanase as compared to that of the control group. Survivors had significantly shorter PPL-ct. Elevated D-dimer was associated with older age. Elevated FM was associated with female gender. Elevated heparanase was independently associated with male gender. Decreased Procoag-PPL clotting time was associated with female gender. One out of four of COVID-19 survivors showed increase at least one biomarker of endothelial cell activation or hypercoagulability. Conclusion(s): Two months after onset of COVID-19, a significant activation of endothelial cells and in vivo thrombin generation persists in at least one out of four survivors of COVID-19. The clinical relevance of these biomarkers in the diagnosis and follow-up of patients with long COVID-19 merits to be evaluated in a prospective clinical study.

Rotational thromboelastometry in heparin monitoring in critical COVID-19 disease

Background: Critically-ill COVID-19 patients on UFH present with heparin resistance (defined as >35.000 IU/day) to reach target aPTT or aXa. APTT is the conventional method as aXa is time consuming and not widely available. APTT or aXa relate differently to doses of UFH, which questions their validity in COVID-19 disease. Whole-blood rotational thromboelastometry (ROTEM) INTEM/HEPTEM clotting time (CT) ratio could be a more functional alternative. Aim(s): Observational study comparing ROTEM INTEM/HEPTEM CT ratio, aPTT and anti-Xa in mechanically ventilated COVID-19 patients on UFH/LMWH. Method(s): We studied a subcohort (n = 106/232) of mechanically ventilated COVID-19 patients of the MaastrICCht cohort. We measured ROTEM, aPTT and aXa on UFH (n = 18) or therapeutic LMWH ( n = 88). Patients received UFH (ECMO/renal replacement therapy) or therapeutic LMWH (venous thromboembolism/cardiac arrhythmia). ROC and Linear regression analyses were assessed for diagnostic characteristics and associations. Result(s): INTEM/HEPTEM CT ratio was significantly higher in UFH-patients (1.4 [1.3-1.4]) compared to LMWH-patients (1.0 [1.0-1.1], p < 0.001) (Table 1). Furthermore, ROC analysis showed an AUC of 0.935 for INTEM/HEPTEM CT ratio, 0.775 for aPTT and 0.632 for aXa, suggesting that INTEM/HEPTEM CT ratio detects presence of UFH more accurately than aPTT and aXa. At an INTEM/HEPTEM CT cut-off of 1.1, sensitivity was 87.5% and specificity was 86.3% for the detection of UFH. Both APTT and aXa were positively associated with INTEM/HEPTEM CT ratio (beta(95%CI) 0.006(0.004-0.008) and 0.153 (0.071-0.234) respectively, Table 2).Only, the association between aXa and INTEM/HEPTEM CT ratio was significantly stronger (p = 0.006) in UFH-patients (beta(95%CI) 0.310 (0.001-0.619), compared to LMWH-patients (beta(95%CI) 0.094 (0.053-0.135). Conclusion(s): ROTEM INTEM/HEPTEM CT ratio can detect the presence of UFH opposed to the presence of therapeutic LMWH in mechanically ventilated COVID-19 patients. INTEM/HEPTEM CT ratio was dependent on aPTT, irrespective of UFH or LMWH use. The association between aXa and INTEM/HEPTEM CT ratio was significantly stronger in patients on UFH than LMWH. (Table Presented).

The assessment of ROTEM test in patients with critical form of COVID-19

Background: Patients with comorbidities could have severe COVID-19 outcomes. Thrombosis or hemorrhages were frequently reported during COVID-19 evolution Aims: The study evaluated the role of ROTEM in the identification of patients with a high risk of vascular complications Methods: We analyzed clinical, ROTEM, and laboratory data of 80 patients admitted to the intensive care unit (ICU) Department of our hospital, during 2020-2021. Result(s): Thrombosis was diagnosed in 26 cases (27.96%) and was more frequent in patients with severe pneumonia and ARDS, Chi-squared 3.55, p = 0.05. Hemorrhages were identified in 21 cases (22.58%) and were less frequent in patients with severe ARDS, Chi-squared = 4.67, p = 0.03. ROTEM analyses were focused on ExTEM, InTEM, and FibTEM clotting time (CT), clot amplitudes after five and ten minutes (A5/A10), maximum clot firmness (MCF), and maximum lysis (ML). In the ExTEM median CT (eCT) value was 90.5 (50-207) s in patients with severe ARDS vs 75(58-571) s in patients with mild/ medium ARDS, p = 0.02;A5 44 (16-68) in patient with thrombosis vs. 53 (12-69) in patient without thrombosis;ML 6 (0-104) in patients with sepsis vs 8 (0-15) in patient without sepsis, p = 0.04.The median FibTEM CT (fCT) was 91 (53-7913) s in patients with severe ARDS vs 76 (52-8203) s in patients with mild/medium ARDS, p = 0.009;there were a significant correlation with ferritin level, rho = 0.22, p = 0.05. An increasing value of fCT during COVID-19 evolution was observed especially in the critical form of COVID-19. Lymphopenia was significant in patient with severe ARDS 0.48 x 1000/uL (0.02-8.03) vs 0.67 x 1000/uL (0.14-9.09) identified in patients with mild/medium ARDS, p = 0.05;it was not obtained significant correlations between lab test and ROTEM results. Conclusion(s): The ROTEM test suggests showing hypercoagulability probably due to severe inflammation. Patients with a critical form of COVID-19 developed thrombotic complications more frequently than hemorrhages. We have to check these results in large cohorts of patients.

Does COVID Affect Clot Formation?

Introduction: Covid infection is associated with an increased rate of thrombosis, up to 50%,1 but the reasons for this are unclear. Rotem is a form of viscoelastic measurement of blood coagulation, allowing graphical representation of the time a clot takes to form (clotting time (CT) and clotting formation time (CFT)), its strength (firmness of the clot after 5 min (A5) and maximum clot firmness (MCF)), and dissolution (maximum lysis (ML). Rotem provides four analyses to each sample: Intem (measure of intrinsic pathway), Extem (extrinsic pathway), Heptem (similar to Intem, excluding heparin effects) and Fibtem (isolating fibrinogen function). In one study, Covid infection was associated with a reduced clotting time and an increased maximum clot firmness overall.2 The same study also suggested a possible increase in Fibtem MCF value for COVID patients, suggesting a strong influence of fibrinogen on the clot. This clinical effectiveness audit was designed to determine if (1) Covid patients in the Royal Surrey County Hospital also displayed increased clotting tendencies compared with patients without, as measured by Rotem and fibrinogen levels, and (2) whether fibrinogen levels correlate with Fibtem MCF levels. Methods: A retrospective data analysis from all Rotem analyses in ICU between June 2020 and February 2021 was performed. To compare coagulation in patients with and without Covid, data were tested for normality using Shapiro-Wilks testing. Normal data were compared by the Student t-test;non-normal data were compared by the Mann-Whitney U-test. A p-value < 0.05 was taken as statistically significant. In the second part, correlation between fibrinogen values and Fibtem MCF values was assessed using a Pearson correlation coefficient. Results: 163 patients had Rotem analyses performed, of whom 12 (7.4%) had a positive Covid test. The median Fibtem A5 in Covid patients was 21 mm compared with 13 mm in non-Covid patients, and median MCF was 25 mm compared with 16 mm, indicating a significant increase in the clot strength by the fibrin component between patients with and without Covid. No other Rotem differences were found. There was also a statistically significant difference between fibrinogen levels in COVID (median 6.2) and non-COVID (median 3.05) patients, suggesting a significant contribution of fibrinogen to clotting in COVID. Finally, the study demonstrated a strong positive correlation between fibrinogen levels, as measured by the laboratory, and corresponding Rotem MCF values in all patients (R2 = 0.8105), and in patients with Covid (R2 = 0.905). Discussion: There was a significant increase in the clot firmness due to the fibrin component, and fibrinogen levels, in patients with Covid infection, compared with those without, consistent with a previous study.2 Rotem measurement of clot strength by fibrin correlated very well with fibrinogen levels. This study suggests that the increase in thrombosis in Covid may be in part related to increased fibrinogen activity. The study may be limited by small numbers of patients, and the heterogeneity of the patients within each group.

Microcirculatory Predictors of Thrombosis in Patients After COVID-19

Introduction: The lack of a history of the course of a new coronavirus infection and the lack of data from randomised trials makes it difficult to choose the right treatment tactics and prescribe adequate prophylaxis in patients who have suffered from COVID-19. Comorbid patients with cardiovascular diseases and endothelial dysfunction have a high risk of a severe course of COVID-19 and subsequent thrombotic complications, which manifest clinically as cardiomyopathy;venous thrombo-embolism (deep vein thrombosis and pulmonary embolism);pulmonary thrombosis in situ;stroke;arterial thrombangiitis;rarely, arterial peripheral thrombosis and microvascular thrombosis, in the lungs, liver, kidneys, brain, etc.;and mild disseminated intravascular coagulation syndrome. The role of endothelial dysfunction in the development of severe complications is underestimated. In the pathogenesis of COVID-19, the defeat of the microcirculatory bed plays a crucial role. The SARS-CoV-2 virus causes associated endotheliitis damage to the endothelium due to virus entry and cytokine storm. Endotheliitis leads to the release of tissue factor, which leads to the formation of an excess of thrombin and fibrin;the body tries to cover the virus with these and prevent its spread, which entails negative side effect such as thrombosis Methods: Sixty-six patients who had COVID-19 were examined (42 women and 24 men;mean age 48 years [range 20 – 80 years]). Patients complained of a feeling of paraesthesia, mainly in the lower extremities, a feeling of heaviness, stiffness in the popliteal region, an increased vascular pattern on the entire surface of the skin, a burning sensation in all vessels, and a feeling of weakness. Ultrasound colour duplex scanning showed no signs of thrombosis in the large vessels. Using a high frequency ultrasound Doppler and a 25 MHz sensor, the nailbed of the first finger of the upper limb was examined. The microcirculatory images were analysed by the shape and spectrum of the curves. Twenty patients received prophylaxis with rivaroxaban 10 mg daily (group 1) and 46 patients did not (group 2). The control examination was carried out four weeks after the start of therapy: sulodexide one capsule twice daily. The coagulogram parameters were also studied. Results: A depletion in spectral characteristics was seen in patients after COVID-19 disease, in comparison to microcirculatory images recorded in healthy individuals. Predominantly, the red part of the spectrum was recorded in patients after COVID-19, the lighter part of the spectrum was not recorded. Group 1 patients had higher amplitude parameters than group 2, but they also registered a depletion in spectral characteristics. Soluble fibrin monomer complexes were increased 4 – 5 times, D-dimer 2 – 2.5 times, and antithrombin III 1.5 times. The international normalised ratio, activated partial thromboplastin time, fibrinogen, prothrombin according to Quick, prothrombin time, clotting time, and bleeding time were within the reference intervals both before and after treatment. Upon repeat examination four weeks after the course of sulodexide therapy, the spectral characteristics were normalised, and the coagulogram parameters were also normalised. Conclusion: The red part of the spectrum, according to the Doppler criteria, corresponds to the fastest particles moving in the middle of the stream. The lighter part of the spectrum corresponds to particles moving more slowly. The reduction in spectral characteristics in patients after COVID-19 disease corresponds to parietal stasis and readiness for thrombosis, which was confirmed by the coagulogram data. Examination of the nailbed using high frequency ultrasound Doppler in patients who have COVID-19 allows the identification of stasis of the parietal blood flow, which corresponds to a prethrombotic state. The prescription of sulodexide allows for an improvement in the condition of patients and normalisation of microcirculation indicators

Comparison of New Viscoelastic Coagulation Monitor with Standard of Care Thromboelastography for Anticoagulation Monitoring during 72-Hour Extracorporeal Life Support

Introduction: Management of coagulation remains the foremost challenge during extracorporeal life support (ECLS). Thromboelastography (TEG) and other viscoelastic clotting tests have shown utility for assessing coagulation status in trauma and ECLS patients and have also been utilized in COVID 19 patients. However, with few exceptions, these methods are performed in a laboratory setting, not at the bedside, and rely on cumbersome, non-portable equipment. The Viscoelastic Coagulation Monitor (VCM;Entegrion;Durham, NC) is a portable device/test developed for use at the bedside and outside hospitals to assess clot formation and lysis using a small sample of whole blood. Blood coagulation is activated by contact with the glass surface on the cartridge, and measurements are derived pertaining to clot formation, stability, and lysis - similar to metrics obtained by the TEG 5000 (Haemonetics;Boston, MA). In a recent study, the relationship of VCM results and heparin dose administered in 36 COVID-19 patients was investigated;however, use of VCM for ECLS with application of heparinase has not been reported. We investigated efficacy of the VCM for coagulation monitoring during 72 hours of continuous ECLS in swine and hypothesized that the VCM with heparinase correlates with TEG heparinase. Methods: Female Yorkshire swine (n=3, 53.4±1.6kg) were anesthetized, mechanically ventilated, and systemically heparinized. Blood samples were collected at baseline, post ECLS, 6, 24, 48, and 72-hours post ECLS initiation. For the VCM, 350μL of whole blood was added to a 0.05 IU heparinase vial, mixed, and then added to a VCM cartridge. For TEG, 340μL of citrated whole blood was added to 20μL 0.2 M CaCl2, and samples were activated with a kaolin reagent. Heparinase cups (Haemonetics;Boston, MA) were used for testing. Spearman correlation was performed to compare standard VCM metrics (clotting time [CT], clot formation time [CFT], alpha, maximum clot firmness [MCF], clot retraction/fibrinolysis [LI30]) to the respective TEG metrics (reaction time [R], clot formation time [K], alpha, maximum amplitude [MA], clot retraction/fibrinolysis [LY30]), and also to other conventional coagulation measurements such as prothrombin time (PT), activated partial thromboplastin time (aPTT), fibrinogen (FIB), and platelet count (PLT) for each timepoint. A p-value of 0.05 was used for significance. Results: All VCM metrics significantly correlated with the respective TEG measurements (see Table 1). Both VCM and TEG show the same positive and negative correlation relationships for clot formation time, clot kinetics, and clot retraction with conventional coagulation tests (see Table 2). Additionally, clotting time and maximum clot firmness did not show moderate or significant correlation with conventional tests. Prothrombin time did not correlate with any values. Conclusion: The VCM is comparable to TEG in assessing coagulation status in heparinized swine and can be used during austere care with ECLS application. In the next round of experiments, we will validate the VCM in clinically-relevant trauma with and without ECLS.

Evaluation of coagulation function by rotation thromboelastometry in critically ill patients with severe Covid-19 Pneumonia

Background-aim: Critically ill patients with COVID-19 pneumonia suffered both high thrombotic and bleeding risk. The effect of SARS-CoV-2 on coagulation and fibrinolysis is not well known. Methods: Retrospective cohort study including 84 patients, during 16 months, divided into two groups: patients with severe SARS-Cov-2 pneumonia (group 1, N=42) and patients with severe non-COVID-19 pneumonia (group 2, N=42). We evaluated coagulation standard parameters (hemoglobin, platelet count and conventional laboratory coagulation tests) in group 1 vs group 2 and coagulation standard parameters on day of admission (T0) and 10 (T10) days after admission to ICU and coagulation function using rotational thromboelastometry (ROTEM) in patients with severe SARS-Cov-2 pneumonia. Results: 84 patients were enrolled into the study. Similar results in conventional laboratory coagulation tests were detected in group 1 and group 2: prothrombin time (15.14s vs 14.76s, p=0.212), international normalized ratio (1.21 vs 1.19, p=0.112), activated partial thromboplastin time (32.17s vs 25.52s, p=0.06), fibrinogen level (6.15 mg/dl vs 3.39 mg/dl, p=0.208), hemoglobin (11.81 g/dl vs 11.20 g/dl, p=0.139) and platelet count (208.98x103/ul vs 288.74 x103/ul, p=0.123). However, a statistically significant difference was observed in the D-dimer count (2442.11 ng/ml vs 370 ng/ml, p=0.03). In addition, statistically significant increase in D-dimer count during Intensive Care Unit (ICU) stay (T0=2442.11 ng/ml vs T10=8564.39 ng/ml, p=0.000) in group 1 were detected. Finally, blood thromboelastometry profiles were consistent with hypercoagulability characterized by higher clot strength (MCF or maximum clot firmness close to upper limit in FIBTEM test, MCF median value= 25.9 mm). Clotting time presented normal results in INTEM (163.41 s) and EXTEM (68.74 s). No sign of secondary hyperfibrinolysis were found during the study period. In six patients a deep vein thrombosis and in six patients a thromboembolic event. Eighteen patients (43%) died during hospitalization due to coagulopathy produced by SARS-Cov-2 pneumonia. Conclusions: The results observed in our study support hypercoagulability in a severe inflammatory state, rather than a Consumption Coagulopathy (DIC) state. More studies are needed to better understanding of coagulopathy produced in patients with severe COVID-19 pneumonia.

Impaired fibrinolysis is implicated in mortality in COVID-19 infection

Introduction: A significant degree of mortality and morbidity in COVID-19 is due to thromboembolic disease. Changes in coagulation markers have been well described in critically unwell patients on ICU. There is less clear evidence regarding these changes at the time of presentation to the Emergency Department and the progression of disease over time. We sought to investigate how coagulation markers change over the course of COVID-19 infection and whether they might predict disease severity. Methods: Patients were recruited from a single University Teaching Hospital ED at the time of presentation. Those with a positive PCR test were followed up throughout their stay. Rotational thromboelastometry (ROTEM) was performed on arrival, after 24 h, 3-5 days and 7 days, alongside routine haematological and biochemical testing. ROTEM values at each of these time points were analysed, and compared. Length of stay and patient outcome were also recorded for subgroup analysis. The ROTEM parameters selected for analysis were both EXTEM and INTEM Clotting Time (CT), Clot Formation Time (CFT), Maximal Clot Firmness (MCF), Alpha Angle (Alpha) and Maximum Lysis Percentage (ML). This reflects clot formation kinetics, mechanical strength and clot breakdown via both extrinsic and intrinsic pathways. Results: EXTEM (7.64 ± 5.53 vs 11.83 ± 6.30) and INTEM ML (4.69 ± 3.55 vs 9.95 ± 5.22) were significantly reduced in those who died vs patients with a prolonged hospital stay. Over time there were no patterns of change to ROTEM values in any outcome group. Conclusions: Comparisons between groups demonstrated that one distinguishing feature between those who require ICU admission or die of COVID-19 compared with those who survive a prolonged hospital stay to discharge was the extent to which fibrinolysis could occur. Failure to break clots down could be a significant mechanism in the mortality and morbidity of COVID-19.

Vaccine-induced thrombotic thrombocytopenia: Novelty testing in the diagnostic workup?

COVID-19 vaccination campagnies with several vaccines types are currently undeway. Recently, the ASTRA ZENECA vaccine has raised public alarm with concerns regarding the development of thrombotic events known as vaccine-induced thrombotic thrombocytopenia (VITT). Early and limited studies have implicated an antibody-mediated platelet activation as the mechanism of the clotting events. Aim of this study was to investigate the platelet and coagulation activation using specialized tests. In this study we enrolled 60 patients (40 men, 20 women;mean age 55±10 years) without cardiovascular risk factors or a history of thrombosis who reported having poplitea deep vein thrombosis (35/60) and pulmonary embolism (25/60) revealed with lower-limb ultrasonography and computed tomography (CT) angiography, respectively, 7 days after vaccination with ASTRA ZENECA. All patients were evaluated for initial testing such as platelet count, prothrombin time (PT), activated partial thromboplastin time (APTT), fibrinogen (Fib) and D-dimer (DD). Platelets were measured by automated analyzer, PT and APTT by coagulometric test, Fib using Clauss method, and DD using ELISA. Complete blood hemostasis was studied by platelet function assay (PFA-100) on Collagen/ADP (CT-ADP) and Collagen/Epinephrine (CT-EPI) cartridges and Thromboelastometry method on Clotting Time (CT), Clotting Formation Time (CFT), Maximum Clot Firmness (MCF), and clot lysis at 30 minutes (LY-30). All patients had thrombocytopenia (60±5x109/L), longer PT (28±10 s) and PTT (50±10 s), lower Fib (80±20 mg/dl), higher DD ((550±100 mg/l). All patients had shorter C/ADP and C/EPI (C/ADP, n.v. 68-121 s (42±10 s) and C/EPI n.v. 84-160 s (38±5 s) and shorter CT (CT, unit: s. n.v. 100-240 s) (INTEM 30±20 s, EXTEM 18±10 s), shorter CFT (CFT, unit: s, n.v. 30-160 s (INTEM 11±10 s, EXTEM 19±10 s), longer MCF (MCF, unit: mm, n.v. 50-72 mm (INTEM 128±10 mm, EXTEM 110±10 mm), and lower LY-30 (LY-30, %: v.n. 15% (INTEM 0.8%, EXTEM 0.7%). These interesting findings may be the novelty in the diagnostic work-up of the VITT. If these tests may aid in the diagnosis of VITT deserve to be confirmed and need reproducing in other studies.

Prospective Assessment of Biomarkers of Hypercoagulability in Oncological Patients and Healthcare Workers Following Vaccination Against Sars-Cov-2 with the mRNA Vaccine. the Roadmap-COVID-19-Vaccin Study

Background: COVID-19 has been associated with hypercoagulability, endothelial cell injury and frequent thrombotic complications resulting both from direct effects of the virus on the endothelium and from the ‘cytokine storm’ resulting from the host's immune response. Since the COVID-19 vaccines have been shown to effectively prevent symptomatic infection including hospital admissions and severe disease, the risk of COVID-19-related thrombosis should be expected to (almost) disappear in vaccinated individuals. However, some rare cases of venous thrombosis have been reported in individuals vaccinated with mRNA vaccines. Thus, there is a sharp contrast between the clinical or experimental data reported in the literature on COVID-19 and on the rare thrombotic events observed after the vaccination with these vaccines. This phenomenon raised some scepticism of even some fear about the safety of these vaccines which could compromise the adhesion of the citizens in the vaccination program. Aims: We conducted a prospective observational study, to explore the impact of vaccination with the BNT162b2 (Pfizer/BioNTech) on blood hypercoagulability and endothelial cell activation and to investigate if this is modified by the presence of active cancer. Methods: In total 229 subjects were prospectively included in the study from April to June 2021. Subjects were stratified in three predefined groups: 127 vaccinated patients with active cancer (VOnco group), 72 vaccinated health care workers (VHcw group) and 30 non vaccinated health individuals (Control group). Blood samples were obtained 2 days after the administration of the first dose of BNT162b2 vaccine and collected in Vacutainer® tubes (0.109 mol/L trisodium citrate). Platelet poor plasma (PPP) was prepared by double centrifugation at 2000 g for 20 minutes at room temperature and plasma aliquots were stored at -80°C until assayed. Samples of PPP were assessed for thrombin generation (TG) with PPP-Reagent® (Thrombogram-Thrombinoscope assay with PPP-Reagent®TF 5pM), E-selectin, D-dimers, (D-Di), Tissue Factor (TFa), procoagulant phospholipid-dependent clotting time (Procag-PPL) and von Willebrand factor (vWF), thrombomodulin (TM), tissue factor pathway inhibitor (TFPI), and platelet factor 4 (PF4). All assays were from Diagnostica Stago (France). The upper and lower normal limits (UNL and LNL) for each biomarker were calculated by the mean±2SD for the control group. Results: All vaccinated subjects showed significantly increased levels of PF4 (71% >UNL, p<0.001), D-Dimers (74% >UNL, p<0.01), vWF (60% >UNL, p<0.01), FVIII (62% >UNL, p<0.01) and shorter Procoag-PPL clotting time (96% <LNL, p<0.001), as compared to controls. Thrombin generation showed significantly higher Peak (60% >UNL, p<0.01), ETP (38% >UNL, p<0.01) and MRI (66% >UNL, p<0.01) but no differences in lag-time in vaccinated subjects as compared to the control group. Vaccinated subjects did not show any increase at the levels of TFa, TFPI, TM and E-selectin in comparison with the control group. The studied biomarkers were not significantly different between the VOnco and VHcw groups. Conclusion: The ROADMAP-COVID-19-Vaccine study shows that administration of the first dose of the BNT162b2 vaccine induced significant platelet activation documented by shorter Procoag-PPL associated with increased levels of PF4. Plasma hypercoagulability was less frequent in vaccinated individuals whereas there was no evidence of significant endothelial cells activation after vaccination. Interestingly, the presence of active cancer was not associated with an enhancement of platelet activation, hypercoagulability, or endothelial cell activation after the vaccination. Probably, the generated antibodies against the spike protein or lead to platelet activation in a FcyRIIa dependent manner that results in PF4 release. The implication of the mild inflammatory reaction triggered by the vaccination could be another possible pathway leading to platelet activation. Nevertheless, vaccination does not provoke endothelial activation even n patients with cancer. The findings of the ROADMAP-COVID-19-Vaccine study support the concept administration of mRNA based vaccines does not directly cause a systematic hypercoagulability. Disclosures: Gligorov: Roche-Genentech: Research Funding;Novartis: Research Funding;Onxeo: Research Funding;Daichi: Research Funding;MSD: Research Funding;Eisai: Research Funding;Genomic Heatlh: Research Funding;Ipsen: Research Funding;Macrogenics: Research Funding;Pfizer: Research Funding. Terpos: Novartis: Honoraria;Janssen: Consultancy, Honoraria, Research Funding;Genesis: Consultancy, Honoraria, Research Funding;Celgene: Consultancy, Honoraria, Research Funding;BMS: Honoraria;Amgen: Consultancy, Honoraria, Research Funding;Takeda: Consultancy, Honoraria, Research Funding;Sanofi: Consultancy, Honoraria, Research Funding;GSK: Honoraria, Research Funding. Dimopoulos: Amgen: Honoraria;BMS: Honoraria;Janssen: Honoraria;Beigene: Honoraria;Takeda: Honoraria.

Novel monitoring and dose adjustment of argatroban to maintain therapeutic anticoagulation

INTRODUCTION: In patients with unreliable activated partial thromboplastin time (aPTT) measurements who require anticoagulation with a direct thrombin inhibitor (DTI), the only reliable alternative measurement at present is a dilute thrombin time (dTT). However, this assay is not always readily available, which limits accurate real-time dose adjustments necessary to maintain therapeutic anticoagulation. DESCRIPTION: A 57 year-old woman with a history of antiphospholipid antibody syndrome, heparin-induced thrombocytopenia, and multiple prior deep venous thromboses and pulmonary emboli was admitted with COVID-19 pneumonia and intubated due to hypoxic respiratory failure. Argatroban was initiated in place of her home medication warfarin. This patient had a prolonged aPTT value at baseline and overnight dTT assay measurements were limited at our institution. To overcome this challenge, a multidisciplinary team of hematology and pharmacy clinicians created a modified aPTT algorithm. By measuring aPTT and dTT simultaneously from patient plasma samples, a patientspecific aPTT target range was derived and argatroban dosing was titrated accordingly. Subsequent aPTT values in the modified target range corresponded to therapeutic dTT values, indicating therapeutic anticoagulation was successfully achieved and maintained. Patient plasma samples were also evaluated retrospectively using a novel point-of-care (POC) coagulation test to detect and quantify the effect of argatroban. A Clotting Time Score (CTS) was derived for each sample tested. Comparison of CTS and dTT values demonstrated moderate positive correlation between test results. All CTS results accurately reflected if argatroban dosing achieved an appropriate level of anticoagulation. DISCUSSION: Therapeutic anticoagulation with a DTI in a patient with unreliable aPTT measurements is challenging but can be achieved with use of a modified aPTT scale, which in our case study was retrospectively confirmed by corresponding dTT measurements. Early validation of a novel test that could offer a rapid, POC alternative to dTT when dTT measurements are necessary but not readily available is promising. Such a technology could dramatically improve rapid accurate titration of DTIs to maintain therapeutic anticoagulation.

Is endothelial dysfunction a driving force of COVID-19 induced coagulopathy?

Background/Introduction: There are numerous reports regarding the direct endothelial damage by the SARS-CoV-2 that can lead to activation of both plasma hemostasis and platelet aggregation. However, the mechanism of interaction between endothelium and haemostasis in COVID-19 remains unclear. Purpose: The aim of our study was to assess the relationship between each link of clot formation process (endothelial function, plasma coagulation, platelet aggregation) with the severity of the disease. Methods: 58 COVID-19 patients were included in our study. Patients were divided into moderate (n=39) and severe (n=18) subgroups. All patients underwent a flow-mediated dilation (FMD) test, impedance aggregation, rotational thromboelastometry, thrombodynamics and von Willebrand factor antigen (vWF: Ag) quantification. All measurements were repeated on days 3 (point 2) and 9 (point 3) of hospitalization. Results: COVID-19 patients demonstrated the enhanced plasma coagulation (clotting time, s 613,0 [480;820], clot growth rate, μm/min 32,75 [29,3;38,7]). At point 1 no significant difference in parameters of plasma coagulation between patients' subgroups was noted. At point 2 a significant decrease in the size (CS, μm 1278.0 [1216.5;1356.5] vs 965.0 [659.8;1098.0], p<0,01) and clot growth rate (μm/min 32,4 [29,2;35,0] vs 17,7 [10,3;24,4], p<0,01) under the influence of anticoagulants in the moderate subgroup compared with point 1 was observed. We didn't observe such phenomenon in severe subgroup. There was no significant difference in platelet aggregation between subgroups at point 1. During the course of the disease the patients in the moderate and severe subgroups demonstrated a significant increase in platelet aggregation induced by arachidonic acid and ADP (severe: AUC ARA 48,0 [25,0;59,0] vs 77,5 [55,8;92,7], p=0,04;AUC ADP 44,0 [41,0;56,0] vs 58,0 [45,5;69,0], p=0,04;moderate: AUC ARA 31,5 [19,8;50,7] vs 56,0 [39,0;76,0], p=0,01;AUC ADP 43,0 [20,0;59,0] vs 56,6 [50,3;70,5], p=0,04;), in moderate subgroup the significant increase in TRAP-induced aggregation was also noted (AUC TRAP 58,0 [41,0;69,5] vs 76,0 [58,3;81,5], p=0,048). There were no significant differences in the FMD-test results between the patient subgroups. FMD-test results were predominantly within the reference ranges (7,1 [4,0;8,8]). Patients in the severe subgroup had significantly higher levels of vWF: Ag (228,0 [205,3;240,7] vs 232,0 [226,0;423,0], p=0,03). Conclusion: SARS-CoV-2 infection was characterized by increased levels of vWF:Ag, that could represent the local endothelial damage, meanwhile there was no generalized endothelial dysfunction assessed via FMD-test in moderate to severe patients. At the same time the enhanced plasma coagulation in COVID-19 patients was observed.