Plasma Fibrinogen Independently Predicts Hypofibrinolysis in Severe COVID-19

High rates of thrombosis are present in patients with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Deeper insight into the prothrombotic state is essential to provide the best thromboprophylaxis care. Here, we aimed to explore associations among platelet indices, conventional hemostasis parameters, and viscoelastometry data. This pilot study included patients with severe COVID-19 (n = 21) and age-matched controls (n = 21). Each patient received 100 mg aspirin therapy at the time of blood sampling. Total platelet count, high immature platelet fraction (H-IPF), fibrinogen, D-dimer, Activated Partial Thromboplastin Time, von Willebrand factor antigen and von Willebrand factor ristocetin cofactor activity, plasminogen, and alpha2-antiplasmin were measured. To monitor the aspirin therapy, a platelet function test from hirudin anticoagulated whole blood was performed using the ASPI test by Multiplate analyser. High on-aspirin platelet reactivity (n = 8) was defined with an AUC > 40 cut-off value by ASPI tests. In addition, in vitro viscoelastometric tests were carried out using a ClotPro analyser in COVID-associated thromboembolic events (n = 8) (p = 0.071) nor the survival rate (p = 0.854) showed associations with high on-aspirin platelet reactivity status. The platelet count (p = 0.03), all subjects. COVID-19 patients presented with higher levels of inflammatory markers, compared with the controls, along with evidence of hypercoagulability by ClotPro. H-IPF (%) was significantly higher among non-survivors (n = 18) compared to survivors (p = 0.011), and a negative correlation (p = 0.002) was found between H-IPF and plasminogen level in the total population. The platelet count was significantly higher among patients with high on-aspirin platelet reactivity (p = 0.03). Neither the ECA-A10 (p = 0.008), and ECA-MCF (p = 0.016) were significantly higher, while the tPA-CFT (p < 0.001) was significantly lower among patients with high on-aspirin platelet reactivity. However, only fibrinogen proved to be an independent predictor of hypofibrinolysis in severe COVID-19 patients. In conclusion, a faster developing, more solid clot formation was observed in aspirin ‘non-responder’ COVID-19 patients. Therefore, an individually tailored thromboprophylaxis is needed to prevent thrombotic complications, particularly in the hypofibrinolytic cluster.

Immature Platelet Fraction in Thrombocytopenic Patients

Bone marrow puncture (BMP), is usually performed to identify thrombopoiesis activity and is still the gold standard to determine the etiology of thrombocytopenia. This diagnostic test is invasive hence it may cause discomfort to the patient. One of noninvasive test to determine the etiology of thrombocytopenia is by measuring immature platelet fraction (IPF). IPF is highly correlated to the activity of thrombopoiesis and by understanding the value, clinicians are able to use it in determining whether or not invasive examination is needed and more importantly avoiding unnecessary platelet transfusion. This research was a cross-sectional descriptive observational study aimed to evaluate IPF value in thrombocytopenic inpatients in the Department of Internal Medicine, Haji Adam Malik Hospital Medan. 83 people were recruited, 48 were female (57.83%) and 35 were male (42.16%). IPF values ranged 0,5-59,6% using Sysmex XN-1000 (reference range 1-4,8%). There were 5 (6.02%) low IPF values, 29 (34.93%) normal IPF values and 49 (59.03%) high IPF values. Evaluation of IPF in thrombocytopenic patients is a promising tool to discriminate central from peripheral thrombocytopenia.

Biomarkers for Thrombosis in COVID-19: A Role for High Sensitivity Troponin-I and Immature Platelet Fraction?

Biomarkers for Thrombosis in COVID-19: A Role for High Sensitivity Troponin-I and Immature Platelet Fraction? Introduction Coronavirus disease 2019 (COVID-19) is associated with increased risk of thrombosis with both venous and arterial thromboembolism observed. While d-dimer elevation has been shown to be associated with thrombosis, this elevation is present in over 50% of COVID-19 infections demonstrating a clear need for more specific biomarkers of thrombosis in this population. While there are a variety of theories to explain the increased risk for thrombosis: all center on Virchow's triad, specifically hypercoagulability and inflammation. Platelets play a significant role in hypercoagulability. Immature platelets, which are thought to be hyper-reactive, may specifically be associated with thrombosis in COVID-19. It would thus be reasonable to expect immature platelet fraction (IPF) and immature platelet count (IPC) to be predictive biomarkers of thrombosis in this population. Beyond hypercoagulability, High-Sensitivity (HS) Cardiac troponin-I can be a biomarker of inflammation and may also be predictive of thrombotic events in COVID-19. The aim of this study was to evaluate the relationship between IPF, IPC, HS cardiac troponin-I and thrombotic events in COVID-19. Methods Using a single center COVID-19 data registry, we extracted all patients with COVID-19 at our single center between May 1, 2020 and January 1, 2021. Patients were stratified into two groups based on thrombotic events during hospitalization, the thrombosis and no thrombosis groups. Biomarker values, including IPF, IPC, platelet counts, d-dimer, and HS cardiac troponin I were extracted. Two-sided Wilcoxon rank test was conducted to test group differences in IPF, IPC, platelet, d-dimer, and HS cardiac troponin-I values. Results There were no significant differences in measurements of IPF at admission, peak IPF , platelet count at admission, peak platelet count, IPC at admission, and peak IPC between the thrombosis and no thrombosis groups. Minimum platelet count values were significantly lower in the thrombosis group compared to the no thrombosis group. D-dimer and troponin values were significantly higher in the thrombosis group than the no thrombosis group. (Table 1) Discussion To our knowledge this is the first study assessing the relationship between IPF, IPC, HS cardiac troponin-I and thrombosis in COVID-19. HS cardiac troponin did appear to be a predictive biomarker for thrombosis in COVID-19. This may be related to vascular inflammation playing a significant role in thrombosis. It may also be secondary to myocardial inflammation associated with severe disease in COVID-19.3 Patients with more severe disease are more prone to thrombosis. Unsurprisingly, our study corroborates evidence in the literature that d-dimer is associated with thrombotic events in COVID-19. On the other hand, IPF and IPC do not appear to be predictive biomarkers for thrombosis in this cohort. This appears consistent with the limited data assessing the relationship between IPF, IPC, and thrombosis outside of COVID-19. This does not dispel the importance of immature platelets in COVID-19, however. IPF and IPC are increased in patients with COVID-19, and our published data indicates they are predictors of COVID-19 severity. However, the relationship between immature platelets and outcomes in acute illness can be complex, as in sepsis, where the trend of IPC is associated with mortality, rather than the initial value. Future studies should delineate the relationship between trends in IPF or IPC and outcomes in COVID-19. Furthermore, it is crucial to define biomarkers of thrombosis and disease severity and mortality in COVID-19 which can potentially guide therapeutic interventions. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

Immature Platelets and Disease Severity in Patients Hospitalized with Acute COVID-19 Who Are Vaccinated Against COVID-19

Introduction: COVID-19 has killed more than four million people worldwide and resulted in strained health resources globally(Dong Lancet ID 2020). There is a critical need for prognostic biomarkers to predict severity and outcomes in COVID-19 patients to allow more efficient resource allocation. Studies using pre-vaccination hospitalized patient data have demonstrated that elevated initial and peak immature platelet fraction (IPF), as well as related platelet indices, were associated with progression to severe COVID-19 outcomes (Welder Br J Haem 2021). This suggests the potential role of immature platelets as a cost-effective biomarker. The underlying pathophysiology of immature platelets in COVID-19 is unclear but these results support the hypothesis of higher inflammatory response, leading to thrombopoiesis mediated by pro-inflammatory cytokines and platelet hyper-reactivity in this population (Manne Blood 2021). It is unclear if this holds true in patients vaccinated against COVID-19. This study aims to assess the relationship between immature platelets in patients vaccinated against COVID-19 and hospitalized with acute COVID-19. Methods: This study used a COVID-19 patient registry established by the University of Texas Southwestern that comprises patients between May 2020 to July 2021. The database was approved by the Institutional Review Board. The study included 22 fully vaccinated adult patients with the mRNA COVID-19 vaccines hospitalized with acute COVID-19 and had IPF measurements during the hospitalization as well as 519 non-intensive care unit (ICU) patients admitted with acute COVID-19 prior to availability of a COVID-19 vaccine (pre-vaccination era). The study conducted non-parametric tests to compare hospital outcomes and covariates of interest between vaccinated patients and pre-vaccination era non-ICU patients. Covariates included for analysis are age, gender, race, length of hospital stay (LOS), dexamethasone use, platelet count, immature platelet count (IPC), IPF, thrombotic events and mortality. Results: All patients vaccinated against COVID-19 were alive without thrombotic events at the time of analysis. One out of 22 vaccinated patients required ICU admission and use of a ventilator. IPF and platelet counts at admission were similar between vaccinated patients and non-ICU patients from pre-vaccine era while IPC was significantly lower in the vaccinated group (Table 1). The vaccinated patient requiring ICU admission and mechanical ventilation was a heavy tobacco user with chronic obstructive pulmonary disease (IPF 6.1%, IPC 4x10 9/L, platelet count 66x10 9/L). There was a disproportionate number of Hispanic patients in the vaccinated cohort (44%, P = 0.02). The LOS was significantly shorter in vaccinated patients compared to pre-vaccination era non-ICU patients by a day and a half (P = 0.047). The admission IPF and IPC were not correlated with increased LOS (IPF Spearman ρ = 0.23, P = 0.31; IPC ρ = -0.34, P = 0.13), while platelet count at admission negatively correlated with LOS (ρ = -0.41, P = 0.06). Discussion: IPF, IPC, and platelet count have previously been demonstrated to be a predictor of increased ICU and hospital LOS, ventilator duration, and in-hospital mortality. To our knowledge, this is the first study assessing the relationship between IPF and platelet indices in hospitalized patients vaccinated against COVID-19. Due to the lack of severe COVID-19 outcomes in these vaccinated patients, LOS was the only variable able to be analyzed and lower platelet count was found to be associated with increased LOS . These preliminary results demonstrated similar initial IPF and platelet counts but lower IPC in a small cohort of vaccinated COVID-19 patients compared with the pre-vaccination era patients with no severe outcomes. This suggests that the predictive value of these biomarkers may also apply to the vaccinated patient population. As IPC in this current study is derived from IPF and platelet counts, independent measure of IPC is needed to confirm this finding in a larger cohort. This study also potentially suggests the protective benefits of COVID-19 vaccines as reported in prior randomized trials (Polack NEJM 2020). Further research is needed to confirm these findings in a larger vaccinated cohort assessing severe outcomes, hospitalization, and death, especially with future infection waves with contagious COVID-19 variants rapidly emerging. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.

Immature Platelet Fraction Does Not Correlate with Treatment Response in Immune Thrombocytopenia

Background: As the treatment options for immune thrombocytopenia (ITP) continue to expand, the choice of which treatment to give to an individual patient has become increasingly complex. Therefore, a laboratory marker to help guide treatment selection would be clinically useful. The immature platelet fraction (IPF), measured clinically by automated Sysmex hematologic analyzers, correlates with bone marrow thrombopoietic activity and may correlate with disease activity in patients with ITP. Objective: To investigate the relationship between pre-treatment immature platelet fraction (IPF) and treatment response among pediatric patients with ITP. Methods: This is an observational cohort study of 148 patients with ITP who received ITP-directed treatment as monotherapy at 3 tertiary academic children's hospitals. Eligibility included a clinical diagnosis of ITP, ITP-directed surgical or pharmacologic treatment given as monotherapy, and available pre-treatment IPF values. The Sysmex XN-series measures IPF by adopted fluorescence flow cytometry using a semiconductor dioxide laser to measure platelets stained with oxazine fluorescent dyes. Demographic and clinical characteristics, laboratory studies, and treatments were collected. Response to treatment was defined as a platelet count ≥30 x 10 9/L and at least 2-fold increase from the baseline platelet count within 3 weeks of first dose of IVIG, Rh(D) immune globulin, or corticosteroid or within 3 months of all other therapies. For second-line treatments, platelet counts within 1 month after a rescue therapy were excluded. Regression analysis was utilized to estimate association between variables; estimated coefficients and p values are reported. Results: The cohort included 148 patients, 52% (n=77) of whom were female. Median age of diagnosis was 8 years (IQR: 3-13). Twenty percent (n=29) of treated patients had secondary ITP including 9 (6%) with Evans syndrome. Median platelet count at time of diagnosis was 5 x 10 9/L, and median IPF at diagnosis was 16.7% (IQR: 7.7-25.8). Median pre-treatment platelet count was 17 x 10 9/L with a median pre-treatment IPF of 16.6% (IQR: 10.0-25.7). There was a significant association between pre-treatment platelet count and pre-treatment IPF (coefficient -0.176, p = 0.003). Increased variation in IPF was seen at lower platelet counts compared to higher platelet counts (p=0.014, Figure 1a). IPF at diagnosis and pre-treatment IPF were not correlated with platelet response to treatment overall (p=0.28 for pre-treatment IPF, p=0.31 for IPF at diagnosis). IPF prior to treatment did not correlate with platelet response to individual medications: IVIG (coefficient 0.001, p=0.78, n=64), corticosteroids (coefficient 0.007, p=0.28, n=43), Rituximab (coefficient -0.01, p=0.52, n=10), and thrombopoietin receptor agonists (coefficient 0.0006, p=0.93, n=26, Figure 1b). Similarly, IPF at diagnosis of ITP did not correlate with platelet response to individual medications. Conclusions: Pre-treatment platelet count and IPF are inversely correlated in children with ITP, although there is significant variability in IPF at low platelet counts. The IPF at ITP diagnosis and prior to treatment does not correlate with platelet response to the most common ITP-directed treatments in children. A normal or elevated IPF should not impact decision-making about initiation of any specific ITP-directed treatments, including thrombopoietin receptor agonists or immunosuppressive therapy, in children with ITP. Figure 1 Figure 1. Disclosures Lambert: Principia: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Bayer: Consultancy; Sysmex: Research Funding; Astra Zeneca: Research Funding; Dova: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; PDSA: Research Funding; Octapharma: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Novartis: Consultancy, Honoraria, Research Funding; ClinGen, ISTH, ASH, GW University: Honoraria; Rigel: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Argenx: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Shionogi: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees. Despotovic: Apellis: Consultancy; UpToDate: Patents & Royalties: Royalties; Novartis: Consultancy, Research Funding; Agios: Consultancy. Kirk: Biomarin: Honoraria. Grace: Agios: Research Funding; Novartis: Research Funding; Dova: Membership on an entity's Board of Directors or advisory committees, Research Funding; Principia: Membership on an entity's Board of Directors or advisory committees.

An Analysis of Immature Platelet Fraction Values

Introduction Immune thrombocytopenia (ITP) in children typically manifests with acute, severe thrombocytopenia leading to variable mucocutaneous and tissue bleeding. Despite gaining insight into disease biology, there is no confirmatory diagnostic test, leaving the clinician to exclude other causes of thrombocytopenia through clinical and laboratory findings. The immature platelet fraction (IPF) is determined by hematologic analyzers via platelet size and RNA content and serves as a measure of bone marrow thrombocyte production. A higher value indicates increased thrombopoiesis. While data exists regarding the validity of IPF as a diagnostic tool for differentiating hypo-productive and consumptive etiologies of thrombocytopenia in the adult population, there is a paucity of data regarding its use in the pediatric population. Objectives This study set out to determine how the IPF value may be utilized to support the diagnosis of ITP among pediatric patients. Methods This is a retrospective cohort study at a single tertiary care children's hospital. The electronic medical record (EMR) system was reviewed to include all patients who had an IPF resulted from January 1, 2016 through December 31, 2021. The investigators utilized REDCap ® (Research Electronic Data Capture) to create a database of pediatric patients with thrombocytopenia including their laboratory findings and final diagnoses. Patients were classified as either ITP or non-ITP. Characteristics of ITP and non-ITP cohorts were compared using Wilcoxon rank sum test and Pearson Chi-square test. An ROC-AUC analysis was conducted to identify the IPF value which best predicted if the patient had ITP or not with a 95% confidence interval (CI). The optimal cutoff value was estimated using Youden's index. Sensitivity and specificity were estimated for the cutoff value. Univariable logistic regression was then used to estimate associations between patient characteristics and diagnosis of ITP. Multivariable logistic regression was used to estimate the association between binary IPF and ITP adjusting for other predictors. Statistical significance was assessed at the 0.05 level. Results Demographics shown in Table 1 are stratified by patients who had their IPF within 30 days of diagnosis of thrombocytopenia and those who did not. The IPF was significantly higher at presentation in all patients with ITP than patients without ITP (median IPF 12.8% vs 10.5%, P=0.018). An optimal IPF cutoff point for ITP vs. not ITP among all patients was determined to be 5.4% with a sensitivity of 0.85, specificity of 0.39, and AUC of 0.623 (95% CI: 0.527, 0.718). The platelet count for patients with ITP was significantly lower at presentation than in patients without ITP (median 16 x 10 9/L vs 86 x 10 9/L, P&lt;0.001). Logistic regression modeling using IPF as a binary variable, either &gt;5.4% or not, showed that IPF &gt;5.4% is significantly associated with ITP (P&lt;0.001). Platelet count at presentation was also significantly associated with a diagnosis of ITP (P&lt;0.001) and a single unit increase in platelet count decreased odds of ITP by a factor 0.98 (95% CI: 0.97, 0.99). After adjusting for platelet count, IPF &gt;5.4% was still significantly associated with ITP (P=0.003). Those with an IPF &gt;5.4% were 3.34 times more likely to have ITP than non-ITP (95% CI: 1.49, 7.31). When the population was adjusted to include only patients with a platelet count of &lt;100 x 10 9/L at presentation and those that had an IPF obtained within 30 days, the optimal cutoff point for IPF was 15% with a sensitivity of 0.46, specificity of 0.80 and AUC of 0.58 (95% CI: 0.458, 0.719). Conclusion The data demonstrates that in thrombocytopenic patients, a high IPF (&gt;5.4%) is associated with ITP, but the sensitivity for predicting ITP is low. Figure 1 Figure 1. Disclosures Despotovic: Apellis: Consultancy; Agios: Consultancy; Novartis: Consultancy, Research Funding; UpToDate: Patents & Royalties: Royalties. Kirk: Biomarin: Honoraria.

Unique characteristics of new complete blood count parameters, the Immature Platelet Fraction and the Immature Platelet Fraction Count, in dengue patients

The advanced platelet parameters Immature Platelet Fraction and Immature Platelet Fraction Count have been implemented in clinical practice as measures of thrombopoietic activity, mainly in hematologic disorders that cause thrombocytopenia. The purpose of this observational study was to examine thrombopoiesis as reflected by these 2 new CBC parameters in patients infected with dengue. The study was conducted in infectious disease referral hospital in Metro Manila, the Philippines. We enrolled hospitalized patients at admission who were diagnosed with acute dengue or community acquired bacterial infection (CABI). Immature Platelet Fraction (IPF) and Immature Platelet Fraction Count were evaluated at admission and during hospitalization. A total of 606 patients were enrolled from May 1, 2017 to June 1, 2018. The participants consisted of 152 patients with dengue infection, 180 confirmed CABI, and 274 suspected CABI patients. At admission, the percent IPF (IPF%) of the patients with dengue was significantly higher than that of the confirmed CABI patients (median 3.7% versus 1.9%; p <0.001). In a time course evaluation, there was no significant difference of IPF% between the patients with dengue infection and the confirmed CABI patients in the febrile phase (median 1.9% versus 2.4%; p = 0.488), however, the IPF% of the patients with dengue infection increased to be significantly higher than that of the confirmed CABI patients in the critical phase (median 5.2% versus 2.2%; p <0.001). Our study elucidated the unique characteristics and time-course trends of IPF percent and number (IPF#) in the patients with dengue infection. IPF% and IPF# are potentially valuable parameters in dengue and further investigation is required for the optimal use in clinical practice.