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.

Analysis of Preplatelets and Their Barbell Platelet Derivatives by Imaging Flow Cytometry

Circulating large ″preplatelets″ undergo fission via barbell platelet intermediates into two smaller, mature platelets. In this study, we determine whether preplatelets and/or barbells are equivalent to reticulated/immature platelets by using ImageStream flow cytometry (ISFC) and super-resolution microscopy. Immature platelets, preplatelets and barbells were quantified in healthy and thrombocytopenic mice, healthy human volunteers, and patients with immune thrombocytopenia (ITP) or undergoing chemotherapy. Preplatelets and barbells were 1.9%±0.18/1.7%±0.48 (n=6) and 3.3%-+1.6/0.5%±0.27 (n=12) of total platelet counts in murine and human whole blood, respectively. Both preplatelets and barbells exhibited high expression of HLA-I with high thiazole orange and mitotracker fluorescence. Tracking dye experiments confirmed that preplatelets transform into barbells and undergo fission ex vivo to increase platelet counts, with dependence upon the cytoskeleton and normal mitochondrial respiration. Samples from antibody-induced thrombocytopenia in mice and patients with ITP had increased levels of both preplatelets and barbells correlating with immature platelet levels. Furthermore, barbells were absent post-chemotherapy in patients. In mice, in vivo biotinylation confirmed that barbells, but not all large platelets, were immature. This study demonstrates that a subpopulation of large platelets are immature preplatelets that can transform into barbells and undergo fission during maturation.

Immature Platelets and Risk of Cardiovascular Events among Patients with Ischemic Heart Disease: A Systematic Review

Background Immature platelets are larger and may be more thrombogenic than mature platelets. This systematic review included studies on the association between mean platelet volume (MPV), immature platelet count (IPC), and immature platelet fraction (IPF) and the risk of major cardiovascular events (MACEs) in patients with acute coronary syndrome (ACS) or stable coronary artery disease (CAD). Methods The literature search included studies in PubMed, Embase, Web of Science, and Cochrane Library. The review was performed according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) statement. Effect estimates that included multivariate adjusted odds ratios, relative risks, or hazard ratios were extracted. Results Forty-two studies were identified. High MPV was positively associated with MACE in 20 of 26 studies of patients with ACS, four of five studies in patients with stable CAD, and in all six studies comprising a combined population with ACS and stable CAD. Using continuous models of MPV in patients with ACS, effect estimates varied from 0.90 (95% confidence interval [CI]: 0.95–1.03) to 1.66 (95% CI: 1.32–2.09). The strength of these associations was broadly similar among patients with stable CAD and in combined populations. Five studies investigated IPC or IPF as exposures and all reported positive associations with MACE among patients with ACS, stable CAD, or in combined populations. Conclusion This review demonstrated clear evidence for positive associations between measures of immature platelets and subsequent risk of MACE in acute and stable ischemic heart disease patients.