BACKGROUND AND OBJECTIVES:Liquid plasma (LQP) stands out as an alternative to thawed plasma (TP) for emergent transfusions due to its longer shelf-life. We aimed to measure fibrinogen, protein C (PC), protein S (PS), factor V (FV), factor VII (FVII) and factor VIII (FVIII) activity in LQP, quantify how these factors' levels change during storage and characterize how they compare in LQP and TP. MATERIALS AND METHODS:Coagulation factor activities were measured on days 15, 26 and 27 for LQP (n = 26) and Day 5 for TP (n = 31). Bayesian statistics was used to compare coagulation factor activity and quantify changes in activity during storage. RESULTS:Fibrinogen and PC activity on Day 26 in LQP (LQP26) was comparable to that on Day 5 in TP (TP5) with posterior mean activity of 257 versus 246 mg/dL and 100.4% versus 108.7%, respectively. FV, FVII and FVIII had lower activity in LQP26 compared to TP5, with posterior mean activities of 42.6% versus 72.0%, 55.0% versus 59.7% and 48.8% versus 59.2%, respectively. PS in LQP26 was low, with posterior mean activity of 28.0%, which was less than half that of TP5 at 66.4%. From Day 15 to Day 26, FVII in LQP decreased at a rate of 3.49% per day, whereas fibrinogen, PC, PS, FV and FVIII activity in LQP remained relatively stable. CONCLUSION:LQP26 has comparable activities of fibrinogen, PC and FVII as TP5, lower activities of FV and PS and slightly lower activity of FVIII. LQP is a viable alternative for use in emergency transfusions and massive transfusion protocols.

Inherited platelet disorders (IPDs) are a heterogeneous group of conditions that present significant challenges in diagnosis and management. Here, we report two cases of patients presenting with clinically significant bleeding but with unclear etiologies by conventional clinical laboratory testing. Further evaluation, utilizing a combination of high-dimensional multiplexed mass cytometry and genetic sequencing, revealed the underlying causes of bleeding in both cases, leading to definitive diagnoses. These cases underscore the potential utility of combined multimodal approaches in evaluating patients with bleeding disorders. Moreover, these high-parameter methods can offer substantial mechanistic insights and can enhance our understanding of the molecular pathogenesis of IPDs. Future studies involving larger patient cohorts are needed to further validate this strategy, directly comparing its diagnostic yield and accuracy with current clinical laboratory testing approaches, which can ultimately improve patient care.

The Heidenhain variant of Creutzfeld-Jakob disease (CJD) is a rare form that initially presents with visual disturbances. In early stages, the presentation can mimic neuromyelitis optica spectrum disorders (NMOSD) and lead to unnecessary treatment modalities. Herein, we describe a case of a 66-year-old man who presented with bilateral vision loss and retro-orbital discomfort. In addition to immunosuppressive therapy, he received 4 rounds of therapeutic plasma exchange after his preliminary diagnosis of NMOSD. We were surprised to note that his condition did not show improvement but deteriorated, with severe neurocognitive symptoms. Eventually, CJD was suspected, and real-time quaking-induced conversion (RT-QuIC) was performed. By the time the diagnosis of Heidenhain variant of CJD was confirmed, the patient was discharged to hospice care and died shortly after.

Abstract Introduction Myeloid neoplasms (MNs) frequently harbor pathogenic mutations not detected by karyotyping and fluorescence in situ hybridization; hence, next‐generation sequencing (NGS) is necessary for diagnosis, risk stratification, and therapy. If, however, NGS is not clinically indicated but still performed, the results may promote futile avenues of investigation, heighten patient distress, and increase cost. Methods We created criteria to approve NGS testing for MN (MN‐NGS) with the goal of maximizing actionable results. These actionable results include making a new MN diagnosis, characterizing a MN with baseline mutational status, and altering treatment plans. Approval criteria included clinical suspicion of new, relapsed, or worsening disease and end‐of‐induction chemotherapy. Cancellation criteria included the suspicion of non‐myeloid disease only, no suspicion of progression of a known MN, no evidence for recurrence post‐transplant, a diagnosis of chronic myeloid leukemia, and cases using blood when a concurrent bone marrow NGS is being performed. We applied these criteria to NGS tests ordered at our institution between August and December 2018 and determined whether any tests meeting our cancelation criteria yielded actionable results. Results Consecutive MN‐NGS orders (n = 174) were retrospectively categorized as appropriate (Group A, n = 115), inappropriate (Group B, n = 29), and appropriately canceled (group C, n = 30). Seventy‐five of the 115 (65%) Group A tests and none of the 29 (0%) Group B tests yielded actionable results (p < 0.0001). Conclusion Approximately one third (59/174) of MN‐NGS test orders can be safely canceled using these criteria, which would result in $150,370 of Centers for Medicare and Medicaid Services‐reimbursed savings annually.

TP53 mutations in patients with AML and MDS frequently portend a poor prognosis, related to both p53 allele status and blast count. In 2022, the ICC and WHO released updated guidelines for classifying p53-mutated AML/MDS. The characteristics of p53 mutations, their associated co-mutations, and their effects on overall survival (OS) are not known in the context of these new guidelines. A retrospective chart review was undertaken for all patients with AML or MDS and at least one TP53 mutation detected on next generation sequencing (NGS) at Yale New Haven Hospital from 2015 to 2023. All patients (N = 210) met criteria for one of the 5 diagnostic classes based on WHO and ICC guidelines. Kaplan-Meier curves with associated log-rank testing and Cox proportional hazards model quantified the effects of clinical and molecular data on survival. Multi-hit pathogenic mutations were related to poorer OS in MDS but not AML using either the WHO (p = .02) or the ICC (p = .01) diagnostic criteria. The most significant predictors of OS in the sample overall were platelet count < 50 K (HR: 2.01, 95

Summary The thrombotic risk with haemoglobin C trait (HbAC) or haemoglobin C disease (HbCC) is unclear. However, individuals with HbCC have demonstrated chronic haemolysis, higher blood viscosity and altered rheology when compared to individuals with wild‐type haemoglobin (HbAA). These physiological alterations may theoretically translate to increased risk of thrombosis; therefore, a systematic literature review was performed to investigate the possible association between HbAC and/or HbCC and thrombosis. Twenty‐two studies met inclusion criteria representing 782 individuals with HbAC ( n = 694) or HbCC ( n = 88). Fifteen studies described the presence/absence of venous thromboembolism (VTE) in patients with HbAC ( n = 685) or HbCC ( n = 79), while seven studies described patients with HbAC ( n = 9) or HbCC ( n = 9) and arterial thrombosis. Most ( n = 20) studies were case reports or case series; however, two studies suggested a potential increased VTE risk with HbAC compared to HbAA in (i) all patients (OR 2.2, 95% CI: 0.9–5.5) and in (ii) pregnant individuals (RR 3.7, 95% CI 0.9–16). This review is the largest assessment of patients with HbC trait or disease and thrombosis to date; despite its limitations, the findings suggest HbC may be a predisposing risk factor to thrombosis. Prospective cohort studies are warranted to definitively elucidate the risk of thrombosis in this population.

Abstract Introduction Hematocrits of red blood cell (RBC) units are generally not measured by blood centers or transfusion services. Red blood cell exchange (RCE) is commonly used to treat sickle cell disease, and a key parameter needed in RCE is the average hematocrit of RBC units. However, there is no consensus on the average hematocrit to use or how it is affected by storage. The goal of our study is to determine the average hematocrit of RBC units by measuring hematologic parameters RBC units during storage and performing statistical modeling of hematocrit changes over time. Methods We sampled aliquots of leukoreduced RBC units in AS-1 (n=10) using the Terumo TSCD-II Sterile Tubing Welder to sterilely collect ~15mL of RBCs in transfer-pack containers. Aliquots were collected weekly from Day 7 to Day 42 of storage. Complete blood counts were performed using the Sysmex XN-3100 Hematology Analyzer. We statistically modeled changes in RBC units during storage by utilizing Bayesian multilevel linear regression models, allowing for capturing effects at the individual RBC-unit level. Posterior distributions of model parameters were estimated and processed using Stan and the rethinking and rstan packages in R. Posterior distributions are summarized with posterior means and 95%-credible intervals in parentheses. Model selection was done by calculating Akaike weights using the widely applicable information criterion (WAIC) to signify statistical support for each model. Results Multilevel linear regression models of hematologic parameters as a function of storage time were developed. Hematocrits increased with a posterior mean of 1.4% (0.98–1.75%) per week of storage, with posterior mean hematocrits of: 61.9% (60.2–63.6%) on Day 7, 63.9% (61.5–66.4%) on Day 21, and 66.0% (62.8–69.7%) on Day 42. These results indicate that a single average cannot be used for RBC units since hematocrits dramatically increase during storage. Mean corpuscular volume (MCV) increased with a posterior mean of 2.31 fL (1.4–3.71) per week whereas hemoglobin remained stable with a posterior mean increase of 0.008 g/dL (-0.016–0.031) per week. Using these results, we subsequently developed a multilevel regression model of hematocrit as a function of MCV. The model using MCV (WAIC=100.2, weight=100%) was favored over the model using time (WAIC=120.0, weight=0%), suggesting that changes in MCV are driving the increasing hematocrits. Conclusion Our study is the first to leverage Bayesian statistics to carefully quantify the average hematocrits of RBC units during storage time and to demonstrate that increasing hematocrits are caused by changes in MCV. These results indicate that transfusion protocols must consider the age of RBC units for RCE calculations, and provisions (e.g., requiring use of fresh units for RCE) may help minimize the variability in post-exchange testing. Moreover, our findings suggest that transfusion services should regularly measure hematocrits of RBC units in their inventory.

Abstract Introduction Red blood cells (RBCs) express anionic phospholipids such as phosphatidylserine (PS) in their inner cell membrane layer. As RBCs age, physiochemical changes in their cell membranes, particularly exposure of PS on the outer membrane layer, lead to phagocytosis and clearance from circulation. PS in RBCs induces activation of the complement system, increasing binding of C3 to RBCs. Thus, quantifying PS-expressing (PS+) RBCs and complement deposition on RBCs (C3+) is a potential measure of RBC injury and may improve transfusion quality when used for predicting post-transfusion RBC recovery. Our goal was to develop a flow cytometry assay that quantifies PS+ and C3+RBCs and to assess how this assay correlates with the direct antiglobulin test for C3 (C3-DAT). Methods Using the Cytoflex S flow cytometer, we measured PS+ and C3+ RBCs using anti-annexin-PE (PS+RBC) and biotin-labeled anti-C3 with streptravidin-PE-Cy5 (C3+RBC). RBCs from healthy donors (NL-RBCs) with negative direct antiglobulin tests for C3 (C3-DAT) (n = 4) were analyzed. To assess the sensitivity of the assay and correlation with C3-DATs, complement control cells (CCC; Immucor) were used as a positive control for both PS+RBCs and C3+RBCs and diluted to different concentrations by mixing CCCs with NL-RBCs. Aliquots of diluted RBCs were analyzed blinded for C3-DAT. Differences between means for PS+RBC and C3+RBC were compared using two-sample t-test, and one-way ANOVA and Tukey’s HSD test were used to determine differences in C3+RBCs among a C3-DAT range of reactivities (0+ to 3+) in diluted CCC samples. P values less than 0.05 were statistically significant. Results In healthy, DAT-negative individuals, the mean positivity for PS+RBCs was 0.28% and for C3+RBCs was 0.34%, indicating that healthy individuals have very low percentages of RBCs with PS and C3 on their cell membrane surface. In CCC diluted samples, the mean C3+RBC percentage was 1.4% in negative C3-DAT samples, 5.9% in weak+ C3-DAT samples, 28.1% in 1+ C3-DAT samples, 67.2% in 2+ C3-DAT samples, and 99.0% in 3+ C3-DAT samples. One-way ANOVA revealed a statistically significant difference in C3+RBCs amongst all C3-DAT reactivities in diluted CCC samples (P<0.001), and Tukey’s HSD test found a mean difference in C3+RBCs percentage of 26.7% between 0+ and 1+ samples (P<0.001), 39.2% between 1+ and 2+ samples (P<0.001), and 31.8% between 2+ and 3+ samples (P<0.01). Undiluted CCC samples had a mean PS+RBC percentage of 7.0% and was statistically different from the PS+RBC percentage in NL-RBCs (P<0.01). Conclusion We describe a feasible assay to measure both PS+RBCs and C3+RBCs using flow cytometry in which CCC can be used as a positive control for PS+RBCs. This assay could serve to evaluate the quality of transfused RBC units and provide insights into the role of phosphatidylserine expression and complement deposition for RBC clearance.

Introduction: Bleeding disorders are a diverse group of conditions characterized by abnormal bleeding tendencies. Platelet dysfunction is a common underlying cause and although traditional platelet aggregometry is important in the clinical work-up, platelet functional assays often provide limited diagnostic insights, prompting the need for more advanced techniques and approaches. This study aims to explore the utility of mass cytometry combined with genetic analysis in assessing platelet function in patients with otherwise unspecified bleeding tendencies. Methods: In a single-center prospective study, enrolled patients with clinical and laboratory suspicion of platelet-related bleeding disorders were subjected to multimodality evaluation. Mass cytometry (CyTOF) allows for simultaneous evaluation of multiple platelet markers on single cells, including those involved in platelet activation, aggregation, and adhesion, through a panel of heavy metal-conjugated antibodies. Whole genome sequencing with targeted analysis was utilized to identify potential genetic variants associated with quantitative and qualitative platelet disorders and correlated with CyTOF findings. Results: CyTOF revealed altered platelet markers associated with activation, aggregation, and adhesion in 40% (n=14) of total participants (n=35) with suspected platelet-related bleeding disorders ( Table 1). CyTOF showed significant concordance with platelet aggregometry in 74% of participants (data not shown). Alterations in specific CyTOF markers (e.g., PAC-1, CD40L) correlated with bleeding risk by univariate analysis of patients stratified to high vs low risk groups based on ISTH bleeding assessment tool (ISTH-BAT) ( Table 1). Multivariate analysis using a random forest machine learning algorithm identified variables most predictive of bleeding risk, including CD40L, PAC-1, P-selectin, and mean platelet volume. Genetic analysis identified one or more variants in genes implicated in qualitative and quantitative platelet disorders in 61% (n=19) of total patients analyzed ( Figure 1). Abnormal CyTOF findings were associated with greater proportion of variants detected (79% with abnormal CyTOF vs 53% with normal CyTOF) ( Figure 1). Direct associations between specific mass cytometry findings and genetic variants (i.e., GPIIb, GPIIIa, GPIb, GPIX, etc.) were discovered for several patients, providing important information in evaluating variant pathogenicity and diagnostic validation. Conclusion: This study highlights the potential of integrating mass cytometry and genetic analysis to assist in the diagnosis of otherwise unspecified bleeding tendencies, bleeding risk stratification, and patient management. Our findings enhance the understanding of platelet function in patients with bleeding disorders and provide valuable insights into the complex relationship between platelet phenotypes and genetic determinants. Further investigations with a larger patient cohort are warranted to validate and expand upon these initial correlations and explore their clinical implications in diagnosis and management of bleeding disorders.

Guillain-Barré syndrome (GBS) is an immune-mediated polyradiculoneuropathy and the most common cause of acute flaccid paralysis worldwide. GBS classically presents with acute, progressive, ascending weakness, reduced to absent reflexes, and albuminocytological dissociation on cerebrospinal fluid (CSF) analysis. Botulism is a neurotoxin-mediated acute descending flaccid paralysis with cranial nerve palsies and dysautonomia. Botulism in adults is caused by ingestion/inhalation of botulinum toxin or wound infection with Clostridium botulinum. Both GBS and botulism can rapidly precipitate respiratory failure; thus, prompt diagnosis and treatment are crucial to mitigate poor outcomes. Herein, we describe a case of botulism initially diagnosed as GBS given classic laboratory features, and describe the importance of careful consideration of the most appropriate therapeutic modalities in cases of acute flaccid paralysis, particularly regarding empiric administration of botulinum antitoxin and use of intravenous immune globulin in lieu of plasma exchange for potential GBS to prevent removal of antitoxin.

Next-generation sequencing (NGS) is used to monitor genetically measurable residual disease (gMRD) following allogeneic stem cell transplantation (aSCT). It is unknown whether an upper limit of chimerism exists such that gMRD NGS testing can be safely forgone. We reviewed 61 patients with acute myeloid leukemia and 24 patients with myelodysplastic syndrome who had at least 1 NGS panel before and after aSCT between 2016 and 2020. Donor chimerism was quantified. Logistic regression characterized which factors predicted gMRD. Receiver operating characteristic (ROC) curves were used to determine the optimal chimerism threshold for which gMRD would not be detected. Data from an additional 22 patients with follow-up NGS testing in 2022 were also analyzed to validate our proposed threshold. A: s expected, donor chimerism was a significant predictor of gMRD (odds ratio, .38; 95% confidence interval, .10 to .62; P = .02). Age, sex, conditioning regimen, presence of a related donor, and diagnosis were not associated with gMRD. A chimerism threshold of 92.5% optimized sensitivity (97.7%) and specificity (95.4%) such that values >92.5% strongly predicted the absence of gMRD (area under the ROC curve [AUC], .986). The validation cohort demonstrated similarly strong predictive capability (AUC, .974) with appropriate sensitivity (100%) and specificity (90.9%). NGS monitoring of gMRD is redundant at chimerism values greater than a more conservative threshold of 92.5% after aSCT. (C) 2023 The American Society for Transplantation and Cellular Therapy. Published by Elsevier Inc. All rights reserved.

OBJECTIVESClinical experts recommend against testing for lupus anticoagulant (LAC) during anticoagulation.METHODSWe quantitated the risk of a single-positive dilute Russell viper venom time (dRVVT) result or partial thromboplastin time-based phospholipid neutralization (PN) result on anticoagulation.RESULTSAny anticoagulation led to a fourfold greater likelihood of single-positive results, primarily by rivaroxaban (odds ratio [OR] = 8.6) and warfarin (OR = 6.6), resulting in a positive dRVVT test with a normal PN test. Heparin and apixaban were twofold more likely to show single-positive results, but enoxaparin did not show significant single positivity.CONCLUSIONSOur results quantitatively support experts' avoidance of LAC testing during anticoagulation.

Introduction: Localized intravascular coagulopathy (LIC) is a well-recognized, though poorly characterized complication of venous malformations (VM) that can lead to bleeding, thrombosis, and phlebolith formation. While LIC has classically been characterized by elevations in D-dimer and reductions in fibrinogen, no comprehensive studies of coagulation parameters in VM have been performed to date. Since 2021, all patients with VM undergoing evaluation for LIC in the Yale Vascular Malformations Program (VaMP) and the Yale Classical Hematology clinic have been subjected to an extensive set of coagulation tests to fully analyze LIC. Our aim was to use these laboratory test results to comprehensively characterize LIC in this population. Methods: We conducted a retrospective chart review of all VM patients presenting to the Yale VaMP and the Yale Classical Hematology clinic for assessment of LIC from 2021 to 2023. All included patients were evaluated for LIC using the following coagulation parameters: von Willebrand Factor (VWF) antigen, VWF activity, factor VIII (FVIII), alpha-2 antiplasmin (A2AP), plasminogen activator inhibitor-1 (PAI-1), thrombin-antithrombin complex (TAT), D-dimer (DD), fibrinogen, prothrombin time (PT), international normalized ratio (INR), and partial thromboplastin time (PTT). Measurements of VWF antigen, VWF activity, FVIII and A2AP were performed using an ACL TOP system (Instrumentation Laboratory; Bedford, MA, USA), while D-dimer and fibrinogen were measured using a BCS XP System (Siemens; Malvern, PA, USA) at our institution's clinical laboratory. PAI-1 and TAT were processed at national Clinical Laboratory Improvement Amendment-certified reference laboratories using ELISA-based assays. Baseline patient characteristics and coagulation test results were extracted via manual chart review. Data analysis was done using IBM SPSS statistics software and GraphPad Prism 9 software. Categorical variables were described using frequency and percentages, while quantitative variables were described using central tendency and dispersion measures. We performed univariate analysis using the Chi-square test. We also analyzed the coagulation tests using a correlation matrix. Statistical significance was set at p<0.05. This project was approved by our Institutional Review Board. Results: A total of 23 patients with VM were included in the analysis (Table 1). The mean age was 38 ± 16 years; the majority (82.6%) were female. The most common anatomic location was the head and neck (52.2%), while the most frequent extent of tissue involvement was muscle (54.5%), with most patients having a single lesion (59.1%) rather than multiple ones. No patients had venous thromboembolism. Normal DD and high TAT levels were observed in most patients (68.2% and 69.6%, respectively). VWF activity, VWF antigen, FVIII, PAI-1, and fibrinogen all were positively correlated (Figure 1). TAT was positively correlated with PAI-1 and inverselycorrelated withPT and INR. DD was positively correlated only with vWF activity. TAT and DD had a poor correlation; among patients with a normal TAT (n=7), 85.7% had a normal DD, while among patients with a normal DD (n=15), only 40% had a normal TAT. Three out of 4 (75%) patients with skin involvement had a high TAT with normal DD, while 2 out of 3 (66.7%) patients with visceral involvement had both high TAT and DD, although these patternswere not statistically significant (P=0.42 and 0.31, respectively). Among patients with muscle involvement (n=11), 45.5% had a high TAT and normal DD while 36.3% had both high TAT and DD. Conclusions: In this first comprehensive hematologic study of VM, we demonstrate that VM-related LIC is characterized by derangements of multiple coagulation parameters. Due to this, measurements of DD and fibrinogen alone may be inadequate for assessing LIC, and the addition of TAT and multiple other coagulation tests may yield a more complete picture of hemostatic derangements in LIC. A lack of correlation between DD and TAT is unexpected and merits further study. Differences in the depth of the VM tissue involvement and TAT/DD correlation may indicate a progression of coagulopathy related to the extent of VM but require further investigation in a larger study.

Objectives We sought to determine risk factors for iv iron infusion-related reactions (IRR), and identify strategies for iron repletion after IRR. Methods We conducted a retrospective chart review of patients treated in the classical hematology clinic at Yale Cancer Center (n = 330 consecutive patients) from 2016 to 2021, who received iv ferumoxytol (60.3%), iron sucrose (14.8%), or iron dextran (10.9%). Results The iv iron IRR was noted in 58 (17.6%) patients, 62.1% of whom had previously tolerated iv iron. The severity of IRR was mild in 22, moderate in 23, and severe in 11 patients. Most (72.4%) patients who experienced IRR tolerated a subsequent iv iron infusion. On multivariable analysis, a history of non-medication allergies was associated with greater odds of IRR (odds ratio [OR] 2.12, 95% confidence interval (CI): 1.16-3.87, p = .01). No patients with type AB blood, and few with type A blood (n = 6), had IRR; compared to type A or AB together, patients with type B (OR 5.00, 95% CI: 1.56-16.06, p = .007) or type O (OR 3.71, 95% CI: 1.44-9.55, p = .007) blood had greater odds of IRR. Conclusions This study highlights a possible association of blood type with iv iron IRR; prospective studies with larger patient numbers are warranted to explore this association.

INTRODUCTION Cirrhosis related coagulation derangements result in altered hemostasis. Endothelial markers such as von Willebrand factor (VWF) antigen and the factor VIII-to-protein C ratio (FVIII/PC) have prognostic significance in patients with cirrhosis. Additional endothelial markers have not been well described in patients with alcohol-associated cirrhosis. We performed plasma proteomic profiling in patients with alcohol-associated cirrhosis to gain insight into the potential roles of endothelial and hemostatic proteins associated with severity of cirrhosis. METHODS Two independent cohorts of patients with Child-Pugh class A (CPA) and C (CPC) alcohol-associated cirrhosis were recruited. Controls were patients without underlying liver disease. For plasma proteomic profiling, plasma supernatant was frozen, then sent to Eve Technologies (Calgary, Alberta, Canada). Linear discrimination analysis (LDA) was applied to the first cohort to identify a set of proteins that clustered patients based on the severity of liver disease (CPA vs CPC) and controls. Identified proteins were then validated using a second independent cohort of patients with alcohol-associated cirrhosis to determine if the protein signature was replicable. Boruta feature selection was performed on cohort 2 to identify candidate proteins to explain the differences in FVIII/PC ratio. A p-value of < 0.05 was considered statistically significant. RESULTS Cohort 1 included 13 patients: 4 CPA, 9 CPC (median age was 57; 4 women; 9 had dyslipidemia, 2 diabetes mellitus, 2 hypertension, and 2 chronic kidney disease). Cohort 2 included 12 patients: 7 CPA, 5 CPC (median age 50; 5 women; 7 had dyslipidemia, 1 diabetes mellitus, 1 hypertension and 0 chronic kidney disease). For cohort 1 LDA clustered patients based on CPA vs CPC with an endothelial, inflammatory and complement signature (fig.1). The proteins set that achieved robust LDA clustering of CPA, CPC, and control patients in cohort 1 included: sVCAM-1 and PAI-1 (endothelial markers); C3, C4, and CFH (complement cascade factors); IL-8 (inflammatory chemokine), NCAM (neural adhesion maker) and BDNF (neurotrophin); MPO (marker of neutrophilic activation); Eotaxin (eosinophil associated chemokine); MIP-1d (activator of T-cells and monocytes). The same protein profile applied to the validation cohort 2 was also able to cluster CPA, CPC, and control patients (fig. 2). Boruta feature selection for proteins of importance for the FVIII/PC ratio included sVCAM-1; CFI, C4b, C4, C5, CFB (complement factors); IL-6 (inflammatory cytokine), and VWF. CONCLUSIONS Using plasma proteomic profiling we describe a signature of endothelial, complement, hemostatic, and inflammatory activation that distinguishes the severity of liver disease in patients with alcohol-associated cirrhosis. Exploratory analyses suggest that endothelial and complement proteins may also contribute to the differences in the FVIII/PC ratio which has prognostic significance in patients with cirrhosis. Future work should continue to highlight the role of endothelium in the progression of liver disease and describe alternative prognostic markers to help guide clinicians. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

This chapter examines the genetic, chemical, and structural regulation of platelet production and kinetics. New insights have been developed in our understanding of the physical development of the megakaryocyte, including the demarcation membrane system, the genetic control of platelet sizing, and the internal and shear-induced regulation of pro-platelet formation and release. The complex pathways for megakaryocyte proliferation, maturation, and movement within the bone marrow are linked to platelet production and key signaling molecules. Mechanisms for both platelet-intrinsic and external regulation of platelet lifespan and survival have contributed to our current understanding of hematopoietic responses to changes in the platelet count.

Platelets have been shown to be associated with pathophysiological processes beyond thrombosis, demonstrating critical additional roles in homeostatic processes, such as immune regulation and vascular remodeling. Platelets themselves can have multiple functional states and can communicate with and regulate other cells, including immune cells and vascular smooth muscle cells, to serve such diverse functions. Although traditional platelet functional assays are informative and reliable, they are limited in their ability to unravel platelet phenotypic heterogeneity and interactions. Developments in methods such as electron microscopy, flow cytometry, mass spectrometry and ‘omics’ studies have led to new insights. In this Review, we focus on advances in platelet biology and function, with an emphasis on current and promising methodologies. We also discuss technical and biological challenges in platelet investigations. Using coronavirus disease 2019 (COVID-19) as an example, we further describe the translational relevance of these approaches and the possible ‘bench-to-bedside’ utility in patient diagnosis and care. In this Review, Hwa and colleagues summarize the latest advances in platelet biology and function and appraise the technical and biological challenges in platelet investigations.