Publication Recommendations to Report Laboratory Data of Neonates - a Modified Delphi Approach.

BACKGROUND: Clinical and analytical information on laboratory data of neonates in scientific publications is sparse and incomplete. Furthermore, interpreting neonatal laboratory data can be complex due to their time-dependent and developmental physiology, and paucity of well-established age-appropriate reference ranges for neonates. This study aims to develop publication recommendations to report laboratory data of neonates to enhance the quality of these data in research and clinical care. METHODS: A modified Delphi approach was used to develop recommendations in cooperation with the International Neonatal Consortium. A Core Group, including different stakeholders, was responsible for developing the recommendations, in collaboration with a Reflection Group, responsible for providing additional input. RESULTS: The recommendations were classified into three categories: 'Clinical Characteristics', 'Bio-analytical Information' and 'Data-analytical Information'. These were each divided into 'Core Data' (always to be reported) and 'Supplemental Considerations' (to be reported when considered relevant to the study). CONCLUSION: Our recommendations provide guidance on standardization of neonatal laboratory data in publications. This will enhance the comparison, replication, and application of study results in research initiatives and clinical practice. Furthermore, these recommendations also serve as foundational work to develop reference ranges for neonatal laboratory values by standardizing the quality of information needed for such efforts. IMPACT: Standardized reporting of neonatal laboratory data in scientific publications will enhance the comparison, replication, and application of study results in research initiatives and clinical practice, as well as improve reporting to regulatory agencies. To integrate multistakeholder perspectives, a modified Delphi approach was used to develop publication recommendations which strengthens the applicability of the recommendations. Implementation of standardization will likely improve the overall quality of neonatal clinical research and neonatal healthcare. In addition, these recommendations are foundational to develop reference ranges for neonatal laboratory values by standardizing the quality of information needed for such efforts.

Rapid Whole-Genomic Sequencing and a Targeted Neonatal Gene Panel in Infants With a Suspected Genetic Disorder.

Importance: Genomic testing in infancy guides medical decisions and can improve health outcomes. However, it is unclear whether genomic sequencing or a targeted neonatal gene-sequencing test provides comparable molecular diagnostic yields and times to return of results. Objective: To compare outcomes of genomic sequencing with those of a targeted neonatal gene-sequencing test. Design, Setting, and Participants: The Genomic Medicine for Ill Neonates and Infants (GEMINI) study was a prospective, comparative, multicenter study of 400 hospitalized infants younger than 1 year of age (proband) and their parents, when available, suspected of having a genetic disorder. The study was conducted at 6 US hospitals from June 2019 to November 2021. Exposure: Enrolled participants underwent simultaneous testing with genomic sequencing and a targeted neonatal gene-sequencing test. Each laboratory performed an independent interpretation of variants guided by knowledge of the patient's phenotype and returned results to the clinical care team. Change in clinical management, therapies offered, and redirection of care was provided to families based on genetic findings from either platform. Main Outcomes and Measures: Primary end points were molecular diagnostic yield (participants with ≥1 pathogenic variant or variant of unknown significance), time to return of results, and clinical utility (changes in patient care). Results: A molecular diagnostic variant was identified in 51% of participants (n = 204; 297 variants identified with 134 being novel). Molecular diagnostic yield of genomic sequencing was 49% (95% CI, 44%-54%) vs 27% (95% CI, 23%-32%) with the targeted gene-sequencing test. Genomic sequencing did not report 19 variants found by the targeted neonatal gene-sequencing test; the targeted gene-sequencing test did not report 164 variants identified by genomic sequencing as diagnostic. Variants unidentified by the targeted genomic-sequencing test included structural variants longer than 1 kilobase (25.1%) and genes excluded from the test (24.6%) (McNemar odds ratio, 8.6 [95% CI, 5.4-14.7]). Variant interpretation by laboratories differed by 43%. Median time to return of results was 6.1 days for genomic sequencing and 4.2 days for the targeted genomic-sequencing test; for urgent cases (n = 107) the time was 3.3 days for genomic sequencing and 4.0 days for the targeted gene-sequencing test. Changes in clinical care affected 19% of participants, and 76% of clinicians viewed genomic testing as useful or very useful in clinical decision-making, irrespective of a diagnosis. Conclusions and Relevance: The molecular diagnostic yield for genomic sequencing was higher than a targeted neonatal gene-sequencing test, but the time to return of routine results was slower. Interlaboratory variant interpretation contributes to differences in molecular diagnostic yield and may have important consequences for clinical management.

What can the plasma proteome tell us about platelets and (vice versa)?

Multi-omics approaches are being used increasingly to study physiological and pathophysiologic processes. Proteomics specifically focuses on the study of proteins as functional elements and key contributors to, and markers of the phenotype, as well as targets for diagnostic and therapeutic approaches. Depending on the condition, the plasma proteome can mirror the platelet proteome, and hence play an important role in elucidating both physiologic and pathologic processes. In fact, both plasma and platelet protein signatures have been shown to be important in the setting of thrombosis-prone disease states such as atherosclerosis and cancer. Plasma and platelet proteomes are increasingly being studied as a part of a single entity, as is the case with patient-centric sample collection approaches such as capillary blood. Future studies should cut across the plasma and platelet proteome silos, taking advantage of the vast knowledge available when they are considered as part of the same studies, rather than studied as distinct entities.

Platelets are key cellular elements of blood with plasma constituting the liquid component. Both platelets and proteins found in plasma rapidly work in unity to prevent/limit blood loss in response to blood vessel damage. Proteomics is the analysis of the entire protein complement of a cell, tissue, or organism under a specific, defined set of conditions. Of note, research to date has shown that platelet and plasma proteomes share many common proteins. In some disease scenarios, plasma proteomes can be used to identify platelet function or dysfunction, while in other scenarios, platelet-specific proteins are needed for physiological assessment. Thus, it may be beneficial to simultaneously study the plasma and platelet proteomes, thereby exploiting the considerable wealth of information provided under such circumstances.

General Anesthesia and the Premature Baby: Identifying Risks for Poor Neurodevelopmental Outcomes.

Preterm birth affects 1 in every 10 infants born in the United States. Importantly, more preterm infants are surviving to discharge from hospital, including those born at the cusp of viability (eg, 22 to 24 wk gestation). Such improvements, however, come at a cost as those delivered at less than 28 weeks gestation have the highest rates of morbidity and mortality. To complicate matters, these extremely preterm infants often require multiple surgical procedures resulting in repeated and prolonged exposures to anesthetic, analgesic, and sedative agents both during procedures and in the neonatal intensive care unit. Consequently, all of these factors, including premature birth itself, correlate with a higher risk for neurodevelopmental disabilities. More studies are needed to address the effects of prematurity-related morbidities and drug exposures on this vulnerable population, with the goal of improving neurodevelopmental outcomes. This brief review will discuss risk factors that impact neurodevelopmental outcomes in premature infants, with a particular focus on anesthetic, analgesic, and sedative agents.

Novel Variant Findings and Challenges Associated With the Clinical Integration of Genomic Testing: An Interim Report of the Genomic Medicine for Ill Neonates and Infants (GEMINI) Study.

Importance: A targeted genomic sequencing platform focused on diseases presenting in the first year of life may minimize financial and ethical challenges associated with rapid whole-genomic sequencing. Objective: To report interim variants and associated interpretations of an ongoing study comparing rapid whole-genomic sequencing with a novel targeted genomic platform composed of 1722 actionable genes targeting disorders presenting in infancy. Design, Setting, and Participants: The Genomic Medicine in Ill Neonates and Infants (GEMINI) study is a prospective, multicenter clinical trial with projected enrollment of 400 patients. The study is being conducted at 6 US hospitals. Hospitalized infants younger than 1 year of age suspected of having a genetic disorder are eligible. Results of the first 113 patients enrolled are reported here. Patient recruitment began in July 2019, and the interim analysis of enrolled patients occurred from March to June 2020. Interventions: Patient (proband) and parents (trios, when available) were tested simultaneously on both genomic platforms. Each laboratory performed its own phenotypically driven interpretation and was blinded to other results. Main Outcomes and Measures: Variants were classified according to the American College of Medical Genetics and Genomics standards of pathogenic (P), likely pathogenic (LP), or variants of unknown significance (VUS). Chromosomal and structural variations were reported by rapid whole-genomic sequencing. Results: Gestational age of 113 patients ranged from 23 to 40 weeks and postmenstrual age from 27 to 83 weeks. Sixty-seven patients (59%) were male. Diagnostic and/or VUS were returned for 51 patients (45%), while 62 (55%) had negative results. Results were concordant between platforms in 83 patients (73%). Thirty-seven patients (33%) were found to have a P/LP variant by 2 or both platforms and 14 (12%) had a VUS possibly related to phenotype. The median day of life at diagnosis was 22 days (range, 3-313 days). Significant alterations in clinical care occurred in 29 infants (78%) with a P/LP variant. Incidental findings were reported in 7 trios. Of 51 positive cases, 34 (67%) differed in the reported result because of technical limitations of the targeted platform, interpretation of the variant, filtering discrepancies, or multiple causes. Conclusions and Relevance: As comprehensive genetic testing becomes more routine, these data highlight the critically important variant detection capabilities of existing genomic sequencing technologies and the significant limitations that must be better understood.

Cangrelor PK/PD analysis in post-operative neonatal cardiac patients at risk for thrombosis.

Essentials An optimal therapeutic strategy has yet to be established to prevent early shunt thrombosis. A phase 1 study of cangrelor was performed in neonates after palliation of congenital heart disease. PD endpoint of >90% platelet inhibition in 60% of patients was achieved at 0.5 µg/kg/min dosing. No serious adverse events related to drug administration were observed, including bleeding. ABSTRACT: Background Systemic-to-pulmonary artery shunt thrombosis is a significant cause of early postoperative mortality in neonates after palliation of congenital heart disease. In the context of thromboprophylaxis, an optimal therapeutic strategy has yet to be established before aspirin administration. Cangrelor, a fast-acting, reversible P2Y12 inhibitor, may fill this unmet need. Objectives To evaluate the pharmacokinetics (PK), pharmacodynamics (PD), and safety of cangrelor in neonates undergoing stage 1 palliation. Methods This prospective, open-label, single-arm study evaluated two cangrelor dosing cohorts following placement of a systemic-to-pulmonary artery shunt, right ventricle-to-pulmonary artery shunt, or ductal stent. Drug concentrations and platelet reactivity, assessed by light transmission aggregometry and in microfluidic assays (MF), were measured. Results Twenty-two patients were consented and 15 received a 1-hour infusion of cangrelor at either 0.5 µg/kg/min (cohort 1) or 0.25 µg/kg/min (cohort 2). Whereas the primary PD endpoint was achieved at the higher dose (ie, reduction in maximal platelet aggregation by ≥90% in 60% of participants), only 29% of those in cohort 2 attained this goal. Comparable and statistically significant results were obtained in MF assays (P < .0001 vs. baseline). Drug levels during infusion were 3-fold higher in cohort 1 vs. cohort 2 (P < .001). Most participants (70%) had undetectable drug levels by 10 minutes postinfusion with full recovery in platelet function at 1 hour. No drug-related bleeding events occurred. Conclusions Favorable PK/PD properties of cangrelor 0.5 µg/kg/min dosing and safety profile warrant further evaluation in neonates following palliative cardiac procedures.

Detection of circulating extracellular mRNAs by modified small-RNA-sequencing analysis.

Extracellular mRNAs (ex-mRNAs) potentially supersede extracellular miRNAs (ex-miRNAs) and other RNA classes as biomarkers. We performed conventional small-RNA-sequencing (sRNA-seq) and sRNA-seq with T4 polynucleotide kinase (PNK) end-treatment of total exRNA isolated from serum and platelet-poor EDTA, ACD, and heparin plasma to study the effect on ex-mRNA capture. Compared to conventional sRNA-seq PNK-treatment increased the detection of informative ex-mRNAs reads up to 50-fold. The exRNA pool was dominated by hematopoietic cells and platelets, with additional contribution from the liver. About 60% of the 15- to 42-nt reads originated from the coding sequences, in a pattern reminiscent of ribosome-profiling. Blood sample type had a considerable influence on the exRNA profile. On average approximately 350 to 1,100 distinct ex-mRNA transcripts were detected depending on plasma type. In serum, additional transcripts from neutrophils and hematopoietic cells increased this number to near 2,300. EDTA and ACD plasma showed a destabilizing effect on ex mRNA and non-coding RNA ribonucleoprotein complexes compared to other plasma types. In a proof-of-concept study, we investigated differences between the exRNA profiles of patients with acute coronary syndrome (ACS) and healthy controls. The improved tissue resolution of ex mRNAs after PNK-treatment enabled us to detect a neutrophil-signature in ACS that escaped detection by ex miRNA analysis.

Outcomes Research in Vulnerable Pediatric Populations.

The Pediatric Anesthesia and Neurodevelopment Assessment (PANDA) study team held its biennial symposium in April 2018 to discuss issues on anesthetic neurotoxicity in the developing brain. One of the sessions invited speakers with different areas of expertise to discuss "Outcomes Research in Vulnerable Pediatric Populations." The vulnerable populations included neonates, children with congenital heart disease, children from low socioeconomic status, and children with incarcerated parents. Each speaker presented some of the ongoing research efforts in these groups as well as the challenges encountered in studying them.

Human plasma and serum extracellular small RNA reference profiles and their clinical utility.

Circulating extracellular RNAs (exRNAs) have the potential to serve as biomarkers for a wide range of medical conditions. However, limitations in existing exRNA isolation methods and a lack of knowledge on parameters affecting exRNA variability in human samples may hinder their successful discovery and clinical implementation. Using combinations of denaturants, reducing agents, proteolysis, and revised organic extraction, we developed an automated, high-throughput approach for recovery of exRNAs and exDNA from the same biofluid sample. We applied this method to characterize exRNAs from 312 plasma and serum samples collected from 13 healthy volunteers at 12 time points over a 2-month period. Small RNA cDNA library sequencing identified nearly twofold increased epithelial-, muscle-, and neuroendocrine-cell-specific miRNAs in females, while fasting and hormonal cycle showed little effect. External standardization helped to detect quantitative differences in erythrocyte and platelet-specific miRNA contributions and in miRNA concentrations between biofluids. It also helped to identify a study participant with a unique exRNA phenotype featuring a miRNA signature of up to 20-fold elevated endocrine-cell-specific miRNAs and twofold elevated total miRNA concentrations stable for over 1 year. Collectively, these results demonstrate an efficient and quantitative method to discern exRNA phenotypes and suggest that plasma and serum RNA profiles are stable over months and can be routinely monitored in long-term clinical studies.

Response biomarkers in neonatal intervention studies.

BackgroundUp to 90% of all drugs used in neonatal intensive care units (NICUs) have not been clinically tested for safety and efficacy. To promote drug development for neonates, the pharmaceutical industry is moving toward rigorous testing, necessitating the need to development, and validating biomarkers in neonates to predict their response. The objective of this review is to evaluate the quality of the response biomarker reporting in neonatal clinical trials.MethodsA validated literature search strategy was applied. Prospective neonatal intervention studies reporting response biomarkers published in 2014 were included. The data were extracted independently and in duplicate using a data-extraction form.ResultsFollowing the full-text review, 167 published prospective neonatal trials were included; 35% (59/167) reported the use of response biomarkers. In these 59 trials, we identified 275 biomarkers used to measure the response (pharmacodynamics and safety) reported as primary or secondary outcomes. Heart rate and oxygen saturation were the most commonly reported. Measurement and instrumentation data were often not provided.ConclusionWe identified a huge variability in the selection, measurement, and reporting of neonatal response biomarkers in prospective intervention studies. Reporting initiatives are needed to reduce research waste and improve the reproducibility of biomarker use in neonatal intervention studies.

A Novel Mutation in Junctional Plakoglobin Causing Lethal Congenital Epidermolysis Bullosa.

We report a case of neonatal generalized erythema and epidermolysis resulting from a novel mutation in the junctional plakoglobin gene causing truncation of the plakoglobin protein. Expedited genetic testing enabled diagnosis while the patient was in the neonatal intensive care unit, providing valuable information for the clinicians and family.

P2Y12 Receptor Function and Response to Cangrelor in Neonates With Cyanotic Congenital Heart Disease.

Shunt thrombosis remains a major cause of morbidity and mortality, especially during the initial palliation for single-ventricle physiology. The authors present evidence that the P2Y12 inhibitor cangrelor may fill a therapeutic void in thromboprophylaxis. They base this theory on results showing that platelets from neonatal patients with cyanotic congenital heart disease have a robust response to adenosine diphosphate and are amenable to P2Y12 inhibition with cangrelor. Unique to this study was their ability to establish drug efficacy in an avatar mouse model that permits the in vivo evaluation of human platelet-mediated thrombus formation illustrating that this P2Y12 inhibitor yields the intended biological response.

PI3Kγ/δ and NOTCH1 Cross-Regulate Pathways That Define the T-cell Acute Lymphoblastic Leukemia Disease Signature.

PI3K/AKT and NOTCH1 signaling pathways are frequently dysregulated in T-cell acute lymphoblastic leukemias (T-ALL). Although we have shown that the combined activities of the class I PI3K isoforms p110γ and p110δ play a major role in the development and progression of PTEN-null T-ALL, it has yet to be determined whether their contribution to leukemogenic programing is unique from that associated with NOTCH1 activation. Using an Lmo2-driven mouse model of T-ALL in which both the PI3K/AKT and NOTCH1 pathways are aberrantly upregulated, we now demonstrate that the combined activities of PI3Kγ/δ have both overlapping and distinct roles from NOTCH1 in generating T-ALL disease signature and in promoting tumor cell growth. Treatment of diseased animals with either a dual PI3Kγ/δ or a γ-secretase inhibitor reduced tumor burden, prolonged survival, and induced proapoptotic pathways. Consistent with their similar biological effects, both inhibitors downregulated genes involved in cMYC-dependent metabolism in gene set enrichment analyses. Furthermore, overexpression of cMYC in mice or T-ALL cell lines conferred resistance to both inhibitors, suggesting a point of pathway convergence. Of note, interrogation of transcriptional regulators and analysis of mitochondrial function showed that PI3Kγ/δ activity played a greater role in supporting the disease signature and critical bioenergetic pathways. Results provide insight into the interrelationship between T-ALL oncogenic networks and the therapeutic efficacy of dual PI3Kγ/δ inhibition in the context of NOTCH1 and cMYC signaling. Mol Cancer Ther; 16(10); 2069-82. ©2017 AACR.

Acalabrutinib (ACP-196) in Relapsed Chronic Lymphocytic Leukemia.

BACKGROUND: Irreversible inhibition of Bruton's tyrosine kinase (BTK) by ibrutinib represents an important therapeutic advance for the treatment of chronic lymphocytic leukemia (CLL). However, ibrutinib also irreversibly inhibits alternative kinase targets, which potentially compromises its therapeutic index. Acalabrutinib (ACP-196) is a more selective, irreversible BTK inhibitor that is specifically designed to improve on the safety and efficacy of first-generation BTK inhibitors. METHODS: In this uncontrolled, phase 1-2, multicenter study, we administered oral acalabrutinib to 61 patients who had relapsed CLL to assess the safety, efficacy, pharmacokinetics, and pharmacodynamics of acalabrutinib. Patients were treated with acalabrutinib at a dose of 100 to 400 mg once daily in the dose-escalation (phase 1) portion of the study and 100 mg twice daily in the expansion (phase 2) portion. RESULTS: The median age of the patients was 62 years, and patients had received a median of three previous therapies for CLL; 31% had chromosome 17p13.1 deletion, and 75% had unmutated immunoglobulin heavy-chain variable genes. No dose-limiting toxic effects occurred during the dose-escalation portion of the study. The most common adverse events observed were headache (in 43% of the patients), diarrhea (in 39%), and increased weight (in 26%). Most adverse events were of grade 1 or 2. At a median follow-up of 14.3 months, the overall response rate was 95%, including 85% with a partial response and 10% with a partial response with lymphocytosis; the remaining 5% of patients had stable disease. Among patients with chromosome 17p13.1 deletion, the overall response rate was 100%. No cases of Richter's transformation (CLL that has evolved into large-cell lymphoma) and only one case of CLL progression have occurred. CONCLUSIONS: In this study, the selective BTK inhibitor acalabrutinib had promising safety and efficacy profiles in patients with relapsed CLL, including those with chromosome 17p13.1 deletion. (Funded by the Acerta Pharma and others; ClinicalTrials.gov number, NCT02029443.).

Exploiting the kinetic interplay between GPIbα-VWF binding interfaces to regulate hemostasis and thrombosis.

Platelet-von Willebrand factor (VWF) interactions must be tightly regulated in order to promote effective hemostasis and prevent occlusive thrombus formation. However, it is unclear what role the inherent properties of the bond formed between the platelet receptor glycoprotein Ibα and the A1 domain of VWF play in these processes. Using VWF-A1 knock-in mice with mutations that enhance (I1309V) or disrupt (R1326H) platelet receptor glycoprotein Ibα binding, we now demonstrate that the kinetic interplay between two distinct contact surfaces influences the site and extent to which platelets bind VWF. Incorporation of R1326H mutation into the major site shortened bond lifetime, yielding defects in hemostasis and thrombosis comparable to VWF-deficient animals. Similarly, disrupting this region of contact with an allosteric inhibitor impaired human platelet accrual in damaged arterioles. In contrast, the I1309V mutation near the minor site prolonged bond lifetime, which was essential for the development of a type 2B-like VWD phenotype. However, combining the R1326H and I1309V mutations normalized both bond kinetics and the hemostatic and thrombotic properties of VWF. These findings broaden our understanding of mechanisms governing platelet-VWF interactions in health and disease, and underscore the importance of combined biophysical and genetic approaches in identifying potential therapeutic avenues for treating bleeding and thrombotic disorders.

A selective microRNA-based strategy inhibits restenosis while preserving endothelial function.

Drugs currently approved to coat stents used in percutaneous coronary interventions do not discriminate between proliferating vascular smooth muscle cells (VSMCs) and endothelial cells (ECs). This lack of discrimination delays reendothelialization and vascular healing, increasing the risk of late thrombosis following angioplasty. We developed a microRNA-based (miRNA-based) approach to inhibit proliferative VSMCs, thus preventing restenosis, while selectively promoting reendothelialization and preserving EC function. We used an adenoviral (Ad) vector that encodes cyclin-dependent kinase inhibitor p27(Kip1) (p27) with target sequences for EC-specific miR-126-3p at the 3' end (Ad-p27-126TS). Exogenous p27 overexpression was evaluated in vitro and in a rat arterial balloon injury model following transduction with Ad-p27-126TS, Ad-p27 (without miR-126 target sequences), or Ad-GFP (control). In vitro, Ad-p27-126TS protected the ability of ECs to proliferate, migrate, and form networks. At 2 and 4 weeks after injury, Ad-p27-126TS-treated animals exhibited reduced restenosis, complete reendothelialization, reduced hypercoagulability, and restoration of the vasodilatory response to acetylcholine to levels comparable to those in uninjured vessels. By incorporating miR-126-3p target sequences to leverage endogenous EC-specific miR-126, we overexpressed exogenous p27 in VSMCs, while selectively inhibiting p27 overexpression in ECs. Our proof-of-principle study demonstrates the potential of using a miRNA-based strategy as a therapeutic approach to specifically inhibit vascular restenosis while preserving EC function.

RUC-4: a novel αIIbß3 antagonist for prehospital therapy of myocardial infarction.

OBJECTIVE: Treatment of myocardial infarction within the first 1 to 2 hours with a thrombolytic agent, percutaneous coronary intervention, or an αIIbß3 antagonist decreases mortality and the later development of heart failure. We previously reported on a novel small molecule αIIbß3 antagonist, RUC-2, that has a unique mechanism of action. We have now developed a more potent and more soluble congener of RUC-2, RUC-4, designed to be easily administered intramuscularly by autoinjector to facilitate its use in the prehospital setting. Here, we report the properties of RUC-4 and the antiplatelet and antithrombotic effects of RUC-2 and RUC-4 in animal models. APPROACH AND RESULTS: RUC-4 was ≈ 20% more potent than RUC-2 in inhibiting human ADP-induced platelet aggregation and much more soluble in aqueous solutions (60-80 mg/mL). It shared RUC-2's specificity for αIIbß3 versus αVß3, did not prime the receptor to bind fibrinogen, or induce changes in ß3 identified by a conformation-specific monoclonal antibody. Both RUC-2 and RUC-4 prevented FeCl3-induced thrombotic occlusion of the carotid artery in mice and decreased microvascular thrombi in response to laser injury produced by human platelets infused into transgenic mice containing a mutated von Willebrand factor that reacts with human but not mouse platelets. Intramuscular injection of RUC-4 in nonhuman primates at 1.9 and 3.85 mg/kg led to complete inhibition of platelet aggregation within 15 minutes, with dose-dependent return of platelet aggregation after 4.5 to 24 hours. CONCLUSIONS: RUC-4 has favorable biochemical, pharmacokinetic, pharmacodynamic, antithrombotic, and solubility properties as a prehospital therapy of myocardial infarction, but the possibility of increased bleeding with therapeutic doses remains to be evaluated.

Analysis of early thrombus dynamics in a humanized mouse laser injury model.

Platelet aggregation and thrombus formation at the site of injury is a dynamic process that involves the continuous addition of new platelets as well as thrombus rupture. In the early stages of hemostasis (within minutes after vessel injury) this process can be visualized by transfusing fluorescently labeled human platelets and observing their deposition and detachment. These two counterbalancing events help the developing thrombus reach a steady-state morphology, where it is large enough to cover the injured vessel surface but not too large to form a severe thrombotic occlusion. In this study, the spatial and temporal aspects of early stage thrombus dynamics which result from laser-induced injury on arterioles of cremaster muscle in the humanized mouse were visualized using fluorescent microscopy. It was found that rolling platelets show preference for the upstream region while tethering/detaching platelets were primarily found downstream. It was also determined that the platelet deposition rate is relatively steady, whereas the effective thrombus coverage area does not increase at a constant rate. By introducing a new method to graphically represent the real time in vivo physiological shear stress environment, we conclude that the thrombus continuously changes shape by regional growth and decay, and neither dominates in the high shear stress region.

Simulation of platelet, thrombus and erythrocyte hydrodynamic interactions in a 3D arteriole with in vivo comparison.

Cylindrical blood vessels, ellipsoid platelets and biconcave-shaped deformable erythrocytes (RBCs) are important participants in hemostasis and thrombosis. However, due to the challenge of combining these components in simulation tools, few simulation studies have included all of them in realistic three-dimensional models. In the present study, we apply a recently developed simulation model to incorporate these components and analyze the flow in a thrombotic tubular arteriole, particularly the detailed hydrodynamic interactions between the thrombus shape, RBCs and platelets. It was found that at certain azimuth positions, the velocity drops in the proximity of both the upstream and downstream edge of the thrombus, which is accompanied by a rapid velocity increase in the narrowed region. The RBCs alter the flow profiles significantly from the typical low Reynolds (Re) number flow, and also enhance the deposition of free flowing platelets onto the thrombus. By evaluating the platelet-thrombus interaction and platelet-RBC interaction together, several mechanisms of platelet deposition augmentation are identified. With in vivo data comparison, our model illustrates the potential of future thrombosis studies that incorporate detailed receptor-ligand adhesion modules.