Platelet physiology and function in neonates and children on extracorporeal membrane oxygenation (ECMO)
AuthorYaw, Hui Ping
Document TypePhD thesis
Access StatusThis item is embargoed and will be available on 2021-10-29. This item is currently available to University of Melbourne staff and students only, login required.
© 2019 Hui Ping Yaw
Extracorporeal membrane oxygenation (ECMO) is a modified form of heart-lung machine that aims to provide short- to-medium length of support to patients with cardiac and/or respiratory dysfunction. Children represent the majority of the ECMO population. While increasing experience and technical improvements for ECMO over the years have seen some improvement in outcomes, the rates of morbidity and mortality remain high in this population and many complications are related to bleeding and thrombosis. Platelets are a key element of the coagulation system. Platelet dysfunction can cause coagulopathy in adults on ECMO, however, the association between modification of platelet function and coagulopathy remains unknown for children. This study hypothesised that there are platelet-specific differences: I.) for paediatric patients on ECMO according to their age, pathway onto ECMO and duration of ECMO that could be associated with the development of bleeding or thrombosis during ECMO and II.) at different sites in a paediatric ECMO system. This study aimed to characterize the molecular indices of circulating platelets in the paediatric ECMO population using whole blood flow cytometry approach: 1) To examine and compare the effect of a patient’s pathway onto ECMO on platelet phenotype and function and their associations with the development of bleeding or thrombosis during ECMO. 2) To examine and compare the effect of a patient’s age on platelet phenotype and function and their associations with the development of bleeding or thrombosis during ECMO. 3) To examine and compare the effect of a patient’s duration of ECMO on platelet phenotype and function and their associations with the development of bleeding or thrombosis during ECMO. 4) To examine and compare the site-specific differences for platelet phenotype and function in the paediatric ECMO system. A total of 22 paediatric patients [median (interquartile range): 0.34 (0.01 – 3.38) years] were included in this study. Citrated whole blood samples were collected and a whole blood flow cytometry method was developed for the evaluation of platelet phenotype and function in the setting of ECMO. The platelet assays were standardized to ensure minimal pre-analytical activation. By using multiple thrombin receptor activator peptide 6 (a thrombin mimic) concentrations, the platelet panels also showed sensitivity to detect subtle changes in platelet response. The multifaceted flow cytometry panels allowed simultaneous evaluation of platelets for their phenotype, function and interactions with monocytes and neutrophils. Such approach to investigate platelet-specific changes from different aspects suits well for the ECMO population, representing a complex group of patients. These results showed that the whole blood flow cytometry assay is a reliable and useful platelet function test for paediatric ECMO patients. Results from the analysis for platelet-specific markers within the first 24 hours showed no difference in platelet phenotype and function between patients from different pathways onto ECMO and different ages. However, the association of platelet-specific changes and the development of clinical events during ECMO were different according to a patient’s age and pathway onto ECMO. Patients who had cardiopulmonary bypass before coming onto ECMO and had bleeding had increased platelet integrin GPIIb/IIIa receptor expression and reduced circulating neutrophil-platelet aggregates level compared to patients who had no bleeding during ECMO. In contrast, patients who had no cardiopulmonary bypass before coming onto ECMO and developed bleeding had reduced platelet response compared to those who had no bleeding during ECMO. Conversely, increased lysosome release was observed for children with thrombosis and may indicate the presence of a protective mechanism against increased thrombus formation. Duration of ECMO had been recognized as an important factor affecting the outcome of paediatric ECMO patients. The results showed an increased level of von Willebrand factor (VWF) receptor and reduced platelet response for granule exocytosis with increasing number of days on ECMO (Day 2 vs. Day 5). Most importantly, such platelet-specific changes that involved GPIb/IX/V receptor and granule release with increasing duration of ECMO were only observed in patients who had bleeding but not in patients without bleeding after five days on ECMO. In addition, elevated circulating monocyte-platelet aggregates level was only observed in patients who had thrombosis but not for those without thrombosis. Together, these results suggested a link between pathway onto ECMO/age/duration of ECMO, bleeding/thrombosis and platelet dysfunction. Hence, markers relevant to the platelet-specific changes could be used as the indicators for increased bleeding or thrombosis risk for paediatric patients during ECMO. Platelet phenotype and function were also compared at different sites in the ECMO system to identify the site-specific differences of platelet-specific changes that have not previously been investigated in a paediatric ECMO system. In the setting of mechanical circulatory support, shear and oxidative stress are known to modify platelet phenotype via integrin GPIIb/IIIa and GPIb/IX/V receptors and increase platelet response via multiple platelet activation pathways. The results demonstrated that platelet phenotype and function were different at different sites in a paediatric ECMO system. The platelet-specific changes observed included the modification of platelet phenotype via increased VWF and integrin GPIIb/IIIa receptor expression, increased platelet activation through the activation of the integrin GPIIb/IIIa receptor and higher platelet responsiveness at the post-oxygenator site compared to the pre-oxygenator site. However, the exact cause of the site-specific differences of platelet phenotype and function remained to be identified. In summary, this study demonstrated the feasibility of using whole blood flow cytometry method with multifaceted platelet-specific panels as a reliable platelet function test in paediatric patients on ECMO. Platelet-specific changes could be associated with the development of bleeding or thrombosis during ECMO. In addition, platelet phenotype and function were different at different sites in a paediatric ECMO system. Together, this study provides new insights for the circuit-related platelet-specific changes and the understanding of how modifications of platelet phenotype and function that are dependent on patient’s factors may be associated with coagulopathy in children on ECMO.
Keywordsplatelets; extracorporeal membrane oxygenation; whole blood flow cytometry; paediatrics; leukocyte-platelet aggregates
- Click on "Export Reference in RIS Format" and choose "open with... Endnote".
- Click on "Export Reference in RIS Format". Login to Refworks, go to References => Import References