Patient selection
This study was approved by The Research and Ethics Committee of Children’s Health Ireland at Crumlin, Dublin, Ireland (GEN/580/17). This study formed part of a larger study on aspirin response in paediatric patients with congenital heart disease. Written informed parental consent was obtained for each participant. All patients were attending cardiac services at Children’s Health Ireland at Crumlin. Recruitment took place between January 2019 and June 2023. Patients were included if they had an in-situ RV-PA xenograft valved-conduit or a bovine jugular venous valve system and were taking aspirin. Exclusion criteria are listed in table 1.
Table 1 – Exclusion Criteria
Patients were prospectively enrolled from both ward and outpatient settings.
Aspirin was dosed at 3-5mg/kg, up to a maximum of 75mg. Institutional practice is to prescribe to the nearest quarter of a 75mg tablet due to the inaccuracy of dispersion techniques with such preparations [18].
Testing
Aspirin response was tested a minimum of two hours post first dose of aspirin, with most patients on established aspirin therapy at time of testing. If non-response was detected, adherence and dose timing were discussed. Participants were invited to provide a confirmatory sample. Dose adjustments, if deemed appropriate, were made in consultation with the primary physician and typically increased in quarter tablet (of 75mg) increments. Aspirin response was measured using two separate tests described below: Thromboelastography with Platelet Mapping (TEGPM) and Light-Transmission Platelet Aggregation (LTA). For patients under the age of 2 years, TEGPM alone was performed due to the phlebotomy requirements for completion of both tests.
Thromboelastography with Platelet Mapping (TEGPM)
TEGPM was performed using a platelet mapping assay on the TEGâ 5000 analyser platform. This gives a quantitative analysis of platelet function based on the formation, strength, and degradation of clots in whole blood. It allows for the contribution of aspirin to be assessed through the addition of arachidonic acid (AA) to determine the response of the TXA2 receptor when compared with standard samples from the index patient. Three assays were performed. The first was performed by adding whole blood to kaolin and measuring TEG on this kaolin-activated blood to derive maximal clot strength (MAThrombin) in the standard TEG fashion. The contribution of fibrin to clot strength (MAFibrin) was assessed through the addition of reptilase and factor XII and measured on a second TEG cup. Finally, arachidonic acid (AA) and Activator F were added to a sample in a third TEG cup to assess the contribution of the COX-1 pathway (MA AA). The percentage platelet inhibition was calculated using the equation 100-{(MA AA –MA Fibrin) / (MA Thrombin –MA Fibrin) X 100}. Non-responsiveness to aspirin was defined as platelet inhibition <50%.
Light-Transmission Platelet Aggregometry (LTA)
LTA was performed on platelet rich plasma samples by placing the sample between the light-source and the photocell. Arachadonic acid was added to the sample in order to activate platelets. Aspirin non-response was defined as platelet aggregation in response to arachidonic acid of >20%.
Statistical analysis
Baseline and demographic data were summarised for the enrolled patients. Normally distributed continuous data are expressed as mean ± SD, non-normally distributed variables were expressed as median (minimum-maximum). Categorical variables were expressed as percentages. Comparison of characteristics between responders and non-responders was performed using the Mann-Whitney U test for continuous variables, and Fisher exact test for categorical data. Statistical analysis was performed using GraphPad Prism version 10.0.0.