Anne-Marie Koop

8 285 valve regurgitation, if present. Two experienced echocardiographers (DvdW and AW) independently performed post-hoc analyses on these images. Blood sampling and biomarker assays Blood samples and echocardiography were obtained on the same day. Blood samples were collected in EDTA or heparin tubes and centrifuged. NT-proBNP and galectin 3 were analyzed immediately. Plasma for measuring GDF-15, MR-proADM, MPO, neprilysin and ET-1 was stored at -80˚C (median storage period: 6.3 months (interquartile range; IQR: 4.4-7.3)), where after analysis took place. All samples were subjected to one freeze-thaw cycle. For detailed description of biomarker assays, see supplemental material. Statistics Data are presented as medians with interquartile ranges (IQR) or frequencies (as percentage). The clinical characteristics, echocardiographic variables and biomarker plasma levels of children with either no residual RV overload, pressure, volume or combined residual RV overload were compared using Kruskal Wallis or Chi-squared test when appropriate. For pairwise testing post-hoc correction was performed with Bonferroni correction. To study differences in biomarker levels between mild- moderate and severe overload, the ratios were calculated of the plasma levels of these biomarkers in patients with mild-to-moderate overload and severe overload, respectively, to the levels in patients with no residual overload. The differences in plasma level of all eight biomarkers in childrenwith good, moderate or poor systolic RV function were analyzed using Kruskal Wallis test. The differences in plasma level of the biomarkers in children with and without RVH and RV dilation were analyzed using Mann Whitney U test. Receiver operating characteristic (ROC) curves were plotted to analyze the ability of the different biomarkers in identifying RV dilatation and/or RVH. To identify a cut-off value the highest sum of specificity and sensitivity were used. Spearman correlation analysis was used to study the correlation between TAPSE and the eight different biomarkers. Analyses were adjusted for age and sex. Statistical analysis was performed using IBM SPSS version 23.0 (Armonk, NY, USA). All statistical tests were two-tailed and a p-value of <0.05 is considered statistically significant.