## 2 Materials and Methods
### TMAD Data Acquisition and Analysis  
TMAD of the tricuspid valve was measured in the RV focused apical four-chamber view, using the 2-dimensional speckle tracking technique. The speckle tracking analysis was performed offline using the QLAB software (QLAB 15.0, Cardiac motion quantification software, Philips Medical Systems, Best, The Netherlands). Three points were selected in a diastolic frame: insertion of the anterior leaflet and the septal leaflet, respectively, into the tricuspid annulus and the right ventricular apex. Further, the lateral (TMADlat) and septal (TMADsept) tricuspid annular points longitudinal displacement were tracked during the cardiac cycle. Midpoint displacement (TMADmid) of the tricuspid valve annulus was also computed by the software. Further, tricuspid annulus midpoint fractional displacement (TMADm%) also known as right ventricular longitudinal shortening fraction (RV-LSF) was calculated automatically by the software, as the maximum displacement of the midpoint throughout the cardiac cycle, as follows: (end diastolic RV length \(-\) end systolic RV length) \(\times\) 100/(end diastolic RV length) (Figure 1) [15].  
\begin{table}
\begin{tabular}{c c c c c c c} \hline \multirow{2}{*}{**Variable**} & \multirow{2}{*}{\begin{tabular}{c} **Control** \\ **(n = 20)** \\ \end{tabular} } & \begin{tabular}{c} **Non-Worsening** \\ **PAH Children** \\ **(n = 11)** \\ \end{tabular} & \begin{tabular}{c} **Worsening** \\ **PAH Children** \\ **(n = 9)** \\ \end{tabular} & \begin{tabular}{c} **P1** \\ **(nw vs. c)** \\ \end{tabular} & \
\begin{tabular}{c} **P2** \\ **(w vs. c)** \\ \end{tabular} &
\begin{tabular}{c} **P3** \\ **(w vs. nw)** \\ \end{tabular} \\ \hline Age (y) & \(10.72\pm 3.65\) & \(10.59\pm 3.33\) & \(10.15\pm 3.80\) & 1.000 & 1.000 & 1.000 \\ Male (\%) & 9 (45\%) & 3 (27.27\%) & 4 (44.44\%) & 0.451 & 1.000 & 0.642 \\ \hline H (m) & 1.45 \(\pm\) 0.20 & \(1.32\pm 0.15\) & 1.31 \(\pm\) 0.23 & 0.239 & 0.267 & 1.000 \\ W (kg) & 40.60 \(\pm\) 18.67 & \(26.95\pm 8.45\) & 29.21 \(\pm\) 10.82 & 0.060 & 0.197 & 1.000 \\ \hline BSA (m\({}^{2}\)) & 1.26 \(\pm\) 0.37 & 0.98 \(\pm\) 0.20 & 1.02 \(\pm\) 0.28 & 0.077 & 0.228 & 1.000 \\ BMI (m/kg\({}^{2}\)) & 17.95 \(\pm\) 3.53 & 15.00 \(\pm\) 2.33 & 16.16 \(\pm\) 2.04 & 0.035 & 0.422 & 1.000 \\ \hline
6 MWD (m) & 473.05 \(\pm\) 46.51 & 419.18 \(\pm\) 54.50 & 406 \(\pm\) 69.50 & 0.036 & 0.012 & 1.000 \\ \hline WHO Functional class 1/2/3/4—at baseline & NA & \begin{tabular}{c} 0/8/3/0 \\ 2.00 (2.00, 3.00) \\ \end{tabular} & \begin{tabular}{c} 1/4/3/1 \\ 2.44 \(\pm\) 0.88 \\ \end{tabular} & NA & NA & 0.656 \\ WHO Functional class 1/2/3/4—after 1 year & NA & \begin{tabular}{c} 0/10/1/0 \\ 2.00 (2.00, 2.00) \\ \end{tabular} & \
\begin{tabular}{c} 0/1/4/4 \\ 3.33 \(\pm\) 0.70 \\ \end{tabular} & NA & NA & 0.001 \\ \hline BNP level (pg/mL) & 9.00 (9.00, 15.60) &
\begin{tabular}{c} 17.90 (10.60, \\ 56.40) \\ \end{tabular} & 53.38 \(\pm\) 27.14 & 0.243 & 0.0001 & 0.018 \\ \hline \end{tabular}  
\begin{tabular}{c c} \hline Data are presented as means \(\pm\) SD or median (25 th, 75 th percentiles): BMI, body mass index; BNP, brain natriuretic peptide; BSA, body surface area; \(c\), control; H, height; HR, heart rate; \(w\), non-worsening; PAH, pulmonary arterial hypertension; p\({}_{1}\), non-worsening group vs. control; p\({}_{2}\), worsening group vs. control; p\({}_{3}\), worsening group vs. non-worsening; W, weight; w, worsening; y, year; WHO, World Health Organization; 6 MWD, 6-min walk distance; variables are expressed as means \(\pm\) SD, medians (25 th, 75 th percentiles) or as numbers (percentages).  
\end{table}
Table 1: General characteristics of the children from the study groups (control, non-worsening PAH, children and worsening PAH children).