2D Echocardiography

For speckle tracking analysis, apical four- and two-chamber views images were obtained using conventional two-dimensional gray scale echocardiography, during breath hold with a stable ECG recording. Particular attention was given to obtain an adequate gray scale image, allowing reliable delineation of myocardial tissue and extracardiac structures. Three consecutive heart cycles were recorded and averaged. The frame rate was set between 60 and 80 frames per second. These settings are recommended to combine temporal resolution with adequate spatial definition, and to enhance the feasibility of the frame-to-frame tracking technique[13 (link)].
Recordings were processed using an acoustic-tracking software (Echo Pac, GE, USA), allowing off-line semi-automated analysis of speckle-based strain[14 (link),15 (link)] (Figure 1). Briefly, LA endocardial surface is manually traced in both four- and two-chamber views by a point-and-click approach. An epicardial surface tracing is then automatically generated by the system, thus creating a region of interest (ROI). After manual adjustment of ROI width and shape, the software divides the ROI into 6 segments, and the resulting tracking quality for each segment is automatically scored as either acceptable or non-acceptable, with the possibility of further manual correction. Segments in which no adequate image quality can be obtained are rejected by the software and excluded from the analysis. Lastly, the software generates strain curves for each atrial segment. In subjects with adequate image quality, a total of 12 segments were then analyzed. To trace the ROI in the discontinuity of LA wall corresponding to pulmonary veins and LA appendage, the direction of LA endocardial and epicardial surfaces at the junction with these structures was extrapolated. Peak atrial longitudinal strain (PALS) was calculated by averaging values observed in all LA segments (global PALS), and by separately averaging values observed in 4- and 2-chamber views (4- and 2-chamber average PALS) (Figure 2). The time to peak longitudinal strain (TPLS) was also measured as the average of all 12 segments (global TPLS) and by separately averaging values observed in the two apical views (4- and 2-chamber average TPLS). In patients in whom some segments were excluded because of the impossibility of achieving adequate tracking, PALS and TPLS were calculated by averaging values measured in the remaining segments.
Reproducibility of PALS and TPLS measurements was assessed in 20 randomly selected subjects. Intra and inter-observer variability coefficients were calculated using images independently recorded in two different occasions by the same investigator or by two different observers.

Baseline clinical information, including age, sex, body mass index (BMI), disease duration, comorbidities, medication, and LV ejection fraction (LVEF) obtained from 2D echocardiography [27 (link)], from each subject was recorded. Participants had blood sampling before the baseline CMR-LGE imaging study and then underwent a graded cardiopulmonary exercise test (CPET). The physical component score (PCS) and mental component score (MCS) of the Medical Outcomes Study Short Form-36 health survey (SF-36) for quality of life (QoL) were assessed before initiating each CPET. The follow-up CMR-LGE, CPET, and blood samplings were performed within 1 week after completing 36 sessions of HIIT. Haematocrit and b-type natriuretic peptide (BNP) were also measured before and after HIIT. After completing the above study, the remaining blood sample was centrifuged at 2500 rpm for 5 min at room temperature for serum preparation. A graphic depicting the experimental procedure is shown in Additional file 1.