*5.1. Transthoracic Echocardiography*

Echocardiography plays the main role in identifying LV myocardial dysfunction and serial evaluation is necessary. Regional abnormalities of LV function may be revealed by early other imaging modalities such as speckle tracking echocardiography or cardiac magnetic resonance, before the overt LV dysfunction, assessed by echocardiography [24].

By baseline echocardiography, a dilated LV has been defined in terms of standard deviations, assessed with Z-scores according to the wide variability in the age and body mass index in DMD patients. In particular, LV dysfunction is defined by a LVEF < 55% and a fractional shortening (FS) < 28% [25,26]. Correlation of 2D and 3D echo techniques for LV diastolic (LVEDV) and systolic volumes (LVESV) was significantly positive, although 3D LVEDV and LVESV were lower when compared to 2D results; meanwhile LV ejection fraction estimation resulted similar by the two methods [27]. However, fractional shortening (FS) has been considered the best surrogate of LV systolic function, with respect to LVEF, for its high reproducibility [28]. FS showed a greater intraclass coefficient (ICC) than LV EF, not depending by age and magnitude of measures [28]. Regarding diastolic function, increased mitral A-wave velocities and lower E/A ratio, lower DTI lateral peak E-wave velocities were observed in DMD patients, compared to age-matched controls [28]. Another early marker of cardiac LV dysfunction in DMD patients is the myocardial performance index (MPI) obtained using both pulse-wave Doppler (PWD) and Doppler tissue imaging (DTI). On the basis of intraclass coefficient correlation (ICC), MPI obtained with DTI was more reproducible. Speckle tracking echocardiography is a technique able to evaluate subclinical LV dysfunction before development of overt LVEF reduction and has been increasingly used in DMD patients. Myocardial strain, obtained by 2D speckle tracking echocardiography is abnormal in nearly 50% of DMD patients, showing lower global longitudinal strain (GLS) values compared to healthy children,

despite a normal LVEF [29,30]. Moreover, a decrease of 0.34% per year of GLS in DMD patients according to age has been recently reported [31]. In this prospective multicenter cross-sectional study, a difference in longitudinal, radial, and circumferential strain, respectively of 3.6%, 9%, and 3.8% between DMD children and matched control subjects was observed with significantly lower values in the inferolateral and anterolateral mid-basal segments [31]. Several retrospective studies previously analyzed circumferential and longitudinal strain in DMD patients, using 2D speckle tracking, with a larger magnitude of difference for these indices [25,28]. However, speckle tracking analysis is often limited in DMD because echocardiographic image quality is poor in these patients and declines by 2.5% for each 1-year increase in age [32]. Poor echocardiographic window is due to chest deformities, lung hyperinflation, and limited mobility. A suboptimal echocardiographic quality, defined as more than 30% of segments inadequately visualized, was found in 50% of 13-years-old DMD patients and 78% of 15-years-old patients [32]. Indeed, LV ejection fraction, obtained by echocardiography, has been demonstrated to correlate poorly with cardiac magnetic resonance (CMR) [26], while two-dimensional fractional shortening and 5/6 area length LVEF correlated strongly with CMR LVEF [25]. Echocardiographic reproducibility of FS and 5/6 area length LVEF has been demonstrated in DMD patients, although seemed to underestimate LV function compared to CMR [25].

Right ventricular (RV) function is often preserved in DMD patients, also in presence of LV dysfunction, probably because of the reduced afterload related to respiratory improvements [33]. However, right ventricle is well studied by CMR because of its high spatial resolution and reproducible data on RV myocardial deformation. In detail, Mehmood et al. [33] reported RV normal values in patients with severe LV dysfunction, and only in few cases advanced RV dysfunction.

#### *5.2. Cardiac Magnetic Resonance*

Cardiac magnetic resonance is assuming an increasingly important role in DMD DCM, for the ability to identify myocardial fibrosis. Both 2D and 3D LV echocardiographic ejection fractions have a low correlation with CMR LVEF [25], and segmental analysis underestimated wall motion abnormalities detected by CMR. Indeed, CMR, not being limited by body habitus, can provide a complete and more accurate three-dimensional analysis of global and segmental LV function if compared to echocardiography with a better reproducibility. Different clinical settings have recently proved the utility of CMR in DMD patients, such as to stratify severity of myocardial involvement, or to assess the efficacy of anti-remodeling therapy in multicenter trials [34,35] to screen asymptomatic DMD female carriers [36,37], to evaluate perioperative cardiac risk. CMR allows a non-invasive myocardial tissue characterization by LGE and T1 mapping techniques, using non-ionizing radiations. The presence of a transmural LGE pattern, often located at the infero-lateral wall, is an independent predictor of adverse cardiac events in DMD patients, also in those with a preserved LVEF [38]. Furthermore, LGE pattern and distribution, ranging from subepicardial to transmural involvement, stratifies the degree of LV dysfunction severity. The presence of subepicardial LGE in the inferolateral free LV wall is a common finding in CMR of DMD patients [39] (Figure 1). In nearly 45% of DMD female carriers, a similar LGE distribution is observed [36,37,40] and it is also associated with a higher clinical class severity and myocardial enzyme release [41]. CMR is also useful to follow the development of LGE over time in DMD patients and carriers able to predict early LVEF decline, considering the higher LVEF reduction described in DMD patients with LGE, independently from age and steroid therapy [42]. However, LGE can detect focal macroscopic fibrosis, while T1 mapping technique preand post-contrast is able to quantify diffuse myocardial fibrosis and extracellular volume expansion (ECV). In DMD patients, T1 mapping has been demonstrated to identify early myocardial fibrosis in absence of LGE [43]. Nevertheless, significant differences in T1 mapping values were observed according to the type of T1 mapping sequence used. Previously, Soslow et al. [44] reported increased ECV values in DMD patients if compared with controls, even in cases of preserved LVEF and in the absence of LGE. Olivieri et al. [43] demonstrated the ability of native T1 mapping, by SASHA and MOLLI sequences, to stratify the presence of fibrosis also in LGE-absence. Thus, T1 mapping in DMD

patients is a surrogate marker of early subclinical involvement detectable before LGE development without the need of contrast. However, T1 mapping has several limitations depending on the type of sequence used, heart rate, different vendors, and inability to discriminate diffuse myocardial fibrosis from inflammation or fat infiltration [45,46]. Finally, myocardial strain analysis may also be obtained by CMR, using feature-tracking technique. Circumferential global myocardial strain has been detected to be more impaired in DMD patients compared to controls, with more pronounced differences in anterolateral, inferolateral, and inferior segments [47]. On the contrary of 2D speckle tracking derived strain, CMR-FT is able to discriminate different values between LGE–positive and LGE-negative areas in DMD patients compared to controls, and also among segments within LGE areas [48].

**Figure 1.** Clinical features of Duchenne muscular distrophy cardiomyopathy (DMD-DCM). Panel (**A**): typical EKG with sinus tachycardia and tall R waves. Panel (**B**): parasternal long axis view of left ventricle (LV). Arrows indicate the presence of posterior wall aneurysm. Panel (**C**): cardiac magnetic resonance: short axis view of the LV. Presence of a transmural late gadolinium enhancement pattern located at the infero-lateral wall (Courtesy of Dr. A. Secinaro).

Cardiac magnetic resonance in DMD DCM is important not only to detect early myocardial changes in case of subtle LV dysfunction, but also to evaluate progression of fibrosis in DMD patients on medical treatment [34,35]. However, the high costs, patient's claustrophobia and the length of the CMR study often limit its use in many clinical centers.
