Myocardial late gadolinium enhancement is associated with clinical presentation in Duchenne muscular dystrophy carriers

DMD-carriers listed in the data base of the Gottfried von Preyer Children’s Hospital, an Austrian tertiary referral centre for neuromuscular diseases in children, were invited to participate in the study after giving written informed consent. Age, body weight and height were recored and the body surface area and body mass index were calculated. The patient’s medical records were used to determine the type of genetic mutation.

The inclusion criteria were: genetic and/or histological identification as a carrier of DMD; aged 18 years and above; provision in writing of informed consent; and a negative pregnancy test in women of childbearing potential. The exclusion criteria were: claustrophobia; excessive obesity to an extent that precluded performance of CMR; chronic renal failure with a GFR <30 ml/min/1,73 m²; implanted pacemakers/defibrillators or other conditions precluding the carriers from CMR assessment; severe arrhythmia; and inability to cooperate during CMR.

Prior to CMR, venous blood was drawn to determine blood count and blood chemistry, especially creatine-kinase (CK), creatine-kinase MB (CK-MB), creatinine, glomerular filtration rate and pro-brain natriuretic peptide (pro-BNP).

Echocardiography was performed using scanners (Vivid 7, GE Healthcare) with a 3.5-MHz transducer in standard imaging planes. Systolic LVF was measured using biplane Simpson’s method, while diastolic LVF was assessed by using the inflow signal at the mitral valve and by tissue Doppler imaging. Diastolic dysfunction was considered to be present if at least half of the following parameters exceeded their respective cut-off value: septal e’ < 7 cm/s, lateral e’ < 10 cm/s, average E/e’ ratio > 14, left atrium volume index > 34 ml/m², tricuscpid regurgitation velocity > 2,8 m/s [20]. Valvular function was evaluated by means of 2D images and colour-coded Doppler.

All patients underwent CMR examinations on a 1.5 T scanner (MAGNETOM Avanto, Siemens Healthcare GmbH, Erlangen, Germany). The tests comprised standard protocols including LGE (0.1 mmol/kg gadobutrol [Gadovist®; Bayer Vital GmbH, Leverkusen, Germany]) if the eGFR was >30 ml/min/1.73 m². Left and right atrial volumes were assessed using the biplane area-length method. At the time of inserting the intravenous cannula, blood was drawn for the measurement of haematocrit and serum creatinine levels. LGE was measured on short axis stacks using a semi-automatic approach set at a threshold of five standard deviations above the mean signal intensity of healthy myocardium.

T1-mapping was performed using electrocardiographically triggered MOLLI based on a 5(3) 3 prototype (five acquisition heartbeats are followed by three recovery heartbeats and a further three acquisition heartbeats) on a short axis mid-cavity slice and a four-chamber view. This method generates an inline, pixel-based T1 map by acquiring a series of images over several heartbeats with shifted T1 times, inline motion correction and inline calculation of the T1 relaxation curve during one breath-hold. The T1 sequence parameters were: starting inversion time (TI) 120 ms; TI increment 80 ms; reconstructed matrix size 256 × 218; and measured matrix size 256 × 144 (phase encoding resolution 66 %, phase encoding field of view 85 %). T1 maps were created both before and 15 min after the application of the contrast agent. For post contrast T1-mapping, a 4(1) 3(1) 2 prototype was used. Regions of interest were defined as left ventricular myocardium without LGE in order to study the prognostic impact of diffuse extracellular matrix expansion that could not be detected during visual assessment. T1 values (ms) of the blood pool were obtained at a sufficient distance from the papillary muscles and the endomyocardial border. Mean T1 values from the short axis and the four-chamber view were calculated CMR-ECV was calculated using the formula [21]:where “T1 myo pre”/“T1 blood pre” indicates myocardial/blood native T1 times and “T1 myo post”/“T1 blood post” indicates T1 times of myocardium/blood 15 min after application of the contrast agent. ECV was considered normal ≤28.5 % [22]. We used a dedicated software (cmr42, Circle Cardiovascular Imaging Inc., Calgary, Alberta, Canada) for further CMR analysis.

The ventricular function was determined on short axis stacks using Simpson’s rule. The epicardial border of the left ventricle was traced in order to determine ventricular mass, multiplying the wall volume by the specific density of the myocardium (1,05 g/cm³).

At each visit blood was drawn to determine the parameters mentioned above and the patients underwent clinical examination. The clinical stage was recorded according to the New York Heart Association (NYHA) classification. Echo (P.W.) and CMR (B.F., S.P.) analyses were performed by experienced observers blinded to the pro-BNP values. All patients were scheduled for a follow-up examination 12 months later. If clinically indicated, additional follow-up examinations (clinical, echocardiography, CMR) were performed. In symptomatic patients with signs of myocardial involvement medical therapy of heart failure was initiated according to the ESC guidelines for the treatment of heart failure.

Statistical analysis was performed using the SPSS package (SPSS 22, IBM Corp., Armonk, New York, USA). In normal distribution, data for continuous variables were expressed as mean ± standard deviation (SD) and intergroup comparisons were carried out using a two-sided unpaired Student’s t-test. Categorical parameters were compared using the chi-square test. Numeric changes over time were assessed by a paired t-test. Differences were considered statistically significant at a p-value < 0.05.

Twenty-two carriers agreed to participate in the study. CMR could not be performed in two carriers for reasons of claustrophobia; so they were excluded from the study. The demographic characteristics of the 20 subjects remaining (mean age 39.47 ± 12.96 years, range 21 to 62 years) are shown in Table 1. Seventeen of the carriers included (85.0 %) were clinically asymptomatic, two were in NYHA class II and one in NYHA III; the latter also displayed neurological symptoms (weakness of the thigh muscles).

Overall, mean ECV was mildly elevated (29.79 ± 2.92 %) with abnormal values in 65.0 % of all cases. These subjects did not differ from those with normal values in regard to clinical and laboratory parameters of cardiac function (data not shown). In nine carriers (45.0 %) LGE was visible in the myocardium of the left ventricle. This group presented higher septum thickness and lower EF in CMR, had higher systolic blood pressure, higher blood levels of CK and CK-MB and shorter walking distances at 6MWT. 90.9 % (n = 10) of the subjects without LGE had normal pro-BNP, whereas 66.7 % (n = 6) of those with LGE had elevated pro-BNP values (p = 0.027). Pro-BNP tended to be higher as well. As a marker of diffuse myocardial fibrosis ECV did not differ significantly between patients with and without LGE (Table 1).

Comparing dyspnea, all 11 individuals without LGE were in NYHA class I, whereas in all subjects in NYHA II (n = 2) and III (n = 1) LGE was present (p = 0.066).

Dividing the cohort in those below (n = 13 [65.0 %]) and those above (n = 7 [25.0 %]) the age-corrected cut-off value of pro-BNP for females, the latter had higher CK and CK-MB levels and shorter distances covered during the 6MWT (421.29 ± 105.20 vs. 508.23 ± 66.03 m, p = 0.035). Septum thickness was higher and ejection fraction in CMR lower. Systolic blood pressure and ECV were comparable (Table 2). LGE was present in 85.7 % of subjects with elevated pro-BNP compared to 23.1 % in those with normal levels (p = 0.027).

The genetic analysis and the distribution of LGE are shown in Table 3.

All subjects were alive 12 months after the initial visit and no major adverse cardiac events were reported. Only eight subjects came to the scheduled follow-up examination, seven of whom had had no LGE visible at the initial CMR examination. The main reasons given for dropping out of the study were social in origin, relating to caring for the diseased child and psychological distress following a son’s death. The data, thus, do not suffice for purposes of statistical analysis. However, none of the women with negative CMR developed LGE during the follow-up period. Overall, no substantial clinically relevant deterioration was observed and ECV remained practically unchanged (−1,84 ± 1.81 %); the subject diagnosed with LGE during the initial examination (#1) walked a shorter distance during the 6MWT conducted at the follow-up examination (163 vs. 210 m), but no change was detected in terms of pro-BNP, LVF or ECV.

DMD is a rare disease; its carriers are regularly detected either fortuitously or in the course of a diseased relative being examined. In addition, the women so affected have to bear the burden of witnessing their sons as they progressively waste away and the responsibility of providing ever-increasing care. Thus, the number of subjects who were willing to participate fully in our study was low. In particular, it proved impossible in most cases to conduct the follow-up examinations scheduled for 12 months later. Such a small cohort also excluded special statistical applications (e.g. multivariate analysis) that would have permitted correction for confounders.

A larger cohort might have also provided a a firmer basis on which to assess a possible correlation between genotype and phenotype. In five carriers no mutation was detected; however, given the histological confirmation of DMD those carriers underwent further analysis.

Progression of myocardial fibrosis might be a slow process and a longer clinical follow-up might be necessary as to permit a prognostic interpretation of our findings and allow for the dynamic changes in diffuse and focal presentations.

To date, no precise normative values have been determined for ECV. However, on applying the highest tertile of Wong et al. [22] we have recently been able to demonstrate a prognostic impact of ECV [38]. That cut-off value was therefore applied throughout the present study.