Background
The Pediatric Heart Network is conducting a large international randomized trial to compare aortic root growth and other cardiovascular outcomes in 608 subjects with Marfan syndrome randomized to receive atenolol or losartan for 3 years. The authors report here the echocardiographic methods and baseline echocardiographic characteristics of the randomized subjects, describe the interobserver agreement of aortic measurements, and identify factors influencing agreement.
Methods
Individuals aged 6 months to 25 years who met the original Ghent criteria and had body surface area–adjusted maximum aortic root diameter (ROOT max ) Z scores > 3 were eligible for inclusion. The primary outcome measure for the trial is the change over time in ROOT max Z score. A detailed echocardiographic protocol was established and implemented across 22 centers, with an extensive training and quality review process.
Results
Interobserver agreement for the aortic measurements was excellent, with intraclass correlation coefficients ranging from 0.921 to 0.989. Lower interobserver percentage error in ROOT max measurements was independently associated (model R 2 = 0.15) with better image quality ( P = .002) and later study reading date ( P < .001). Echocardiographic characteristics of the randomized subjects did not differ by treatment arm. Subjects with ROOT max Z scores ≥ 4.5 (36%) were more likely to have mitral valve prolapse and dilation of the main pulmonary artery and left ventricle, but there were no differences in aortic regurgitation, aortic stiffness indices, mitral regurgitation, or left ventricular function compared with subjects with ROOT max Z scores < 4.5.
Conclusions
The echocardiographic methodology, training, and quality review process resulted in a robust evaluation of aortic root dimensions, with excellent reproducibility.
Marfan syndrome (MFS) is a systemic disorder of connective tissue caused by mutations in FBN1 , the gene encoding fibrillin-1. The leading cause of mortality in MFS is cardiovascular disease, including aortic root dilation and aortic dissection. The National Heart, Lung, and Blood Institute–funded Pediatric Heart Network is conducting a randomized trial to compare the rates of aortic root enlargement and other short-term cardiovascular outcomes in children and young adults with MFS randomized to receive atenolol or losartan for 3 years. The rationale and design of this randomized trial have been reported.
The primary outcome measure of this trial is the change over time (slope) in maximum aortic root diameter (ROOT max ) Z score measured by echocardiography at the level of the sinuses of Valsalva. Accurate and reproducible measurement of the aortic root diameter is particularly important in patients with MFS because aortic root size is one of the best predictors of cardiovascular outcome. Factors that may adversely affect intraobserver and interobserver agreement of aortic measurements in patients with MFS include increasing age and body size, pectus abnormality or scoliosis, and severity of aortic root dilation.
We report here the specifics of the echocardiographic methods used in this trial, including the training and quality review process, a detailed analysis of the interobserver and intraobserver agreement in baseline aortic measurements, and the factors influencing agreement in this cohort. The large sample size, measurement of all aortic dimensions in systole and diastole, as well as the analysis of multiple beats are distinguishing features of this trial. A detailed description of the baseline clinical characteristics of the screened population and enrolled subjects is being reported separately.
Methods
Patients
Subjects enrolled in this trial were individuals aged 6 months to 25 years who met the original Ghent criteria for MFS, with a body surface area (BSA)–adjusted ROOT max Z score > 3 and absolute ROOT max < 5 cm. Patients with prior aortic surgery were excluded. A total of 608 subjects were enrolled between February 2007 and February 2011. The study protocol was approved by the institutional review board or institutional ethics board at each participating center, and informed consent and assent were obtained, depending on age, from patients and their parents or legal guardians before trial enrollment.
Echocardiographic Protocol
The echocardiographic technical protocol and study manual were developed after consensus was reached among several experts in the field. The detailed protocol is provided in the online Appendix (available at www.onlinejase.com ). Echocardiograms were performed on study subjects at baseline and at 6 months, 1 year, 2 years, and 3 years after randomization to either atenolol or losartan. Before baseline echocardiography, all patients on prophylactic therapy for aortic root dilation underwent taper and washout according to protocol. The data analyses presented here are based on the baseline echocardiographic studies only.
Patient length in centimeters and weight in kilograms were measured by an MFS trial study coordinator at the time of echocardiography and were used to calculate BSA. An automated vital sign device (Dinamap; GE Medical Systems, Waukesha, WI) was used to record multiple right brachial blood pressures and heart rates during echocardiographic assessment. Blood pressure and heart rate were recorded after the patient had been in a recumbent position for 5 to 10 min, during or immediately after recording of aortic images for calculation of stiffness indices. Four samples were obtained, the first of which was discarded (because it is the least reliable). The other samples were averaged.
Complete echocardiographic studies were performed using local instrumentation, transducer selection, and machine settings that provided the optimal images on the basis of the judgment of the ultrasonographer. Harmonic imaging and lateral gain were used when they helped define structures such as endocardial borders. Specific instructions for image optimization were given to the centers, including the following: use of zoom mode to optimize screen resolution when anatomic structures were to be measured, recording of transitions from full screen to zoom and from two-dimensional (2D) to spectral Doppler to enable the core laboratory readers to identify structures and sample locations, use of full-screen M mode to optimize distance-axis screen resolution, use of the highest sweep speed on M-mode and spectral Doppler recordings to optimize time-axis screen resolution, use of baseline velocity and wall-filter adjustment on spectral Doppler, adequate electrocardiographic strip for documentation of heart rate, and recording without the local measurements, if possible, to permit independent review by the core laboratory readers. For each structural measurement, 6 to 10 cardiac cycles were recorded, and for color Doppler evaluation of the valves, 10 to 15 cardiac cycles were recorded. ROOT max was measured locally in triplicate and averaged to calculate a Z score to determine eligibility before randomization.
Core Laboratory Analysis
Measurements were performed in the core laboratory on a microcomputer-based workstation custom programmed for electronic caliper overlay of captured digital images for recording (EchoTrace; Marcus Laboratories, Boston, MA). The aortic diameters were measured at their maximum and minimum dimensions in systole and diastole from inner edge to inner edge at the aortic valve annulus, aortic root at the sinuses of Valsalva, sinotubular junction, and ascending aorta ( Figure 1 ). Each of these eight measurements was performed in triplicate by two independent observers.
The anteroposterior diameter of the distal thoracic aorta was measured at the level of the diaphragm. The lateral diameter of the main pulmonary artery (MPA) was measured from a parasternal imaging window. The durations of diastolic antegrade and retrograde flow in the proximal and distal descending thoracic aorta were measured. Left ventricular (LV) size and functional parameters were calculated according to American Society of Echocardiography pediatric guidelines in the apical and parasternal short-axis views.
Mitral valve prolapse (MVP) or tricuspid valve prolapse (TVP) was classified as present if all or part of a valve leaflet was observed to pass through the plane of the valve annulus in the parasternal long-axis imaging plane. Prolapse was classified as borderline if one of the valve leaflets was observed to manifest posterior motion relative to the other leaflet without passing through the plane of the valve annulus. The degree of mitral regurgitation (MR) and aortic regurgitation (AR) were categorized qualitatively as mild or more, trivial, or none.
The eight averaged aortic measurements were reported for each of the two independent observers. Z scores were available for the maximum dimensions only. Raw values and Z scores were calculated for LV dimensions, volumes, mass, shortening fraction, and ejection fraction.
Percentage duration of diastolic flow reversal in the proximal and distal thoracic aorta was calculated. The arterial pressure-strain elastic modulus and stiffness index were calculated for the aortic root at the sinuses of Valsalva and the ascending aorta. The averages of the systolic, diastolic, and mean blood pressures along with corresponding Z scores (Boston Children’s Hospital normative database) were calculated.
Training and Quality Control
The training process for site investigators included a face-to-face session with site investigators and a webinar session during which samples of each recording were reviewed in detail. An instructional CD-ROM was also distributed to all sites. As part of quality control, each site went through a certification process whereby three local aortic root measurements in addition to three full study protocol echocardiograms were reviewed at the core laboratory. A detailed review letter with constructive feedback was provided to each site.
Two core laboratory readers blinded to treatment assignment independently reviewed all echocardiograms. Each study was graded for image quality (excellent, good, fair, or unacceptable) and variation from protocol. A random subset (8%) of the studies was reviewed twice by each core laboratory reader to assess intrareader agreement.
Statistical Methods
Summary statistics are presented as mean ± SD or as median (interquartile range [IQR]). Continuous variables were compared by treatment assignment and other characteristics (e.g., gender, family history of MFS) using Student’s t tests or Wilcoxon’s rank-sum tests for approximately normally distributed and skewed variables, respectively. Categorical variables were compared by treatment assignment and other subgroup factors using χ 2 or Fisher’s exact tests. To examine the associations of age and most echocardiographic indices, we compared growing children (male < 16.0 years of age, female < 15.0 years of age) with all others (male ≥ 16.0 years of age, female ≥ 15.0 years of age). For aortic elastic modulus and stiffness index, we examined differences in outcomes by age using age quartiles. Group comparisons that required age adjustment used continuous age and were performed using analysis of covariance and logistic regression for continuous and dichotomous measures, respectively. Echocardiographic and blood pressure Z scores for BSA and/or age were derived from the Boston Children’s Hospital normative database and compared with a healthy population mean of zero using a one-sample t test or Wilcoxon’s signed-rank test. Interobserver and intraobserver agreement was estimated using intraclass correlation coefficients (ICCs) and 95% confidence intervals. Agreement was also estimated by a percentage error measurement, defined as the absolute value of the difference in the two measurements divided by the mean of the two measurements multiplied by 100. To identify factors that might influence interobserver agreement, a multivariate model using percentage error was constructed (defined as the absolute difference between raters divided by the mean across raters multiplied by 100). To assess the robustness of the findings, a secondary analysis of group differences and ICC estimates was conducted using weighting according to image quality, with higher quality images having greater weight. This secondary analysis had little impact on statistical inferences. Because of the large number of comparisons, P values ≤ .01 were considered statistically significant. Analyses were conducted using R version 2.14.1 (R Foundation for Statistical Computing, Vienna, Austria) and SAS version 9.3 (SAS Institute Inc., Cary, NC).
Results
Imaging of the aortic root was graded as excellent in 203 (33%), good in 357 (59%), and fair in 47 (8%) patients. ROOT max was measurable at least once in all echocardiograms and in triplicate in 95%.
Agreement of the Echocardiographic Measurements
Interobserver Agreement
Interobserver agreement for the aortic measurements was excellent, with ICCs between the two core laboratory readers ranging from 0.921 to 0.989 ( Table 1 ). Bland-Altman plots for interobserver agreement on all aortic measurements depict excellent agreement, with a systematic bias wherein the primary reader obtained lower values than the secondary reader ( Figures 2 A and 2 B). This difference was slight, with mean interobserver differences ranging from 0.5 to 2.0 mm. However, the plots do not reveal any other systematic trends; specifically, the degree of agreement does not depend on the magnitude of the measurement. The Bland-Altman plots were similar when absolute dimensions, percentage error, and Z scores (not shown) were used.
| Variable | n | ICC (95% CI) |
|---|---|---|
| Minimum aortic annular diameter | 592 | 0.962 (0.955–0.967) |
| Maximum aortic annular diameter | 589 | 0.921 (0.907–0.933) |
| Minimum aortic root diameter | 600 | 0.987 (0.985–0.989) |
| ROOT max | 607 | 0.989 (0.987–0.990) |
| Minimum sinotubular junction diameter | 487 | 0.952 (0.942–0.960) |
| Maximum sinotubular junction diameter | 538 | 0.948 (0.939–0.956) |
| Minimum ascending aortic diameter | 483 | 0.972 (0.966–0.976) |
| Maximum ascending aortic diameter | 525 | 0.971 (0.966–0.976) |
| Ascending aorta elastic modulus ∗ | 480 | 0.626 (0.533–0.688) |
| Ascending aorta stiffness index ∗ | 480 | 0.589 (0.509–0.657) |
| Aortic root elastic modulus ∗ | 598 | 0.708 (0.658–0.752) |
| Aortic root stiffness index ∗ | 598 | 0.676 (0.619–0.724) |
The univariate analysis showed that the interobserver percentage error in ROOT max measurements was significantly lower by later read date and differed by center ( P = .003 and P < .001, respectively). The other variables examined include image quality, age at echocardiography, BSA, absolute aortic root dimension, presence of pectus deformity or scoliosis, number of Ghent criteria met, family history of MFS, and presence of FBN1 mutation. The multivariate model showed that lower interobserver percentage error in ROOT max measurements was only independently associated (model R 2 = 0.15) with better image quality ( P = .002) and later study reading date ( P < .001).
Stiffness Indices
Interobserver agreement for aortic stiffness indices was moderate, with ICCs between the two core laboratory readers ranging from 0.59 to 0.71 for the ascending aorta and aortic root ( Table 1 ).
One Beat versus Three Beats
In 95% of the echocardiographic examinations, the primary reader obtained three measurements of the primary outcome (ROOT max ). Percentage error was significantly lower for all averaged aortic measurements compared with single-beat measurements, except for maximum aortic annulus and maximum sinotubular junction ( P ≤ .01; data not shown). For example, median percentage error for three-beat versus single-beat ROOT max dimension was 3.6 ± 2.6% versus 3.9 ± 3.0% ( P = .0002).
Intraobserver Agreement of the Aortic Root and MPA Measurements
Intraobserver agreement for aortic root ( Figure 3 ) and MPA measurements was excellent for both primary and secondary readers, with all ICCs > 0.90.
Interobserver Agreement between the Local Echocardiographic Laboratory and the Core Laboratory
The only reported measurement from the local laboratory was the ROOT max used to determine eligibility. Interobserver agreement for the ROOT max measurement between the local and core laboratories was near perfect (ICC, 0.988; 95% confidence interval, 0.986–0.990; 3.36 ± 0.71 vs 3.35 ± 0.70; P = .55).
Echocardiographic Characteristics of the Randomized Cohort
The echocardiographic characteristics of the randomized subjects did not differ by treatment arm ( Table 2 ). By design, randomized subjects had larger aortic dimensions ( Z score = 4.04; IQR, 3.41 to 4.92) than healthy children (mean Z score = 0). MPA dimension Z scores were also larger but to a lesser degree (mean Z score = 2.22 ± 1.34). All median and mean values for LV size and function were within 1 standard deviation of mean values for healthy children, except for diastolic septal and posterior wall thickness Z scores (−1.20 [IQR, −1.83 to 0.45] and −1.22 ± 1.02, respectively).
| Characteristic | Aggregate ( n = 608) | Treatment A ( n = 303) | Treatment B ( n = 305) | P | |||
|---|---|---|---|---|---|---|---|
| n | Value | n | Value | n | Value | ||
| Aortic annular maximum diameter Z score | 589 | 1.76 ± 1.30 † | 296 | 1.71 ± 1.24 | 293 | 1.81 ± 1.36 | .32 |
| Aortic annular maximum diameter (cm) | 589 | 2.01 ± 0.42 | 296 | 2.01 ± 0.42 | 293 | 2.00 ± 0.41 | .79 |
| ROOT max Z score (sinuses of Valsalva) | 607 | 4.04 (3.41 to 4.92) † | 303 | 4.04 (3.47 to 4.82) | 304 | 4.04 (3.34 to 5.01) | .68 ∗ |
| ROOT max (cm) (sinuses of Valsalva) | 607 | 3.36 ± 0.71 | 303 | 3.37 ± 0.72 | 304 | 3.36 ± 0.71 | .96 |
| Aortic sinotubular junction maximum diameter Z score | 552 | 1.98 (1.32 to 2.72) † | 276 | 1.94 (1.35 to 2.58) | 276 | 2.04 (1.30 to 2.85) | .14 ∗ |
| Aortic sinotubular junction maximum diameter (cm) | 552 | 2.41 ± 0.53 | 276 | 2.40 ± 0.54 | 276 | 2.43 ± 0.52 | .51 |
| Ascending aortic maximum diameter Z score | 543 | 0.88 (0.38 to 1.47) † | 263 | 0.80 (0.37 to 1.38) | 280 | 0.91 (0.40 to 1.56) | .13 ∗ |
| Ascending aortic maximum diameter (cm) | 543 | 2.30 (1.94 to 2.60) | 263 | 2.30 (1.93 to 2.56) | 280 | 2.30 (1.97 to 2.63) | .62 ∗ |
| Descending aortic maximum diameter (cm) | 592 | 1.31 ± 0.33 | 296 | 1.32 ± 0.34 | 296 | 1.31 ± 0.32 | .92 |
| Percentage duration of diastolic flow reversal in PTA | 319 | 17.8 (10.3 to 23.4) | 158 | 17.3 (4.7 to 23.2) | 161 | 18.3 (11.7 to 23.4) | .28 ∗ |
| Percentage duration of diastolic flow reversal in DTA | 513 | 0.0 (0.0 to 9.4) | 259 | 0.0 (0.0 to 9.0) | 254 | 0.0 (0.0 to 10.6) | .58 ∗ |
| MPA maximum diameter (cm) | 512 | 2.36 ± 0.46 | 261 | 2.37 ± 0.47 | 251 | 2.34 ± 0.45 | .47 |
| MPA maximum diameter Z score | 512 | 2.22 ± 1.34 † | 261 | 2.19 ± 1.36 | 251 | 2.26 ± 1.32 | .55 |
| Presence of AR | 607 | 67 (11%) | 302 | 35 (12%) | 305 | 32 (11%) | .70 |
| Among subjects with AR | 67 | 35 | 32 | .29 | |||
| Trivial | 47 (70%) | 27 (77%) | 20 (62%) | MH = .19 | |||
| Mild or more | 20 (30%) | 8 (23%) | 12 (38%) | ||||
| Presence of MR | 597 | 384 (64%) | 298 | 195 (65%) | 299 | 189 (63%) | .61 |
| Among subjects with MR | 384 | 195 | 189 | .09 | |||
| Trivial | 278 (72%) | 149 (76%) | 129 (68%) | MH = .07 | |||
| Mild or more | 106 (28%) | 46 (24%) | 60 (32%) | ||||
| MVP | 596 | 298 | 298 | .38 | |||
| None | 230 (39%) | 114 (38%) | 116 (39%) | MH = .45 | |||
| Borderline | 140 (24%) | 64 (22%) | 76 (25%) | ||||
| Present | 226 (38%) | 120 (40%) | 106 (36%) | ||||
| TVP | 551 | 276 | 275 | .18 | |||
| None | 281 (51%) | 143 (52%) | 138 (50%) | MH = .66 | |||
| Borderline | 149 (27%) | 66 (24%) | 83 (30%) | ||||
| Present | 121 (22%) | 67 (24%) | 54 (20%) | ||||
| LV size and function | |||||||
| LV end-diastolic volume Z score | 521 | −0.23 ± 1.38 † | 260 | −0.25 ± 1.36 | 261 | −0.21 ± 1.39 | .69 |
| LV end-systolic volume Z score | 521 | −0.42 ± 1.36 † | 259 | −0.42 ± 1.40 | 262 | −0.43 ± 1.31 | .93 |
| LV ejection fraction (%) | 521 | 64.6 ± 6.4 | 260 | 64.4 ± 6.1 | 261 | 64.9 ± 6.6 | .44 |
| LV mass Z score | 518 | −0.25 (−0.90 to 0.68) † | 259 | −0.22 (−0.99 to 0.66) | 259 | −0.27 (−0.87 to 0.71) | .79 ∗ |
| LV mass/volume ratio (g/mL) | 519 | 0.92 ± 0.15 | 260 | 0.91 ± 0.15 | 259 | 0.92 ± 0.15 | .83 |
| LV end-diastolic dimension Z score | 585 | 0.70 (−0.14 to 1.65) † | 291 | 0.71 (−0.30 to 1.63) | 294 | 0.70 (0.01 to 1.72) | .28 ∗ |
| LV end-systolic dimension Z score | 585 | 0.35 (−0.53 to 1.22) † | 291 | 0.31 (−0.71 to 1.28) | 294 | 0.39 (−0.44 to 1.19) | .33 ∗ |
| LV mass Z score (by M-mode echocardiography) | 585 | −0.65 (−1.52 to 0.34) † | 291 | −0.74 (−1.51 to 0.27) | 294 | −0.59 (−1.56 to 0.39) | .46 ∗ |
| LV shortening fraction (%) | 587 | 37.2 ± 5.1 | 292 | 37.2 ± 5.3 | 295 | 37.2 ± 5.0 | .98 |
| LV diastolic septal thickness Z score | 585 | −1.20 (−1.83 to −0.45) † | 291 | −1.15 (−1.79 to −0.54) | 294 | −1.23 (−1.87 to −0.37) | .78 ∗ |
| LV diastolic posterior wall thickness Z score | 585 | −1.22 ± 1.02 † | 291 | −1.23 ± 1.01 | 294 | −1.21 ± 1.03 | .84 |
| Stiffness indices | |||||||
| Ascending aortic elastic modulus (mm Hg) | 492 | 318 (231 to 447) | 237 | 319 (231 to 430) | 255 | 317 (231 to 463) | .60 ∗ |
| Ascending aortic stiffness index | 492 | 4.95 ± 2.78 | 237 | 4.81 ± 2.62 | 255 | 5.08 ± 2.92 | .29 |
| Aortic root elastic modulus (mm Hg) | 598 | 618 (407 to 914) | 299 | 602 (386 to 887) | 299 | 644 (431 to 970) | .112 ∗ |
| Aortic root stiffness index | 598 | 8.2 (5.5 to 12.2) | 299 | 7.9 (5.2 to 11.8) | 299 | 8.5 (5.9 to 12.4) | .110 ∗ |
| Systolic blood pressure (mm Hg) | 606 | 97.2 ± 14.3 | 303 | 97.2 ± 13.4 | 303 | 97.2 ± 15.1 | .98 |
| Systolic blood pressure Z score | 604 | −0.65 ± 0.98 † | 303 | −0.69 ± 0.99 | 301 | −0.61 ± 0.97 | .30 |
| Diastolic blood pressure (mm Hg) | 606 | 58.6 ± 9.9 | 303 | 59.1 ± 9.9 | 303 | 58.1 ± 9.9 | .22 |
| Diastolic blood pressure Z score | 604 | 0.32 ± 0.94 † | 303 | 0.34 ± 0.97 | 301 | 0.29 ± 0.91 | .57 |
| Mean blood pressure (mm Hg) | 606 | 69.9 ± 15.4 | 303 | 70.5 ± 15.1 | 303 | 69.3 ± 15.8 | .33 |
| Mean blood pressure Z score | 593 | −0.25 ± 0.96 † | 298 | −0.24 ± 1.01 | 295 | −0.26 ± 0.92 | .81 |
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