J Cardiol 2004 Jan; 43（1）: 23 – 29

Experimental Study of Quantitative
Assessment of Left Ventricular
Mass With Contrast Enhanced
Real-Time Three-Dimensional
Echocardiography
Lei

ZHUANG,

Xin-Fang

WANG, MD

Ming-Xing

XIE, MDMD

Li-XinHong-Wen

CHEN, MD
FEI, MD

Ying

YANG, MD

Jing

WANG, MD

Run-Qing

HUANG, MD

Ou-Di

CHEN, MD

Abstract ─────────────────────────────────────────────────────────────────────────────────────────────────────────────────────────────────────────────────────────────────────
Objectives. To evaluate the feasibility and accuracy of measurement of left ventricular mass with intravenous contrast enhanced real-time three-dimensional（RT3D）echocardiography in the experimental setting.
Methods. RT3D echocardiography was performed in 13 open-chest mongrel dogs before and after intravenous infusion of a perfluorocarbon contrast agent. Left ventricular myocardium volume was measured
according to the apical four-plane method provided by TomTec 4D cardio-View RT1.0 software, then theleft ventricular mass was calculated as the myocardial volume multiplied by the relative density of
myocardium. Correlative analysis and paired t-test were performed between left ventricular mass obtained
from RT3D echocardiography and the anatomic measurements.
Results. Anatomic measurement of total left ventricular mass was 55.6 ± 9.3 g, whereas RT3D echocardiographic calculation of left ventricular mass before and after intravenous perfluorocarbon contrast agent
was 57.5 ± 11.4 and 55.5 ± 9.3 g, respectively. A significant correlation was observed between the RT3D
echocardiographic estimates of total left ventricular mass and the corresponding anatomic measurements（r ＝ 0.95）. A strong correlation was found between RT3D echocardiographic estimates of left ventricularmass with perfluorocarbon contrast and the anatomic results（r ＝ 0.99）. Analysis of intraobserver andinterobserver variability showed strong indexes of agreement in the measurement of left ventricular mass
with pre and post-contrast RT3D echocardiography.
Conclusions. Measurements of left ventricular mass derived from RT3D echocardiography with andwithout intravenous contrast showed a significant correlation with the anatomic results. Contrast enhanced
RT3D echocardiography permitted better visualization of the endocardial border, which would provide amore accurate and reliable means of determining left ventricular myocardial mass in the experimental setting.
──────────────────────────────────────────────────────────────────────────────────────────────────────────────────────────J Cardiol 2004 Jan ; 43（1）: 23−29

Key Words
Contrast echocardiography three-dimensional
Experimental medicine

Cardiac volume left ventricular mass──────────────────────────────────────────────Department of Echocardiography, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan,
China
Address for correspondence : WANG X-F, MD, Department of Echocardiography, Union Hospital, Tongji Medical College, Huazhong
University of Science and Technology, 1277 Jiefang Avenue, Wuhan, 430022, PR China
Manuscript received March 24, 2003 ; revised July 24 and September 24, 2003 ; accepted September 24, 2003

23

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Zhuang, Wang, Xie et al

INTRODUCTION
Accurate measurement of left ventricular mass isimportant to assess left ventricular remodeling during the progress of cardiovascular disease1）. Up tonow, two-dimensional（2D）echocardiography has
been essential for assessment of left ventricularmass, but the usefulness of routine echocardiography is limited by many factors such as poor
acoustic windows in 10−15% of patients, andendocardial dropout caused by body habitus andlung diseases, with the anterior wall and apex mostfrequently affected2）. Dynamic three-dimensional（3D）echocardiography may provide more accurate
and repeatable estimations of cardiac structure sizeand function than conventional 2D echocardiography imaging does3）. However, off-line dynamic 3D
echocardiography is not regarded as feasible inevery clinical practice because these techniques arelimited by lengthy data acquisition times, computer
processing times, and extensive operator interactionthat precludes the imaging of dynamic structureseven with gating techniques. Recently, real-time
three-dimensional（RT3D）echocardiography has
been developed to acquire more stereotype anatomic information4）. The advantage over 2D echocardiography is the effective and accurate way toassess left ventricular mass. This study assessed theaccuracy of measurement of left ventricular mass
with RT3D echocardiography before and after
intravenous administration of the perfluorocarbon
contrast agent in the experimental setting.
SUBJECTS AND METHODS
Animal preparation
Thirteen mongrel dogs were studied, weight 11
to 15 kg. All operative and animal management
procedures were approved by the Animal Care and
Use Committee of the National Heart Institute. The
dogs underwent intravenous sedation with sodium
pentobarbital（25 mg/kg）and subsequent endotracheal intubation and ventilation with a volumecycled ventilator. A Swan-Ganz catheter was positioned in the pulmonary artery via the femoral vein,
and another catheter was positioned in the rightcommon femoral artery for monitoring systemicarterial pressure and blood gases. Arterial bloodgases and pH were maintained within physiologic
ranges. Then a lateral thoracotomy was performed
and the heart was suspended in an epicardial cradle.
A warm water bath was placed over the anterior

surface of the heart for ultrasound scanning.
Real-time three-dimensional echocardiography
RT3D echocardiography was performed with a
Philips Sonos 7500 ultrasonographic machine（Philips Co.）. This unit uses a 2−4 MHz matrixarray transducer for rapid beam forming to scan apyramidal volume. Real-time volumetric scanningrequires no off-line reconstruction techniques,enabling dynamic three-dimensional visualizationand quantification of the heart in real time using atransthoracic approach. Images can be viewed in alive 3D or full volume mode. Usually we
perform a two-dimensional study first and select theregion of interest on the middle line, then press thefull volume button, try to include all targets andthen touch the acquire button. Triggered byelectrocardiography, the four 0−15 °, 15−30 °,30−45 °and 45−60 °adjacent 15 × 60 °narrowangle display stereo datasets are acquired in orderin the first, third, fifth and seventh cardiac cycles.
These images are combined to produce the 60 ×
60 ° pyramidal image . One full volume acquisition will take 5−8 sec. The pyramidal dataset
obtained from the apical four-chamber view cancontain the whole heart structure. During the procedure ventilation to the dog should be paused to
avoid image distortion due to respiration andmotion. Data post-process was performed by aworkstation TomTec 4D cardio-View RT1.0（TomTec, Inc.）.
Preparation of perfluorocarbon contrast agent
Perfluorocarbon-exposed sonicated dextrosealbumin was manufactured according to theapproach described previously 5）. One part 5%
human serum albumin and three parts 5% dextrose
were drawn into a 10 ml syringe. The 10 ml syringe
was attached to a three-way stopcock that was also
attached to a syringe containing 2 ml of perflurocarbon gas. Then the dextrose-albumin solutions weresterile sonicated（Sonicator, Heat Systems Inc.）at a
frequency of 20 kHz lasting for 60 sec. During sonication, the 2 ml of perfluorocarbon in syringe wasslowly introduced into the dextrose-albumin bymeans of the stopcock. After sonication, themicrobubble solution was placed in a vertical position in a 10 ml syringe until a distinct upper whitelayer and a lower clear layer formed. The lower
clear layer was decanted, and the upper white layer
was used for intraveous injections. The concentraJ Cardiol 2004 Jan; 43（1）: 23 – 29

Quantitative Assessment of Left Ventricular Masstion of microbubbles measured by Coulter counterwas 1.8 × 109/ml, the mean size of microbubbles
was 3.4 ± 1.3μm, and the volume of one bolus
injection was 3−4 ml. RT3D echocardiography
examination was performed before and after intravenous infusion of perfluorocarbon contrast agent.
Measurements of left ventricular mass with realtime three-dimensional echocardiography
The pyramidal dataset derived from the fullvolume mode permits simultaneous, multiple
plane display of two sector arcs（B-scans）and C
scan by a TomTec workstation with 4D cardioView RT 1.0 software. Calculation of the left ventricular mass from the RT3D echocardiographic
images was performed by an interactive-aided manual tracing technique to outline left ventricularmyocardium volume between the epicardial andendocardial borders according to the four-plane
method（Fig. 1）. Left ventricular mass was calculated as the myocardial volume multiplied by the
relative density of myocardium（1.05）. Usually
image acquisition under full volume mode took
less than 5 min and the procedure to calculate left
ventricular mass with 4D cardio-View RT 1.0 software needed no more than 10 min, so the wholeprocess takes 15 min. RT3D echocardiographic
images（10 dogs）pre and post contrast enhancement
were analyzed by two independent observers
unaware of the anatomic measurements. In addition, all left ventricular mass measurements wererepeated at least 1 week apart by the same observer
to allow subsequent assessment of intraobserver
variability.
Measurements of anatomic left ventricular mass
After the RT3D echocardiographic examination
was completed, the dogs were euthanized with an
overdose of pentobarbital and potassium chloride
and the heart was excised. Then the left ventricle
was isolated and cut off and weighed.
Statistical methods
All data are expressed as mean ± SD. To determine whether the difference in the values between
RT3D echocardiographic imaging and anatomic
measurements was statistically significant, a paired
t-test was performed ; the level of significance was
set to p ＜ 0.05. Linear regression analysis was used
to determine the correlation between left ventricular mass from excised heart and the results derived
J Cardiol 2004 Jan; 43（1）: 23 – 29

25

from RT3D echocardiographic images before andafter injection of ultrasonic contrast medium. Bland
and Altman analysis was used to assess agreement
between two measurements.
RESULTS
Immediately after intravenous injection, perfluorocarbon contrast microbubbles were observed in
the left ventricular cavity, and the visualization of
the left ventricular endocardial border was perfect（Fig. 2）. Anatomic measurements of total left ventricular mass ranged from 38.9 to 71.1 g（mean ±SD : 55.6 ± 9.3 g）, whereas the RT3D echocardiographic calculation of left ventricular mass ranged
from 39.3 to 82.3 g（ mean ± SD : 57.5 ± 11.4 g ;p ＝ 0.10 vs anatomic measurements）. RT3D
echocardiographic calculation of the left ventricularmass after intravenous perfluorocarbon contrast
agent ranged from 37.7 to 70.5 g（ mean ± SD :55.5 ± 9.3 g ; p ＝ 0.57 vs anatomic measurements）. There was a significant correlation between
the RT3D echocardiographic estimates of total left
ventricular mass and the corresponding anatomic
measurements（ r ＝ 0.95, mean difference 1.9 ±3.8 g ; Fig. 3）. A strong correlation was foundbetween the RT3D echocardiographic calculationof left ventricular mass with perfluorocarbon contrast and the anatomic measurements（ r ＝ 0.99,
mean difference 0.1 ± 0.9 g ; Fig. 4）.
There was a high index of interobserver（r ＝0.88, mean difference 1.6 ± 3.7 g）as well as
intraobserver（ r ＝ 0.92, mean difference 1.4 ±3.2 g）agreement for the RT3D echocardiographic
measurements of left ventricular mass. There was ahigher index of interobserver（r ＝ 0.96, mean difference − 0.5 ± 1.9 g）
as well as intraobserver（r ＝0.98, mean difference 0.4 ± 1.5 g）agreement forthe contrast enhanced RT3D echocardiographic
measurements of left ventricular mass.
DISCUSSION
Left ventricular remodeling determined byechocardiography is recognized as one of the mostpowerful predictors of morbidity and mortality inpatients with severe cardiovascular diseases. Up to
now conventional 2D echocardiography hasremained the main method for calculation of left
ventricular mass, but 2D echocardiography provides only tomographic images of the object. In
order to appreciate the three-dimensional structural
relationships, mental reconstruction of these views

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Zhuang, Wang, Xie et al

Fig. 1 Measurement of left ventricular myocardium volume from real-time three-dimensional
echocardiographic images
Myocardium volume was manually outlined along the endocardial and epicardial border on two long-axis
and one short-axis views extracted from the pyramidal dataset according to the apical four-plane method
provided by TomTec 4D cardio-View RT 1.0 software package.

Fig. 2 Measurement of left ventricular myocardium volume by real-time three-dimensional
echocardiography
After intravenous infusion of perfluorocarbon contrast, the endocardial border was visualized clearly.
A : Real-time three-dimensional echocardiographic data in the apical four-chamber view.
B : Short-axis view of ventricles.

is required by an experienced observer which limits
the ability to analyze structures of unknown orcomplex shapes. Various computer based threedimensional reconstruction techniques have beendeveloped to replace the subjective mental

reassembly of complex cardiac geometry 6）.
Dynamic 3D echocardiography could produce moreaccurate and repeatable left ventricular volume estimations without the geometric assumptions
required by conventional two-dimensional
J Cardiol 2004 Jan; 43（1）: 23 – 29

Quantitative Assessment of Left Ventricular Mass

27

Fig. 3 Relationships between real-time three-dimensional echocardiographic and anatomic measurements of left ventricular mass measured in 13 dogs
Correlation between measurements obtained by the two methods（A）. Bland and Altman plot of the difference between real-time three-dimensional echocardiographic and anatomic measurements of left ventricular mass
（B）. The dashed and solid lines indicate the mean ± 2SD.
RT3DE ＝ real-time three-dimensional echocardiography ; LV ＝ left ventricular.

Fig. 4 Relationships between contrast enhanced real-time three-dimensional echocardiographic andanatomic measurements of left ventricular mass measured in 13 dogs
Correlation between measurements obtained with the two methods（A）. Bland and Altman plot of the difference between contrast enhanced real-time three-dimensional echocardiographic and anatomic measurements of left ventricular mass
（B）. The dashed and solid lines indicate the mean ± 2 SD.
Abbreviations as in Fig. 3.

imaging7）, but the early reconstruction of threedimensional imaging requires a method for registering the spatial location of multiple two-dimensional images and combination in three dimensions
with electrocardiography and respiratory gating
during the different cardiac cycles8）, so clinicalapplication is greatly hampered by the complicatedprocedure, slow frame rate and artifacts due to respiratory and cardiac motion.
The matrix array transducer used in real-time
J Cardiol 2004 Jan; 43（1）: 23 – 29

three-dimensional imaging was proposed by Duke
University in the United States and made available
commercially by Philips Corporation recently9）.
The transducer crystal is equally divided intominute square elements by many longitudinal andhorizontal lines. The elements are arrayed in amatrix of 60 × 60 ＝ 3,600 elements（or 80 × 80 ＝
6,400）. By delaying the firing time to each element
controlled by computer, the ultrasonic wave beams
have different phases, and the direction of the wave

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Zhuang, Wang, Xie et alfront can be altered. The Philips Sonos 7500 ultrasonographic machine uses a dedicated three-dimensional echo probe for rapid beam forming to scan apyramidal volume. Real-time volumetric scanningrequires no off-line reconstruction techniques,enabling live three-dimensional visualization andquantification of the heart. RT3D echocardiography
was used to measure phantoms of different size andshape, and demonstrated the validation of this system to assess left ventricular volume 10）. RT3D
echocardiography and 2D echocardiography ofseven aneurysmal balloons, 15 sheep（5 with chronic left ventricular aneurysms and 10 without left
ventricular aneurysms）during 60 different hemodynamic conditions and 29 patients（13 with chronic
left ventricular aneurysms and 16 with normal left
ventricle）and demonstrated that RT3D echocardiography could accurately quantify left ventricularvolumes for geometrically asymmetric left ventricles associated with ventricular aneurysms11）.
One of the main limitations of RT3D echocardiography is the diminished image resolution compared to state-of-the-art 2D echocardiography. On
the other hand, irregular respiration rate, arrhythmia and body motion may affect the combinationof full volume imaging during several cardiac
cycles leading to artifacts. Contrast agents may
facilitate endocardial definition and increase theaccuracy of volume measurements obtained fromthree-dimensional images12）.
Recently, numerous advances have been made incontrast echocardiography, particularly in the formulation of agents that cross the pulmonary circulation to opacify the left cardiac chambers andmyocardium. The new type of ultrasound contrast
agent is usually composed of albumin or phospholipid shell containing perfluorocarbon or hexafluoride gas, with an average bubble diameter of 4 to
6μm13）. These microbubbles behave as intravenous
tracers with rheological properties similar to red
blood cells and can resonate in an ultrasound fieldand give particularly bright signals at frequencies at

or above their natural frequencies 14）. Contrast
enhancement in a group of patients with poor
images resulted in a significantly higher segmental
salvage rate compared to harmonic image alone15）.
Measurement of left ventricular volumes and ejection fraction before and after intravenous contrast
echocardiography and comparison of these measurements obtained by magnetic resonance imaging
showed the use of contrast improved the accuracyof echocardiographic assessment of left ventricularvolumes and ejection fraction16）. Evidence suggeststhat harmonic contrast echocardiography gives thebest signal to noise ratio for left ventricular opacification.
The present study found that left ventricularmass derived from RT3D echocardiography correlated well with the anatomic measurements andimproved after injection of intravenous contrast
agent. Contrast enhanced RT3D echocardiography
also seemed to have a higher index of intraobserver
and interobserver agreement. RT3D echocardiography with perfluorocarbon microbubbles contrast
medium provides better visualization of the endocardial border and a more accurate and repeatablemeasurement of total left ventricular mass, especially if the technician has less experience of imaging analysis or in patients with poor acoustic windows.
CONCLUSIONSThis study describes a novel methodology for thequantitative assessment of left ventricular massusing RT3D echocardiographic images. Left ventricular mass measurement derived from RT3D
echocardiography with and without contrast medium injection were correlated well with anatomic
measurements in an animal study, but contrast
enhanced RT3D echocardiography permitted
immediate excellent visualization of endocardial
border in any plane, which would provide a moreaccurate and reliable means of determining left
ventricular mass.

J Cardiol 2004 Jan; 43（1）: 23 – 29

Quantitative Assessment of Left Ventricular Mass

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