Chinese Medical Journal 2005;118(14):1175-1181
Assessment of left ventricular segmental function after autologous bone marrow stem cells transplantation in patients with acute myocardial infarction by tissue tracking and strain imaging

RUAN Wen,  PAN Cui-zhen,  HUANG Guo-qian,  LI Yan-lin,  GE Jun-bo,  SHU Xian-hong

RUAN Wen (Department of Cardiology, Zhongshan Hospital of Fudan University, Shanghai 200032, China)

PAN Cui-zhen (Department of Cardiology, Zhongshan Hospital of Fudan University, Shanghai 200032, China)

HUANG Guo-qian (Department of Cardiology, Zhongshan Hospital of Fudan University, Shanghai 200032, China)

LI Yan-lin (Department of Cardiology, Zhongshan Hospital of Fudan University, Shanghai 200032, China)

GE Jun-bo (Department of Cardiology, Zhongshan Hospital of Fudan University, Shanghai 200032, China)

SHU Xian-hong (Department of Cardiology, Zhongshan Hospital of Fudan University, Shanghai 200032, China)

Correspondence to:SHU Xian-hong,Department of Cardiology, Zhongshan Hospital, Fudan University, Shanghai 200032, China (Tel: 86-21-64041990-2530. Fax:. E-mail:sxh@zshospital.com)
Keywords
echocardiography;tissue Doppler imaging;acute myocardial infarction;bone marrow stem cells;cell transplantation
Abstract
Background Emerging evidence suggests that stem cells can be used to improve cardiac function in patients after acute myocardial infarction. In this randomized trial, we aimed to use Doppler tissue tracking and strain imaging to assess left ventricular segmental function after intracoronary transfer of autologous bone-marrow stem cells (BMCs) for 6 months’ follow up.
Methods Twenty patients with acute myocardial infarction and anterior descending coronary artery occlusion proven by angiography were double-blindedly randomized into intracoronary injection of bone-marrow cell (treated, n=9) or diluted serum (control, n=11) groups. GE vivid 7 and Q-analyze software were used to perform echocardiogram in both groups 1 week, 3 months and 6 months after treatment. Three apical views of tissue Doppler imaging were acquired to measure peak systolic displacement (Ds) and peak systolic strain (εpeak) from 12 segments of LV walls. Left ventricular ejection fraction (LVEF), end-diastolic volume (EDV) and end-systolic volume (ESV) were obtained by Simposon’s biplane method.
Results (1) 3 months later, Ds and εpeak over the infract-related region clearly increased in the BMCs group [Ds: (4.49±2.71) mm vs (7.56±2.95) mm, P<0.01; εpeak: (-13.40±6.00)% vs (-17.06±6.05)%, P<0.01], but not in the control group [Ds: (4.74±2.67) mm vs (5.01±3.23) mm, P>0.05; εpeak : (-13.84±6.05)% vs (-15.04±6.75)%, P>0.05]. At the same time, Ds over the normal region also increased, but the Ds enhancement was markedly higher in the BMCs group than that in the control group [(3.21±3.17) mm vs (0.76±1.94) mm, P<0.01]. Parameters remained steady from the 3rd to 6th month in either group (P>0.05). (2) LVEF in treated and control groups were almost the same at baseline (1st week after PCI) [(53.37±8.92)% vs (53.51±5.84)%, P>0.05]. But 6 months later, LVEF in the BMCs group were clearly higher than that in the control group [(59.33±12.91)% vs (50.30±8.30)%, P<0.05]. (3)There were no evident difference in EDV or ESV between two groups at baseline [EDV:(113.74±23.24) ml vs (129.94±32.72) ml , P>0.05; ESV: (57.12±18.66) ml vs (62.09±17.68) ml, P>0.05]. Three months later, EDV and ESV in the control group were markedly greater than those in the BMCs group [EDV: (154.89±46.34) ml vs (104.85±33.21) ml, P<0.05; ESV :(82.91±35.79) ml vs (49.54±23.32) ml, P<0.05]. But EDV and ESV did not change much from 3rd to 6th month in either group (P>0.05).
Conclusions Emergency transplantation of autologous BMCs in patients with acute myocardial infarction helps to improve global and regional contractility and attenuate post-infarction left ventricular remodeling. Tissue tracking and strain imaging provide quick, simple and noninvasive methods for quantifying left ventricular segmental function in humans.
Recent experimental and clinical studies suggest that stem cell transplantation into damaged myocardium may have the potential to restore myocardial viability and improve cardiac function.[1-4] Therefore, it has become a new focus of interest in treating congestive heart failure and death in patients after myocardial infarction. Currently, however, the clinical application of stem cell therapy is still in its preliminary stage.[2-4] In order to screen appropriate candidates for the therapy, more detailed information concerning the position and time of early myocardial functional recovery is needed. Echocardiography, the most commonly used noninvasive assessment, plays an important role in achieving this goal. Nevertheless, the traditional echocardialgraphic methods to evaluate segmental myocardial function were limited by their subjectiveness and semi-quantitativeness, which makes it necessary to find a more objective and excise measurement.

Doppler tissue imaging (DTI) is a newly emerged technique of echocardiograph for quantitative assessment of segmental cardiac function. Recently, many new refinements of DTI have been developed, among which tissue tracking (TT) and strain (ε) imaging are two promising ones. Tissue tracking measures the longitudinal myocardial motion amplitude in each region during systole, visualized by a graded display of 7 color bands indicating different distances of systolic motion. Strain reflects the extent of myocardial fiber shortening, expressed as the percent change from the original dimension. To this day, the normal value of TT and ε have been determined,[5-7] and their ability to detect abnormal regional function after myocardial ischemia has been confirmed.[8-11] It has also been validated that in normal myocardium, changes in ε is closely linked with changes in contractility,[12] and systolic displacement of the atrioventricular plane toward the apex can provide important information about LV systolic function.[13,14] But up to now, there is little study especially designed to evaluate the usefulness of TT and ε in detecting early changes of damaged myocardial function after stem cell therapy.

Thus, in this work we sought to apply TT and ε imaging in the following up of AMI patients after BMCs or pure percutaneous transluminal coronary angioplasty (PTCA) therapy in order to evaluate their abilities to detect early changes in myocardial segmental contractility and, secondly, to assess the effectiveness of BMCs therapy in improving regional myocardial function.

METHODS

Patients
A total of 20 patients with acute myocardial infarction [mean age (58±7) years; 19 men and 1 woman] were included in this randomized, double-blinded study. Patients were selected prospectively and consecutively from those who were admitted to emergency department of Zhongshan Hospital from July 2003 to Aug 2004 after a chest pain of (12.1±12.6) hours prior. Infarction was diagnosed by medical history, physical examination, electrocardiaogram (ECG), positive cardiac enzyme, and coronary angiography. In all, angiography showed a >90% coronary stenosis or occlusion in left anterior descending artery, among them 9 had more than one criminal coronary arteries (5 from treated group,4 from control group). Within 2 hours after successful PTCA, all patients were randomized to be administered with intracoronary injection of BMCs (n=9), or diluted serum (n=11). Except for the grouping factor, there was no statistical difference between the two groups in age, gender, risk factors (hypertension, diabetes mellitus and hyperlipidemia), hours of chest pain before PTCA, position and degree of coronary occlusion. Besides, after treatment, patients in both groups showed a TIMI Ⅲ coronary artery flow, and all received aspirin, nitrate, angiotensin-converting enzyme (ACE) inhibitor, β-blocker, statin and clopidogrel for routine treatment. Both groups underwent 1-week, 3-month, and 6-month follow-up by 2D and tissue Doppler echocardiogram. The ethics review board of the Zhongshan Hospital of the Fudan University of Shanghai, China, approved the protocol, and all procedures conformed to institutional guidelines.

Echocardiography
All echocardiography examinations were performed on an ultrasound machine (GE Vivid Seven, Horten, Norway) with a 1.5-4.3 MHz transducer. Echocardiograms were stored digitally and analysed offline.

The end-diastolic and end-systolic volume (EDV, ESV) ejection fractions (LVEF) were estimated using Simposon’s modified biplane method on the basis of 3 measurements.

Doppler tissue imaging
DTI was obtained from the apical 4- and 2-chamber and apical long-axis views by 2 investigators blind to
clinical and angiographic information. The sample region was 5×5 pixels and Doppler scanning frame rates were kept between 104-180 frames/s. A simultaneous electrocardiogram registration was required to define the beginning and end of the systole (The machine automatically sets the systole from the peak of the R wave to the end of the T wave). TT and ε analysis were performed offline as special modalities of DTI by Q-analysis software (GE).

For quantitative analysis of the wall motion, peak systolic displacement (Ds) and peak systolic strain (εpeak) were measured on 12 segments of the LV wall. Normal, hyperkinetic or hypokinetic motion was expressed by a positive Ds or negative εpeak value, while akinetic or dyskinetic motion was demonstrated by zero or reverse Ds or εpeak value. The mean Ds and εpeak were calculated from 3 consecutive heart beats. In this study, each patient had a total or subtotal occlusion in left anterior descending coronary artery, thus we define the lateral and anterior wall, anterior and posterior interventricular septum as the infract-related region, the post and inferior wall as the normal region, Ds and εpeak results from the infarcted or normal region were obtained by averaging data from the related LV walls.

Statistics
All data are presented as mean±standard deviation (SD). Student’s t test was used for the inter-group comparison of LVEF, EDV and ESV. However, because of the abnormal distribution in some of the DTI data, nonparametric Mann-Whitney test was used for the inter-group and intra-group comparisons of Ds and εpeak . Statistical analysis was performed with SPSS for Windows (Version 10.0). Statistical significance was accepted when P<0.05.

RESULTS

Clinical data between the two groups did not differ significantly ( Table 1 ). Conventional 2D assessment of EDV, ESV and LVEF was successful in all the patients. Quantitative analysis was possible in 100% of the segments by tissue tracking, but only 90% of the segments by strain imaging because of noise and artifacts.

Analysis of LV segmental function
The results of peak systolic displacement (Ds) and strain (εpeak) at 1-week, 3-month, and 6-month follow-up are exemplified in Figs. 1 , 2, 3 and 4 , and summarized in Table 2.

Baseline (1 week after PCI) parameters in both groups did not differ significantly over any region (P>0.05). Three month later, Ds and εpeak at the infract-related region clearly increased in the BMCs group [Ds: (4.49±2.71) mm vs (7.56±2.95) mm, P<0.01; εpeak: (-13.40±6.00)% vs (-17.06±6.05)%, P<0.01], but not in the control group [Ds: (4.74±2.67) mm vs (5.01±3.23) mm, P>0.05; εpeak : (-13.84±6.05)% vs (-15.04±6.75)%, P>0.05], indicating significant improvement of damaged myocardial function after stem cell implantation.

Moreover, Ds (but not εpeak) over the normal region also increased owing to the passive tethering from the recovered myocardial contraction in the neighbouring segments. But the Ds enhancement was significantly higher in the BMCs group than that in the control group [(3.21±3.17) mm vs (0.76±1.94) mm, P<0.01], still reflecting better recovery of myocardial contractility after BMCs therapy than after the routine therapy. There was no evident difference in either Ds or εpeak between 3-month and 6-month follow-up in each group (P>0.05).

Analysis of LV global function and volume
Comparisons of LVEF, EDV and ESV in the two groups are presented in Table 3.

LVEF in the treated and control groups were almost the same at baseline ( 1-week after PCI) [(53.37±8.92)% vs (53.51±5.84)%, P>0.05]. After 6 months, LVEF in the BMCs group was significantly higher than that in the control group [(59.33±12.91)% vs (50.30±8.30)%, P<0.05], demonstrating better retoration of global function after early enhancement of regional function after cell therapy.There was no evident difference in EDV and ESV between two groups at baseline [EDV: (113.74±23.24) ml vs (129.94±32.72) ml, P>0.05, ESV: (57.12±18.66) ml vs (62.09±17.68) ml, P>0.05]. Three months later, EDV and ESV in the control group were markedly greater than those in the BMCs group [EDV: (154.89±46.34) ml vs (104.85±33.21) ml, P<0.05; ESV: (82.91±35.79) ml vs (49.54±23.32) ml, P<0.05], showing the effect of BMCs transplantation in attenuating left ventricular remodeling process. But EDV and ESV did not change much from the 3rd to 6th month in either group (P>0.05).

DISCUSSION

In this work, we proved that intracoronary transplantation of BMCs was effective in improving the segmental and global cardiac function, as well as alleviating LV remodeling in patients after AMI. Besides, we demonstrated the ability of tissue tracking and strain Doppler echocardiography to quantify regional myocardial contractility in patients with different degrees of myocardial functional recovery. We also found that TT was more repeatable and less noise influenced, while strain was independent of passive tethering effect. Thus, the conjunction of the two modalities would be promising in future clinical application.

TT/ε and segmental LV function
As is known to all, regional cardiac function changes earlier than the global function, thus quick and accurate assessment of the LV segmental motion is extremely important in judging weather the ischemic myocardium revitalized after BMCs therapy. Tissue tracking reflects the myocardial contractility by tracing the largest segmental displacement toward the apex at systole, in normal people the greatest and smallest displacement locates in LV basal and apical segment respectively.[7] As for systolic strain, it represents the contractile function by calculating the percentage of myocardial deformation, and studies have shown that strains in the normal human heart were homogeneously distributed from base to apex of LV, which facilitated the differentiation of normal from abnormal contractility in different regions of the heart.[5-7]

During myocardial infarction, both the scale and velocity of cardiac contraction decreases which is interpreted as the decrease, diminishing, or even reverse of peak systolic displacementing (Ds) and strain (εpeak) on TT and ε imaging.[8-11] As a result, increase in the TT or ε value suggests the recovery of contractility in this area. In this study, we found that segmental displacement and deformation in the infracted regions enhanced significantly at 3-month follow-up in BMCs group,but did not in the control group, indicating much earlier rehabilitation of segmental contractility after transplantation of BMCs. Moreover, Ds (not εpeak) in the normal region also improved after 3 months because of the passive tethering from neighbouring myocardium whose contractile function was enhanced, but the amount of enhancement was much greater in the treated group than in the control group, which also suggests greater degree of recovery after BMCs implantation. However, the parameters in both groups did not present statistically evident difference between the 3rd and 6th month follow-up, demonstrating that the improvement entered into a plateau after 3 months.

Global LV function
As an organic system, all parts of the heart work harmoniously to produce an effective cardiac output. Under the condition of myocardial infarction, the damaged myocardium lost their activation and contraction, leading to the deterioration of LV global function. But if saved in time, the ischemic myocardium would regain their vitality, making the whole heart contraction more powerfully and synchronously, which benefits the ejection fraction in the end. In this study, at the 3rd month, segmental function of the infarcted region improved evidently in the BMCs group (P<0.05), but did not show much enhancement in the control group (P>0.05). Correspondently, LVEF increased in the BMCs group (P>0.05), but slipped in the control group (P>0.05), however, there was no significant difference between the two groups in LVEF at 3-month. At the 6th months, LVEF of the transplantation group became markedly higher than that of the control group (P<0.05). The results suggested that it took some time for the left ventricular function to recover after infarction, and it was based on an in-time rescue of the dying cardiocmyocytes, and restoration of segmental contractility. BMCs therapy provided a good way to achieve this goal. Besides, the study also reminded us that TDI was not only more sensitive than the traditional echocardiography in assessing LV regional function, it could also serve as a predictor of the LVEF and patient’s prognosis.

LV remodeling
After myocardial infarction, the surviving cardiomyocytes bordering the infarct zone become hypertrophied as part of an adaptive mechanism to compensate for the loss of myocardium.[2] However, the self-renewal mechanism is insufficient to prevent the apoptosis of these hypertrophied cardiomyocytes, which leads to left ventricular remodeling in the end. It is believed that the heart muscle capillary bed is best penetrated at the accurate stage of myocardial infarction, during which different inflammatory factors were released owing to the injury, making this time the best chance for BMCs transplantation.[3,15]

In this study of early BMCs transplantation within 2 hours after PCI, we observed that the LV end-systolic and end-diastolic volume remained steady in BMCs group, but gradually increased in the control group, which made us believe that the repopulation of viable cardiomyocytes and neovascularization at an early stage accelerates the healing of injured myocardium and attenuates the left ventricular remodeling.

Limitations
The study was limited by the small population of patients, relatively short time of total follow-up, and lacking of comparison with the data before or within 3 month after the treatment, besides, the multiple transplantation techniques of the BMCs over the world also made it difficult to judge the earliest time of cardiac recovery post-transplantation. Therefore, further study from large, randomized and controlled trials is needed to clarify the short- and long-term effects of this therapy.

As for the DTI modalities, both tissue tracking and strain measurements are dependent on the clear 2D imaging and direction of the Doppler angle in relation to myocardial motion.[16] To overcome this, care was taken to keep the angle between the direction of the Doppler beam and that of tissue deformation as small as possible.

Conclusions
In the present study we proved that balloon catheter delivery of autolougous bone marrow cells to the infarcted region immediately after PCI can significantly improve segmental and global LV function, as well as inhibit left ventricular remodeling. We also demonstrated the ability of tissue tracking and strain imaging in detecting different degrees of segmental myocardial function in a clinical setting. The novel techniques may help to improve noninvasive quantification of regional myocardial function in humans.

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