Comments for:

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Response to Ballo et al.

Submitter: Ehud Razin | ehudr@cc.huji.ac.il

isreal

Published December 19, 2006

In response to the critique raised in the letter from Ballo and colleagues, please find below our reply.
+ The correspondents state that "many of the observed between-groups differences in cardiac mass were not sustained by corresponding differences in LV end-diastolic diameter and/or wall thickness...” The cardiac mass was directly evaluated by weighing, and the heart weight to body weight ratio (HW/BW) was used. These measurements represent an evaluation the "whole heart", a three-dimensional assessment, whereas in comparison the echo measurements represent a two-dimensional evaluation. Thus the presence of differences in cardiac mass based on weight measurements with no significant differences in dimensions measured by echo does not exclude the presence of differences that would have been seen if it were possible to make a three-dimensional evaluation by echo.
+ Regarding the correspondents’ comment: “the use of unadjusted systolic indices calculated at the level of the endocardium such as shortening fraction ... overestimates the real performance of LV circumferential fibers …” We agree that the calculated shortening fraction might overestimate the real performance of circumferential fibers. But the repeated use in all mice of these two-dimensional measurements at a constant level in the LV increased the ability of this measurement to provide a good comparison between mice in each of the groups and changes over time and age was based on accepted criteria1.
+ We also agree that “the use of Doppler-derived cardiac output…” should be “also interpreted with caution”. Such caution is clearly required to be exercised when addressing the relative limitations of this method, due to its dependence on loading conditions. Nevertheless, the use of methods less dependent on these conditions is more limited in small animals, even though these have been performed recently2. Stress-adjusted midwall fractional shortening often used in research studies, as suggested by the correspondents, is clearly one of the options, but not the only one. The use of tissue-Doppler methods and myocardial strain imaging for this purpose in small animal research is of growing importance, but clearly not common practice. However, we are convinced, as suggested by the comments to our paper, that their role will be demonstrated and strengthened in future studies.
+ The correspondents also address the issue of the “importance of long-axis mitral annulus excursion, obtained in human …” in the assessment of LV pump function. We could not agree more. However, we elected not to use it in our present study. Its use in small animals will certainly be considered as an important tool in our future studies, as well as in studies by others.
1.Schiller, NB et al. 1989. Recommendation for quantification of the left ventricle by two-dimensional echocardiography. American Society of Echocardiography Committee on Standards, Subcommittee on quantification of Two-Dimensional Echocardiograms. J. Am. Soc. Echocardiography; 2: 358-367
2. Sebag IA, et al. 2005. Quantitative assessment of regional myocardial function in mice by tissue Doppler imaging: comparison with hemodynamics and sonomicrometry. Circulation May 24; 111(20): 2611-6.
Sincerely yours,
Ehud Razin, PhD
Sagi Tshori, MD
Dan Gilon, MD

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Cardiac hypertrophy and systolic function: what is the effective role of transcription factor MITF?

Submitter: Piercarlo Ballo | pcballo@tin.it

Cardiology Operative Unit, S. Andrea Hospital, La Spezia, Italy

Published December 11, 2006

To the Editor:
We have read with interest the recent article by Tshori et al. regarding the role of microphthalmia transcription factor (MITF) in the genesis of cardiac hypertrophy (1). The authors found that mouse strains with two different MITF mutations showed lower heart weight/body weight (HW/BW) ratio, reduced hypertrophic response to beta-adrenergic stimulation by isoproterenol, and impaired left ventricular (LV) systolic function in comparison with their normal littermates. These findings support the intriguing hypothesis that MITF may play a key role in modulating beta-adrenergic-induced cardiac hypertrophy and LV systolic performance.
The following aspects should be discussed in this study. It seems somewhat surprising that many of the observed between-group differences in cardiac mass were not sustained by corresponding differences in LV end- diastolic diameter and/or wall thicknesses. For instance, sp/sp normal littermates showed a 19% increase in the HW/BW ratio following isoproterenol administration for 7 days, but also showed no differences with sp/sp mice treated with saline as regards end-diastolic LV diameter (3.07 ± 0.17 vs 3.05 ± 0.26 mm), septal thickness (0.51 ± 0.05 vs 0.52 ± 0.03 mm), and posterior wall thickness (0.56 ± 0.02 vs 0.58 ± 0.05 mm). Similarly, a 7% increase in the HW/BW ratio was observed in ce/ce MITF- mutated mice treated with isoproterenol, but there were no differences in either end-diastolic LV diameter or end-diastolic LV wall thicknesses with ce/ce MITF-mutated mice treated with saline. In addition, 5-week-old ce/ce and tg/tg MITF-mutated mice had lower heart weight and HW/BW ratio, but also similar end-diastolic LV diameter and wall thicknesses in comparison with their normal littermates. The presence of between-group differences in cardiac mass with no significant differences in any LV dimension is difficult to explain. By definition, any increase in LV mass is characterized by enlargement of diastolic LV cavity size and/or diastolic LV wall thickening, based on the presence of an eccentric or concentric pattern of LV geometrical remodeling (2,3). Also, echocardiographic measurements of LV dimensions and mass in mice have been shown to correlate well with autopsy findings (4,5). The possibility that the mismatch between cardiac mass and LV dimensions in this study could depend on differences in atrial and/or right ventricular mass is reasonably unlikely. Moreover, a specific reduction in both LV and right ventricular mass after normalization to body weight was found in 15-month-old ce/ce MITF-mutated mice in comparison with their normal littermates, despite no differences in end-diastolic LV diameter and wall thicknesses.
Other issues derive from the methods used for the evaluation of cardiac function. The use of unadjusted systolic indices calculated at the level of endocardium – such as the shortening fraction used by the authors – overestimates the real performance of LV circumferential fibers, that are predominantly distributed within the midwall layers (6). The calculation of fractional shortening at the level of midwall represents a more accurate method to assess effective LV systolic function, and its use in mice has been previously validated (7,8). The use of Doppler-derived cardiac output – the other systolic index measured in this study – should also be interpreted with caution, due to its strong dependence on loading conditions (9). The negative effect of LV afterload on systolic shortening and ejection – particularly important when pharmacological manipulation of haemodynamic status is performed, e.g. by cathecolamine infusion (10) – was not taken into account by the authors. Adjustment to end-systolic LV wall stress is strictly required to provide a load-independent index of LV systolic performance (11). To date, stress-adjusted midwall fractional shortening is commonly used in research studies as a reliable measure of effective circumferential LV myocardial contractility (12). Furthermore, it should be pointed out that simple determination of circumferential systolic indices does not provide a full quantification of complex LV systolic dynamics. The assessment of longitudinal LV function by echocardiographic tissue Doppler and/or myocardial strain imaging has recently gained ground as a key analysis for the quantification of effective LV systolic performance (13), and its feasibility and accuracy in mice have been demonstrated (14). Notably, a depression in longitudinal LV function is frequently observed in the presence of normal circumferential function (15), and has been shown to detect early contractile impairment and to predict mortality in mice exposed to pharmacological cardiac injury better than conventional circumferential indices (16). Furthermore, data obtained in humans suggest that long-axis mitral annulus excursion is the principal contributor to LV pump function, accounting for approximately 60% of LV stroke volume (17). Further studies are needed to investigate the effective role of MITF in affecting both cardiac mass and LV systolic function.
REFERENCES
1. Tshori S, Gilon D, Beeri R, Nechushtan H, Kaluzhny D, Pikarsky E, Razin E. Transcription factor MITF regulates cardiac growth and hypertrophy. J Clin Invest 2006;116:2673-81.
2. Ballo P, Mondillo S, Guerrini F, Barbati R, Picchi A, Focardi M. Midwall mechanics in physiologic and hypertensive concentric hypertrophy. J Am Soc Echocardiogr 2004;17:418-27.
3. Takimoto E, Champion HC, Li M, Ren S, Rodriguez ER, Tavazzi B, Lazzarino G, Paolocci N, Gabrielson KL, Wang Y, Kass DA. Oxidant stress from nitric oxide synthase-3 uncoupling stimulates cardiac pathologic remodeling from chronic pressure load. J Clin Invest 2005;115:1221-31.
4. Manning WJ, Wei JY, Katz SE, Litwin SE, Douglas PS. In vivo assessment of LV mass in mice using high-frequency cardiac ultrasound: necropsy validation. Am J Physiol 1994;266:H1672-5.
5. Gardin JM, Siri FM, Kitsis RN, Edwards JG, Leinwand LA. Echocardiographic assessment of left ventricular mass and systolic function in mice. Circ Res 1995;76:907-14.
6. Ballo P, Mondillo S, Galderisi M. Determinants of discrepancy between left ventricular chamber systolic performance and effective myocardial contractility in subjects with hypertension. J Hum Hypertens 2006;20:701-3.
7. Ding B, Price RL, Goldsmith EC, Borg TK, Yan X, Douglas PS, Weinberg EO, Bartunek J, Thielen T, Didenko VV, Lorell BH. Left ventricular hypertrophy in ascending aortic stenosis mice: anoikis and the progression to early failure. Circulation 2000;101:2854-62.
8. Mori T, Chen YF, Feng JA, Hayashi T, Oparil S, Perry GJ. Volume overload results in exaggerated cardiac hypertrophy in the atrial natriuretic peptide knockout mouse. Cardiovasc Res 2004;61:771-79.
9. Kulandavelu S, Qu D, Adamson SL. Cardiovascular function in mice during normal pregnancy and in the absence of endothelial NO synthase. Hypertension 2006;47:1175-82.
10. Hoit BD, Khan ZU, Pawloski-Dahm CM, Walsh RA. In vivo determination of left ventricular wall stress-shortening relationship in normal mice. Am J Physiol 1997;272:H1047-52.
11. Williams RV, Lorenz JN, Witt SA, Hellard DT, Khoury PR, Kimball TR. End-systolic stress-velocity and pressure-dimension relationships by transthoracic echocardiography in mice. Am J Physiol 1998;274:H1828-35.
12. Ballo P, Bocelli A, de Martino M, Mondillo S. Is left ventricular performance effectively preserved in children with growth hormone deficiency? J Clin Endocrinol Metab 2006 Feb 17; [Epub ahead of print].
13. Mondillo S, Galderisi M, Ballo P, Marino PN; Study Group of Echocardiography of the Italian Society of Cardiology. Left ventricular systolic longitudinal function: comparison among simple M-mode, pulsed, and M-mode color tissue Doppler of mitral annulus in healthy individuals. J Am Soc Echocardiogr 2006;19:1085-91.
14. Sebag IA, Handschumacher MD, Ichinose F, Morgan JG, Hataishi R, Rodrigues AC, Guerrero JL, Steudel W, Raher MJ, Halpern EF, Derumeaux G, Bloch KD, Picard MH, Scherrer-Crosbie M. Quantitative assessment of regional myocardial function in mice by tissue Doppler imaging: comparison with hemodynamics and sonomicrometry. Circulation 2005;111:2611-6.
15. Ballo P, Quatrini I, Giacomin E, Motto A, Mondillo S. Circumferential versus longitudinal function in patients with hypertension: a nonlinear relation. J Am Soc Echocard 2007; in press.
16. Neilan TG, Jassal DS, Perez-Sanz TM, Raher MJ, Pradhan AD, Buys ES, Ichinose F, Bayne DB, Halpern EF, Weyman AE, Derumeaux G, Bloch KD, Picard MH, Scherrer-Crosbie M. Tissue Doppler imaging predicts left ventricular dysfunction and mortality in a murine model of cardiac injury. Eur Heart J 2006;27:1868-75.
17. Carlsson M, Ugander M, Mosen H, Buhre T, Arheden H. Atrioventricular plane displacement is the major contributor to left ventricular pumping in healthy adults, athletes and patients with dilated cardiomyopathy. Am J Physiol Heart Circ Physiol 2006 Nov 10; [Epub ahead of print].