Biophysical Journal Volume 106 May 2014 2267–2274
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New Insights on Human Skeletal Muscle Tissue Compartments Revealed by In Vivo T2 NMR Relaxometry Ericky C. A. Araujo,†‡* Yves Fromes,§ and Pierre G. Carlier†§{* † Institute of Myology, Paris, France; ‡University Paris-Sud (Paris 11), Orsay, France; §University Pierre and Marie Curie (Paris 06), Paris, ´ nergie Atomique et aux E ´ nergies Alternatives, Institut d’Imagerie Biome´dicale, Molecular Imaging Research France; and {Commissariat a` l’E Center, IdM Nuclear Magnetic Resonance Laboratory, Paris, France
ABSTRACT The spin-spin (T2) relaxation of 1H-NMR signals in human skeletal muscle has been previously hypothesized to reveal information about myowater compartmentation. Although experimental support has been provided, no consensus has yet emerged concerning the attribution of specific anatomical compartments to the observed T2 components. Potential application of a noninvasive tool that might offer such information urges the quest for a definitive answer to this question. The purpose of this work was to obtain new information that might help elucidate the mechanism of T2 distribution in muscle. To do so, in vivo T2 relaxation data was acquired from the soleus of eight healthy volunteers using a localized Carr-Purcell-Meiboom-Gill technique. Each acquisition contained 1000 echoes with an interecho spacing of 1 ms. Data were acquired from each subject under different vascular filling preparations expected to change exclusively the extracellular water fraction. Two exponential components were systematically observed: an intermediate component (T2 ~ 32 ms) and a long component (100 < T2 < 210 ms). The relative fraction and T2 value characterizing the long component systematically increased after progressive augmentation of extracellular water volume. Characteristic relaxation behavior for each vascular filling condition was analyzed with a two-site exchange model and a three-site two-exchange model. We show that a two-site exchange model can only predict the observations for small exchange rates, much more representative of transendothelial than transcytolemmal exchange regimes. The three-site two-exchange model representing the intracellular, interstitial, and vascular spaces was capable of precisely predicting the observations for realistic transcytolemmal and transendothelial exchange rates. The estimated intrinsic relative fractions of each of these compartments corroborate with estimations from previous works and strongly suggest that the T2 relaxation from water within the intracellular and interstitial spaces is described by the intermediate component, whereas the long component represents water within the vascular space.
INTRODUCTION Alterations of intra- and extracellular myowater content and exchange kinetics are known to reflect exercise- and pathology-induced changes in tissue (1,2). A noninvasive technique giving access to such information in vivo would have important applications in the fields of exercise physiology and neuromuscular disorders. Nuclear magnetic resonance (NMR) is sensitive to these changes (3,4) and is a valuable tool for addressing these questions. Proton (1H) spin-spin (T2) relaxation has been shown to be multiexponential in many biological tissues. In skeletal muscle tissue, at least three exponential components were systematically observed in ex vivo and in vivo studies (5–10): a short one (T2 < 10 ms) accounting for 100 ms) accounting for components characterized by T2 values and relative fractions that correspond to the intermediate and long components also observed in previous studies (Fig. 3). This demonstrated the reproducibility of the methodology and supported the approach of a biexponential fit to analyze our data. Additionally, the results obtained from both methods were quite similar (Table 1). The vascular filling preparations exclusively influenced the long T2 component, whose relative fraction and value systematically increased after progressive augmentation of the extracellular space. These were strong evidences in favor of the anatomical compartmentation hypotheses. Deeper analysis of the compartmental exchanges is needed to elucidate the histological interpretation of this compartmentation. Our analyses showed that a 2SX model is only capable of explaining the observed relaxation data for slow exchange regimes (see Fig. 4). In previous studies investigating transcytolemmal water exchange, the intracellular residence time in muscle has been estimated between 550 and 1100 ms (24,25). In our analysis, when setting the intracellular residence time to 1000 ms, the results show that the observed T2-relaxation data cannot be explained by a 2SX model representing an intra-/extracellular compartmentation for realistic extracellular T2 values (T2b < 500 ms) (see Table 2). However, setting the
TABLE 2 Extracted intrinsic parameters of the 2SX model for the three vascular filling conditions at intermediate and slow exchange regimes; SE is presented within parentheses ti [s] Vascular condition 1
10
Vascular draining Free perfusion Vascular filling Vascular draining Free perfusion Vascular filling
Pa [%]
T2a [ms]
T2b [ms]
92.7(2.5) 30.0(0.5) 500(1216) 90.2(1.0) 30.1(0.3) 500(312) 85.2(0.5) 30.5(0.2) 500(81) 95.4(0) 31.8(0) 195.2(0) 92.8(0) 32.2(0) 199.5(0) 86.5(0) 32.7(0) 204.6(0)
R2 ( 103) 999.8 999.6 999.4 1000 1000 1000
R2 is the square of the correlation coefficient between predicted and experimental data.
FIGURE 4 Characteristic relaxation curves of the vascular draining (triangles), normal (squares), and vascular filling (circles) conditions, and the corresponding fitted curves characterized by the extracted intrinsic parameters of the 2SX model for both intermediate (crosses) and slow (lines) exchange regimes. Biophysical Journal 106(10) 2267–2274
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Araujo et al.
TABLE 3 Extracted intrinsic parameters of the 3S2X system characterizing the intracellular, interstitial, and vascular spaces for the three vascular filling conditions; standard error is presented within parentheses Vascular condition Pc [%]
Pi/Pe
T2i [ms] T2e [ms]
tc [ms]
R2
have studied a system composed by the intracellular and interstitial spaces according to a 2SX model. The analytical b 2b , b 2a and T solution of Eq. 5, for the apparent T2 values, T in the 2SX model has been previously presented by Hazlewood et al. (6), and is given by
Vascular draining 4.6(0) 12.7(0.3) 31.5(0) 35.8(1.0) 535(5) 1 Free perfusion 7.3(0) 6.73(0.04) 31.5 37.4(0.3) 1122(3) 1 Vascular filling 13.8(0) 6.18(0.04) 31.5 41.1(0.5) 9813(333) 1 R2 is the square of the correlation coefficient between predicted and experimental data.
intracellular residence time to 10 s avoids the need for abnormally high extracellular T2 values to explain the data (see Table 2). Intravascular residence times have been estimated between 0.3 and 3 s (24,26,27), and vascular water fraction shall be ~4% (28). For these values, the extravascular residence time at homeostasis must be between 7 and 72 s (Eq. 6), which characterizes a slow exchange regime. Theses evidences supported preferably an extra-/ intravascular compartmental interpretation, suggesting that relaxation of transverse magnetization from both intracellular and interstitial spaces are described by the intermediate component. This implies that the apparent intracellular and interstitial T2 values, Tic and Tint, shall be close enough to the observed intermediate T2. To investigate the intrinsic system parameters that might satisfy this condition, we
FIGURE 5 (a) Characteristic relaxation curves of the vascular draining (triangles), normal (squares), and vascular filling (circles) conditions, and the corresponding fitted curves (lines) characterized by the extracted intrinsic parameters of the 3S2X model. (b) Obtained T2 spectra from regularized inversion of the fitted curves resulting from the 3S2X model. Biophysical Journal 106(10) 2267–2274
with C1 ¼ 1 C2 ¼ 2
b 2a ¼ T
1 ; ðC1 þ C2 Þ
(9)
b 2b ¼ T
1 ; ðC1 C2 Þ
(10)
1 1 1 1 1 þ þ þ ; 2 T2a T2b t a t b
(11)
1 1 1 1 þ T2a t a T2b t b
2
4 þ ðt a t b Þ
!12 :
(12)
The average observed interstitial to intracellular water volume ratio is ~0.13 (28,29). Consequently, an intracellular residence time of 1 s would imply an interstitial residence time of 130 ms, at homeostasis. By applying these values into Eqs. 9–12, one may verify that for Tint > 80 ms, b 2a and T b 2b , would differ by the apparent T2 values, T >20 ms, which should be easily resolved in the obtained T2 spectra, considering the precision of our results for the observed intermediate T2 peaks (Fig. 3 b and Table 1). Usually, Tint is assumed to be considerably lengthened relative to the intracellular T2 due to the lower protein concentration in the interstitium. However, the macromolecules constituting the extracellular matrix have been shown to have nonnegligible relaxivities (30), and values as short as Tint ¼ 25 ms have been observed for synthetic interstitial models (9). This was further confirmed by the observed short T2 value in muscle sites with abnormally high endomysial accumulation such as fibrosis (31), and the long T2 values characteristic of interstitial edema (3). The 3S2X model has been previously shown to allow more accurate estimates of extracellular and vascular spaces and blood-to-tissue contrast agent transfer in pharmacokinetic studies (32). In this work, this model was shown to be capable of precisely predicting the characteristic relaxation curves for realistic physiological residence times (24– 27) (Fig. 5 and Table 3). The estimated interstitial T2 value was within the range, Tint < 80 ms, predicted by our 2SX analysis, and increased after progressive augmentation of its water content, as observed by the variations of the estimated intracellular to interstitial volume fraction ratio Pi/Pe (Table 3). The estimated compartmental relative fractions in normal conditions were in good agreement with results of previous studies (28,29). Estimated relative fraction of the vascular space increased after augmentation of extracellular volume. The
Interpretation of Muscle T2 Relaxation
augmentation of the intravascular residence, tc, must reflect attainment of quasi-normal equilibrium transendothelial exchange after fluid redistribution within vascular and interstitial spaces at each vascular condition. The observed nonlinear increase of tc with the vascular volume must reflect contributions of small veins and arteries within the VOI that could not be seen in the localization images, because they present a much higher compliance than capillaries and experience negligible exchange with the parenchyma. Vascular and interstitial water fractions varied between the imposed vascular filling conditions. While the vascular space varied progressively from one condition to another, the interstitial water fraction doubled when passing from the vascular draining to the vascular filling condition and presented a very small reduction (8%) when passing from the vascular filling to the normal condition. The rapid variation must be a result of the manual compression, which leads to interstitial space draining, whereas the slower variation reflects interstitial volume inertia as a consequence of the interstitial edema-preventing mechanisms (33). Furthermore, it has been previously shown that interstitial space volume varies at a much slower rate than the vascular space after imposed variations on vascular hydrostatic pressure (34). Finally, extracted T2 spectra from the fitted characteristic relaxation curves demonstrated that indeed only two T2 components may be obtained from the proposed 3S2X model characterized by the extracted intrinsic parameters (Fig. 5 b). In a previous study by Noseworthy et al. (12), the long T2 value resulting from biexponential fitting of relaxation data obtained within the soleus of healthy volunteers increased with hyperoxia exposure. This result was explained as reflecting the well-known blood-oxygenation-level-dependent effect (35) and the long T2 component was attributed to vascular water. In this work, the observed increase of the long T2 value after vascular filling was explained by an increase of the intravascular residence time. It is possible that blood O2 saturation was lower at vascular draining condition due to the imposed muscular ischemia of the procedure. However, the 3S2X model has too many parameters and vascular T2 had to be set constant to constrain the 3S2X model and obtain stable parameter estimations. Two-dimensional correlation NMR experiments have been previously proposed as a means for studying exchange dynamics (36); these methods allow simultaneous and independent estimation of compartment-intrinsic T2 value and exchange rates (13). The results from this work encourage the application of such methods for more precise parameter estimation of compartmental exchange models of myowater distribution. The interpretation of relaxometry studies relies on the accuracy and precision of the extracted parameters characterizing the multiexponential model that explains the observed relaxation. Such extraction constitutes an inverse
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Laplace transform of noisy data, to which there is no unique solution. Different methods for solving this problem without the need for imposing the number of T2 components characterizing the model exist (37–41) and are still the matter of research for the optimization of inversion methods. Our choice for a regularized inversion method was based on the fact that it yields nonnegative solutions and allows regularization, which has been shown by our results to ensure the reproducibility of the solutions. CONCLUSION 1. The biexponential T2 relaxation behavior of 1H-NMR signals observed in skeletal muscle reflects an anatomical compartmentation of myowater. 2. Water in the vascular space has nonnegligible contributions to observed muscle T2 values as obtained from monoexponential fitting of T2 relaxation data. 3. The biexponential T2 relaxation behavior observed in skeletal muscle cannot be explained with a 2SX model representing an intra-/extracellular anatomical compartmentation of myowater for realistic transcytolemmal exchange rates. However, for the slower transendothelial exchange rates, a 2SX model representing an extra-/ intravascular compartmentation allows precise prediction of the observations. 4. A 3S2X model representing water exchange dynamics among intracellular, interstitial, and vascular spaces is capable of explaining T2 relaxation behavior observed in skeletal muscle tissue for realistic transmembrane exchange rates and allowed estimating interstitial T2 value, which has been shown to be close to the intracellular T2 for healthy tissue in normal conditions. 5. For healthy skeletal muscle tissue in normal conditions, the observed intermediate T2 component reflects the relaxation behavior of water within the intracellular and interstitial spaces, and the long T2 component reflects the relaxation of water within the vascular space. The authors thank Dr. Alexandre Vignaud for his technical support during the development of the NMR sequences and the Drs. Paulo L. de Sousa, Noura Azzabou, Nicolas Decorte and Pierre-Yves Baudin for their critical reading of the manuscript.
REFERENCES 1. Louie, E. A., D. F. Gochberg, ., B. M. Damon. 2009. Transverse relaxation and magnetization transfer in skeletal muscle: effect of pH. Magn. Reson. Med. 61:560–569. 2. Arpan, I., S. C. Forbes, ., K. Vandenborne. 2013. T2 mapping provides multiple approaches for the characterization of muscle involvement in neuromuscular diseases: a cross-sectional study of lower leg muscles in 5–15-year-old boys with Duchenne muscular dystrophy. NMR Biomed. 26:320–328. 3. Ploutz-Snyder, L. L., S. Nyren, ., R. A. Meyer. 1997. Different effects of exercise and edema on T2 relaxation in skeletal muscle. Magn. Reson. Med. 37:676–682. Biophysical Journal 106(10) 2267–2274
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4. Saab, G., R. T. Thompson, and G. D. Marsh. 2000. Effects of exercise on muscle transverse relaxation determined by MR imaging and in vivo relaxometry. J. Appl. Physiol. 88:226–233.
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28. Robertson, J. D. 1965. Studies on the chemical composition of muscle tissue. III. The mantle muscle of cephalopod mollusks. J. Exp. Biol. 42:153–175. 29. Ling, G. N., and M. H. Kromash. 1967. The extracellular space of voluntary muscle tissues. J. Gen. Physiol. 50:677–694. 30. Laurens, E., E. Schneider, ., A. Calabro. 2012. A synthetic cartilage extracellular matrix model: hyaluronan and collagen hydrogel relaxivity, impact of macromolecular concentration on dGEMRIC. Skeletal Radiol. 41:209–217. 31. de Jong, S., J. J. Zwanenburg, ., P. R. Luijten. 2011. Direct detection of myocardial fibrosis by MRI. J. Mol. Cell. Cardiol. 51:974–979. 32. Paudyal, R., H. Poptani, ., J. D. Glickson. 2013. Impact of transvascular and cellular-interstitial water exchange on dynamic contrastenhanced magnetic resonance imaging estimates of blood to tissue transfer constant and blood plasma volume. J. Magn. Reson. Imaging. 37:435–444. 33. Aukland, K., and G. Nicolaysen. 1981. Interstitial fluid volume: local regulatory mechanisms. Physiol. Rev. 61:556–643. 34. Schu¨tze, H., W. Hildebrandt, and J. Stegemann. 1991. The interstitial fluid content in working muscle modifies the cardiovascular response to exercise. Eur. J. Appl. Physiol. Occup. Physiol. 62:332–336. 35. Wright, G. A., B. S. Hu, and A. Macovski. 1991. 1991 I. I. Rabi Award. Estimating oxygen saturation of blood in vivo with MR imaging at 1.5 T. J. Magn. Reson. Imaging. 1:275–283. 36. Lee, J., C. Labadie, ., G. S. Harbison. 1993. Two-dimensional inverse Laplace transform NMR: altered relaxation times allow detection of exchange correlation. J. Am. Chem. Soc. 115:7761–7764. 37. Lanczoz, C. 1957. Applied Analysis. Prentice Hall, Englewood Cliffs, NJ. 38. Yeramian, E., and P. Claverie. 1987. Analysis of multiexponential functions without a hypothesis as to the number of components. Nature. 326:169–174. 39. Jibia, A. U., and M.-J. E. Salami. 2012. An appraisal of Gardner transform-based methods of transient multiexponential signal analysis. Int. J. Comput. Theory Eng. 4:16–25.
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40. Clayden, N. 1992. Pade-Laplace analysis in the fitting of multi-exponential nuclear magnetic resonance relaxation decay curves. J. Chem. Soc., Faraday Trans. 88:2481–2486.
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Biophysical Journal 106(10) 2267–2274
2267
New Insights on Human Skeletal Muscle Tissue Compartments Revealed by In Vivo T2 NMR Relaxometry Ericky C. A. Araujo,†‡* Yves Fromes,§ and Pierre G. Carlier†§{* † Institute of Myology, Paris, France; ‡University Paris-Sud (Paris 11), Orsay, France; §University Pierre and Marie Curie (Paris 06), Paris, ´ nergie Atomique et aux E ´ nergies Alternatives, Institut d’Imagerie Biome´dicale, Molecular Imaging Research France; and {Commissariat a` l’E Center, IdM Nuclear Magnetic Resonance Laboratory, Paris, France
ABSTRACT The spin-spin (T2) relaxation of 1H-NMR signals in human skeletal muscle has been previously hypothesized to reveal information about myowater compartmentation. Although experimental support has been provided, no consensus has yet emerged concerning the attribution of specific anatomical compartments to the observed T2 components. Potential application of a noninvasive tool that might offer such information urges the quest for a definitive answer to this question. The purpose of this work was to obtain new information that might help elucidate the mechanism of T2 distribution in muscle. To do so, in vivo T2 relaxation data was acquired from the soleus of eight healthy volunteers using a localized Carr-Purcell-Meiboom-Gill technique. Each acquisition contained 1000 echoes with an interecho spacing of 1 ms. Data were acquired from each subject under different vascular filling preparations expected to change exclusively the extracellular water fraction. Two exponential components were systematically observed: an intermediate component (T2 ~ 32 ms) and a long component (100 < T2 < 210 ms). The relative fraction and T2 value characterizing the long component systematically increased after progressive augmentation of extracellular water volume. Characteristic relaxation behavior for each vascular filling condition was analyzed with a two-site exchange model and a three-site two-exchange model. We show that a two-site exchange model can only predict the observations for small exchange rates, much more representative of transendothelial than transcytolemmal exchange regimes. The three-site two-exchange model representing the intracellular, interstitial, and vascular spaces was capable of precisely predicting the observations for realistic transcytolemmal and transendothelial exchange rates. The estimated intrinsic relative fractions of each of these compartments corroborate with estimations from previous works and strongly suggest that the T2 relaxation from water within the intracellular and interstitial spaces is described by the intermediate component, whereas the long component represents water within the vascular space.
INTRODUCTION Alterations of intra- and extracellular myowater content and exchange kinetics are known to reflect exercise- and pathology-induced changes in tissue (1,2). A noninvasive technique giving access to such information in vivo would have important applications in the fields of exercise physiology and neuromuscular disorders. Nuclear magnetic resonance (NMR) is sensitive to these changes (3,4) and is a valuable tool for addressing these questions. Proton (1H) spin-spin (T2) relaxation has been shown to be multiexponential in many biological tissues. In skeletal muscle tissue, at least three exponential components were systematically observed in ex vivo and in vivo studies (5–10): a short one (T2 < 10 ms) accounting for 100 ms) accounting for components characterized by T2 values and relative fractions that correspond to the intermediate and long components also observed in previous studies (Fig. 3). This demonstrated the reproducibility of the methodology and supported the approach of a biexponential fit to analyze our data. Additionally, the results obtained from both methods were quite similar (Table 1). The vascular filling preparations exclusively influenced the long T2 component, whose relative fraction and value systematically increased after progressive augmentation of the extracellular space. These were strong evidences in favor of the anatomical compartmentation hypotheses. Deeper analysis of the compartmental exchanges is needed to elucidate the histological interpretation of this compartmentation. Our analyses showed that a 2SX model is only capable of explaining the observed relaxation data for slow exchange regimes (see Fig. 4). In previous studies investigating transcytolemmal water exchange, the intracellular residence time in muscle has been estimated between 550 and 1100 ms (24,25). In our analysis, when setting the intracellular residence time to 1000 ms, the results show that the observed T2-relaxation data cannot be explained by a 2SX model representing an intra-/extracellular compartmentation for realistic extracellular T2 values (T2b < 500 ms) (see Table 2). However, setting the
TABLE 2 Extracted intrinsic parameters of the 2SX model for the three vascular filling conditions at intermediate and slow exchange regimes; SE is presented within parentheses ti [s] Vascular condition 1
10
Vascular draining Free perfusion Vascular filling Vascular draining Free perfusion Vascular filling
Pa [%]
T2a [ms]
T2b [ms]
92.7(2.5) 30.0(0.5) 500(1216) 90.2(1.0) 30.1(0.3) 500(312) 85.2(0.5) 30.5(0.2) 500(81) 95.4(0) 31.8(0) 195.2(0) 92.8(0) 32.2(0) 199.5(0) 86.5(0) 32.7(0) 204.6(0)
R2 ( 103) 999.8 999.6 999.4 1000 1000 1000
R2 is the square of the correlation coefficient between predicted and experimental data.
FIGURE 4 Characteristic relaxation curves of the vascular draining (triangles), normal (squares), and vascular filling (circles) conditions, and the corresponding fitted curves characterized by the extracted intrinsic parameters of the 2SX model for both intermediate (crosses) and slow (lines) exchange regimes. Biophysical Journal 106(10) 2267–2274
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Araujo et al.
TABLE 3 Extracted intrinsic parameters of the 3S2X system characterizing the intracellular, interstitial, and vascular spaces for the three vascular filling conditions; standard error is presented within parentheses Vascular condition Pc [%]
Pi/Pe
T2i [ms] T2e [ms]
tc [ms]
R2
have studied a system composed by the intracellular and interstitial spaces according to a 2SX model. The analytical b 2b , b 2a and T solution of Eq. 5, for the apparent T2 values, T in the 2SX model has been previously presented by Hazlewood et al. (6), and is given by
Vascular draining 4.6(0) 12.7(0.3) 31.5(0) 35.8(1.0) 535(5) 1 Free perfusion 7.3(0) 6.73(0.04) 31.5 37.4(0.3) 1122(3) 1 Vascular filling 13.8(0) 6.18(0.04) 31.5 41.1(0.5) 9813(333) 1 R2 is the square of the correlation coefficient between predicted and experimental data.
intracellular residence time to 10 s avoids the need for abnormally high extracellular T2 values to explain the data (see Table 2). Intravascular residence times have been estimated between 0.3 and 3 s (24,26,27), and vascular water fraction shall be ~4% (28). For these values, the extravascular residence time at homeostasis must be between 7 and 72 s (Eq. 6), which characterizes a slow exchange regime. Theses evidences supported preferably an extra-/ intravascular compartmental interpretation, suggesting that relaxation of transverse magnetization from both intracellular and interstitial spaces are described by the intermediate component. This implies that the apparent intracellular and interstitial T2 values, Tic and Tint, shall be close enough to the observed intermediate T2. To investigate the intrinsic system parameters that might satisfy this condition, we
FIGURE 5 (a) Characteristic relaxation curves of the vascular draining (triangles), normal (squares), and vascular filling (circles) conditions, and the corresponding fitted curves (lines) characterized by the extracted intrinsic parameters of the 3S2X model. (b) Obtained T2 spectra from regularized inversion of the fitted curves resulting from the 3S2X model. Biophysical Journal 106(10) 2267–2274
with C1 ¼ 1 C2 ¼ 2
b 2a ¼ T
1 ; ðC1 þ C2 Þ
(9)
b 2b ¼ T
1 ; ðC1 C2 Þ
(10)
1 1 1 1 1 þ þ þ ; 2 T2a T2b t a t b
(11)
1 1 1 1 þ T2a t a T2b t b
2
4 þ ðt a t b Þ
!12 :
(12)
The average observed interstitial to intracellular water volume ratio is ~0.13 (28,29). Consequently, an intracellular residence time of 1 s would imply an interstitial residence time of 130 ms, at homeostasis. By applying these values into Eqs. 9–12, one may verify that for Tint > 80 ms, b 2a and T b 2b , would differ by the apparent T2 values, T >20 ms, which should be easily resolved in the obtained T2 spectra, considering the precision of our results for the observed intermediate T2 peaks (Fig. 3 b and Table 1). Usually, Tint is assumed to be considerably lengthened relative to the intracellular T2 due to the lower protein concentration in the interstitium. However, the macromolecules constituting the extracellular matrix have been shown to have nonnegligible relaxivities (30), and values as short as Tint ¼ 25 ms have been observed for synthetic interstitial models (9). This was further confirmed by the observed short T2 value in muscle sites with abnormally high endomysial accumulation such as fibrosis (31), and the long T2 values characteristic of interstitial edema (3). The 3S2X model has been previously shown to allow more accurate estimates of extracellular and vascular spaces and blood-to-tissue contrast agent transfer in pharmacokinetic studies (32). In this work, this model was shown to be capable of precisely predicting the characteristic relaxation curves for realistic physiological residence times (24– 27) (Fig. 5 and Table 3). The estimated interstitial T2 value was within the range, Tint < 80 ms, predicted by our 2SX analysis, and increased after progressive augmentation of its water content, as observed by the variations of the estimated intracellular to interstitial volume fraction ratio Pi/Pe (Table 3). The estimated compartmental relative fractions in normal conditions were in good agreement with results of previous studies (28,29). Estimated relative fraction of the vascular space increased after augmentation of extracellular volume. The
Interpretation of Muscle T2 Relaxation
augmentation of the intravascular residence, tc, must reflect attainment of quasi-normal equilibrium transendothelial exchange after fluid redistribution within vascular and interstitial spaces at each vascular condition. The observed nonlinear increase of tc with the vascular volume must reflect contributions of small veins and arteries within the VOI that could not be seen in the localization images, because they present a much higher compliance than capillaries and experience negligible exchange with the parenchyma. Vascular and interstitial water fractions varied between the imposed vascular filling conditions. While the vascular space varied progressively from one condition to another, the interstitial water fraction doubled when passing from the vascular draining to the vascular filling condition and presented a very small reduction (8%) when passing from the vascular filling to the normal condition. The rapid variation must be a result of the manual compression, which leads to interstitial space draining, whereas the slower variation reflects interstitial volume inertia as a consequence of the interstitial edema-preventing mechanisms (33). Furthermore, it has been previously shown that interstitial space volume varies at a much slower rate than the vascular space after imposed variations on vascular hydrostatic pressure (34). Finally, extracted T2 spectra from the fitted characteristic relaxation curves demonstrated that indeed only two T2 components may be obtained from the proposed 3S2X model characterized by the extracted intrinsic parameters (Fig. 5 b). In a previous study by Noseworthy et al. (12), the long T2 value resulting from biexponential fitting of relaxation data obtained within the soleus of healthy volunteers increased with hyperoxia exposure. This result was explained as reflecting the well-known blood-oxygenation-level-dependent effect (35) and the long T2 component was attributed to vascular water. In this work, the observed increase of the long T2 value after vascular filling was explained by an increase of the intravascular residence time. It is possible that blood O2 saturation was lower at vascular draining condition due to the imposed muscular ischemia of the procedure. However, the 3S2X model has too many parameters and vascular T2 had to be set constant to constrain the 3S2X model and obtain stable parameter estimations. Two-dimensional correlation NMR experiments have been previously proposed as a means for studying exchange dynamics (36); these methods allow simultaneous and independent estimation of compartment-intrinsic T2 value and exchange rates (13). The results from this work encourage the application of such methods for more precise parameter estimation of compartmental exchange models of myowater distribution. The interpretation of relaxometry studies relies on the accuracy and precision of the extracted parameters characterizing the multiexponential model that explains the observed relaxation. Such extraction constitutes an inverse
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Laplace transform of noisy data, to which there is no unique solution. Different methods for solving this problem without the need for imposing the number of T2 components characterizing the model exist (37–41) and are still the matter of research for the optimization of inversion methods. Our choice for a regularized inversion method was based on the fact that it yields nonnegative solutions and allows regularization, which has been shown by our results to ensure the reproducibility of the solutions. CONCLUSION 1. The biexponential T2 relaxation behavior of 1H-NMR signals observed in skeletal muscle reflects an anatomical compartmentation of myowater. 2. Water in the vascular space has nonnegligible contributions to observed muscle T2 values as obtained from monoexponential fitting of T2 relaxation data. 3. The biexponential T2 relaxation behavior observed in skeletal muscle cannot be explained with a 2SX model representing an intra-/extracellular anatomical compartmentation of myowater for realistic transcytolemmal exchange rates. However, for the slower transendothelial exchange rates, a 2SX model representing an extra-/ intravascular compartmentation allows precise prediction of the observations. 4. A 3S2X model representing water exchange dynamics among intracellular, interstitial, and vascular spaces is capable of explaining T2 relaxation behavior observed in skeletal muscle tissue for realistic transmembrane exchange rates and allowed estimating interstitial T2 value, which has been shown to be close to the intracellular T2 for healthy tissue in normal conditions. 5. For healthy skeletal muscle tissue in normal conditions, the observed intermediate T2 component reflects the relaxation behavior of water within the intracellular and interstitial spaces, and the long T2 component reflects the relaxation of water within the vascular space. The authors thank Dr. Alexandre Vignaud for his technical support during the development of the NMR sequences and the Drs. Paulo L. de Sousa, Noura Azzabou, Nicolas Decorte and Pierre-Yves Baudin for their critical reading of the manuscript.
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Biophysical Journal 106(10) 2267–2274
