Box plots of the model parameter <i>v</i>_<i>b</i> obtained with the different models and methods in the different skeletal muscle regions of the pigs’ hind legs.

<p>Box plots of the model parameter <i>v</i>_<i>b</i> obtained with the different models and methods in the different skeletal muscle regions of the pigs’ hind legs.</p

Validation of Blood Volume Fraction Quantification with 3D Gradient Echo Dynamic Contrast-Enhanced Magnetic Resonance Imaging in Porcine Skeletal Muscle

<div><p>The purpose of this study was to assess the accuracy of fractional blood volume (<i>v</i>_<i>b</i>) estimates in low-perfused and low-vascularized tissue using dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI). The results of different MRI methods were compared with histology to evaluate the accuracy of these methods under clinical conditions. <i>v</i>_<i>b</i> was estimated by DCE-MRI using a 3D gradient echo sequence with k-space undersampling in five muscle groups in the hind leg of 9 female pigs. Two gadolinium-based contrast agents (CA) were used: a rapidly extravasating, extracellular, gadolinium-based, low-molecular-weight contrast agent (LMCA, gadoterate meglumine) and an extracellular, gadolinium-based, albumin-binding, slowly extravasating blood pool contrast agent (BPCA, gadofosveset trisodium). LMCA data were evaluated using the extended Tofts model (ETM) and the two-compartment exchange model (2CXM). The images acquired with administration of the BPCA were used to evaluate the accuracy of <i>v</i>_<i>b</i> estimation with a bolus deconvolution technique (BD) and a method we call equilibrium MRI (EqMRI). The latter calculates the ratio of the magnitude of the relaxation rate change in the tissue curve at an approximate equilibrium state to the height of the same area of the arterial input function (AIF). Immunohistochemical staining with isolectin was used to label endothelium. A light microscope was used to estimate the fractional vascular area by relating the vascular region to the total tissue region (immunohistochemical vessel staining, IHVS). In addition, the percentage fraction of vascular volume was determined by multiplying the microvascular density (MVD) with the average estimated capillary lumen, , where <i>d</i> = 8<i>μ</i>m is the assumed capillary diameter (microvascular density estimation, MVDE). Except for ETM values, highly significant correlations were found between most of the MRI methods investigated. In the cranial thigh, for example, the <i>v</i>_<i>b</i> medians (interquartile range, IQRs) of IHVS, MVDE, BD, EqMRI, 2CXM and ETM were <i>v</i>_<i>b</i> = 0.7(0.3)%, 1.1(0.4)%, 1.1(0.4)%, 1.4(0.3)%, 1.2(1.8)% and 0.1(0.2)%, respectively. Variances, expressed by the difference between third and first quartiles (IQR) were highest for the 2CXM for all muscle groups. High correlations between the values in four muscle groups—medial, cranial, lateral thigh and lower leg - estimated with MRI and histology were found between BD and EqMRI, MVDE and 2CXM and IHVS and ETM. Except for the ETM, no significant differences between the <i>v</i>_<i>b</i> medians of all MRI methods were revealed with the Wilcoxon rank sum test. The same holds for all muscle regions using the 2CXM and MVDE. Except for cranial thigh muscle, no significant difference was found between EqMRI and MVDE. And except for the cranial thigh and the lower leg muscle, there was also no significant difference between the <i>v</i>_<i>b</i> medians of BD and MVDE. Overall, there was good <i>v</i>_<i>b</i> agreement between histology and the BPCA MRI methods and the 2CXM LMCA approach with the exception of the ETM method. Although LMCA models have the advantage of providing excellent curve fits and can in principle determine more physiological parameters than BPCA methods, they yield more inaccurate results.</p></div

Light microscopy image of a histological preparation from the medial thigh muscle with vascular endothelium stained brown by isolectin.

<p>The selection was performed by semiautomatic analysis of the images with the microscope’s morphometry software to determine the area, number and diameter of the vessels. The outer borders of the segmented vessels are labeled in green.</p

Spearman correlation coefficients of the individual results of the four MRI methods for the muscle groups in the hind legs of the pigs and Wicoxon rank sum test results between the different MRI methods and between MRI and histology.

<p>Spearman correlation coefficients of the individual results of the four MRI methods for the muscle groups in the hind legs of the pigs and Wicoxon rank sum test results between the different MRI methods and between MRI and histology.</p

BPCA data recorded from CA-induced time-dependent relaxation rate changes in a) the aorta (related to the arterial hematocrit level Hct_<i>a</i>)) and b) the medial thigh muscle (black circles) of the same experiment as the examples in Fig 2.

<p>Additionally, in a) the time delay and bolus-dispersion-corrected AIF and in b) one example of the fitting results with the BD method (red solid lines) is shown. As indicated by the statistical results (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0170841#pone.0170841.t001" target="_blank">Table 1</a>), <i>v</i>_<i>b</i> is determined lower by BD compared with 2CXM (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0170841#pone.0170841.g002" target="_blank">Fig 2</a>) and EqMRI (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0170841#pone.0170841.g004" target="_blank">Fig 4</a>).</p

Median values of the model parameter <i>v</i>_<i>b</i> with first (Q1) and third quartiles (Q3) and interquartile ranges (IQR) obtained with the different models and methods in the different skeletal muscle regions of the pigs’ hind legs.

<p>Also shown are the mean values of the contrast-to-noise ratio (CNR) of the CA time courses for each muscle region. The mean CNRs of the AIFs were 80 ± 40 (92 ± 37) for the BPCA (LMCA) measurements, respectively.</p

For the EqMRI method, the same BPCA relaxation rate changes as in Fig 3 are shown as black circles.

<p>No correction with respect to the arterial hematocrit level was performed at this point. Red solid lines represent the estimated average equilibrium states of a) the AIF in the aorta and in b) the medial thigh muscle (black circles) according to <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0170841#pone.0170841.e018" target="_blank">Eq (14)</a> are shown.</p

Comparison of blood and plasma volume fractions of skeletal muscle of different species reported in the literature.

<p>Comparison of blood and plasma volume fractions of skeletal muscle of different species reported in the literature.</p