Diffusion tensor imaging and histology of developing hearts

Abstract

Diffusion tensor imaging (DTI) has emerged as a promising method for noninvasive quantification of myocardial microstructure. However, the origin and behavior of DTI measurements during myocardial normal development and remodeling remain poorly understood. In this work, conventional and bicompartmental DTI in addition to three-dimensional histological correlation were performed in a sheep model of myocardial development from third trimester to postnatal 5 months of age. Comparing the earliest time points in the third trimester with the postnatal 5 month group, the scalar transverse diffusivities preferentially increased in both left ventricle (LV) and right ventricle (RV): secondary eigenvalues D2 increased by 54% (LV) and 36% (RV), whereas tertiary eigenvalues D3 increased by 85% (LV) and 67% (RV). The longitudinal diffusivity D1 changes were small, which led to a decrease in fractional anisotropy by 41% (LV) and 33% (RV) in 5 month versus fetal hearts. Histological analysis suggested that myocardial development is associated with hyperplasia in the early stages of the third trimester followed by myocyte growth in the later stages up to 5 months of age (increased average myocyte width by 198%, myocyte length by 128%, and decreased nucleus density by 70% between preterm and postnatal 5 month hearts.) In a few histological samples (N = 6), correlations were observed between DTI longitudinal diffusivity and myocyte length (r = 0.86, P < 0.05), and transverse diffusivity and myocyte width (r = 0.96, P < 0.01). Linear regression analysis showed that transverse diffusivities are more affected by changes in myocyte size and nucleus density changes than longitudinal diffusivities, which is consistent with predictions of classical models of diffusion in porous media. Furthermore, primary and secondary DTI eigenvectors during development changed significantly. Collectively, the findings demonstrate a role for DTI to monitor and quantify myocardial development, and potentially cardiac disease. Copyright © 2016 John Wiley & Sons, Ltd.

Keywords: DTI; fractional anisotropy; helix angle; mean diffusivity; preterm lamb heart; principal diffusivities; sheet angle.

Figure 1

From top to lower row, representative gross pictures and DTI maps of developing lamb hearts, including scalar fractional anisotropy (FA), mean diffusivity (MD), fiber helix angle (HA) and E2A (obtained from the secondary eigenvector). Fetal hearts at 120 (D120), 130 (D130), 140 days (D140) post gestation, and term 150 (D150) and 5-month (5m) old hearts are shown. Units of MD (×10⁻³ mm²/s), and HA and E2A (degrees).

Figure 2

Groups averaged DTI scalar measurements (mean±SEM) taken from regions of interest in the left (LV, top row) and right ventricles (RV, bottom row). Asterisks (*) denote Bonferroni corrected P<0.05 compared to 5-months group. Primary (D₁), secondary (D₂), tertiary (D₃), and fractional anisotropy (FA) are shown. Units of diffusivities (×10⁻³ mm²/s).

Figure 3

Group-averaged histograms of myocardial HA (a, b) and E2A (c, d) angles measured from 5 slices in each heart for the LV and RV, respectively. Lines and error-bars represent (mean±SEM) of the respective bin. For clarity of presentation, only the averaged histograms from 3 groups are shown for D120 (N=6), D150 (N=6), and 5m (N=8).

Figure 4

Normalized diffusion-induced signal attenuation in regions of interest taken from the circumferential fiber region of preterm 130-days (D130) and term 5-months (5m) hearts for b-value up to 5,000 s/mm². Bi-exponential fitting (solid line) is shown for 2 encoding directions (a) longitudinal, and (b) transverse to myocytes.

Figure 5

Representative maps obtained from bicompartment DTI showing maps of fast (a, b) and slow (c, d) fractional anisotropy, fast (e, f) and slow (g, h) mean diffusivity (MD), and fast (i, j) and slow (k, l) helix angle (HA) maps from 130-days preterm and 5-month old hearts, respectively. Units of MD (×10⁻³ mm²/s), and HA (degrees).

Figure 6

Confocal images from fetal lambs at 100- and 130-days, full term (at 150-days, or D150), and postnatal term 5-months (5m) hearts. Composite tile-scans for DAPI (top row, nuclei stain in blue), WGA second row (mainly extracellular space stains in green) are shown for 1 mm × 1 mm tissue area, in addition to zoomed longitudinal (third row) and transverse (lower row) views (overlaid DAPI and WGA) taken from 3D acquisitions with an image resolution of 0.2 μm × 0.2 μm × 0.2 μm. Scale bar is 50 μm.

Figure 7

Scatter plots of DTI scalar longitudinal (D₁) vs. myocyte length (a) and nucleus density (b), transverse diffusivities (average D₂ and D₃) vs. microstructural myocyte width (c) and nucleus density (d). Data points represent the mean and SEM of 3 measurements from each heart that underwent both DTI and confocal microscopy. Linear regression (solid lines) and the 95% confidence interval (dashed lines) are shown in addition to the coefficient of determination (R²) and its associated P value.