Ferumoxytol-enhanced cardiovascular magnetic resonance detection of early stage acute myocarditis

Figures

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  Fig. 1. Study design The schema shows the timeline for each group and the CMR protocol

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  Fig. 2. Histological assessment of myocarditis Representative pictures of H&E staining (left column) and Masson Trichrome staining (the second column from the left) from Day 21 (upper row), Day 14 (middle row), and Control (lower row) groups are shown in the panels (a). The middle panels (a) show segmentations of inflammatory lesions in LV on ImageJ software. The nuclei are highlighted in red in those pictures. The right panels (a) show the segmented necrotic area (the second column from the right) and fibrotic area (right column). The necrotic area (b) and fibrotic area (c) of Day 21, Day 14, and Control hearts were quantified and displayed in the graphs

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  Fig. 3. Detection of myocarditis by FE-CMR, early gadolinium enhancement (EGE), and late gadolinium enhancement (LGE) Representative images of FE-CMR performed at 6 h (6 h-FE-CMR) and 24 h (24 h-FE-CMR) after ferumoxytol administration, EGE, and LGE, are shown (a) (upper row: day 21, middle row: day14, lower row: control). Enhanced areas suggesting inflammation are pointed by arrows. The right panels show inflammatory lesions (both left ventricle (LV) and right ventricle (RV)) segmented on 6 h-FE-CMR in light blue. The box and whisker graph (b) shows the regions detected as myocardial inflammation by 6 h-FE-CMR, 24 h-FE-CMR, EGE, and LGE. At day 14, FE-CMR detected more extensive myocardial inflammation compared with EGE (p = 0.02) and LGE (p < 0.01)

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  Fig. 4. T2* and delta R2* of the myocardium The representative mapping images (a) and graphs (b) compare the pre- and post-contrast (6 h and 24 h) T2* and delta R2* of LV between the control, day 14, and day 21 groups. The images were obtained from the same rats used in Fig. 3. The day 21 images are magnified to highlight the region with especially high T2* (septum and RV), which showed less ferumoxytol accumulation on 6 h-FE-CMR but high gadolinium accumulation on EGE and LGE (c). Histologically, this region corresponded with myocardial necrosis (right panel, arrows). The scatter plot (d) shows a positive linear correlation between the necrotic area size (%LV) and pre-contrast T2* of LV (r = 0.86, p < 0.001). [statistical analysis; * p = 0.01, †p < 0.01, ‡p < 0.001, compared to Control]

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  Fig. 5. Comparison of iron particle distribution between necrotic and non-necrotic inflammation Iron distribution in the myocardial inflammatory regions was compared between necrotic and non-necrotic tissues on Prussian blue staining (a). Necrotic area is highlighted by the dotted line. Prussian blue and H&E staining of the necrotic region (b & c, respectively) and the non-necrotic regions (d & e, respectively) clearly show the different distribution of ferumoxytol between those regions. The number of iron particles was counted in the high-power field and compared between the non-necrotic and necrotic inflammatory areas (e)

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  Fig. 6. Immunohistochemistry for macrophages Macrophage were detected as CD68 positive cells and visualized with DAB substrate. Macrophage infiltration in the myocardium was confirmed both at Day 21 (a) and Day 14 (b) but not in Control (c). Dual staining of iron particles (Prussian blue) and macrophages (DAB) shows their different distributions between 24 h (d) and 6 h (e) post-ferumoxytol injection

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  Fig. 7. Enhanced vascular permeability and FE-CMR. Inflammatory areas on 6 h-FE-CMR (left panel) and areas with Evans blue distribution (right), which represents enhanced vascular permeability, were compared in the hearts with myocardial inflammation (upper row) and without inflammation (lower row)