Tracking iron in multiple sclerosis: a combined imaging and histopathological study at 7 Tesla

Abstract

Previous authors have shown that the transverse relaxivity R 2 * and frequency shifts that characterize gradient echo signal decay in magnetic resonance imaging are closely associated with the distribution of iron and myelin in the brain's white matter. In multiple sclerosis, iron accumulation in brain tissue may reflect a multiplicity of pathological processes. Hence, iron may have the unique potential to serve as an in vivo magnetic resonance imaging tracer of disease pathology. To investigate the ability of iron in tracking multiple sclerosis-induced pathology by magnetic resonance imaging, we performed qualitative histopathological analysis of white matter lesions and normal-appearing white matter regions with variable appearance on gradient echo magnetic resonance imaging at 7 Tesla. The samples used for this study derive from two patients with multiple sclerosis and one non-multiple sclerosis donor. Magnetic resonance images were acquired using a whole body 7 Tesla magnetic resonance imaging scanner equipped with a 24-channel receive-only array designed for tissue imaging. A 3D multi-gradient echo sequence was obtained and quantitative R 2 * and phase maps were reconstructed. Immunohistochemical stainings for myelin and oligodendrocytes, microglia and macrophages, ferritin and ferritin light polypeptide were performed on 3-to 5-mm thick paraffin sections. Iron was detected with Perl's staining and 3,3 0 -diaminobenzidine-tetrahydrochloride enhanced Turnbull blue staining.In multiple sclerosis tissue, iron presence invariably matched with an increase in R 2 *. Conversely, R 2 * increase was not always associated with the presence of iron on histochemical staining. We interpret this finding as the effect of embedding, sectioning and staining procedures. These processes likely affected the histopathological analysis results but not the magnetic resonance imaging that was obtained before tissue manipulations. Several cellular sources of iron were identified. These sources included oligodendrocytes in normal-appearing white matter and activated macrophages/microglia at the edges of white matter lesions. Additionally, in white matter lesions, iron precipitation in aggregates typical of microbleeds was shown by the Perl's staining. Our combined imaging and pathological study shows that multi-gradient echo magnetic resonance imaging is a sensitive technique for the identification of iron in the brain tissue of patients with multiple sclerosis. However, magnetic resonance doi:10.1093/brain/awr278 imaging-identified iron does not necessarily reflect pathology and may also be seen in apparently normal tissue. Iron identification by multi-gradient echo magnetic resonance imaging in diseased tissues can shed light on the pathological processes when coupled with topographical information and patient disease history.

<i>In Vivo</i> Brain MR Imaging at Subnanoliter Resolution: Contrast and Histology

“…63,70 Accordingly, layer IV in the human visual cortex, well known as the stria of Gennari for its pronounced myelination, can be clearly delineated by MR imaging 1,2 based on a locally increased T 2 relaxation rate. The effect is supposed to be caused by iron rather than macromolecules because the corresponding ex vivo MR imaging contrast 68,71 is better co-localized with the iron distribution than with myelin or cell assemblies and is reduced after chemical extraction of tissue iron. 68 …”

“…Microglial cells are able to transform toxic Fe 2+ ions into more stable Fe 3+ ions by binding them to ferritin. 71 The resulting accumulation of activated microglias may therefore shorten the T 1 and T 2 relaxation times 64 of affected brain tissue. Microglial cells may also express a high oxygen superoxide scavenging potential related to the presence of MnSOD.…”

“…91 Reduced T 2 * in ex vivo brain MR imaging of a patient with multiple sclerosis co-localized with both oligodendrocytes and activated microglial cells that had taken up iron released by tissue damage. 71 Accordingly, subtle MTR reductions in otherwise normal-appearing WM are associated with marked microglial activation in the brain of patients with multiple sclerosis 92 because intracellular paramagnetic ions are known to reduce MT saturation in vivo. 30,32 One MR imaging study 90 using rats observed an accumulation of activated microglia after transient ischemia as a source of T 1 contrast in vivo in line with a case report in humans.…”

“…1, 2, 4, 5, 6), [28][29][30][31] there may be a particular inflammatory condition in which the MTR is more sensitive than T 1 W or T 2 W MR imaging. In other words, longer T 1 and T 2 caused by increased free water content may be counterbalanced by the astrocytic, microglial, macrophagic, or hemorrhagic accumulation of iron and manganese, 71,75,80,87,91 whereas the MTR is reduced by each of these processes (Table 2). For example, an increase in astrocytes and activated microglia appears to outweigh the decrease in oligodendrocytes in plaques, 107 which may offset prolonged relaxation times due to increased free water.…”

“…In fact, a number of investigations reported a significantly reduced MTR in otherwise normal-appearing WM and GM, 92,106,[108][109][110][111][112][113][114] a close correlation between the MTR and clinical disability, 114 -117 and a potential prognostic value of magnetization transfer MR imaging. 114,118 -120 More recent studies suggest that iron-laden/activated microglia 71,121 and/or aggregates typical of small bleeding 71 cause focally increased T 2 relaxation rates and/or reduced MTR 92 within normal-appearing WM. Conflicting results 122 may be explained by heterogeneous and variable patterns in pathology, e.g., the extent of tissue damage and remyelination.…”