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. 2023 Nov;33(6):e13150.
doi: 10.1111/bpa.13150. Epub 2023 Jan 31.

Quantitative magnetic resonance imaging reflects different levels of histologically determined myelin densities in multiple sclerosis, including remyelination in inactive multiple sclerosis lesions

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Quantitative magnetic resonance imaging reflects different levels of histologically determined myelin densities in multiple sclerosis, including remyelination in inactive multiple sclerosis lesions

Vanessa Wiggermann et al. Brain Pathol. 2023 Nov.

Abstract

Magnetic resonance imaging (MRI) of focal or diffuse myelin damage or remyelination may provide important insights into disease progression and potential treatment efficacy in multiple sclerosis (MS). We performed post-mortem MRI and histopathological myelin measurements in seven progressive MS cases to evaluate the ability of three myelin-sensitive MRI scans to distinguish different stages of MS pathology, particularly chronic demyelinated and remyelinated lesions. At 3 Tesla, we acquired two different myelin water imaging (MWI) scans and magnetisation transfer ratio (MTR) data. Histopathology included histochemical stainings for myelin phospholipids (LFB) and iron as well as immunohistochemistry for myelin proteolipid protein (PLP), CD68 (phagocytosing microglia/macrophages) and BCAS1 (remyelinating oligodendrocytes). Mixed-effects modelling determined which histopathological metric best predicted MWF and MTR in normal-appearing and diffusely abnormal white matter, active/inactive, inactive, remyelinated and ischemic lesions. Both MWI measures correlated well with each other and histology across regions, reflecting the different stages of MS pathology. MTR data showed a considerable influence of components other than myelin and a strong dependency on tissue storage duration. Both MRI and histology revealed increased myelin densities in inactive compared with active/inactive lesions. Chronic inactive lesions harboured single scattered myelin fibres indicative of low-level remyelination. Mixed-effects modelling showed that smaller differences between white matter areas were linked to PLP densities and only to a small extent confounded by iron. MWI reflects differences in myelin lipids and proteins across various levels of myelin densities encountered in MS, including low-level remyelination in chronic inactive lesions.

Keywords: MRI; MTR; histopathology; iron; multiple sclerosis; remyelination.

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Conflict of interest statement

The authors have no conflicts to disclose related to the manuscript. Romana Höftberger has received speaker honoraria from Euroimmun, Novartis and Biogen.

Figures

FIGURE 1
FIGURE 1
Overview of processing pipeline. Differently stained histology sections were aligned with 2D FSL FLIRT. To register all quantitative images, first a pipeline was established based on the 3D T2 scans, which were reoriented and resliced to match the histology. Registration between histology and magnetic resonance imaging (MRI) was performed using NiftyReg. Quantitative maps were then registered to the 3D T2 and the already established pipeline and registration matrices were applied
FIGURE 2
FIGURE 2
Qualitative comparison of the histology and co‐registered MRI data for four sections from four cases (A: Case 4, B: Case 7, C: Case 1, D: Case 2). The histological images were contrast‐inverted to visually match the magnetic resonance imaging (MRI) contrasts so that lower intensities represent lower myelin or iron concentrations. Co‐registration achieved good matching of the MRI to the LFB stains, which were chosen as histology reference. Region of interests (ROIs) were centred in representative regions, yielding good local correspondence of ROIs on visual inspection. The ROIs were drawn with a gap to avoid partial volume effects at anatomical or lesion borders. AcInL, active/inactive lesions; DAWM, diffusely abnormal WM; InaL, inactive lesions; IsL, ischemic lesions; NAWM, normal‐appearing WM; SdP, shadow plaques
FIGURE 3
FIGURE 3
Histological sections of a double‐hemispheric coronal slice level of Case 7. (A) CD68 immunohistochemistry revealed accumulated activated microglia and macrophages at the edge of an active/inactive periventricular lesion (left black square, enlarged in C), but not at the inactive periventricular lesion edge (right black square, enlarged in D). (B) Similarly, TBB staining showed iron accumulation at the active/inactive (E), but not at the inactive lesions edge (F). Both lesions had iron loss in their centres. In WM, iron staining was highest subcortically, gradually decreased in DAWM and towards lesions, and was lowest within lesion centres. (G) Luxol fast blue–periodic‐acid Schiff (LFB‐PAS) myelin staining visually distinguished areas of normal‐appearing white matter (NAWM) (mainly subcortically and in the capsula interna), diffusely abnormal white matter (DAWM) and multiple sclerosis (MS) lesions. Black arrows indicate the ill‐defined DAWM, characterised by reduced myelin intensity. Both periventricular MS lesions (enlarged in I and J) showed myelin loss. Notably, scattered thin myelin sheaths were present within some inactive lesion centres (J, high magnification inset with arrows), but never in active/inactive lesions (I). Occasional macrophages with PAS‐positive cytoplasmic inclusions, indicative of remote demyelination, were found. (H) BCAS1 immunohistochemistry revealed moderate immunoreactivity in myelin, rendering exquisite histological myelin contrast. Strongly BCAS1‐positive glial cells without processes and dystrophic morphology accumulated in areas of WM damage, such as DAWM (see quantitative data) and WM abutting active/inactive lesion edges (K). Process‐bearing actively remyelinating oligodendrocytes could be found exclusively in few inactive lesions (L) and shadow plaques
FIGURE 4
FIGURE 4
Correlation of all MRI metrics. Both myelin water fraction (MWF) estimates showed excellent agreement with a minimal intercept (A), whereas correlations between MWF and magnetisation transfer ratio (MTR) depended on storage times (B,C). Correlations between MWF and MTR were significant in tissues with storage times of <1 year (dotted line) and were non‐significant for tissues with extended storage time >9 years (dashed line). Note that CPMG data were only available for samples with extended storage time (C). Symbols indicate different cases; colours distinguish different tissue types. AcInL, active/inactive lesions; DAWM, diffusely abnormal WM; InaL, inactive lesions; IsL, ischemic lesions; NAWM, normal‐appearing WM; SdP, shadow plaques
FIGURE 5
FIGURE 5
Comparison of quantitative MRI values and optical densities across regions of interest. Panels on the left display the histological density measurements [LFB (A), PLP (C), TBB (E)], panels on the right the magnetic resonance imaging (MRI) myelin measurements (GraSE myelin water fraction (MWF) (B), magnetisation transfer ratio (MTR) (D)). Panel F displays the separately obtained BCAS1 counts. LFB and proteolipid protein (PLP) differences between region of interests (ROIs), including in lesions, were well reflected in both MWF measurements. By contrast, MTR changes appeared more closely linked to the Turnbull Blue (TBB) staining intensities and MTR values overlapped between regions. Note that the data collection for Carr–Purcell–Meiboom–Gill (CPMG) and MTR were incomplete, thus not all ROIs are present in the MTR plot. AcInL, active/inactive lesions; DAWM, diffusely abnormal WM; InaL, inactive lesions; IsL, ischemic lesions; NAWM, normal‐appearing WM; SdP, shadow plaques
FIGURE 6
FIGURE 6
Within‐case correlations between LFB and GraSE MWF across all ROIs. In order to suppress between‐section deviations, all correlations shown are weighted averages of the individual within‐section correlations. Different symbols distinguish data from different sections of a case. Different colours indicate the different region of interest (ROI) types. The slopes of the average correlations ranged from 0.06% to 0.15% MWF per optical density unit. All correlations were significant. Case 5 is not shown as the GraSE data were deemed unreliable for analysis. AcInL, active/inactive lesions; DAWM, diffusely abnormal WM; InaL, inactive lesions; IsL, ischemic lesions; NAWM, normal‐appearing WM; SdP, shadow plaques

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