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. 2016 Jul 5;12:219–226. doi: 10.1016/j.nicl.2016.07.003

Diffusivity in multiple sclerosis lesions: At the cutting edge?☆

Alexander Klistorner a,b,⁎, Chenyu Wang c, Vera Fofanova b, Michael H Barnett c, Con Yiannikas d, John Parratt d, Yuyi You a,b, Stuart L Graham b
PMCID: PMC4950592  PMID: 27489769

Abstract

Background

Radial Diffusivity (RD) has been suggested as a promising biomarker associated with the level of myelination in MS lesions. However, the level of RD within the lesion is affected not only by loss of myelin sheaths, but also by the degree of tissue destruction. This may lead to exaggeration of diffusivity measures, potentially masking the effect of remyelination.

Objective

To test the hypothesis that the T2 hyperintense lesion edge that extends beyond the T1 hypointense lesion core is less affected by tissue loss, and therefore a more appropriate target for imaging biomarker development targeting de- and re-myelination.

Method

Pre- and post-gadolinium (Gd) enhanced T1, T2 and DTI images were acquired from 75 consecutive RRMS patients. The optic radiation (OR) was identified in individual patients using a template-based method. T2 lesions were segmented into T1-hypointense and T1-isointense areas and lesion masks intersected with the OR. Average Radial, Axial and Mean diffusivity (RD, AD and MD) and fractional anisotropy (FA) were calculated for lesions of the entire brain and the OR. In addition, Gd enhancing lesions were excluded from the analysis.

Results

86% of chronic T2 lesions demonstrated hypointense areas on T1-weighted images, which typically occupied the central part of each T2 lesion, taking about 40% of lesional volume. The T1-isointense component of the T2 lesion was most commonly seen as a peripheral ring of relatively constant thickness (“T2-rim”). While changes of diffusivity between adjacent normal appearing white matter and the “T2-rim” demonstrated a disproportionally high elevation of RD compare to AD, the increase of water diffusion was largely isointense between the “T2-rim” and T1-hypointense parts of the lesion.

Conclusion

Distinct patterns of diffusivity within the central and peripheral components of MS lesions suggest that axonal loss dominates in the T1 hypointense core. The effects of de/remyelination may be more readily detected in the “T2-rim”, where there is relative preservation of structural integrity. Identifying and separating those patterns has an important implication for clinical trials of both neuroprotective and, in particular, remyelinating agents.

Highlights

  • •

    Distinct patterns of diffusivity within the central and peripheral components of MS lesions were identified.

  • •

    Axonal loss is likely to dominate the T1 hypointense core.

  • •

    The effects of de/remyelination may be more readily detected in the “T2-rim”.

1. Introduction

Recent interest in the development of remyelinating therapies has increased demand for reliable in vivo surrogate markers of remyelination. Quantification of the diffusion characteristics of brain tissue, in particular Radial Diffusivity, has been suggested as a promising imaging biomarker associated with the level of tissue myelination. Experimental models of demyelination have demonstrated a close correlation between degree of myelin loss and alterations in RD (Song et al., 2005; Janve et al., 2013). In post-mortem studies of human MS brains, elevation of RD was topographically linked to areas of histologically identified demyelination (Schmierer et al., 2007; Schmierer et al., 2008; Wang et al., 2015). A close relationship between increase in RD and electrophysiological measures of demyelination was also reported in patients with MS (Alshowaeir et al., 2014). However, some recent studies have failed to demonstrate an unequivocal relationship between increased RD and the degree of demyelination, suggesting that this measure is not pathologically specific (Klawiter et al., 2012).

Currently, conventional MRI is the gold standard to identify focal inflammatory demyelination in MS. Typically, acute T2 lesions comprise an area of demyelination and surrounding edema. Resolution of acute inflammation and edema are probably responsible for reduction in lesion size, with a permanent residual lesion that includes demyelinated (and partially remyelinated) axons (Schmierer et al., 2009) and areas of expanded extracellular space. The widening of extracellular space is believed to be caused by tissue destruction and may occupy up to 87% of the lesion volume (Barnes et al., 1991; Miller, 2008). The majority of chronic T2 lesions are also seen as hypointense areas on T1-weighted images, although typically the change of the signal intensity on the T1-weighted image occurs only in part of the T2 lesion (Barkhof and van Walderveen, 1999). The level of T1 hypointensity varies significantly from slightly hypointense in comparison to surrounding NAWM to approaching the intensity of CSF, reflecting a variable degree of widening of the extracellular space (Loevner et al., 1995; Van Waesberghe et al., 1998; Miller, 2008).

The extent of lesional tissue loss is also closely linked to DTI measures, since expansion of extra-cellular space dramatically increases isotropic diffusion of water molecules (Rovaris et al., 2005). Therefore, it is highly likely that the level of RD within the lesion is affected not only by loss of myelin sheath, but also by the level of tissue destruction. Tissue destruction may therefore ‘exaggerate’ both RD and AD (Wang et al., 2011), masking the potential effect of remyelination on diffusivity measures.

The purpose of this study was to examine diffusivity indices in T2 and T1 lesions of MS patients. Since T1 changes, which are closely relate to loss of tissue matrix, constitute only part of a T2 lesion, we hypothesized that the T2 hyperintense lesion edge that extends beyond the T1 hypointense lesion core (which we called “T2-rim” area) may be less affected by tissue loss, and therefore be a better target for studying the effects of de/remyelination.

The specificity of altered diffusion for pathologic changes is limited by the wide spectrum of normal anisotropy indices in the brain (Bammer et al., 2000). We studied lesions in the optic radiations, highly organized fibre tracts that are a frequent site of MS pathology, to facilitate accurate measurement of relative diffusivity change along axonal bundles (Mädler et al., 2008). In addition, internal structure of the OR does not contain a significant number of crossing fibers, which can potentially (and sometimes paradoxically) alter diffusivity (Yeatman et al., 2012; Winston, 2012). This point is especially pertinent considering the issues that surround misalignment between corresponding eigenvectors with the underlying tissue structures (Wheeler-Kingshott and Cercignani, 2009).

2. Material and methods

The study was approved by University of Sydney and Macquarie University Human Research Ethics Committees. All procedures followed the tenets of the Declaration of Helsinki and written informed consent was obtained from all participants.

2.1. Subjects

Seventy-five consecutive patients with Relapsing-Remitting MS (RRMS) and no history of clinical optic neuritis (ON) in at least one eye were enrolled. RRMS was defined according to standard criteria (Polman et al., 2011). A history of ON was based on the patient's clinical notes and the absence of previous symptoms. Patients with any other systemic or ocular disease, in particular those that could potentially affect our measurement parameters were excluded.

2.2. MRI protocol

The following sequences were acquired using a 3T GE Discovery MR750 scanner (GE Medical Systems, Milwaukee, WI):

1. Pre- and postcontrast (gadolinium) Sagittal 3D T1: GE BRAVO sequence, FOV 256 mm, Slice thickness 1 mm, TE 2.7 ms, TR 7.2 ms, Flip angle 12°, Pixel spacing 1 mm. Acquisition Matrix (Freq. × Phase) is 256 × 256, which results in 1 mm isotropic acquisition voxel size. The reconstruction matrix is 256 × 256.

2. FLAIR CUBE; GE CUBE T2 FLAIR sequence, FOV 240 mm, Slice thickness 1.2 mm, Acquisition Matrix (Freq. × Phase) 256 × 244, TE 163 ms, TR 8000 ms, Flip angle 90°, Pixel spacing 0.47 mm. The reconstruction matrix is 512 × 512.

3. Whole brain 64-directions diffusion weighted imaging with 2 mm isotropic acquisition matrix (TR/TE = 8325/86 ms, b = 1000 s/mm2, number of b0s = 2).

2.3. Reconstruction of individual optic radiations

Individual optic radiation masks was reconstructed from in-house optic radiation template through non-linear registration as follow:

  • 1.

    3DT1 images were lesion-inpainted (FSL) using T2 lesion masks,

  • 2.

    individual brain masks were derived using Brain Extraction Tool from FSL with manual quality control,

  • 3.

    deformation maps were obtained from non-linear registration between individual brain and ICBM 2009a standard brain template (Fonov et al., 2011),

  • 4.

    in-house optic radiation template was mapped into individual patient's space through the deformation map.

2.4. Lesion identification and analysis

Whole brain T2 lesions were identified on the co-registered T2 FLAIR images and segmented semi-automatically using ITK-SNAP 3 software (http://www.itksnap.org) by a trained analyst. To minimize partial volume effect, only T2 lesions with a volume larger than 100 mm3 were evaluated.

T1 lesions were identified on pre-contrast 3D-T1-weighted images. They were defined as regions with low signal intensity relative to the surrounding white matter and corresponded with an area of high signal intensity on T2. The lesion (hypo-) intensity was not quantified. T2 lesions demonstrating Gd-enhancement were excluded from analysis. Similar to T2 lesions, T1 lesions were segmented semi-automatically using ITK-SNAP 3 software by a trained analyst (VF). Both T2 and T1 lesion segmentation was verified by two more persons, who have extensive experience in lesion segmentation (AK, CW). Occasional discrepancies were resolved by consensus.

Each T1 hypointense lesion typically occupied the central area of a T2 lesion, leaving the peripheral part of the T2 lesion T1-isointense. We called this part of the T2 lesion the “T2-rim” (Fig. 1). A minority of T2 lesions did not display T1 hypointensity; these lesions were typically small or had a linear, elongated shape (blue arrowheads in Fig. 1), contributing little to the total T2 lesion volume. Therefore, for the purpose of analysis, T1-hypointense and T2-rim areas of T2 lesions were evaluated separately.

Fig. 1.

Fig. 1

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Examples of lesion segmentation. Green-T1 hypointense area, Red-“T2-rim” area. Blue arrowheads denote T2 lesions without corresponding T1 hypointensity. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)

Since the majority of lesions had a spherical shape, we calculated radius of T2 and T1 lesions by dividing the total lesion load by the lesion number. In addition, the width of the T2-rim area was calculated as a difference between the radii of T2 and T1 lesions.

Lesion masks were then intersected with the OR to identify and measure the volume of T2, T1 and T2-rim lesions within the OR, as described in detail elsewhere (Klistorner et al., 2015).

For single lesion analysis, only well demarcated T2 lesions with no visible abnormality in the corresponding part of contralateral OR on both T2 and T1 images were selected. The region of the contralateral OR corresponding in volume and position to T2 lesion was used as a reference representing NAWM (Fig. 2).

Fig. 2.

Fig. 2

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Analysis of single optic radiation lesion.

3. Results

Seventy-five consecutive RRMS patients (age: 41.6 ± 10.1, disease duration: 4.9 ± 3.6 years, 25M/50F, EDSS score: 1.42 ± 1.38) were enrolled in the study. Patients with any other systemic or ocular diseases were excluded. All patients were relapse-free for at least 3 months prior to the enrolment. Seventy patients (93%) were receiving disease-modifying therapy at the time of enrolment (7-Betaferon, 20-Copaxon, 25-Gylenia, 6-Tysabril, 8-Avonex, 2-Tecfidera, 2-Rebif).

A history of optic neuritis (ON) in one eye was not an exclusion criteria, however, none of the patients had ON or visual disturbances within 6 months of baseline assessment. All patients with history of ON received steroid as part of ON treatment.

There were total of 849 lesions identified on T2-FLAIR scans (Table 1). Of these, 726 lesions (86%) demonstrated (partial) hypointensity on T1 images. The mean T1 lesion volume constituted approximately 41% of T2 lesion volume. There was highly significant correlation between T2 and T1 lesion number, lesion volume and average single lesion radius (r2 = 0.96, 0.92 and 0.86 respectively, p < 0.001 for all) (Fig. 3 below). However, while the volume of the T2-rim area correlated significantly with total T2 lesion volume (r2 = 0.95, p < 0.001), there was no correlation between width of T2-rim ring and T2 lesion radius (r2 = 0.049, p > 0.05) indicating that the width of the T2-rim is independent of the volume of T2 lesion, and remains relatively constant.

Table 1.

Whole brain lesions (Mean ± SD).

Number of lesions Lesion volume mm3 Average volume of single lesion, mm3 Average radius of single lesion, mm
T2 lesions 849 5546 ± 6001 430 ± 211 4.64 ± 0.75
T1 lesions 726 2191 ± 2798 187 ± 119 3.45 ± 0.74
T2-rim area 849 3852 ± 3544 267 ± 119 1.2 ± 0.29 (width)
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Fig. 3.

Fig. 3

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Correlation between T2 lesion radius and: a) T1 lesion radius, b) T2-rim area width.

Of 75 patients, 63 (84%) demonstrated T2 lesions within the ORs. The mean volume of OR T2, T1 and T2-rim area was 1303 ± 1269, 460 ± 558 and 836 ± 771 mm3 respectively. There was a highly significant correlation between OR T2 and T1 lesion volumes (R2 = 0.88) (Fig. 4). OR lesion volume also correlated significantly with whole brain lesion volume (r2 = 0.74 and r2 = 0.70 for T2 and T1 lesions respectively).

Fig. 4.

Fig. 4

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Correlation between OR T2 and T1 lesion volume.

3.1. DTI analysis

Diffusivity coefficients of entire OR and OR lesions (total T2 lesions, T1 and T2-rim areas) are presented in Table 2.

Table 2.

Diffusivity coefficients of OR and OR lesions (Mean (SD)).

AD RD FA MD
Entire OR 1.17 (0.04) 0.68 (0.05) 0.35 (0.03) 0.85 (0.05)
T2 lesions 1.47 (0.15) 0.87 (0.10) 0.33 (0.04) 1.07 (0.11)
T1 area 1.62 (0.22) 1.00 (0.14) 0.31 (0.05) 1.22 (0.14)
T2-rim area 1.39 (0.17) 0.81 (0.08) 0.34 (0.05) 1.01 (0.08)
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Both RD and AD of the OR demonstrated highly positive correlations with OR T2 lesion load (r2 = 0.69 and r2 = 0.46 respectively, p < 0.001 for both) and OR T1 lesion load (r2 = 0.59 and r2 = 0.48 respectively, p < 0.001 for both), while OR FA was negatively correlated with volume of OR lesions (r2 = − 0.53 and r2 = − 0.40 for T2 and T1 lesions respectively, p < 0.001 for both) (Fig. 5).

Fig. 5.

Fig. 5

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Correlation between OR T2 lesion volume and OR diffusivity coefficients including AD, RD and FA.

Compartmentalization of T2 lesions revealed that both AD and RD were significantly higher in T1 areas compared to the T2-rim area, producing a 21% rise in MD value between the two. While absolute change in diffusivity between T2-rim and T1 areas was very similar for both RD and AD (0.20 and 0.24), RD demonstrated slightly larger relative increase compare to AD (24% vs 17% for RD and AD respectively), which resulted in a small, but significant drop of FA between T2-rim and T1 areas (− 0.03 or 8.8%). In addition, AD and RD were highly correlated between each other in T1 lesional areas (r2 = 0.63, p < 0.001), suggesting predominantly isotropic diffusivity, while in T2-rim areas the correlation was less apparent (r2 = 0.31, p < 0.01) (Fig. 6).

Fig. 6.

Fig. 6

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Correlation between AD and RD in: a) T1 area, b) “T2-rim” area.

We have recently reported a very different pattern of diffusivity change between OR lesional (T2) tissue and OR NAWM in a similar cohort of MS patients (Klistorner et al., 2015), which demonstrated more significant increase of RD compare to AD, resulting in substantial drop of FA (18%) (Fig. 7). Since in the previous study the entire T2 lesional area (including the T1 component) was analysed, it is reasonable to assume that the difference between increase of RD and AD at the NAWM/T2-rim border (and, as a result, the drop in FA) is even higher.

Fig. 7.

Fig. 7

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Change of diffusivity coefficients at the borders of NAWM-“T2-rim” area and “T2-rim”-T1 area. While RD increases to exactly the same amount between NAWM and “T2-rim” area and between “T2-rim” area and T1 area, increase of AD is much more pronounced between “T2-rim” area and T1 area compare to border between NAWM and “T2-rim” area. This results in significantly larger drop of FA between NAWM and “T2-rim” area compare to border between “T2-rim” area and T1 area. Vertical axis represents change in diffusivity for RD, AD (μm2/ms for both) and FA (arbitrary units).

To confirm this we performed the analysis of diffusivity in individual OR lesions. Twenty-seven subjects had isolated T2 lesions on one side of the OR and normal looking white matter in the corresponding part of the contralateral OR (which was used as a reference area analogous to NAWM). Diffusivity of T1 and T2-rim areas of the lesions were measured separately.

The result of this single lesion analysis (Table 3) demonstrated that RD continued to increase from NAWM to T2-rim area and then to T1 area with equal increments (0.20 and 0.20) The gradient of AD rise, however, increased from 0.10 at the border between NAWM and T2-rim area to 0.20 at the border between T2-rim and T1 areas. This was reflected in a more substantial drop of FA at NAWM/T2-rim border (− 0.12) compared to T2-rim/T1 border (− 0.05).

Table 3.

Diffusivity coefficients of OR single lesions (Mean (SD)).

AD RD FA MD
NAWM 1.39 (0.11) 0.61 (0.07) 0.49 (0.05) 0.87 (0.06)
T2-rim area 1.49 (0.12) 0.82 (0.08) 0.37 (0.05) 1.04 (0.07)
T1 area 1.69 (0.15) 1.02 (0.17) 0.32(0.07) 1.25 (0.15)
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In addition, a significant positive correlation was found between the level of diffusivity in T2 lesions and the proportion of the lesion occupied by T1 hypointensity (Fig. 8). Linear Regression analysis indicated that not only > 65% of AD variability, but also more than half of RD variability in T2 lesions can be explained by the relative size of T1 hypointensity and, therefore, is related to the extent (and probably severity) of axonal loss.

Fig. 8.

Fig. 8

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Correlation between lesional diffusivity and proportional size of the T1 hipointense area within T2 lesion: a) RD, b) AD.

4. Discussion

In the current study we examined the directional diffusivity in OR lesions of RRMS patients. Application of novel compartmentalized approach suggested new way of assessing lesional diffusivity which may facilitate the measurement of demyelination/remyelination in deep brain lesions and potentially serve as an outcome measure in therapeutic trials of emerging pro-reparative treatment.

We demonstrated a high correlation of both T2 and T1 OR lesion volume with whole brain lesional load, supporting the use of the OR as a model of pathological activity in MS. We also confirmed our previous observation that the major part of OR DTI variability is associated with lesional damage (Klistorner et al., 2014). However, while RD is more dependent on lesion volume, the variation of AD is probably impacted by other factors, including severity of the structural damage within the lesions and Wallerian degeneration (Klistorner et al., 2015).

Various patterns of abnormal diffusivity in T2 vs T1 lesions have been previously described in MS. In chronic lesions the most common finding was increased water diffusion (ADC or MD) and reduced FA in lesional tissue compared to NAWM; and more marked diffusion alterations were present in T1 hypointense lesions compared to T1 isointense lesions (Filippi et al., 2001; Droogan et al., 1999; Bammer et al., 2000; Scanderbeg et al., 2000; Werring et al., 1999; or see Sbardella et al., (2013) for review).

Since primary identification of the lesions was always based on T2-weighted (and proton density) images, in earlier studies the term “T1 isointense lesion” simply meant “pure” (i.e. not detectable on T1 images) T2 lesions, while the term “T1 hypointense lesion” was used to mark those T2 lesions which contained areas of low signal intensity on T1 non-contrast images. The relative number of reported hypointense T1 lesions varied between studies from as low 10% to as high as 70% of total lesion number (Filippi et al., 2001; Werring et al., 1999; Bammer et al., 2000; Nusbaum et al., 2000). This disagreement can potentially be related to the different definitions of T1 hypointensity. Thus, in some studies the intensity of the lesions on T1 images was compared to the surrounding NAWM (Filippi et al., 2001; Scanderbeg et al., 2000; Werring et al., 1999), while in others-to the grey matter (Nusbaum et al., 2000), while in others several levels of hypointensity were used(Loevner et al., 1995).

Here we used the most frequently reported definition of T1 hypointensity: an area of low T1 signal intensity in comparison to the surrounding NAWM. We demonstrated that the vast majority (86%) of T2 lesions correspond with areas of T1 hypointensity. The high rate of detection of the T1 hypointensity found in our study may partially be explained by the better resolution of our imaging technique, which employed 1 mm slices, compared to 5 mm slice thickness used in earlier studies and use of 3D T1 sequence. In addition, inclusion of slightly hypointense lesions may also increase the rate of T1 lesions reported here.

Segmentation of the T2 lesions was also performed differently from some of the earlier studies. Previously the entire area of T2 lesions displaying abnormal T1 signal was marked and analysed as “T1 hypointense lesion”, even if it was only partially occupied by T1 hypointensity (as was frequently the case) (Loevner et al., 1995; Filippi et al., 2001; Bammer et al., 2000; Werring et al., 1999). In the current study, however, every lesion displaying T1 signal abnormality was segmented (compartmentalized) into two areas: genuine T1 area (which represented a part of T2 lesion seen as hypointense area on T1-weighted images) and “T2-rim” area, which corresponded with the component of the T2 lesion not visible (i.e. isointense) on T1-weighted images. Diffusivity of both areas was analysed separately, an approach that we hypothesized would provide better insight into the nature of diffusivity and its relationship with demyelinating and destructive components of lesional pathology.

In addition, the use of primary eigenvalues (AD and RD) rather than composite indices (ADC, MD or FA) employed in earlier studies, may also be beneficial in understanding mechanisms of diffusivity change(Aung et al., 2014; Klistorner et al., 2015; Schmierer et al., 2007).

Segmentation of T2 lesions demonstrated that the area of T1 hypointensity typically occupies the central area of the lesion, leaving the periphery of the lesion T1-isointense, presumably reflecting distinct pathological changes occurring within the lesion. T2-hyperintensity, while pathologically non-specific, marks the external border of the entire chronically demyelinated area (Schmierer et al., 2009). Conversely, T1 hypointensity is linked to more severe tissue damage caused by axonal loss and resulting widening of the extracellular space (Loevner et al., 1995; Van Waesberghe et al., 1998; van Walderveen et al., 1998; Brex et al., 2000). Therefore, destructive changes are likely to primarily impact MRI appearance and diffusion metrics in the central part of the lesion, masking to a large degree any effect of demyelination. In contrast, relative preservation of axons in the peripheral part of the lesion would render demyelination the key pathological feature. As a consequence, the rim of the lesion, while isointense on T1, remains visible on T2-weighted images.

This hypothesis is supported by OR diffusivity data. While the current study revealed a significant increase of diffusivity (both RD and AD) toward the center of the lesion, RD demonstrated a disproportionally high (in relation to AD) increase between NAWM and T2-rim areas in comparison with the changes observed between T2-rim and T1 areas. This resulted in a reduced rate of FA decline, which changed from 0.12 at the NAWM/T2-rim border to only 0.05 at T2-rim/T1 border.

Since an increase of RD (but not AD) is believed to be related to the degree of demyelination (Schmierer et al., 2007; Klawiter et al., 2011; Song et al., 2005), our results, which show a much larger change of RD compared to AD at the NAWM/T2-rim border, suggest significant loss of myelin in T2-rim area. In contrast, similar changes of RD and AD at T2-rim/T1 border imply more isotropic diffusion in the central (T1) part of the lesion, which is indicative of severe tissue destruction and widening of the extracellular space, in agreement with the previously reported association between lesion T1 hypointensity and the degree of diffusion (Scanderbeg et al., 2000; Werring et al., 1999; Filippi et al., 2000; Rovaris et al., 2005). Differences in directional diffusivity between T2-rim and T1 areas are also reflected in a larger reduction of FA at NAWM/T2-rim compare to T2-rim/T1 borders.

The mostly isotropic diffusion in T1 lesions also reflects a very high correlation between AD and RD within this area, while in T2-rim areas the correlation was less apparent (r2 = 0.63 vs 0.31). This also supports the assumption that a mechanism responsible for the comparable increase of both RD and AD (such as enlargement of an extra-cellular space due to axonal loss) dominates the changes in diffusivity in the T1 area.

Compared to T1 lesions the standard deviation of RD in T2-rim areas was notably smaller, possibly reflecting similar degree of demyelination, whereas the severity of tissue destruction and axonal loss, which is an additional factor contributing to the magnitude of diffusivity in the T1 area, may vary considerably (Kolasinski et al., 2012; van Walderveen et al., 1998; Lassmann et al., 1994).

An alternative explanation for the different patterns of diffusivity in the peripheral vs central part of the lesion could be partial remyelination, which is typically observed at the border of the chronic MS lesion and is likely to be visible on T2-weighted images as a mildly hyperintense area (Nijeholt et al., 2001)(van Walderveen et al., 1998). It is conceivable that more severe loss of myelin in the central area of the lesion contributes to widening of the extracellular space (particularly considering the large relative volume of myelin sheath compare to axonal volume (Mohammadi et al., 2015)). However, based on the notion that an increase in RD is linked to loss of myelin, while change of AD is largely associated with axonal degeneration, severe demyelination alone could not explain the mostly isotropic nature of diffusivity increase toward the center of the lesion.

Regardless of the origin of the expanded extra-cellular space (which may be a combined effect of axonal loss, severe demyelination and post-inflammatory cavitation), increased tissue water content alone will result in an isotropic rise in diffusivity, reducing the potential sensitivity of RD to de/remyelination (Klawiter et al., 2011; Wang et al., 2015; Wang et al., 2011). This is consistent with our finding that isotropic increase of diffusivity in the T1 lesion area significantly contributes to RD of the T2 lesion area.

Another important observation in the current study relates to the width of T2-rim area, which remained constant regardless of the lesion volume. While the significance of this fact remains obscure, it may reflect relative preservation of axons at the lesion edge. Conversely, it may be related to the partial remyelination, which is often limited to the lesion border. We previously demonstrated that the degree of remyelination following optic neuritis, is relatively constant and does not correlate with lesion size (Klistorner et al., 2010). However, since axonal survival and remyelination are intimately linked (preserved axons are needed to initiate remyelination, while remyelinated fibers are better preserved), a combination of both factors may be responsible for the phenomenon observed in this study. The constant T2-rim width may also explain why small T2 lesions with a linear, elongated shape do not always display associated T1 hypointensity; T2-rim areas from opposite sides of the lesion may simply fill its entire cross-section. This is supported by the observation that conversion of T1 acute lesions to chronic black holes is size-dependent (Van Waesberghe et al., 1998).

In conclusion, our results demonstrate different patterns of diffusivity between central and peripheral parts of the MS lesions, and we propose that axonal loss dominates the former, while the effect of de/remyelination may potentially be better detected in the latter. Identifying and separating those patterns has important implications for clinical trials of both neuroprotective and, in particular, remyelinating agents. Disentangling the effects of multiple mechanisms may considerably increase the sensitivity of MRI techniques to monitor subtle changes of axonal loss and myelination.

a) axial view, b) coronal view, c) sagittal view.

Red-T1 hypointense area, Green-“T2-rim” area, Blue-corresponding NAWM area. T1 volume 122 mm2, T2-rim volume 143 mm2, contralateral ROI volume-220 mm2. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)

Footnotes

☆

Supported by grant from National Multiple Sclerosis Society (NMSS), Novartis Save Neuron Grant, Sydney Eye Hospital foundation grant YY is National Health and Medical Research Council (NHMRC) fellow.

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