Application of the Technique of Magnetic Resonance Voxel-based

Morphometry in Patients with Multiple Sclerosis before and after

using High-dose Immunosuppressive Therapy with Autologous

Hematopoietic Stem Cell Transplantation: Preliminary Results

E. A. Gorbunova

, G. N. Bisaga

, A. G. Trufanov

, M. P. Topuzova

, A. V. Fokin

, A. G. Levchuk

A. A. Medenikov

, E. D. Vyshedkevich

, T. A. Bukkieva

, A. Yu.

Efimtsev

and I. A. Mashchenko

MRI Department, Almazov National Medical Research Centre, Akkuratova str., 2, Saint-Petersburg, Russian Federation

Neurology Department, Almazov National Medical Research Centre, Akkuratova str., 2, Saint-Petersburg,

Russian Federation

Neurology Department, Military Medical Academy n.a. S. M. Kirov, Lebedeva str., 6, Saint-Petersburg,

Russian Federation

Keywords: Multiple Sclerosis, Morphometry, Voxel-based Morphometry, Autologous Haematopoietic Stem Cell

Transplantation.

Abstract: Correlation analysis of magnetic resonance voxel-based morphometric parameters of patients with multiple

sclerosis before and after high-dose immunosuppressive therapy with transplantation of autologous

hematopoietic stem cells (HDIT + AHSCT). The use of MR-morphometry methods makes it possible to

quantitatively and objectively changes of the volume and size of the structures of the brain and cerebellum in

patients with MS. We observed 10 patients with MS (3 men, 7 women) who underwent HDIT + AHSCT, and

MRI studies were performed before and after transplantation. After postprocessing of MRI data and statistic

analyses of morphometry parameters some changes were accrued: some patients showed negative dynamics

in white and grey mater volumes and decrease of the absolute volume of the thalamus. Also, study showed

positive dynamics in reducing the number of new MS lesions. These results can be associated with the result

of treatment, with a local decrease in edema and inflammation. Study showed the need for dynamic MR

control of the grey and white matter of the brain and subcortical structures with the help of MR morphometry.

ABBREVIATIONS

MS – multiple sclerosis

VBM – voxel-based morphometry

GM – grey matter

WM – white matter

HDIT – high-dose immunosuppressive therapy

AHSCT – autologous stem cell transplantation.

1 INTRODUCTION

Multiple sclerosis (MS) is an autoimmune disease of

the central nervous system characterized by frequent

episodes of inflammation with demyelination and

neurodegeneration (Lipp, 2018). MS usually begins

at a young age (20–40 years) and is more common in

women. (Lublin, 2014). MS is characterized by an

immune attack on the myelin surrounding the axons

of neurons. This inflammation can damage the axons.

Evidence of neurodegeneration that extends beyond

inflammatory foci is the primary neurodegenerative

component of MS followed by secondary

inflammation (Lipp, 2018; Trapp, 2008; Stys 2012).

Traditionally lesions in multiple sclerosis are the

focus of diagnosis, prognosis and evaluation for

treatment. More recently, studies in multiple sclerosis

have focused on abnormalities in a brain and brain

volume loss, not only as predictors, but also as a result

of clinical trials evaluating the effectiveness of the

treatment (Lipp, 2018).

According to statistics, the prevalence of MS is

50–100 cases per 100 thousand population. There are

more than 2 million patients with multiple sclerosis in

the world. In addition, the average age of patients

Gorbunova, E., Bisaga, G., Trufanov, A., Topuzova, M., Fokin, A., Levchuk, A., Medenikov, A., Vyshedkevich, E., Bukkieva, T., Eﬁmtsev, A. and Mashchenko, I.

Application of the Technique of Magnetic Resonance Voxel-based Morphometry in Patients with Multiple Sclerosis before and after using High-dose Immunosuppressive Therapy with

Autologous Hematopoietic Stem Cell Transplantation: Preliminary Results.

DOI: 10.5220/0010389902430248

In Proceedings of the 14th International Joint Conference on Biomedical Engineering Systems and Technologies (BIOSTEC 2021) - Volume 1: BIODEVICES, pages 243-248

ISBN: 978-989-758-490-9

243

with this disease has dropped significantly and the

disease usually affects the working-age population.

There are different types of MS: a type with

repeated exacerbations and periods of remission, the

so-called "relapsing-remitting type", which can later

develop into a "secondary progressive type" or may

also have persistent progression from the onset of the

disease with no relapses or remissions - "primary

progressive type" (Silva, 2018; Lublin, 2014).

MS is characterized by a progressive course.

Current disease-modifying therapy inevitably leads to

disability, loss of the ability to provide self-care and

loss of cognitive functions.

Today, different neuroimaging techniques such as

magnetic resonance spectroscopy - quantitative

determination of metabolites in tissues, diffuse tensor

MRI (DTI) (Ontaneda, 2017; Das, 2019), magnetic

resonance morphometry have the potential to reveal

the pathogenic mechanisms of the disease and

determine the markers of the neurodegenerative and

atrophic process in multiple sclerosis (Bogachev,

2014).

The method of high-dose immunosuppressive

therapy with transplantation of autologous

hematopoietic stem cells (HDIT + AHSCT) is

intended to provide specialized medical care to

patients with MS. HDIT + AHSCT is considered by

the American Society for Blood and Bone Marrow

Transplantation (ASBMT) as the “standard”

treatment for MS in the nearest future, because this

course of treatment can possibly regenerate the cells

of immune system repertoire and enhance the

immune tolerance (Cohen 2019; Mancardi, 2017,

Gholamzad, 2018). This method of treatment has a

long-term immunomodulation effect, as a result of

which 85% of patients after AHSCT maintain a stable

remission from 5 years and more. This method is

relatively safe, but unfortunately, it is expensive and

is not included in the standards of care for MS

patients.

Atrophy of the grey (GM) and white matter (WM)

of the brain and subcortical structures is manifested

in the early stages of the disease, and its degree

correlates with physical and cognitive impairment.

The effect of treatment on cerebral atrophy predicts

further disability (Ontaneda, 2017; Righart, 2017;

Ghione, 2018). The annual decrease of brain volume

occurs 4 times more often in patients with

apolipoprotein E-e4 in the genotype, than in patients

without it. However, according to other studies, the

genotype with the presence of apolipoprotein E-e4

does not affect the degree of atrophy (Fernandez,

2015). Initially, the presence of cerebral atrophy in

patients with MS was qualitatively identified like an

expansion of the cerebral ventricles and subarachnoid

spaces and a decrease in the volume of brain matter.

The next step was the automatic quantitative

assessment of brain atrophy using voxel magnetic

resonance morphometry (Krotenkova, 2014).

Magnetic resonance morphometry is an accurate

quantitative technique that allows to study changes in

volumetric parameters of brain structures in different

diseases, including those with MS. In addition, MR-

morphometry can be informative in assessing the

treatment and dynamics of the disease development.

2 PURPOSE

To establish significant changes in the structures of

the brain of the patients with MS as a result HDIT +

AHSCT treatment using magnetic resonance voxel-

based morphometry.

3 MATERIALS AND METHODS

3.1 Study Population

An open, single-center, uncontrolled study of the

results of magnetic resonance voxel-based

morphometry of patients with MS before and after

HDIT + AHSCT (at two time points with an interval

of about 12 months).

The HDIT + AHSCT is conducted in several

stages, which occurred sequentially and included

mobilization and procurement of hematopoietic stem

cells (HSC), cryopreservation of HSC,

immunosuppression state by therapy with doses of

cyclophosphamide and rituximab, and transplantation

of HSC with subsequent support therapy with post-

transplant rehabilitation and assessment of

effectiveness.

The mobilization and procurement of HSC was

carried out in two stages:

1) Granulocyte colony-stimulating factor (G-

CSF) - dose 10 μg / kg / day (4 days)

2) Leukocytapharesis (5th day) when the total

number of leukocytes reaches more than 10x10 ^ 9 /

The number of CD34 + cells in leukopheresis

products should be 2-4x106 / kg of body weight.

Cryopreservation of the HSC was carried out with

10% dimethyl sulfoxide, followed by its storage at

-180°C in liquid nitrogen until transplantation.

Conditioning was carried out using high-dose

immunosuppressive therapy with cyclophosphamide

(50-200 mg/kg) with rituximab (500 mg/m^2).

NDNSNT 2021 - Special Session on Non-invasive Diagnosis and Neuro-stimulation in Neurorehabilitation Tasks

244

AHSCT was performed after thawing the frozen

graft in a water bath at a temperature of 37°C. The

transplantation of AHSC was carried out through a

central venous catheter.

Support therapy was aimed at symptomatic

treatment and treatment of AHSCT complications.

We observed 10 patients with MS (3 men, 7

women) who underwent HDIT + AHSCT, and MRI

studies were performed in 2 time points: before and

after transplantation. In this study, the small sample

of patients is due to the small number of patients who

underwent transplantation. The average age of the

patients was 41.6 ± 8.9 years.

The inclusion criteria of the patients for HDIT +

AHSCT were:

1. Age 18-65 years; verified diagnosis of MS;

EDSS 1.0-6.5;

2. The presence of confirmed progression of the

disease against the background of standard therapy;

3. Deterioration in EDSS by 1 point or more at

baseline level <5 points, deterioration in EDSS by 0.5

point or more at baseline level> 5 points;

4. The emergence of new (including Gd +) MS

lesions;

5. Absence of severe concomitant pathology; no

treatment with interferon drugs and

immunosuppressants in the last 3 months.

3.2 MR Imaging Protocol

MRI studies were carried out on a high-field magnetic

resonance imager "Siemens Magnetom Symphony"

with a magnetic field induction of 1.5 T using a head

coil.

All patients underwent structural MRI with

obtaining T1 and T2 weighted images and FLAIR

(Fluid attenuated inversion). Pulse sequence data of a

T1-weighted gradient echo (MP-RAGE -

Magnetization Prepared Rapid Acquired Gradient

Echoes) was collected to combine MRI data of

anatomical structures of the brain, (slice thickness -

4.5 mm, number of slices - 29, the number of

repetitions - 120, scan time - 6 minutes). This

sequence has high resolution with 0.8 mm isotropic

voxeles.

3.3 Image Analyses

At present, mathematical models have been

developed, with the help of which we can analyse

“thin” (not visible on MRI tomograms)

morphological changes, including secondary atrophic

changes in MS, which can be quantified and

presented topographically. One of the new and very

promising methods used to quantify GM, WM and

lesion volume is voxel-based morphometry (VBM).

The VBM workflow includes three main

preprocessing steps:

(1) Tissue classification, which is based on

intensity values and mainly serves to segment the

brain into grey matter, white matter, cerebrospinal

fluid after "removal" of the skull bones and other

structures of the head.

(2) Spatial normalization (linear and non-linear) -

needed to provide matching across brain voxels, in

accordance with the individual brain anatomy of each

person.

(3) Spatial smoothing, followed by statistical

analysis. Smoothing, since spatial normalization is

not ideal and small individual differences in the

anatomy of the brain remain. Spatial smoothing takes

into account these residual differences in local

anatomy. Therefore, after smoothing, each voxel is a

sphere, which is similar to the smoothing kernel, or,

in other words, the weighted average of the values

and values of adjacent voxels (Kurth, 2015; Gaser,

2016).

For morphometric evaluation, T1-WI images

were converted from the standard digital imaging and

communication (DICOM) format (.dcm) to the

Neuroimaging Informatics Technology Initiative

(NIfTI) (.nii) format suitable for analysis and post-

processing using the SPM12 - CAT extension.

Volumetric analysis using CAT allows to accurately

assess the structure of the brain and to avoid operator

errors when carrying out "manual" segmentations.

3.4 Data Analysis

Longitudinal evaluation of the results of

neuroimaging studies of each patient individually, as

well as their group totality (analysis of morphometry

volumetric data) were carried out.

Correlations between results of MR-morphometry

in 2 time points (before and after stem cell

transplantation) were established. Statistical

processing of the received data were carried out using

the package Statistica by StatSoft, (Mann- Whitney

test). Differences were considered significant at p

<0.005. Correlations were calculated using

Spearman's test.

3.5 Results

When assessing the dynamics of morphometric

indicators of the same patients at two time points

(before and after HDIT + AHSCT), 70% of patients

showed negative dynamics in white matter atrophy,

Application of the Technique of Magnetic Resonance Voxel-based Morphometry in Patients with Multiple Sclerosis before and after using

High-dose Immunosuppressive Therapy with Autologous Hematopoietic Stem Cell Transplantation: Preliminary Results

245

while other 20% showed positive dynamics

(increased white matter volume), 10% showed no

change). Besides, 70% of patients have had negative

dynamics in grey matter atrophy (other 30% ща

patients showed positive dynamics in grey matter

atrophy).

Average

GM volume

(cm

/%)

Average

WM volume

(cm

/%)

Before HDIT+

AHSCT

735,150

±72,379

487,896

±91,716

After HDIT+

AHSCT

720,432

±54,347

459,019

±90,089

In addition, 62.5% of patients showed positive

dynamics in reducing the number of new MS lesions,

which also can be associated with the result of

treatment, with a local decrease in edema and

inflammation.

a b c

Figure 1: (a - axial, b - coronal, c - sagittal coronal). The

results of MRI morphometry of the lesions of 42 y.o. female

patient before HDIT + AHSCT. Red colour indicates MS

lesions.

a b c

Figure 2: (a - axial, b - coronal, c - sagittal coronal).The

results of MRI morphometry of the lesions of 42 y.o. female

patient after HDIT + AHSCT. Red colour indicates MS

lesions.

In 84% of the cases, there is a decrease of the

absolute volume of the thalamus (in cm ^ 3/%), while

in 50% of the cases the decrease was below the

average age norm. All these patients had a secondary

progressive type of MS, while statistically the

decrease of the volume of thalamus was not

associated with the duration of the disease or EDSS

score.

When discussing central nervous system atrophy

and MRI morphometry data in MS, it should be

mentioned that, in addition to the gradually increasing

loss of brain matter in MS, short-term brain volume

fluctuations may also occur. Inflammation and edema

as a result of the formation of new lesions lead to a

temporary increase of the brain volume, and vice

versa, for example, taking corticosteroids leads to a

short-term decrease of it - pseudoatrophy. The

mechanism of this process is not entirely clear, but it

is believed that this occurs as a result of a decrease in

inflammation in the central nervous system and

associated edema (Krotenkova, 2014). As a

rconsequence of the ongoing treatment of

HDIT+AHSCT and the anticipated results of this

treatment, it is expected that the decrease in the

absolute volumes of GM and WM is associated with

a decrease of edema and inflammation of the brain.

However, further dynamic observation is necessary,

due to the likelihood of an incorrect interpretation of

the results, since a decrease of brain volumes may be

associated with the ongoing process of

neurodegeneration. It is well known, that cerebral

atrophy occurs at all stages of MS, since at the

preclinical stages of the disease, and progresses

throughout the disease at a faster rate than during the

normal aging process (Inglese, 2018; Fox, 2016).

A. Cifelli et al. It was shown that, compared with

the control group, of patients with a secondary

progressive type of the disease, the volume of the

thalamus decreased by 17%, and the transverse size

of the third ventricle doubled, and a clear inverse

relationship between their volumes was revealed. The

above MRI data are confirmed by histological

studies: a decrease of the number of neurons in the

medial posterior thalamic nucleus and a decrease of

the total volume of the thalamus by 22% was revealed

(Cifelli, 2002). It has also been proven that the reduce

of thalamus volume annually increases by -0.71% per

year (Azevedo, 2018).

4 CONCLUSIONS

The use of MR-morphometry methods makes it

possible to quantitatively and objectively detect

changes of the volume and size of the structures of the

brain and cerebellum in patients with MS. The most

significant results were obtained for the amount of

WM atrophy in patients with MS. The changes

identified in our study correspond to the data of some

other studies: the process of neurodegeneration can

NDNSNT 2021 - Special Session on Non-invasive Diagnosis and Neuro-stimulation in Neurorehabilitation Tasks

246

last up to 1-2 years after the start of therapy with HSC.

This indicates the need for dynamic MR control of the

grey and white matter of the brain and subcortical

structures using MR morphometry to assess the

effectiveness of treatment and the patient’s life

prognosis. Besides, these results can be used for

assessing the prediction of the further course of

multiple sclerosis for patients who underwent

HDIT+AHSCT.

CONFLICT OF INTERESTS

The authors declare no conflict of interest

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