Content uploaded by Anne-Marie Van Dam
Author content
All content in this area was uploaded by Anne-Marie Van Dam on Aug 05, 2015
Content may be subject to copyright.
Ann. N.Y. Acad. Sci. ISSN 0077-8923
ANNALS OF THE NEW YORK ACADEMY OF SCIENCES
Issue: Neuroimmunomodulation in Health and Disease
Pathological differences between white and grey matter
multiple sclerosis lesions
Marloes Prins,1Emma Schul,1Jeroen Geurts,1Paul van der Valk,2Benjamin Drukarch,1
and Anne-Marie van Dam1
1Department of Anatomy and Neurosciences, and 2Department of Pathology, VU University Medical Center, Neuroscience
Campus Amsterdam, Amsterdam, The Netherlands
Address for correspondence: Anne-Marie van Dam, Ph.D., VU University Medical Center, Department of Anatomy and
Neurosciences, Van der Boechorststraat 7, 1081 BT Amsterdam, The Netherlands. amw.vandam@vumc.nl
Multiple sclerosis (MS) is a debilitating disease characterized by demyelination of the central nervous system (CNS),
resulting in widespread formation of white matter lesions (WMLs) and grey matter lesions (GMLs). WMLs are
pathologically characterized by the presence of immune cells that infiltrate the CNS, whereas these immune cells are
barely present in GMLs. This striking pathological difference between WMLs and GMLs raises questions about the
underlying mechanism. It is known that infiltrating leukocytes contribute to the generation of WMLs; however, since
GMLs show a paucity of infiltrating immune cells, their importance in GML formation remains to be determined.
Here, we review pathological characteristics of WMLs and GMLs, and suggest some possible explanations for
the observed pathological differences. In our view, cellular and molecular characteristics of WM and GM, and local
differences within WMLs and GMLs (in particular, in glial cell populations and the molecules they express), determine
the pathway to demyelination. Further understanding of GML pathogenesis, considered to contribute to chronic MS,
may have a direct impact on the development of novel therapeutic targets to counteract this progressive neurological
disorder.
Keywords: multiple sclerosis; white matter lesion; grey matter lesion; inflammation
Introduction
Multiple sclerosis (MS) is a chronic inflammatory,
demyelinating disease of the central nervous sys-
tem (CNS) and is the most common cause of non-
traumatic neurological disability in young adults.1
It is pathologically characterized by disruption of
the blood–brain barrier (BBB), an influx of leuko-
cytes into the CNS (leading to a local inflamma-
tory environment), and demyelination resulting in
a loss of conductance velocity within axons.2Ulti-
mately, this leads to a loss of function for relevant
axonal tracts, accounting for the neurological symp-
toms of MS,3which can vary during the course
of the disease from sensory (e.g., disturbed vision
and sensation4–7) and mobility-related symptoms
(e.g., spasticity, ataxia, or tremor8–12), to fatigue13–15
and cognitive dysfunction.16,17 The disease course is
variable among affected subjects, but the majority of
MS patients have a biphasic disease course, starting
with relapsing-remitting MS (RRMS), during which
patients experience alternating episodes of clini-
cal symptoms and complete or incomplete recov-
ery. Subsequently, the disease may transform into
secondary progressive MS (SPMS) characterized by
gradual worsening of the neurological symptoms.1
Traditionally, MS was regarded as a demyelinat-
ing autoimmune disorder resulting from a lack of
discrimination between self-antigens (i.e., myelin)
and foreign antigens and would subsequently lead to
demyelination and death of oligodendrocytes medi-
ated by infiltrating leukocytes.18,19 Recent observa-
tions, however, have challenged this idea because
demyelinated white matter (WM) areas are not
always associated with infiltrating leukocytes in
early MS20,21 and in a subset of MS patients
with oligodendrogliopathy.22,23 Moreover, no
doi: 10.1111/nyas.12841
1
Ann. N.Y. Acad. Sci. xxxx (2015) 1–15 C2015 New York Academy of Sciences.
WM versus GM pathology of multiple sclerosis Prins et al.
specific autoantigen has yet been found, and thus
far only modest success has been made in delay-
ing the progression of disability with established
-interferon therapy, which suppresses the adaptive
and innate autoimmune response.24 These obser-
vations have led to the hypothesis that a mecha-
nism other than, or in addition to, autoimmunity
is involved in the pathogenesis of MS. It has been
proposed that inflammation in MS is a secondary
phenomenon that occurs as a response to primary
oligodendrocyte dysfunction.25,26 In favor of this
hypothesis is the recent observation that pharma-
cological protection of oligodendrocytes is benefi-
cial in the clinical outcome of an experimental MS
model.27 Still, this challenging hypothesis needs fur-
ther supporting research.
Further complicating the understanding of MS
pathogenesis is the variety of lesion types seen
throughout the brains of those afflicted with
MS. While white matter lesions (WMLs) can be
classified on the basis of their immunological
activity,28–30 grey matter lesions (GMLs) are char-
acterized by only minor infiltration of immune
cells.31,32 Whether WMLs and GMLs each repre-
sent a distinct type of pathology with a unique ori-
gin, or sequential stages in the evolution of a single
type of MS, has not yet been resolved. Although
both WMLs and GMLs are characterized by areas of
focal demyelination, their histopathological features
differ. The most prominent difference is the lower
number of infiltrating immune cells in GMLs.31,32
In this review, we summarize current knowledge
on the pathological status of WMLs and GMLs, the
cell types involved, and several underlying factors
possibly explaining the histopathological differences
between these lesions.
White and grey matter lesions in multiple
sclerosis
White matter lesions
The location, size, and shape of WMLs vary among
patients. The anatomical localization of WMLs
can explain certain neurological symptoms, such
as visual dysfunction, which correlate with WMLs
within the optic nerves and tracts within the frontal
lobe.5,33 In addition, lesions within WM tracts
of the brainstem account for impaired auditory
functioning.7Lesions within WM tracts of the
brainstem and cerebellum correlate with bowel and
bladder dysfunction,34 and lesions within the cor-
ticospinal tract34 and sensory tract35 in the spinal
cord are most likely the underlying cause of motor
and sensory deficits, respectively.3,10
Several ways of pathologically classifying WM
lesions have been described. The staging system
defined by Bø and Trapp29,30 distinguishes active,
chronic active, and inactive lesions. Active lesions
are hypercellular lesions characterized by relative
axonal preservation, massive infiltration of lympho-
cytes, major histocompatibility complex (MHC)-
II+cells and/or myelin-laden macrophages that are
evenly distributed throughout the lesion. Chronic
active lesions also present with relative axonal
preservation, but the myelin-laden macrophages
accumulate at the edges of the lesion. Inactive lesions
are hypocellular lesions characterized by a substan-
tial loss of axons and oligodendrocytes, astrogliosis,
and minor infiltration by macrophages/microglia
and lymphocytes.28–30,36 Added to this staging
system is another type of lesion, the preac-
tive lesion, characterized by clusters of MHC-II+
microglia without demyelination and the absence
of lymphocytes.28,37–39
An additional way of characterizing active WMLs
is based on their pathological profiles; four dis-
tinct patterns of demyelination have been described.
Pattern I is characterized by T cell infiltration
and active demyelination with many activated
microglia and myelin-laden macrophages. However,
no immunoglobulin (Ig) and complement deposi-
tion are present. Moreover, the demyelination pro-
cess is characterized by a simultaneous loss of all
myelin proteins from damaged myelin sheaths. Pat-
tern II lesions, the most frequently seen pattern, are
similar to pattern I lesions, but additionally show
Ig and complement deposition. Pattern III lesions
also present with infiltrated inflammatory cells and
microglia and macrophage activation, without Ig
and complement deposition, but there is clear oligo-
dendrocyte apoptosis, with a preferential loss of the
protein myelin-associatedg lycoprotein (MAG). Pat-
tern IV lesions are extremely rare and are associated
with nonapoptotic death of oligodendrocytes in a
small rim of periplaque WM.22,36 Whether these
different patterns represent different subtypes of
WMLs or different stages within the formation of
WMLs is still debated.20,36
Grey matter lesions
Although MS was frequently considered a WM dis-
ease and certain clinical deficits can be attributed
2Ann. N.Y. Acad. Sci. xxxx (2015) 1–15 C2015 New York Academy of Sciences.
Prins et al. WM versus GM pathology of multiple sclerosis
to WMLs in functionally relevant tracts, WM
did not always explain or predict the clini-
cal symptoms and radiological observations in
patients.40 This clinical–radiological paradox has
been largely solved by accumulating evidence from
histopathological41–43 and high-resolution imag-
ing studies44–48 showing that the CNS GM is also
affected in MS patients. Moreover, the presence of
GMLs was associated with clinical disability.45,48
GMLs can occur in various brain regions of
MS patients, such as the cerebral cortex,49–51 deep
grey matter structures such as the thalamus49,51
and hippocampus,52,53 the cerebellum,49,54 and the
GM of the spinal cord.49 The presence of these
GMLs may explain certain cognitive impairments;
for example, lesions in the hippocampus and amyg-
dala are associated with impaired memory,55,56 and
psychiatric problems such as depression correlated
with temporal lobe lesions,57 which occur in a large
number of MS patients early in the disease.17,58–60
Until now three pathological patterns of GMLs
have been described, but only for cortical
demyelination.32 Type I lesions are leukocortical
lesions that include both subcortical WM and cor-
tex. Type II lesions are located within the cortex
without extending to the surface of the brain or to
subcortical WM, and type III lesions are subpial,
extending from the pial surface into the cortex.32
Bø et al. added a fourth category, type IV lesions,
which extend throughout the full width of the
cerebral cortex without affecting the WM.61 These
lesions are not defined by their immunological
activity.
The cells involved in MS pathology
Infiltration of immune cells
The general view has been that MS pathology
starts with the development of acute inflammatory
lesions and damage to the BBB. This idea is
mainly substantiated by the observation that
active lesions are characterized by perivascular
infiltration of leukocytes62–65 and by data derived
from animal models, notably experimental autoim-
mune encephalomyelitis (EAE).66–69 Indeed, when
peripheral monocytes and macrophages were
depleted, clinical symptoms hardly developed in
animals suffering from EAE.70,71 The involvement
of macrophages in the pathogenesis of MS is further
substantiated by the observation that macrophages
are present within WMLs of MS patients. Two
phenotypesofmacrophageshavebeenidentified:
classically activated M1 macrophages and alterna-
tively activated M2 macrophages. M1 macrophages
typically express proinflammatory and cytotoxic
factors that contribute to demyelination and
axonal damage. M2 macrophages, on the other
hand, contribute to a protective environment by
secreting anti-inflammatory and growth factors.72
Interestingly, in human postmortem active and
chronic active WMLs, activated macrophages
were observed, the majority of which displayed
an M1 activation status. However, a substantial
part of the activated macrophages in these WMLs
also expressed M2 characteristics, suggesting an
intermediate activation state of macrophages in MS
lesions.73
In addition to macrophages, different subsets of
CD4+and CD8+T cells and T helper (TH)1 and
TH17 cells have been identified in EAE and MS
lesions.74–77 Studies in EAE show that CD4+Tcells
become activated in the periphery before the onset
of clinical symptoms of EAE.78,79 However, accu-
mulating evidence suggests an additional role for
CD8+T cells. This type of T cell has been described
to be cytotoxic, and it has been proposed that lesion
formation in EAE is initiated by CD4+T cells, while
amplification of the immune response and damage
is mediated by CD8+T cells.66,75
T cells enter the CNS by passing the BBB and
glia limitans.80 The BBB is a selective functional
barrier composed of endothelial cells, astrocyte
endfeet, and pericytes. Between adjacent cerebral
endothelial cells, tight junctions are present, of
which the three main family proteins are claudin,
occludin, and junction adhesion molecules.81,82
T cells pass the BBB through a sequence of
interactions with brain endothelial cells, involving
leukocyte rolling, adhesion, activation, arrest, and
eventually transendothelial migration. These com-
plex interactions depend on the interaction between
L-selectin present on T cells and E- and P-selectin
present on inflamed endothelial cells and the expres-
sion of integrins and chemokines.83 During MS,
several factors (e.g., increased presence of inflam-
matory cytokines, such as interleukin (IL)-17 and
IL-2284) can inflict damage to tight junctions. Once
passing the glia limitans, regulated by macrophage-
derived matrix metalloproteinases (MMPs),85 into
the CNS, T cells are reactivated by local and infiltrat-
ing activated antigen presenting cells (APCs), which
3
Ann. N.Y. Acad. Sci. xxxx (2015) 1–15 C2015 New York Academy of Sciences.
WM versus GM pathology of multiple sclerosis Prins et al.
present self-antigens.66,75 Subsequently, inflamma-
tory processes lead to damaged myelin and axons.
T cell infiltration and reactivity may not be the
only crucial step during MS pathogenesis. When
T cell infiltration was prevented by using autologous
hematopoietic stem cell transplantation, demyeli-
nation and axonal damage still occurred in regions
with active macrophages and/or microglia.86 In
addition, cortical GMLs show significantly less infil-
tration of T cells compared to WMLs.31,32 The
paucity of T cells in GMLs questions their impor-
tance in GML formation; thus, although T cell
infiltration appears to play a crucial role in the
pathogenesis of MS, lesion formation can still occur
in the absence of T cells. In addition to T cells, glial
cells are known to be present in MS lesions, suggest-
ing that glial cells also play an important role during
lesion formation.
Activated microglia and astrocytes
Microglia are parenchymal tissue macrophages and
the primary responding cells when the homeostasis
of the brain is challenged by infection or injury.87
Upon activation, microglia undergo a morpholog-
ical transformation from a ramified to an amoe-
boid phenotype. They can either adapt a classically
activated or M1 phenotype or an alternatively acti-
vated or M2 phenotype. M1 microglia are phago-
cytic and produce proinflammatory cytokines, such
as IL-1, IL-6, IL-12, tumor necrosis factor (TNF)-
␣, and cytotoxic substances (e.g., nitric oxide
(NO) and reactive oxygen species (ROS). M2
microglia are also phagocytic, but promote tissue
regeneration by producing growth factors and/or
anti-inflammatory cytokines, such as transforming
growth factor (TGF)-, insulin-like growth factor
(IGF)-1, and IL-10. M2 microglia are involved in
downregulating the production of proinflammatory
cytokines, increasing anti-inflammatory molecules,
and facilitating tissue repair.72,88,89 In addition to
microglia, astrocytes function as immunocompe-
tent cells by secreting neurotrophic and/or neu-
rotoxic factors90,91 or augmenting the immune
response by attracting immune cells from the blood
circulation into the CNS.92,93
The exact role of glial cells in the pathogenesis
of MS remains elusive. Studies in EAE animals sug-
gest that their activation occurs in the early phase of
lesion formation, even before infiltration of immune
cells.94–96 In addition, glial cells have been suggested
to be involved in the attraction and migration of
leukocytes toward sites of inflammation. Microglia
and astrocyte signaling contributes to the initiation
and progression of lesion formation and to clinical
symptoms in animal models of MS. Indeed, paralyz-
ing microglia in CD11b-HSVTK transgenic mice97
or preventing microglia signaling98 resulted in a
delay in disease onset and suppression of clinical
signs in EAE. In addition, in a mouse model of
MS using cuprizone-induced demyelination, spe-
cific ablation of astrocytes resulted in a significant
reduction in demyelination in both WM and GM,
which was accompanied by a decrease in the number
of activated microglia present at lesion sites.99
Despite the apparent detrimental role of glial cells
in MS, a beneficial role of these cells has also been
described. M2 microglia and/or macrophages and
astrocytes are crucial for efficient remyelination in
animal models of MS.100–103 In addition, an inter-
mediate microglia phenotype was recently described
in both preactive and remyelinating MS lesions (i.e.,
microglia expressing M1 as well as M2 markers104),
underscoring the dual role of glial cells in MS.
Activated microglia. Activated microglia have
been identified postmortem in WMLs and
GMLs.32,105 In addition, an in vivo study using
positron emission tomography (PET) showed that
activated microglia are present in cortical GM of
MS patients.106 Microglia present in human MS
WMLs were found to express vascular cell adhesion
molecule (VCAM)-1, which binds leukocytes.107
Moreover, activated microglia are associated with
lymphocyte and plasma cell infiltration of the
meninges.108 In addition to their involvement in the
attraction of leukocytes through the BBB, several
lines of evidence point toward a role for microglia
in demyelination and axonal damage. For exam-
ple, microglia are located more so at the border of
the lesion, the site where extensive oligodendrocyte
damage occurs.32 Recently, it was shown that acti-
vated microglia form perivascular clusters at sites
of BBB leakage, with fibrinogen deposition even
before the onset of demyelination in EAE, which
was associated with axonal damage.109 In addition,
an in vitro study showed that the expression of
proinflammatory cytokines, inducible nitric oxide
synthase (iNOS), and ROS by activated microglia
results in a significant increase in damaged myelin
and axons.110 Activation of microglia is not only
4Ann. N.Y. Acad. Sci. xxxx (2015) 1–15 C2015 New York Academy of Sciences.
Prins et al. WM versus GM pathology of multiple sclerosis
associated with tissue damage, but also with
increased clinical disability of MS patients. A PET
study using the peripheral benzodiazepine recep-
tor ligand PK11195 showed a significant increase in
microglial activation in the cortical GM of patients
with RRMS or SPMS, which correlated with clinical
disability.106 Moreover, immunohistochemical data
from postmortem cortical tissue from MS patients
showed a less favorable disease course when cortical
lesions presented with a rim of activated microglia at
their borders.111 In general, less activated microglial
cells are present in GMLs compared to WMLs.
Of interest is a recent study that showed that
MS patients having the HLA-DRB1*1501 genotype
present with greater microglial activation within
cortical lesions than do MS patients without this
genotype,112 suggesting a genotype-specific com-
ponent contributing to microglial activation in, at
least, GMLs.
In contrast to a detrimental clinical outcome asso-
ciated with microglia activation, a protective role
of microglia has also been described (reviewed in
Napoli and Neumann113 ). Microglia in the normal-
appearing WM of MS patients have an alerted but
immunosuppressed phenotype, suggesting a pro-
tective role for microglia in the early phase of lesion
formation.114 During the active phase of demyeli-
nation, amoeboid microglia containing ingested
myelin-derived lipids have been shown to produce
anti-inflammatory cytokines and growth factors
that promote regeneration.115 Moreover, phagocytic
microglia have been observed in MS lesions contain-
ing the triggering receptor expressed on myeloid
cells (TREM)-2. The TREM-2 receptor plays a cru-
cial role in the process of myelin debris clearance.116
Blocking TREM-2 in EAE results in more severe
EAE, reflected by higher clinical scores.117 In addi-
tion, since myelin debris is an inhibitor of remyeli-
nation, removal of myelin debris is essential for
optimal remyelination.118
Activated astrocytes. Not only activated micro-
glia, but also hypertrophic astrocytes and astroglio-
sis have been identified in WMLs, and to a lesser
extent in GMLs, of MS patients.119 In addition
to their ability to provide metabolic support for
neurons, take up and release neurotransmitters,
and maintain BBB function, astrocytes have also
been described as immunocompetent cells. Astro-
cytes express various pattern recognition receptors
(PRRs), such as Toll-like receptors( TLRs), and upon
activation, astrocytes contribute to the immune
response of the CNS, by secreting either proin-
flammatory cytokines or neurotrophic factors, such
as brain-derived neurotrophic factor (BDNF).91 In
addition, astrocytes are able to augment the immune
response by attracting immune cells from the blood
circulation into the CNS, either by increasing BBB
permeability or through the release of chemokines
(e.g., monocyte chemoattractant protein (MCP)-
1, also known as CCL2).92,93 In a later stage of
MS,astrocytesbecomemorehelpfulbypromot-
ing remyelination and tissue repair.103 On the other
hand, scar formation by astrocytes is known to
inhibit remyelination by preventing the migration
of oligodendrocyte precursor cells (OPCs) toward
the lesion.120
To conclude, both activated microglia and astro-
cytes are present in WMLs and, to a lesser extent,
in GMLs. The exact functions of microglia and
astrocytes depend on the timing of their activa-
tion and the subsequent production of pro- or
anti-inflammatory factors, which can either cre-
ate conditions that can be detrimental and lead to
demyelination or that promote remyelination and
prevent axonal loss.
Pathological cellular differences between
WMLs and GMLs
Although studies on biopsy material showed infil-
trated leukocytes within early GMLs,42,121 several
histopathological postmortem studies demonstra-
ted far fewer infiltrated leukocytes within GMLs
than in WMLs. Immunohistochemical analyses
showed that CD3+T cells and CD68+microglia
and/or macrophages in WMLs significantly out-
numbered the cells observed in cortical GMLs of
MS patients. This observation is best illustrated in
leukocortical lesions encompassing WM and GM
(type I lesions) where WM areas encompass higher
levels of T cells and microglia/macrophages and
GM areas are almost devoid of these inflamma-
tory cells.32 In addition, the WM part of type I
lesions presented with a significantly higher number
of LN3+and CD68+cells compared to the GM part
of the same type I lesion,122 which was also observed
in hippocampal lesions with respect to numbers
of LN3+cells (Fig. 1).123 Moreover, numbers of
CD3+T cells were significantly lower in intracor-
tical lesions compared to type I lesions.31 Similarly,
5
Ann. N.Y. Acad. Sci. xxxx (2015) 1–15 C2015 New York Academy of Sciences.
WM versus GM pathology of multiple sclerosis Prins et al.
Figure 1. MHC-II+cells in a mixed hippocampal WM/GM lesion. MS lesions are recognized by the loss of myelin basic protein
(MBP) immunoreactivity in WM and GM areas. (A) Loss of MBP immunoreactivity in a hippocampal lesion of an MS patient. Some
MBP immunoreactivity is still present in the non-lesioned area (lower left corner). (B) A higher magnification shows the absence of
MBP immunoreactivity in the WM and GM lesion area. (C) The WM part of the lesion is characterized by the substantial presence
of MHC-II+cells, while in the GM part of the lesion, lower levels of MHC-II+cells can be found. (D) At a higher magnification,
it becomes clear that the MHC-II+cells in the WMLs have mostly an amoeboid shape, whereas in the GMLs they show a more
ramified morphology. The dotted line indicates the border between WMLs and GMLs. The frame in A refers to the areas shown in
B–D. Scale bar (A) =1 mm; scale bar (B, D) =50 m; scale bar (C) =100 m. For detailed description of immunohistological
methods, we refer the reader to Prins et al.123
CD8+,CD4
+, and CD45RO+T cells were most fre-
quently detected in WMLs, less in type I lesions, and
the least in intracortical lesions.31 Interestingly, deep
GM (DGM) lesions are characterized by an inter-
mediate inflammatory phenotype, as the number of
CD3+T cells in this type of lesion was significantly
higher compared to control DGM, although not
as high as in WMLs.124 Similarly, in vivo microglia
activation in DGM (e.g., thalamus) was higher com-
pared to in cortical areas but lower compared to in
WM areas in patients suffering from clinical isolated
syndrome (CIS), a first clinical symptom suggestive
of MS.125
Explanations for pathological cellular
differences between WMLs and GMLs
The underlying mechanisms resulting in the appar-
ent pathological cellular differences between WMLs
and GMLs remain to be explained. Thus far, several
options have come into focus, such as temporal,
local, cellular, and molecular differences in white
compared to grey matter (summarized in Table 1)
that ultimately, however, still result in similar out-
comes (i.e., demyelinating activity and formation of
WMLs and GMLs).
Temporal difference between the presence
of inflammatory cells in WMLs versus GMLs
Although an immunohistochemical study on cor-
tical demyelination found, using autopsy material,
that demyelination in the cerebral cortex was mostly
seen in MS patients in the chronic progressive
phases of the disease and only rarely in patients
in the acute or relapsing phase,41 several studies
have now reported that GMLs are already present
during the early stages of MS, even when WM
6Ann. N.Y. Acad. Sci. xxxx (2015) 1–15 C2015 New York Academy of Sciences.
Prins et al. WM versus GM pathology of multiple sclerosis
Tab l e 1. Pathological characteristics of WMLs and cortical GMLs in postmortem material of MS patients
WML Cortical GML
Cell influx Infiltrating T cells and macrophages
observed by postmortem
immunohistopathological
analysis62–65,73–77
Little or no inflammatory cells present as
observed by postmortem
immunohistopathological
analysis,31,32,123 but observed in biopsy
material42,121
Local characteristics Myelin debris inducing an inflammatory
response122,155
Neuronal dampening of the immune
response by membrane-bound
molecules136,137 and by secreting
neurotransmitters151,152
Tcellsiteofentry Blood–brain barrier22 Meninges,130,131 choroid plexus,135,163 or
subarachnoid space164
Glial cell characteristics:
Number of microglia Higher than in GM156 Lower than in WM156
Inflammatory profile of
microglia
More prone to be proinflammatory;158,159
complement deposition128
Less prone to be proinflammatory;158,159
no complement deposition128
Expression of molecules that
regulate leukocyte migration
High microglial MRP-14161 and astrocytic
CCL-2 expression123,162
Little microglial MRP-14161 and
astrocytic CCL-2 expression123,162
pathology is very limited.48,126 Moreover, GM
demyelination in early active MS was recently
reported using an immunohistochemical approach
with biopsy material.42 This finding is supported
by a case study showing demyelination in biopsy
material of an early MS patient, even before
radiological evidence of WMLs became visible
on magnetic resonance imaging (MRI) scans.121
Although no comparison was done with WMLs in
biopsy material, infiltrating CD3+and CD8+Tcells
were frequently present in this biopsy material of
cortical lesions,42,121 ascanbeobservedinpost-
mortem WMLs. In contrast, immunohistochemical
studies showed a paucity of infiltrating immune
cells in postmortem GM cortical and hippocampal
lesions.31,32,123 This contrast in cellular pathology
of postmortem WMLs and GMLs warrants further
research. However, we cannot exclude the possibil-
ity that lesions found in biopsy material are in an
earlier stage of demyelination compared to lesions
in postmortem tissue and that infiltrating immune
cells are only present during the early stages of
GML formation.
Other T cell entry sites
BBB damage is one of the main characteristics of
WMLs in MS. This, together with passage of the glia
limitans, gives T cells access to the CNS. Thus far,
no BBB leakage has been demonstrated in GMLs127
and no significant complement deposition in GMLs
has been observed.128 Additionally, the diameter of
the blood vessels within lesions was found to be
increased in WMLs, whereas this increase was less
within GMLs.129
Instead of infiltration of immune cells through
the BBB, several studies have suggested a causal
link between meningeal inflammation and corti-
cal GM demyelination.130,131 In addition, a sig-
nificant interdependence between the presence of
activated microglia and meningeal T cell infiltration
was observed, suggesting that microglial activation
in the cortex of MS patients is, at least in part, driven
by the meningeal inflammatory response.108
Although this may hold true for cortical lesions in
MS, this does not necessarily explain the occurrence
of subcortical GMLs in, for example, the hippocam-
pus or thalamus. Another pathway through which
T cells can enter the CNS is via the intrathecally
localized choroid plexus (CP).132,133 Arecentstudy
showed that the epithelial cell layer of the CP is
populated by a distinct population of CD4+T cells,
which differ from those in the blood circulation,
with T cell receptors specific to CNS antigens.134 Of
interest is the observation that during EAE, CCR6+
T cells enter the CNS via the CP epithelial cell layer
by a CCL20-dependent pathway.135
Thus, BBB damage was not detected within
GMLs; however, T cells derived from the meninges
or the CP epithelial cell layer can enter the cere-
brospinal fluid (CSF) and thereby disseminate at the
7
Ann. N.Y. Acad. Sci. xxxx (2015) 1–15 C2015 New York Academy of Sciences.
WM versus GM pathology of multiple sclerosis Prins et al.
pial surface or subventricular regions, respectively.
Nevertheless, DGM structures that are not directly
located next to the meninges or CP-containing ven-
tricles probably lack T cell infiltration via these
routes.
Local differences
Immunosuppressive properties of neurons. In
contrast to WM, which is devoid of neurons but
has numerous oligodendrocytes as well as microglia
and astroglia, GM is mainly composed of neurons
surrounded by GM astrocytes and microglia and
fewer oligodendrocytes. The presence of neurons
might be causing the quick resolution of inflam-
mation in GMLs by creating an anti-inflammatory
environment. Neurons are able to inhibit immune
responses by constitutively expressing a wide array
of membrane-bound molecules, such as CD200 and
CD47, and by releasing factors, such as chemokines
and neurotransmitters.136,137 The CD200 receptor
(CD200R) is expressed on macrophages, microglia,
and T cells. Upon CD200R–CD200 signaling, the
production of proinflammatory cytokines by pri-
mary microglia (e.g., IL-1and IL-6) is decreased,
while the production of the anti-inflammatory
cytokine IL-10 is increased in a microglia–neuron
coculture system. Here, IL-10 protects neurons
from inflammatory damage,138 suggesting that
the expression and activation of the CD200R on
macrophages/microglia by neuronal CD200 is anti-
inflammatory and neuroprotective. In addition,
CD47 expressed by neurons interacts with its recep-
tor signal-regulatory protein-␣(SIRP-␣)present
on microglia and/or macrophages and inhibits
TNF-␣expression and phagocytosis.139,140 How-
ever, CD200141 and CD47142 are also known to be
expressed by oligodendrocytes, exerting the same
immunosuppressive function within WM. Inter-
estingly, CD200 and CD47 expression is signifi-
cantly reduced in and around WMLs,143 suggesting
a reduced immunosuppressive environment within
WMLs. However, whether CD200 and CD47 are
also downregulated within GMLs remains to be
determined. One membrane-bound molecule that
is exclusively expressed by neurons is the neuronal
cell adhesion molecule ICAM-5, which downregu-
lates T cell activation.144 In addition to membrane-
bound molecules, neurons secrete several factors,
such as chemokines and neurotransmitters. Neu-
rons constitutively express CX3CL1, which interacts
with the CX3CR1 receptor present on microglia145
and T cells,146 thereby suppressing the produc-
tion of proinflammatory cytokines and nitric oxide
(NO) and neuronal cell death induced by activated
microglia.147,148
Neurotransmitters. The two major neurotrans-
mitters in the brain, glutamate and gamma-
aminobutyric acid (GABA), are not only functional
neurotransmitters, but also act as immunomod-
ulators. Several glutamate receptors have been
described, including ionotropic and metabotropic
receptors (mGLuRs);149,150 the latter type is clas-
sified into three subgroups. Glutamate exerts dual
actions on immune activity; depending on which
group of mGLuRs is activated, glutamate can
be immunosuppressive or stimulate the release
of proinflammatory cytokines. By the activation
of group III mGLuRs, glutamate attenuates the
neurotoxicity of microglia,151 while activation of
group II mGLuRs can induce TNF-␣expression by
microglia.152 Since TNF-␣, among other proinflam-
matory factors, is involved in the influx of immune
cells through the BBB, glutamate can indirectly
affect this event during MS. Glutamate can exert its
effectsinWMaswellasGM,sinceglutamatecanbe
released within the synaptic cleft, but axonal release
within WM is also possible.153 However, GABA lev-
els are significantly higher within human cortical
GM compared to WM.154 GABA is known for its
immunosuppressive effects; for example, IL-6 and
IL-20p40 release by microglia is attenuated by
GABA,152 suggesting that GABA contributes to a
more immunosuppressive environment within GM
than within WM. Thus, GABA indirectly reduces
immune cell infiltration by inducing immunosup-
pressive factors.
Myelin-induced immune reaction. WM is
mainly composed of axons enwrapped with myelin,
formed by oligodendrocytes. A higher amount of
myelin within WM than GM results in more myelin
debris in WMLs than in GMLs during MS lesion
formation. Thus, the increased presence of myelin
debris could contribute to the significantly higher
numbers of activated immune cells within WMLs.
A histopathological study in cuprizone-induced
demyelination showed that lesions in GM areas with
inherently lower levels of myelin present with less
activated microglia and astrogliosis compared to
GM areas with a higher density of myelin or WM
8Ann. N.Y. Acad. Sci. xxxx (2015) 1–15 C2015 New York Academy of Sciences.
Prins et al. WM versus GM pathology of multiple sclerosis
Figure 2. CCL2 and CCR2 immunoreactivity in WM and GM bordering a hippocampal lesion. (A) WM bordering a lesion shows
CCL2 immunopositive cells and (B) numerous CCR2 immunopositive cells. In contrast, (C) GM bordering the same lesion is devoid
of CCL2-expressing cells, but it does show CCR2 immunoreactivity (D), although to a lesser extent compared to WM. Scale bar
(A–D) =25 m. For detailed description of immunohistochemical methods, we refer the reader to Prins et al.123
areas,122,155 while there is no evidence of a differ-
ence between the total numbers of glial cells in WM
and GM in mice.122 Similar results were observed in
type I leukocortical lesions of MS patients; specif-
ically, the myelin-rich cortical layer showed more
LN3+cellscomparedtoothercorticallayersencom-
passing less myelin.122 Moreover, when WM and
GM areas in the mouse brain were injected with
similar amounts of myelin debris, both areas pre-
sented with a similar number of activated microglia
and astrocytes.122 This suggests that the amount of
myelin debris, which is higher in WM compared to
GM, is a crucial factor for activation of local immune
cells (i.e., glia).
Glial cell differences in white and grey matter
Besides the observed difference in infiltrating T cells
in WMLs compared to GMLs, a difference between
these types of lesions with respect to activated glial
cells was observed as well. This questions whether
glial cells in WM and GM differ. In humans,
microglial cells in WM exceed those in GM,156
although this is the reverse in rodents where more
microglia can be found in GM than in WM.157 In
addition, a constitutively higher number of HLA-
DR+microgliahasbeenfoundinWMthaninGM
postmortem normal human brain tissue.158 More-
over, a recent study showed that WM microglia
showed more age-related phenotypic changes than
GM microglia; specifically, increased expression of
functional markers such as CD68, F4/80, and CD11b
(the -integrin marker of microglia) was shown
specifically in WM of aged mice,159 suggesting a
higher reactivity to disturbed homeostasis within
WM compared to GM. We consider the local differ-
ences in molecules involved in leukocyte attraction
and migration to be of importance. The expression
of the macrophage early activation marker migra-
tion inhibitory factor–related protein-14 (MRP-14),
known to regulate leukocyte migration,160 by
perivascular macrophages and/or microglial cells
was found to be significantly lower in the neocortex
9
Ann. N.Y. Acad. Sci. xxxx (2015) 1–15 C2015 New York Academy of Sciences.
WM versus GM pathology of multiple sclerosis Prins et al.
compared to WM in the marmoset EAE model of
MS.161 Moreover, astrocytic expression of CCL2,
known for its role in the process of attracting
peripheral immune cells, was found to be clearly
expressed in WMLs but barely present in GMLs of
MS patients123 and in the cuprizone mouse model
of MS.162 Of interest is that the receptor for CCL2
(CCR2) was present on microglia in both WMLs and
GMLs (Fig. 2).123 These observations may explain
the pathological difference found between WMLs
and GMLs, in that a factor of importance for attract-
ing immune cells, CCL2, is barely present in GMLs,
resulting in the absence of those cells in GMLs,
in contrast to WMLs where CCL2 is abundantly
expressed in MS.
Conclusion
MS was long considered a WM disease of the CNS,
driven by T cells reactive against myelin antigens,
resulting in demyelination, oligodendrocyte death,
and axonal damage. However, more recent imaging
and immunohistochemical observations recognize
the formation of lesions in GM as an important
pathological and clinically relevant process as well.
In the present review, we focus on the observed
pathological difference between WMLs and GMLs.
The low number of leukocytes in GMLs implies that
either demyelination does not require the presence
of leukocytes or that the pathogenesis of WML for-
mation differs from that of GMLs. Evidence already
points toward a possible difference in the path-
ways underlying demyelination in WM and GM,
such as the finding that there is no, or only a
low, correlation between the extent of demyelina-
tion in the WMLs and GMLs.41,43,51 We prop o s e
that cellular and molecular differences within glial
cell populations in WM and GM, and in particu-
lar the molecules expressed by them, and the local
cellular environment determine the pathway to
demyelination, which in GML formation is largely
irrespective of immune cell infiltration. Future stud-
ies should focus on further understanding the dif-
ferences between WML and GML formation. In our
view, a greater understanding of GML pathogenesis,
which is considered to underlie disability in chronic
MS, may have a direct impact on the development
of novel therapeutic targets to counteract this pro-
gressive neurological disorder.
Conflicts of interest
The authors declare no conflicts of interest.
References
1. Noseworthy, J., C. Lucchinetti, M. Rodriguez, et al. 2000.
Multiple sclerosis. N.Engl.J.Med.372: 1502–1517.
2. Lassmann, H. 2011. Review: the architecture of inflam-
matory demyelinating lesions: implications for studies on
pathogenesis. Neuro pat hol. Appl . Neurobiol. 37: 698–710.
3. Tallantyre, E.C., L. Bø, O. Al-Rawashdeh, et al. 2010.
Clinico-pathological evidence that axonal loss underlies
disability in progressive multiple sclerosis. Mult. Scler. 16:
406–411.
4. McDonald, W.I. & D. Barnes 1992. The ocular manifesta-
tions of multiple sclerosis. 1. Abnormalities of the afferent
visual system. J. Neurol. Neurosurg. Psychiatry 55: 747–752.
5. Balcer, L.J. 2006. Optic Neuritis.N.Engl.J.Med.354: 1273–
1280.
6. Cruccu, G., A. Biasiotta, S. Di Rezze, et al. 2009. Trigeminal
neuralgia and pain related to multiple sclerosis. Pain 143:
186–191.
7. Coelho, A., B. Ceranic, D. Prasher, et al. 2007. Auditory
efferent function is affected in multiple sclerosis. Ear Hear.
28: 593–604.
8. Prosperini, L., A. Kouleridou, N. Petsas, et al. 2011. The
relationship between infratentorial lesions, balance deficit
and accidental falls in multiple sclerosis. J. Neurol. Sci. 304:
55–60.
9. Papadopoulos, A., S. Gatzonis, A. Gouliamos, et al. 1994.
Correlation between spinal cord MRI and clinical features
in patients with demyelinating disease. Neuroradiology 36:
130–133.
10. Zackowski,K.M.,S.A.Smith,D.S.Reich,et al. 2009. Sen-
sorimotor dysfunction in multiple sclerosis and column-
specific magnetization transfer-imaging abnormalities in
the spinal cord. Brain 132: 1200–1209.
11. Kalkers, N.F., R.L.M. Strijers, M.M.S. Jasperse, et al. 2007.
Motor evoked potential: a reliable and objective measure to
document the functional consequences of multiple sclero-
sis? Relation to disability and MRI. Clin. Neurophysiol.118:
1332–1340.
12. Bjartmar, C., G. Kidd, S. M¨
ork, et al. 2000. Neurologicaldis-
ability correlates with spinal cord axonal loss and reduced
N-acetyl aspartate in chronic multiple sclerosis patients.
Ann. Neurol. 48: 893–901.
13. Bakshi, R. 2003. Fatigue associated with multiple sclerosis:
diagnosis, impact and management. Mult. S cler. 9: 219–227.
14. Comi, G., L. Leocani, P. Rossi, et al. 2001. Physiopathology
and treatment of fatigue in multiple sclerosis. J. Neurol.
248: 174–179.
15. Induruwa, I., C.S. Constantinescu & B. Gran 2012. Fatigue
in multiple sclerosis: a brief review. J. Neurol. Sci. 323: 9–15.
16. Chiaravalloti, N.D. & J. DeLuca 2008. Cognitive impair-
ment in multiple sclerosis. Lancet Neurol.7: 1139–1151.
17. Rao,S.,G.Leo,L.Bernardin,et al. 1991. Cognitive dys-
function in multiple sclerosis. I. Frequency, patterns, and
prediction. Neurolog y 41: 685–691.
10 Ann. N.Y. Acad. Sci. xxxx (2015) 1–15 C2015 New York Academy of Sciences.
Prins et al. WM versus GM pathology of multiple sclerosis
18. Amor, S., F. Puentes, D. Baker, et al. 2010. Inflammation in
neurodegenerative diseases. Immunology 129: 154–169.
19. Stadelmann, C., C. Wegner & W. Br¨
uck 2011. Inflamma-
tion, demyelination, and degeneration: recentinsig hts from
MS pathology. Biochim. Biophys. Acta 1812: 275–282.
20. Barnett, M.H. & J.W. Prineas 2004. Relapsing and remitting
multiple sclerosis: pathology of the newly forming lesion.
Ann. Neurol. 55: 458–468.
21. Henderson, A.P., M.H. Barnett, J.D. Parratt, et al. 2009.
Multiple sclerosis: distribution of inflammatory cells in
newly forming lesions. Ann Neurol.66: 739–753.
22. Lucchinetti, C., W. Br¨
uck, J. Parisi, et al. 2000. Hetero-
geneity of multiple sclerosis lesions: implications for the
pathogenesis of demyelination. Ann. Neurol. 47: 707–717.
23. Marik, C., Pa. Felts, J. Bauer, et al. 2007. Lesion genesis in a
subset of patients with multiple sclerosis: a role for innate
immunity? Brain 130: 2800–2815.
24. Dhib-Jalbut, S. & S. Marks 2010. Interferon-beta mecha-
nisms of action in multiple sclerosis. Neurolog y 74(Suppl
1): S17–24.
25. Trapp, B.D. & K.-A. Nave 2008. Multiple sclerosis: an
immune or neurodegenerative disorder? Annu.Rev.Neu-
rosci. 31: 247–269.
26. Stys, P.K., G.W. Zamponi, J. van Minnen, et al. 2012. Will
the real multiple sclerosis please stand up? Nat. Rev. Neu-
rosci. 13: 507–514.
27. Way, S.W., J.R. Podojil, B.L. Clayton, et al. 2015. Pharma-
ceutical integrated stress response enhancement protects
oligodendrocytes and provides a potential multiple sclero-
sis therapeutic. Nat. Commun. 6: 6532.
28. Van der Valk, P.. & C.J. De Groot 2000. Staging of multiple
sclerosis (MS) lesions: pathology of the time frame of MS.
Neuro pat hol. Appl . Neurobiol.26: 2–10.
29. B¨
o, L., S. M¨
ork, Pa. Kong, et al. 1994. Detection of MHC
class II-antigens on macrophages and microglia, but not
on astrocytes and endothelia in active multiple sclerosis
lesions. J. Neuroimmunol. 51: 135–146.
30. Trapp, B.D., J.Peterson, R.M. Ransohoff, et al. 1998. Axonal
transection in the lesions of multiple sclerosis. N. Engl.
J. Med. 338: 278–285.
31. Bø, L., C. Vedeler, H. Nyland, et al. 2003. Intracortical
multiple sclerosis lesions are not associated with increased
lymphocyte infiltration. Mult. Scler. 9: 323–331.
32. Peterson, J.W., L. B¨
o, S. M¨
ork, et al. 2001. Transected neu-
rites, apoptotic neurons, and reduced inflammation in cor-
tical multiple sclerosis lesions. Ann. Neurol. 50: 389–400.
33. Kolbe, S.C., M. Marriott, A. Van Der Walt, et al. 2012.
Diffusion tensor imaging correlates of visual impairment
in multiple sclerosis and chronic optic neuritis. Invest.
Ophthalmol. Vis. Sci. 53: 825–832.
34. Charil, A., A.P. Zijdenbos, J. Taylor, et al. 2003. Statistical
mapping analysis of lesion location and neurological dis-
ability in multiple sclerosis: application to 452 patient data
sets. Neuroimage 19: 532–544.
35. De Groot, V., H. Beckerman, B.M. Uitdehaag, et al. 2009.
Physical and cognitive functioning after 3 years can be
predicted using information from the diagnostic process
in recently diagnosed multiple sclerosis. Arch. Phys. Med.
Rehabil.90: 1478–1488.
36. Popescu, B.F.G. & C.F. Lucchinetti 2012. Pathology of
Demyelinating Diseases. Annu.Rev.Pathol.7: 185–217.
37. Van Waesberghe, J.H., W. Kamphorst, C.J. De Groot, et al.
1999. Axonal loss in multiple sclerosis lesions: magnetic
resonance imaging insights into substrates of disability.
Ann. Neurol. 46: 747–754.
38. Van der Valk, P. & S. Amor 2009. Preactive lesions in mul-
tiple sclerosis. Curr. Opin. Neurol.22: 207–213.
39. Singh, S., I. Metz, S. Amor, et al. 2013. Microglial nodules
in early multiple sclerosis white matter are associated with
degenerating axons. Acta Neuropathol.125: 595–608.
40. Barkhof, F. 2002. The clinico-radiological paradox in
multiple sclerosis revisited. Curr. Opin. Neurol. 15: 239–
245.
41. Kutzelnigg, A., C.F. Lucchinetti, C. Stadelmann, et al. 2005.
Cortical demyelination and diffuse white matter injury in
multiple sclerosis. Brain 128: 2705–2712.
42. Lucchinetti, C.F., B.F.G. Popescu, R.F. Bunyan, et al. 2011.
Inflammatory cortical demyelination in early multiple scle-
rosis. N.Engl.J.Med.365: 2188–2197.
43. B¨
o, L., J.J.G. Geurts, P. van der Valk, et al. 2007. Lack of cor-
relation between cortical demyelination and white matter
pathologic changes in multiple sclerosis. Arch. Ne urol. 64:
76–80.
44. Bagnato, F., J.A. Butman, S. Gupta, et al. 2006. In vivo
detection of cortical plaques by MR. Am. J. Neuroradiol.
27: 2161–2167.
45. Calabrese,M.,M.A.Rocca,M.Atzori,et al. 2009. Cortical
lesions in primary progressive multiple sclerosis A 2-year
longitudinal MR study. Neurolog y 72: 1330–1336.
46. Filippi, M., P. Preziosa, E. Pagani, et al. 2013. Microstruc-
tural magnetic resonance imaging of cortical lesions in mul-
tiple sclerosis. Mult. Sc ler. 19: 418–426.
47. Sethi, V., T.A. Yousry, N. Muhlert, et al. 2012. Improved
detection of cortical MS lesions with phase-sensitive inver-
sion recovery MRI. J.Neurol.Neurosurg.Psychiatry83:877–
882.
48. Calabrese,M.,N.DeStefano,M.Atzori,et al. 2007. Detec-
tion of cortical inflammatory lesions by double inversion
recovery magnetic resonance imaging in patients with mul-
tiple sclerosis. Arch. Neurol. 64: 1416.
49. Gilmore, C.P., I. Donaldson, L. B¨
o, et al. 2009. Regional
variations in the extent and pattern of grey matter demyeli-
nation in multiple sclerosis: a comparison between the
cerebral cortex, cerebellar cortex, deep grey matter nuclei
and the spinal cord. J.Neurol.Neurosurg.Psychiatry80:
182–187.
50. Kutzelnigg, A., C.F. Lucchinetti, C. Stadelmann, et al. 2005.
Cortical demyelination and diffuse white matter injury in
multiple sclerosis. Brain 128: 2705–2712.
51. Vercellino, M., F. Plano, B. Votta, et al. 2005. Grey matter
pathology in multiple sclerosis. J. Neuropathol. Exp. Neurol.
64: 1101–1107.
52. Geurts, J.J., L. Bo, S.D. Roosendaal, et al. 2007. Extensive
hippocampal demyelination in multiple sclerosis. J. Neu-
ropathol. Exp. Neurol. 66: 819–827.
53. Papadopoulos, D., S. Dukes, R. Patel, et al. 2009. Substan-
tial archaeocortical atrophy and neuronal loss in multiple
sclerosis. Brain Pathol.19: 238–253.
11
Ann. N.Y. Acad. Sci. xxxx (2015) 1–15 C2015 New York Academy of Sciences.
WM versus GM pathology of multiple sclerosis Prins et al.
54. Kutzelnigg, A., J.C. Faber-Rod, J. Bauer, et al. 2007.
Widespread demyelination in the cerebellar cortex in mul-
tiple sclerosis. Brain Pathol.17: 38–44.
55. Benedict, R.H.B., D.Ramasamy, F. Munschauer,et al. 2009.
Memory impairment in multiple sclerosis: correlation with
deep grey matter and mesial temporal atrophy. J. Neurol.
Neurosurg. Psychiatry 80: 201–206.
56. Hulst, H.E., M.M. Schoonheim, S.D. Roosendaal, et al.
2012. Functional adaptive changes within the hippocampal
memory system of patients with multiple sclerosis. Hum.
Brain Mapp. 33: 2268–2280.
57. Feinstein,A.,P.Roy,N.Lobaugh,et al. 2004. Structural
brain abnormalities in multiple sclerosis patients with
major depression. Neurolog y 62: 586–590.
58. Feuillet,L.,F.Reuter,B.Audoin,et al. 2007. Early cognitive
impairment in patients with clinically isolated syndrome
suggestive of multiple sclerosis. Mult. Scler. 13: 124–127.
59. Schulz, D., B.Kopp, A. Kunkel, et al. 2006. Cognition in the
early stage of multiple sclerosis. J. Neurol. 253: 1002–1010.
60. Haase, C.G., M. Tinnefeld, M. Lienemann, et al. 2003.
Depression and cognitive impairment in disability-free
early multiple sclerosis. Behav. Neurol. 14: 39–45.
61.Bø,L.,Ca.Vedeler,H.I.Nyland,et al. 2003. Subpial
demyelination in the cerebral cortex of multiple sclerosis
patients. J. Neuropathol. Exp. Neurol. 62: 723–732.
62. Booss,J.,M.M.Esiri,W.W.Tourtellotte,et al. 1983.
Immunohistological analysis of T lymphocyte subsets in
the central nervous system in chronic progressive multiple
sclerosis. J. Neurol. Sci. 62: 219–232.
63. Wucherpfennig, B.K.W., J. Newcombe, H. Li, et al. 1992.
T Cell receptor Valpha-Vbeta repertoire and cytokine gene
expression in active multiple sclerosis lesions. J. Exp. Med.
175: 993–1002.
64. Disanto, G., J.M. Morahan, M.H. Barnett, et al. 2012. The
evidence for a role of B cells in multiple sclerosis. Neurol ogy
78: 823–832.
65. Kaur, G., J. Trowsdale & L. Fugger 2013. Natural killer cells
and their receptors in multiple sclerosis. Brain 136: 2657–
2676.
66. Sospedra, M. & R. Martin 2005. Immunology of multiple
sclerosis. Annu.Rev.Immunol.23: 683–747.
67. Bar-Or, A. & P.J. Darlington 2011. “The immunology of
multiple sclerosis.” In Multiple Sclerosis Therapeutics,4th
ed. J.A. Cohen & R.A. Rudick, Eds.: 20–34. Cambridge:
Cambridge University Press.
68. Fletcher, J.M., S.J. Lalor, C.M. Sweeney, et al. 2010. T
cells in multiple sclerosis and experimental autoimmune
encephalomyelitis. Clin. Exp. Immunol. 162: 1–11.
69. Martin, R., H.F. McFarland & D.E. McFarlin 1992.
Immunological aspects of demyelinating diseases. Annu.
Rev. Immunol. 10: 153–187.
70. Tran, E.H., K. Hoekstra, N. VanRooijen, et al. 1998.
Immune invasion of the central nervous system
parenchyma and experimental allergic encephalomyelitis,
but not leukocyte extravasation from blood, are prevented
in macrophage-depleted mice. J. Immunol.161: 3767–3775.
71. Huitinga, I., N. van Rooijen, C.J. de Groot, et al. 1990.
Suppression of experimental allergic encephalomyelitis in
Lewis rats after elimination of macrophages. J. Exp. Med.
172: 1025–1033.
72. Mosser, D.M. & J.P. Edwards 2008. Exploring the full spec-
trum of macrophage activation. Nat. Rev. Immunol. 8:
958–969.
73. Vogel, D.Y.S., E.J.F. Vereyken, J.E. Glim, et al. 2013.
Macrophages in inflammatory multiple sclerosis lesions
have an intermediate activation status. J. Neuroinflamma-
tion 10: 35.
74. Traugott, U., E.L. Reinherz& C.S. R aine1983. Multiple scle-
rosis. Distribution of T cells, T cell subsets and Ia-positive
macrophages in lesions of different ages. J. Neuroimmunol.
4: 201–221.
75. McFarland, H.F. & R. Martin 2007. Multiple sclerosis: a
complicated picture of autoimmunity. Nat. Immunol. 8:
913–919.
76. Weiss, Ha., J.M. Millward & T. Owens 2007. CD8+Tcells
in inflammatory demyelinating disease. J. Neuroimmunol.
191: 79–85.
77. O’Connor,R.A.,C.T.Prendergast,A.Catherine,etal. 2008.
Cutting edge: Th1 cells facilitate the entry of Th17 cells to
the central nervous system during experimental autoim-
mune encephalomyelitis. J. Immunol. 181: 3750–3754.
78. Furtado, G.C., M.C.G. Marcondes, J. Tsai, et al. 2008. Swift
entry of myelin-specific T lymphocytes into the central ner-
vous system in spontaneous autoimmune encephalomyeli-
tis. J. Immunol. 181: 4648–4655.
79. Murphy, ´
A.C., S.J. Lalor, M.A. Lynch, et al. 2010. Infiltra-
tion of Th1 and Th17 cells and activation of microglia in
the CNS during the course of experimental autoimmune
encephalomyelitis. Brain. Behav. Immun. 24: 641–651.
80. Owens, T., I. Bechmann & B. Engelhardt 2008. Perivas-
cular spaces and the two steps to neuroinflammation.
J. Neuropathol. Exp. Neurol. 67: 1113–1121.
81. Ballabh, P., A. Braun & M. Nedergaard 2004. The blood-
brain barrier: an overview: structure, regulation, and clin-
ical implications. Neurobiol. Di s. 16: 1–13.
82. Zlokovic, B.. V 2008. The blood-brain barrier in health
and chronic neurodegenerative disorders. Neuron 57: 178–
201.
83. Ley,K.,C.Laudanna,M.I.Cybulsky,et al. 2007. Getting
tothesiteofinflammation:theleukocyteadhesioncascade
updated. Nat. Rev. Immunol. 7: 678–689.
84. Kebir, H., K. Kreymborg, I. Ifergan, et al. 2007. Human
TH17 lymphocytes promote blood-brain barrier disrup-
tion and central nervous system inflammation. Nat. Med.
13: 1173–1175.
85. Agrawal, S., P. Anderson, M. Durbeej, et al. 2006. Dystro-
glycan is selectively cleaved at the parenchymal basement
membrane at sites of leukocyte extravasation in experi-
mental autoimmune encephalomyelitis. J. Exp. Med. 203:
1007–1019.
86. Metz, I., C.F. Lucchinetti, H. Openshaw, et al. 2007. Autolo-
gous haematopoietic stem cell transplantation fails to stop
demyelination and neurodegeneration in multiple sclero-
sis. Brain 130: 1254–1262.
87. Nimmerjahn, A., F. Kirchhoff & F. Helmchen 2005. Resting
microglial cells are highly dynamic surveillants of brain
parenchyma in vivo. Science 308: 1318–1318.
88. Aguzzi, A., Ba. Barres & M.L. Bennett 2013. Microglia:
scapegoat, saboteur, or something else? Science 339: 156–
161.
12 Ann. N.Y. Acad. Sci. xxxx (2015) 1–15 C2015 New York Academy of Sciences.
Prins et al. WM versus GM pathology of multiple sclerosis
89. Benarroch, E.E. 2013. Microglia: Multiple roles in surveil-
lance, circuit shaping, and response to injury. Neurolog y
81: 1079–1088.
90. Jack, C., J. Antel & W.B.R. Uck 2007. Contrasting potential
of nitric oxide and peroxynitrite to mediate oligodendro-
cyte injury in multiple sclerosis. Glia 55: 926–934.
91. Stadelmann, C., M. Kerschensteiner, T. Misgeld, et al. 2002.
BDNF and gp145trkB in multiple sclerosis brain lesions:
neuroprotective interactions between immune and neu-
ronal cells? Brain 125: 75–85.
92. Farina, C., F. Aloisi & E. Meinl 2007. Astrocytes are active
players in cerebral innate immunity. Tre nds Im muno l .28:
138–145.
93. Dong, Y. & E.N. Benveniste 2001. Immune function of
astrocytes. Glia 190: 180–190.
94. Ponomarev, E.D., L.P. Shriver, K. Maresz, et al. 2005.
Microglial cell activation and proliferation precedes the
onset of CNS autoimmunity. J. Neurosci. Res. 81: 374–
389.
95. D’Amelio, F.E., M.E. Smith & L.F. Eng 1990. Sequence of
tissue responses in the early stages of experimental aller-
gic encephalomyelitis EAE: immunohistochemical, light
microscopic, and ultrastructural observations in the spinal
cord. Glia 3: 229–240.
96. Morcos, Y., S.M. Lee & M.C. Levin 2003. A role for hyper-
trophic astrocytes and astrocyte precursors in a case of
rapidly progressive multiple sclerosis. Mult. Scler. 9: 332–
341.
97. Heppner, F.L., M. Greter, D. Marino, et al. 2005.
Experimental autoimmune encephalomyelitis repressed by
microglial paralysis. Nat. Med. 11: 146–152.
98. Xiao, Y., J. Jin, M. Chang, et al. 2013. Peli1 promotes
microglia-mediated CNS inflammation by regulating Traf3
degradation. Nat. Me d. 19: 595–602.
99. Skripuletz, T., D. Hackstette, K. Bauer, et al. 2013. Astro-
cytes regulate myelin clearance through recruitment of
microglia during cuprizone-induced demyelination. Brain
136: 147–167.
100. Kotter, M.R., C. Zhao, N. van Rooijen, et al. 2005.
Macrophage-depletion induced impairment of experimen-
tal CNS remyelination is associated with a reduced oligo-
dendrocyte progenitor cell response and altered growth
factor expression. Neurobiol. Dis. 18: 166–175.
101. Voss, E.V., J. ˇ
Skuljec,V.Gudi,et al. 2012. Characterisation
of microglia during de- and remyelination: can they create
a repair promoting environment? Neurobiol. Dis. 45: 519–
528.
102. Rawji, K.S. & V.W. Yong 2013. The benefits and detriments
of macrophages/microglia in models of multiple sclerosis.
Clin.Dev.Immunol.2013: 1–13.
103. Miljkovi´
c, D., G. Timotijevi´
c&M.MostaricaStojkovi
´
c
2011. Astrocytes in the tempest of multiple sclerosis. FEBS
Lett.585: 3781–3788.
104. Peferoen, L.A.N., D.Y.S.Vogel, K. Ummenthum,et al. 2015.
Activation status of human microglia is dependent on
lesion formation stage and remyelination in multiple scle-
rosis. J. Neuropathol. Exp. Neurol. 74: 48–63.
105. Bø, L. 2009. The histopathology of grey matter demyelina-
tion in multiple sclerosis. Acta Neurol. Scand. 120: 51–57.
106. Politis, M., P. Giannetti, P. Su, et al. 2012. Increased
PK11195 PET binding in the cortex of patients with MS
correlates with disability. Neurology 79: 523–530.
107. Peterson,J.W., L. B¨
o, S. M¨
ork, et al. 2002. VCAM-1-positive
microglia target oligodendrocytes at the border of multi-
ple sclerosis lesions. J. Neuropathol. Exp. Neurol. 61: 539–
546.
108. Dal Bianco, A., M. Bradl, J. Frischer, et al. 2008. Multiple
sclerosis and Alzheimer’sdisease. Ann. Neurol. 63: 174–183.
109. Davalos, D., J.K. Ryu, M. Merlini, et al. 2012. Fibrinogen-
induced perivascular microglial clustering is required for
the development of axonal damage in neuroinflammation.
Nat. Commun. 3: 1227.
110. Di Penta, A., B.Moreno, S. Reix, et al. 2013. Oxidative stress
and proinflammatory cytokines contribute to demyelina-
tion and axonal damage in a cerebellar culture model of
neuroinflammation. PLoS One 8: e54722.
111. Kooi, E.-J.,E.M.M. Str ijbis, P. van der Valk, et al. 2012. Het-
erogeneity of cortical lesions in multiple sclerosis: clinical
and pathologic implications. Neurology 79: 1369–1376.
112. Yates, R.L., M.M. Esiri, J. Palace, et al. 2015. The influence
of HLA-DRB1*15 on motor cortical pathology in multiple
sclerosis. Neuropathol. Appl. Neurobio l. 41: 371–384.
113. Napoli, I. & H. Neumann 2010. Protective effects of
microglia in multiple sclerosis. Exp. Neurol. 225: 24–28.
114. Melief, J., K.G. Schuurman, M.D.B. van de Garde, et al.
2013. Microglia in normal appearing white matter of mul-
tiple sclerosis are alerted but immunosuppressed. Glia 61:
1848–1861.
115. Boven, L.A., M. Van Meurs, M. Van Zwam, et al. 2006.
Myelin-laden macrophages are anti-inflammatory, consis-
tent with foam cells in multiple sclerosis. Brain 129: 517–
526.
116. Takahashi, K., C.D.P. Rochford & H. Neumann 2005.
Clearance of apoptotic neurons without inflammation by
microglial triggering receptor expressed on myeloid cells-2.
J. Exp. Med. 201: 647–657.
117. Piccio, L., C. Buonsanti, M. Mariani, et al. 2007. Block-
ade of TREM-2 exacerbates experimental autoimmune
encephalomyelitis. Eur. J. Immunol. 37: 1290–1301.
118. Kotter, M.R., W.-W. Li, C. Zhao, et al. 2006. Myelin impairs
CNS remyelination by inhibiting oligodendrocyte precur-
sor cell differentiation. J. Neurosci. 26: 328–332.
119. Petzold, A., M.J.Eikelenboom, D. Gveric, et al. 2002. Mark-
ers for different glial cell responses in multiple sclerosis:
clinical and pathological correlations. Brain 125: 1462–
1473.
120. Bannerman, P., A. Hahn, A. Soulika, et al. 2007. Astroglio-
sis in EAE spinal cord: derivation from radial glia, and
relationships to oligodendroglia. Glia 55: 57–64.
121. Popescu, B.F.G., R.F. Bunyan, J.E. Parisi, et al. 2011. A case
of multiple sclerosis presenting with inflammatory cortical
demyelination. Neurol ogy 76: 1705–1710.
122. Clarner, T., F. Diederichs, K. Berger, et al. 2012. Myelin
debris regulates inflammatory responses in an experimen-
tal demyelination animal model and multiple sclerosis
lesions. Glia 60: 1468–1480.
123. Prins, M., R. Dutta, B. Baselmans, et al. 2014. Discrep-
ancy in CCL2 and CCR2 expression in white versus grey
13
Ann. N.Y. Acad. Sci. xxxx (2015) 1–15 C2015 New York Academy of Sciences.
WM versus GM pathology of multiple sclerosis Prins et al.
matter hippocampal lesions of multiple sclerosis patients.
Acta Neuropathol. Commun. 2: 98.
124. Haider, L., C. Simeonidou, G. Steinberger, et al. 2014.
Multiple sclerosis deep grey matter: the relation between
demyelination, neurodegeneration, inflammation and
iron. J. Neurol. Neurosurg. Psychiatry 85: 1386–1395.
125. Giannetti, P., M. Politis, P. Su, et al. 2015. Increased
PK11195-PET binding in normal-appearing white matter
in clinically isolated syndrome. Brain 138: 110–119.
126. Calabrese, M., M. Battaglini, A. Giorgio, et al. 2010. Imag-
ing distribution and frequency of cortical lesions in patients
with multiple sclerosis. Neuro log y 75: 1234–1240.
127. Van Horssen, J., B.P. Brink, H.E. de Vries, et al. 2007. The
blood-brain barrier in cortical multiple sclerosis lesions. J.
Neuro pat hol. Exp. Ne urol. 66: 321–328.
128. Brink, B.P., R. Veerhuis, E.C.W. Breij, et al. 2005. The
pathology of multiple sclerosis is location-dependent: no
significant complement activation is detected in purely cor-
tical lesions. J. Neuropathol. Exp. Neurol. 64: 147–155.
129. Pham, H., A.A. Ramp, N. Klonis, et al. 2009. The
astrocytic response in early experimental autoimmune
encephalomyelitis occurs across both the grey and white
matter compartments. J. Neuroimmunol. 208: 30–39.
130. Howell, O.W., Ca. Reeves, R. Nicholas, et al. 2011.
Meningeal inflammation is widespread and linked to cor-
tical pathology in multiple sclerosis. Brain 134: 2755–2771.
131. Choi, S.R., O.W. Howell,D. Carassiti, et al. 2012. Meningeal
inflammation plays a role in the pathology of primary pro-
gressive multiple sclerosis. Brain 135: 2925–2937.
132. Schwartz, M., J. Kipnis, S. Rivest, et al. 2013. How do
immune cells support and shape the brain in health, dis-
ease, and aging? J. Neurosci. 33: 17587–17596.
133. Dragunow, M. 2013. Meningeal and choroid plexus cells—
novel drug targets for CNS disorders. Brain Res.1501: 32–
55.
134. Baruch, K., N. Ron-Harel, H. Gal, et al. 2013. CNS-specific
immunity at the choroid plexus shifts toward destructive
Th2 inflammation in brain aging. Proc. Natl. Acad. Sci.
U.S.A. 110: 2264–2269.
135. Reboldi, A., C. Coisne, D. Baumjohann, et al. 2009. C-C
chemokine receptor 6-regulated entry of TH-17 cells into
the CNS through the choroid plexus is required for the
initiation of EAE. Nat. Immunol. 10: 514–523.
136. Biber, K., H. Neumann, K. Inoue, et al. 2007. Neuronal
“On” and “Off” signals control microglia. Tre nds N eur o s c i.
30: 596–602.
137. Wright, G.J., M. Jones, M.J. Puklavec, et al. 2001. The
unusual distribution of the neuronal/lymphoid cell sur-
face CD200 OX2. glycoprotein is conserved in humans.
Immunology 102: 173–179.
138. Hernang´
omez, M., L. Mestre, F.G. Correa, et al. 2012.
CD200-CD200R1 interaction contributes to neuropro-
tective effects of anandamide on experimentally induced
inflammation. Glia 60: 1437–1450.
139. Smith, R.E., V. Patel, S.D. Seatter, et al. 2003. A novel
MyD-1 SIRP-1alpha. signaling pathway that inhibits LPS-
induced TNFalpha production by monocytes. Blood 102:
2532–2540.
140. Oldenborg, Pa., H.D. Gresham & F.P. Lindberg 2001.
CD47-signal regulatory protein alpha SIRPalpha regulates
Fcgamma and complement receptor-mediated phagocyto-
sis. J. Exp. Med. 193: 855–862.
141. Koning,N., D.F.Swaab, R.M. Hoek, et al. 2009. Distribution
of the immune inhibitory molecules CD200 and CD200R
in the normal central nervous system and multiple sclero-
sis lesions suggests neuron-glia and glia-glia interactions.
J. Neuropathol. Exp. Neurol. 68: 159–167.
142. Gitik, M., S. Liraz-Zaltsman, P.-A. Oldenborg, et al. 2011.
Myelin down-regulates myelin phagocytosis by microglia
and macrophages through interactions between CD47 on
myelin and SIRP␣signal regulatory protein-␣;. on phago-
cytes. J. Neuroinflammation 8: 24.
143. Koning, N., L. B¨
o, R.M. Hoek, et al. 2007. Downregulation
of macrophage inhibitory molecules in multiple sclerosis
lesions. Ann. Neurol. 62: 504–514.
144. Tian, L., J. Lappalainen, M. Autero, et al. 2008. Shedded
neuronal ICAM-5 suppresses T-cell activation. Blood 111:
3615–3625.
145. Harrison, J.K., Y. Jiang, S. Chen, et al. 1998. Role for
neuronally derived fractalkine in mediating interactions
between neurons and CX3CR1-expressing microglia. Proc.
Natl. Acad. Sci. U.S.A. 95: 10896–10901.
146. Smolders, J., E.B.M. Remmerswaal, K.G. Schuurman, et al.
2013. Characteristics of differentiated CD8+. and CD4 +.
Tcellspresentinthehumanbrain.Acta Neuropathol. 126:
525–535.
147. Cardona, A.E., E.P. Pioro, M.E. Sasse, et al. 2006. Control
of microglial neurotoxicity by the fractalkine receptor. Nat.
Neuro sci. 9: 917–924.
148. Mizuno, T., J. Kawanokuchi, K. Numata, et al. 2003. Pro-
duction and neuroprotective functions of fractalkine in the
central nervous system. Brain Res.979: 65–70.
149. Traynelis, S.F., L.P. Wollmuth, C.J. Mcbain, et al. 2010. Glu-
tamate receptor ion channels: structure, regulation, and
function. Pharmacol. Rev. 62: 405–496.
150. Niswender, C.M. & P.J. Conn 2010. Metabotropic glu-
tamate receptors: physiology, pharmacology, and disease.
Annu.Rev.Pharmacol.Toxicol.50: 295–322.
151. Taylor, D.L., L.T. Diemel & J.M. Pocock 2003. Activation of
microglial group iii metabotropic glutamate receptors pro-
tects neurons against microglial neurotoxicity. J. Neurosci.
23: 2150–2160.
152. Pocock, J.M. & H. Kettenmann 2007. Neurotransmitter
receptors on microglia. Tre n ds Ne uro s c i .30: 527–535.
153. Kukley, M., E. Capetillo-Zarate & D. Dietrich 2007. Vesic-
ular glutamate release from axons in white matter. Nat.
Neuro sci. 10: 311–320.
154. Jensen, J.E.,B.D.B. Frederick & P.F. Renshaw 2005. Grey and
white matter GABA level differences in the human brain
using two-dimensional, J-resolved spectroscopic imaging.
NMR Biomed.18: 570–576.
155. Gudi, V., D. Moharregh-Khiabani, T. Skripuletz, et al. 2009.
Regional differences between grey and white matter in cup-
rizone induced demyelination. Brain Res.1283: 127–138.
156. Mittelbronn, M., K. Dietz, H.J. Schluesener, et al. 2001.
Local distribution of microglia in the normal adult human
14 Ann. N.Y. Acad. Sci. xxxx (2015) 1–15 C2015 New York Academy of Sciences.
Prins et al. WM versus GM pathology of multiple sclerosis
central nervous system differs by up to one order of mag-
nitude. Acta Neuropathol.101: 249–255.
157. Lawson, L.J., V.H. Perry, P. Dri, et al. 1990. Hetero-
geneity in the distribution and morphology of microglia
in the normal adult mouse brain. Neurosc ience 39:
151–170.
158. Gehrmann, J., R.B. Banati & G.W. Kreutzberg 1993.
Microglia in the immune surveillance of the brain: human
microglia constitutively express HLA-DR molecules.
J. Neuroimmunol. 48: 189–198.
159. Hart, A.D., A. Wyttenbach, V.H. Perry, et al. 2012. Age
related changes in microglial phenotype vary between CNS
regions: grey versus white matter differences. Brain. Behav.
Immun. 26: 754–765.
160. Nacken, W., J. Roth, C. Sorg, et al. 2003. S100A9/S100A8:
Myeloid representatives of the S100 protein family as
prominent players in innate immunity. Microsc. Res. Tech.
60: 569–580.
161. Merkler, D., R. B¨
oscke, B. Schmelting, et al. 2006. Differ-
ential macrophage / microglia activation in neocortical eae
lesions in the marmoset monkey. Brain Pathol.16: 117–
123.
162. Buschmann, J.P., K. Berger, H. Awad, et al. 2012. Inflam-
matory response and chemokine expression in the white
matter corpus callosum and gray matter cortex region dur-
ing cuprizone-induced demyelination. J. Mol. Neurosci. 48:
66–76.
163. Murugesan, N., D. Paul, Y. Lemire, et al. 2012. Activeinduc-
tion of experimental autoimmune encephalomyelitis by
MOG35-55 peptide immunization is associated with differ-
ential responses in separate compartments of the choroid
plexus. Fluids Barriers CNS 9: 15.
164. Howell,O.W.,E.K. Schulz-Trieglaff, D. Carassiti, et al.2015.
Extensive grey matter pathology in the cerebellum in multi-
ple sclerosis is linked to inflammation in the subarachnoid
space. Neuropathol. Appl. Neurobiol. 1–16.
15
Ann. N.Y. Acad. Sci. xxxx (2015) 1–15 C2015 New York Academy of Sciences.
