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Do oligodendrocytes mediate iron regulation in the human brain.

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the adult human brain has not been investigated to our
knowledge. Ferritin is a 24 subunit protein that forms
a protomer of relative molecular weight 480,000. Its
role as an iron storage protein has been established [7],
but its presence and function in the CNS have received little attention to date.
Do Oligodendrocytes
Mediate Iron Regulation
in the Human Brain?
Megan R. Gerber, BA, and James R. Connor, PhD
We used immunohistochemical studies to demonstrate
that transferrin (the iron mobilization protein) and ferritin (the iron storage protein) are specifically localized in
oligodendrocytes in gray and white matter of the human central nervous system. In addition, iron is also
localized predominantly in oligodendrocytes. Oligodendrocytes have been well established as the cells responsible for myelin production in the central nervous system.
The results of this study suggest that oligodendrocytes
(or a subpopulation of oligodendrocytes) might have the
additional function of mediating iron mobilization and
storage in the central nervous system.
Gerber MR, Connor JR. Do oligodendrocytes
mediate iron regulation in the human brain? Ann
Neurol 1989;26:95-98
The importance of iron for normal functioning of the
central nervous system (CNS) is well established 111.
A number of neurological diseases are thought to be
associated with disruptions in iron metabolism; these
include Alzheimer's, Hallervorden-Spatz, Pick's, Huntington's chorea, and tardive dyskinesia 121. Changes
in iron levels in the brain have been demonstrated to
be associated with behavioral and mood disorders 121.
Dietary deficiencies in iron during development result
in motor and behavioral dysfunctions that persist into
adulthood 111. Yet, even in adults with severe anemia,
iron content in the brain is relatively unaffected. This
latter observation suggests that there is local storage of
CNS iron, which is readily mobilized.
Our laboratory is investigating iron regulation in the
CNS by focusing on transferrin and ferritin. Transferrin is an 80,000 molecular weight glycoprotein that is
the major iron-binding and transport protein in vertebrates. Although some controversy exists regarding
the cellular distribution of transferrin during development, it is now generally accepted that transferrin persists in the CNS in oligodendrocytes in adult chickens
[3] and rats 14-61. The distribution of transferrin in
From the Department of Anatomy and Center for Neuroscience,
Milton S. Hershey Medical Center, Pennsylvania State University,
Hershey, PA.
Received Oct 7, 1988, and in revised form Dec 9. Accepted for
publication Dec 11, 1988.
Address correspondence to Dr Connor, Department of Anatomy,
Milton S . Hershey Medical Center, Pennsylvania State Univetsity,
Hershey, PA 17033.
Materials and Methods
Autopsy specimens were obtained from the brains of 7 adult
patients in whom neuropathological changes were absent.
Postmortem time prior to fixation ranged from 5% to 16%
hours. The following brain regions were chosen for analysis:
cerebellum, motor cortex (precentrzl gyrus), superior temporal gyrus, superior frontal gyrus, and occipital cortex
(cuneus). These areas were removed and immersion-fixed in
formalin-ethanol-acetic(FEA) acid [SJ or 10% neutral buffered formalin (NBF) for 4 to 22 hours and then embedded
in paraffin and cut on a rotary microtome at thicknesses of 8
pm. The tissue was mounted on a slide, rehydrated, and
processed for imrnunohistochemical study. Prior to incubation in the primary antibody, the slides were treated with
0.3% hydrogen peroxide, 1% dimethyl sulfoxide, and
blocked in either goat (for ferritin) or rabbit (for transferrin)
serum (Sigma, St Louis, MO; 1:30). In a cold room with the
temperature at 4°C the sections were incubated overnight in
antiserum to human transferrin (Cappel, Malvern, PA,
1:500) or human ferritin (ICN Immunobiologicals, Costa
Mesa, CA; 1:500, 1.250). Control sections were incubated
overnight in rabbit (for ferritin) or goat (for transferrin)
serum instead of primary antibody. On the following day,
the sections were developed according to the peroxidaseantiperoxidase method of Sternberger 191. Each step was
separated by two washes in phosphate-buffered saline solution (PBS; 0.1 M) of 10 minutes each. The reaction product
was visualized using 3 '3'-diaminobenzidine (DAB) as the
chromogen. Finally, the tissue was osmicated (0.2% osmium
tetraoxide) for 2 minutes.
Ferric iron (Fe3+)localization was analyzed using Perk'
histochemical procedure [lo] intensified by DAB {I 11.
Some tissue used for iron localization was cut at thicknesses
of 50 pm using a vibratome. The histochemical staining procedure for iron was identical regardless of tissue preparation.
The results were not affected by furation time or the
postmortem period. The cell type subject to immunostaining was not influenced by the type of fixative;
however, the FEA fix was found to provide a more
intense reaction. Oligodendrocytic immunostaining
was not seen in the control sections.
The predominant cell type that exhibits immunoreactivity to transferrin antiserum is a small, round cell
that is often found in pairs or clusters of 3 or 4 cells in
the gray matter (Fig 1). The reaction product is localized in a cap-like fashion in the soma and usually
does not extend into the processes. The nuclei, which
are generally eccentric, do not contain reaction product. In the white matter, immunoreactive cells are es-
Copyright 0 1989 by the American Neurological Association 95
Fig 1. A paraffin-embeddedsection from the gray matter of the
motor cortex (precentralgyrusj of an adult human, which has
been imnaunoreactedwith antiserum t o transfewin. The cells containing transfew;n are generally round and the reaction product
is mostly confined to the soma. The immunoreactive cells occasionally occur in pairs (arrow). A perivascular immunoreactive cell
is visible (arrowhead). (x 1,000.)
Fig 3. Ferritin-containing celb in the gray matter of motor cortex. The immunoreactive celb are small and round, are frequentlyfound in a perineuronalposition, occasionally occur in
pairs (arrow), and are also seen along blood vessels (clear arrow).
( x 850.)
Fig 4. In the white matter of the motor cortex, cells containing
ferritin are round and do not have immunoreactive proceues (arrow). ( X 900.)
Fig 2. Transfewin-containingcells (arrow) in the white matter
from the motor cortex of an adult human, shown in a parafinembedded section. Almost none of the transferrin-containingcells
have immunoreactiveprocesses. ( x 900.)
sentially identical in morphology to their gray matter
counterparts (Fig 2). They are round, contain reaction
product only in the soma, and possess a nonimmunoreactive eccentric nucleus.
The cells chat exhibited positive immunostaining for
ferritin were similar morphologically to those that
stained for transferrin. The cells containing ferritin,
found in both gray and white matter, are small, round
cells with a cap-like reaction product. Most of the ferritin-positive cells in the gray matter were associated
96 Annals of Neurology Vol 26 No 1 July 1787
either with blood vessels (perivascular) or with neurons (perineuronal) (Fig 3). The reaction product is restricted to the soma, in which the generally eccentric
nuclei remain unreacted (Fig 4). Finally, the cells staining subsequent to the Perls’ reaction for ferric iron
possessed a morphology similar to those exhibiting immunoreactivity to the transferrin and ferritin antisera
(Figs 5 and 6).
The results of this study demonstrate that transferrin
and ferritin are present in the white and gray matter of
the human CNS. The cells that were labeled in each of
the three separate reactions (immunohistochemistry
for transferrin and for ferritin, and the Perls’ reaction)
were small and round with eccentric nuclei that are
characteristic of oligodendrocytes C12). The appearance of the transferrin immunostaining is consistent
with previous descriptions of oligodendrocytic transferrin staining in the rat brain 15, 6) and of histochemical staining in the oligodendrocyte for iron 1131. The
localization of transferrin, ferritin, and iron in oligodendrocytes suggests that these cells might be the mediators of iron metabolism in the CNS. In further
support of this notion, it has been previously demonstrated that carbonic anhydrase, the enzyme that
catalyzes the conversion of carbon dioxide and water
to bicarbonate ion, is found in oligodendrocytes and
indeed might be a specific marker for this cell 114, 151.
Bicarbonate ion is required for the binding of iron to
transferrin 116). Thus, all of the compounds necessary
for iron storage and mobilization are found in oligodendrocytes.
Fig 5 . The Pwls’ reaction for iron (intensijiedby 3’-3’-diaminobenzidine) on a vtbratome section from the human cerebral cortex
wveals that most of the stained celh are round with eccentric
nuclei (dark arrow) and occasionally occur in clusters of three or
more (arrowhead). The iron-containingcells are also found in a
permeuronal position (clear arrow). ( X 1,000.)
The importance of iron at the level of cell function is
that iron is necessary for oxidative metabolism; it is an
essential component of enzyme systems in the Krebs
cycle and cytochrome oxidase system. It is also a cofactor in the synthesis of the monoamines, serotonin,
dopamine, and norepinephrine, as well as in the function of dopamine receptors 1171. The importance of
iron in CNS function is perhaps underscored by reports that iron levels in the globus pallidus and putamen exceed even hepatic iron levels 1171.
The role of transferrin in iron mobilization in the
CNS has recently been elucidated by the demonstration that neurons in culture receive iron via transferrin
118-203 and that iron enters the brain by way of a
transferrin receptor-mediated process in both rat and
human [21,22). Whether transferrin in the oligodendrocyte is endogenous or derived from the plasma is
unknown, but the messenger RNA for transferrin is
present in the brain, specifically in oligodendrocytes
Fig 6. Iron staining in the white matter ofa paraffin-embedded
section reveals celh that are round and have no visible processes,
and the reaction product is confined to one pole athe perikaryal
cytoplasm (arrow). The background staining ofthe white matter
following a histochemical reaction for iron has a patchy appearance where some areas are stained more intensely than others.
The significance of the patchy reaction is unclear but the pattern
is consistent. ( x 900.)
The possibility that oligodendrocytes are involved in
mediating iron levels in the CNS represents, to our
knowledge, a novel concept in neurobiology. There
are three subpopulations of oligodendrocytes: interfascicular (white matter), perineuronal satellite, and perivascular oligodendrocytes. It has been reported that
oligodendrocytes that produce myelin are supporting
three times their weight in myelin membrane 1241.
Clearly the iron requirement for these cells must be
great. It has also been found, however, that perineuronal oligodendrocytes have oxidative metabolism that is
an order of magnitude higher than neighboring neurons 1251, and these cells do not normally make myelin. The function of perivascular and perineuronal
oligodendrocytes is not clear, but it has long been suspected that there is a trophic interaction between glia
and neurons. Given the present findings, oligodendrocytes (particularly satellite cells) in the gray matter may
be the mediators of iron availability for neurons.
Brief Communication: Gerber and Connor: Oligodendrocytes and Iron Regulation
Supported by United States Public Health Service grants HL07477
(to M. R. G.), NS22671 (J. R. C.) and funds provided by Alzheimer’s Disease Research, a program of the American Health Assistance Foundation, Rockville, MD 0. R. C.).
We thank Dr J. Towfighi for his assistance in obtaining autopsy
material for this study, D. Hinton and S. Menzies for their expert
technical support, and T. Segneri who provided secretarial assistance.
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Tomaculous Neuropathy
Presenting as Acute
Recurrent Polyneuropathy
Juan L. Joy, MD, and Shin J. Oh, MD
Tomaculous neuropathy (TN) is classically associated
with the inherited, recurrent focal neuropathies. We report a case of T N manifesting as an acute recurrent
polyneuropathy. A 28-year-old woman had 2 episodes
of acute, ascending, symmetrical sensorimotor deficit.
Teased nerve-fiber preparation confirmed the presence
of TN. Extensive investigations failed to reveal other
cause for her symptoms. We believe that this case is
unique and broadens the clinical spectrum of TN.
Joy JL, Oh SJ. Tomaculous neuropathy presenting
as acute recurrent polyneuropathy.
Ann Neurol 1989;26:98-100
The term tomaculons nenropatby (TN), from the Latin
word tomacnlum, meaning “sausage,” was coined in
1975 by Madrid and Bradley { 11 to describe focal sausage-shaped swellings of myelin sheaths. This histopathological finding has been classically associated with
the inherited, recurrent focal neuropathies: hereditary
neuropathy with liability to pressure palsies (HNPP)
From the Neurology Department, University of Alabama at Birmingham, Birmingham, AL.
Received Apr 12, 1988,and in revised form Nov 14, 1988,and Jan
11, 1989.Accepted for publication Jan 18, 1989.
Address correspondence to Dr Joy, Neurology Department, University of Alabama at Birmingham, UAB Station, Birmingham, AL
98 Copyright 0 1989 by the American Neurological Association
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oligodendrocytes, iron, brain, regulation, human, mediated
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