| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
ARTICLES |
-Induced Injury1
Department of Pediatrics, CB 7220, The University of North Carolina at Chapel Hill, Chapel Hill, North Carolina 27599-7220
Address all correspondence and requests for reprints to: Dr. A. J. D’Ercole, Department of Pediatrics, CB 7220, The University of North Carolina at Chapel Hill, Chapel Hill, North Carolina 27599-7220.
 |
Abstract |
|---|
 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
(TNF-) has been causally implicated in
several demyelinating disorders, including multiple sclerosis. Because
insulin-like growth factor I (IGF-I) is a potent stimulator of
myelination, we investigated whether it can protect oligodendrocytes
and myelination from TNF--induced damage using mouse glial cultures
as a model. Compared with controls, TNF- decreased oligodendrocyte
number by approximately 40% and doubled the number of apoptotic
oligodendrocytes and their precursors. Addition of
Boc-aspartyl(Ome)-fluoromethyl ketone (BAF), an inhibitor of
interleukin-1ß converting enzyme (ICE)/caspase proteases, blocked
TNF--induced reductions in oligodendrocytes, indicating that the
TNF--induced reduction in oligodendrocytes is, at least in part, due
to apoptosis, and that ICE/caspases are one of TNF- action
mediators. Simultaneous addition of IGF-I to TNF--treated cultures
negated these TNF- effects nearly completely. Furthermore, IGF-I
promoted oligodendrocyte precursor proliferation and/or differentiation
in TNF--treated cultures. To analyze TNF- and IGF-I actions on
oligodendrocyte function, we measured the abundance of messenger RNAs
(mRNAs) for two major myelin-specific proteins, myelin basic protein
(MBP) and proteolipid protein (PLP). While TNF- decreased MBP and
PLP mRNA abundance by 5- to 6-fold, IGF-I abrogated TNF--induced
reductions in a dose- and time-dependent manner. The changes in MBP and
PLP mRNA abundance could not be completely explained by the changes in
oligodendrocyte number, indicating that myelin protein gene expression
is regulated by both TNF- and IGF-I. These data support the
hypothesis that TNF- can mediate oligodendrocyte and myelin damage,
and indicate that IGF-I protects oligodendrocytes from TNF- insults
by blocking TNF--induced apoptosis, and by promoting oligodendrocyte
and precursor proliferation/differentiation and myelin protein gene
expression. |
Introduction |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
(TNF-), a
17-kDa cytokine, has been implicated in the mechanism of several
demyelinating disorders, including multiple sclerosis (MS) and its
animal model experimental autoimmune encephalomyelitis [EAE, see
review (1)]. TNF- abundance is greatly increased in the areas
surrounding damaged regions of the central nervous system (CNS) and in
the cerebral-spinal fluid (CSF) of patients with MS and EAE rats. In
organotypic rat spinal cord culture, TNF- selectively damages
oligodendrocytes and myelin, resulting in swelling of myelin sheaths
and oligodendrocyte death (2). In the cultures of purified human (3)
and rat (4) oligodendrocytes, as well as CG4 cells (5), a
nontransformed rat oligodendroglial cell line, TNF- has been shown
to induce cell death with characteristics of apoptosis (programmed cell
death). TNF-’s effects on myelination also have been validated
using genetic methods. Overexpression of TNF- in the brains of
transgenic (Tg) mice results in severe CNS hypomyelination during
development and early death of the affected animals (6, 7), which, at
least in part, results from oligodendrocyte apoptosis (8). These Tg
mice, however, can be rescued by a monoclonal antibody against TNF-.
Most recently, Taupin et al. (9) generated several
additional lines of TNF- Tg mice, and showed that when EAE is
induced in these Tg mice, clinical abnormalities and morphological
changes become more severe, compared with those of normal littermate
controls. Conversely, while the course of EAE is not altered in the
mice carrying null mutation for either TNF- or TNF-ß (10, 11),
induction of EAE is completely blocked in mice in whom the expression
of both TNF- and TNF-ß is ablated (12). These data suggest that
either TNF- or TNF-ß can mediate the pathophysiology of EAE, and
each can subserve the role of the other when one is not expressed. Insulin-like growth factor I (IGF-I), a member of the insulin superfamily, is widely expressed in CNS during development (13, 14). Several lines of evidence indicate that IGF-I promotes the development and survival of oligodendrocytes and their precursors, as well as myelination. For example, using Tg mouse models of IGF-I overexpression and of decreased IGF-I bioavailability, we have demonstrated that IGF-I increases myelination by promoting oligodendrocyte proliferation and/or survival and myelin-specific protein gene expression, resulting in an increased number of myelinated axons and in thickened myelin sheaths (15, 16). Experimental data also suggest that IGF-I protects the CNS from demyelinating injuries and promotes recovery after such damage: 1) IGF-I expression increases in astrocytes located near lesioned areas in animal CNS injury models, such as EAE, an animal model of MS (17), cryogenic injury (18), and chemical (19) and hypoxic-ischemic (20) insults; 2) IGF-I prevents cultured oligodendrocytes and their precursors from apoptosis induced by growth factor withdrawal (21); 3) IGF-I inhibits demyelination induced by anti-white matter antiserum and complement in organotypic nerve tissue culture (22); and 4) systemic injection of IGF-I into EAE rats reduces lesion severity and clinical deficits, and increases the expression of myelin-related protein genes (23).
We postulated that IGF-I protects oligodendrocytes and myelination from
TNF- damage. To test our hypothesis and determine the possible
mechanisms of IGF-I’s actions, we employed mixed glial cultures
derived from neonatal brains of C57/B6 mice, a strain that is highly
susceptible to EAE induction. We selected the study of mixed glial
cultures as our first step to investigate IGF-I’s actions because it
permits examination of both oligodendrocytes and their precursors under
the same conditions. Herein we provide evidence that IGF-I protects
oligodendrocytes from TNF--induced damage by blocking its promotion
of apoptosis, promoting oligodendrocyte proliferation and maturation,
and stimulating myelin-specific protein gene expression.
|
Materials and Methods |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
For immunohistochemical staining and labeling studies (see below),
cells (1–1.5 x 106/well) were seeded into 12-well
plates containing polylysine-coated glass coverslips. After cultured
overnight in DMEM supplemented with 10% FCS, cells were washed with
serum-free medium 3x and switched to a defined DMEM (24), containing
20 nM progesterone, 100 µM putrescine, 30
nM triiodothyronine, 50 µg/ml transferrin, 100 ng/ml
biotin, 5 ng/ml selenium, 4.5 g/liter glucose, and 1% heat-inactivated
FCS. Neuronal cells do not survive in this defined medium. The defined
medium alone or together with recombinant human IGF-I (Genentech, Inc., South San Francisco, CA), mouse TNF- (Gibco BRL, Gaithersburg, MD), and/or
(Boc-aspartyl(Ome)-fluoromethyl ketone (BAF, 100 µM,
Enzyme Systems Products, Dublin, CA), was changed daily.
BAF was dissolved in DMSO such that the final concentration of DMSO in
cultures was 0.1%. This concentration of DMSO had no apparent effect
on glial cultures.
For messenger RNA (mRNA) analysis, 4 x 106 cells were
seeded into a 60-mm Petri dish. One day later, cultures were washed
with serum-free medium 3x, and defined DMEM alone or with the addition
of IGF-I, and/or TNF- or rat interferon-
(IFN-, Gibco BRL) was added and changed daily.
Northern blot hybridization analysis
Total RNA was extracted from cultures using the acidic
guanidinium thiocyanate-phenol-chloroform method (25). Aliquots of 20
µg total RNA were electrophoresed, transferred onto nylon membranes
(DuPont NEN, Boston, MA) and UV cross-linked. Membranes
were stained with 0.02% methylene blue and photographed to quantify
the amount of RNA transferred. Membranes were hybridized with
radiolabeled single stranded DNA probes (see below) in Church’s buffer
(0.5 M sodium phosphate, pH 7.1/7% SDS/0.1 mM
EDTA) and washed at high stringency with 40 mM sodium
phosphate and 0.2% SDS at 55 C for 60 min. The position of specific
mRNAs was detected by autoradiography. After autoradiography, membranes
were stripped in 20 mM sodium phosphate with 0.5
mM EDTA at 80 C for 60 min, and hybridized with a second
probe.
Quantification was performed using a computer-assisted image analysis system (Image-Pro, Media Cybermetics, Silver Spring, MD). To ensure the accuracy of the changes in mRNA abundance and equal loading of RNA, the mRNA levels were normalized to cyclophilin mRNA abundance or to the amount of 18S rRNA on the membrane. The abundance of cyclophilin mRNA closely paralleled the amount of 18S rRNA transferred, as estimated by methylene blue staining.
Probes
Proteolipid protein (PLP), myelin basic protein (MBP), and
cyclophilin DNA fragments were amplified by PCR and used as templates
for probes, as described in a previous report (15). The PLP DNA
fragment corresponded to base pairs (bp) 268–903 of mouse PLP
complementary DNA (cDNA) (26), the MBP DNA fragment corresponded to bp
58–490 of the mouse MBP cDNA (27), and the cyclophilin DNA fragment
corresponded to bp 106–517 of the rat cyclophilin cDNA (28).
Single-stranded DNA hybridization probes were generated from these
templates by linear PCR (15, 29) using their respective 3' end primers
and 32P-labeled dCTP (Amersham Corp.,
Arlington Heights, IL).
Immunohistochemical staining
After fixation with 4% paraformaldehyde in PBS, coverslips with
adhered cultured brain cells were washed with PBS 3x. Oligodendrocytes
were identified by incubation with polyclonal antibodies against either
MBP (1:250, Chemicon International, Temecula, CA) or galactocerebroside
(GC, 1:50, Chemicon International). Oligodendrocyte precursors were
recognized by a monoclonal antibody A2B5 (1:25, Boehringer Mannheim, Indianapolis, IN). Antibody-antigen complexes were
detected by peroxidase- or alkaline phosphatase-conjugated secondary
antibodies supplied in ABC Kits (Vector Laboratories, Inc., Burlingame, CA) and visualized by incubation with DAB or
Vector Red (Vector Laboratories, Inc.), respectively.
Double immunohistochemical staining and TdT-mediated dUTP-Dig nick
DNA end-labeling (TUNEL)
After fixation with 4% paraformaldehyde and washing with PBS,
oligodendrocytes and their precursors were labeled with an anti-MBP
antibody and A2B5 antibody, respectively, and visualized with Vector
Red, as described above in the Immunohistochemical Staining section.
Cells then were fixed again with 4% paraformaldehyde for 10 min,
treated with 0.1% Triton X-100 in PBS for 10 min at room temperature,
and washed with PBS 3x. Apoptotic cells were labeled with dUTP-Dig
(supplied in the ApopTag kit; Oncor, Gaithersburg, MD) and an anti-Dig
antibody conjugated with peroxidase according to the manufacture’s
protocol, and subsequently visualized by incubation with DAB and
nickel. TUNEL and immunostained double-labeled cells were counted.
Bromo-2-deoxyuridine (BrdU) labeling and detection
To label proliferating cells, BrdU (1 µg/ml) was added into
culture medium every 12 h for 3 days. In other experiments,
cultures were pulse-labeled with 3 µg/ml BrdU for 24 h at the
end of a 3-day culture period. In each case, cultures were fixed with
4% paraformaldehyde for 10 min, washed with PBS and treated with 0.1%
Triton X-100. Oligodendrocytes and their precursors were identified
with anti-MBP antibody and A2B5 antibody, respectively, and visualized
with Vector Red. BrdU-labeled cells were detected using a Cell
Proliferation Kit (Amersham Corp.) and visualized by
incubation with DAB and nickel, according the manufacturer’s
protocol.
Statistics
One-way ANOVA was used to test statistic significance among
groups and followed by comparison for the group means according to
Newman-Keuls’ method.
|
Results |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
and IGF-I, and
their interactions, on oligodendrocytes and myelination, mixed glial
cultures treated for 3 days with TNF- and/or IGF-I were
immunochemically stained using anti-MBP and A2B5 antibodies to detect
oligodendrocytes and their precursors, respectively. Consistent with
other reports (30, 31, 32), oligodendrocytes in untreated control cultures
were scattered on the top of an astrocyte layer (Fig. 1
), and many A2B5 positive cells
exhibited bipolar or unipolar morphology, whereas some were round
apolar (data not shown). Most MBP-labeled oligodendrocytes had long
processes and produced large sheet-like myelin membranes (Fig. 1A).
Treatment with IGF-I (30 ng/ml) appeared to increase the number of
labeled oligodendrocytes, the size of their sheet-like membranes and
the intensity of their immunostaining (Fig. 1B). In contrast, TNF-
(100 ng/ml) appeared to reduce the number of oligodendrocytes and the
length of their processes (Fig. 1C). In addition, sheet-like membranes
of many oligodendrocytes were disrupted (Fig. 1C). In cultures treated
with both IGF-I and TNF- (Fig. 1D), the number of oligodendrocytes
appeared to be greater than in the cultures treated with TNF- alone,
although the morphology of oligodendrocytes did not appear markedly
different from those that were TNF--treated. No obvious changes in
oligodendrocyte precursor morphology were observed among the cultures
(data not shown).
|
). This finding
is consistent with the observation that IGF-I increases the rat
oligodendrocyte number in culture (33, 34). A higher dose of IGF-I (100
ng/ml) did not further increase oligodendrocyte number (data not
shown), and therefore, 30 ng/ml was the maximum concentration used in
all subsequent studies. In contrast, TNF--treated cultures had an
approximately 40% decrease in oligodendrocyte number. Addition of
IGF-I to TNF--treated cultures negated the effects of TNF- (Fig. 2). Similar results were also obtained when an antibody for
galactocerebroside (GC), another oligodendrocyte-specific marker, was
used to identify oligodendrocytes (data not shown).
|
). In contrast to its
effects on oligodendrocyte number, TNF- increased the number of
A2B5-positive oligodendrocyte precursors by 10%. This finding is
similar to the data of others showing that TNF- increases cultured
rat oligodendrocyte precursors (35). Simultaneous addition of IGF-I and
TNF- to cultures did not further increase the number of
oligodendrocyte precursors.
To determine whether the increases in oligodendrocyte precursor number
induced by IGF-I and TNF- were due to increased proliferation, we
exposed cultures to BrdU, a thymidine analog that can be taken up
during DNA synthesis phase of cell cycle, for the last 24 h of a
3-day treatment period. BrdU-labeled A2B5-positive precursors were then
quantified (Fig. 3A). In control
cultures, approximately 3% of total A2B5-positive cells were labeled
with BrdU. Compared with untreated cultures, cultures treated with
either IGF-I, TNF-, or both exhibited approximately 42%
(P < 0.01), approximately 11% (not significant) and
approximately 23% (P < 0.05) increases in the number
of BrdU-labeled A2B5-positive cells, respectively. These data, together
with the data showing that IGF-I and TNF- increase the number of
total A2B5 cells, strongly suggest that each IGF-I and TNF- is
capable of stimulating oligodendrocyte precursor
proliferation.
|
influence on the proliferation and/or
differentiation of oligodendrocytes, we: 1) labeled cultures with BrdU
every 12 h for 3 days and identified MBP-positive cells labeled
with BrdU, and 2) compared the number of oligodendrocytes before and
after the treatment period. In untreated cultures, approximately 6% of
oligodendrocytes were labeled with BrdU (Fig. 3A). Compared with
untreated cultures, the number of BrdU-labeled oligodendrocytes in
TNF--treated cultures was decreased by approximately 32%. Although
IGF-I treatment only modestly increased oligodendrocyte number
(approximately 11%), it normalized oligodendrocyte number in TNF-
treated cultures (I + T vs. control, no significant
difference). Compared with the beginning of experiments,
oligodendrocyte number increased approximately 26% after 3 days in
untreated cultures (Fig. 3B), indicating that oligodendrocytes continue
to mature in our culture conditions. Addition of IGF-I further
increased the number of oligodendrocytes by approximately 88%. In
contrast, TNF- slightly decreased the number of oligodendrocytes
(
5%, not significant). Addition of IGF-I to TNF--treated
cultures restored the number of oligodendrocytes to those in untreated
control cultures.
We next asked whether the reductions in oligodendrocyte number in
TNF--treated cultures were due to apoptosis, and if so, whether
IGF-I inhibits TNF--induced death of oligodendrocytes and their
precursors. By combining the TUNEL method to identify apoptotic cells
and immunostaining with anti-MBP antibody and A2B5 antibody to identify
oligodendrocytes and their precursors, we quantified the number of
apoptotic oligodendrocytes and their precursors (Fig. 4, A and B). In untreated cultures,
approximately 2% and 0.45% of total oligodendrocytes and their
precursors, respectively, were found to be apoptotic (Fig. 4C). In
multiple experiments treatment with IGF-I for 2 days decreased the
number of apoptotic oligodendrocytes by approximately 50% and their
precursors by approximately 36%. However, because the absolute changes
in number were small, these differences did not meet tests of
statistical significance. In contrast, treatment with TNF- for 2
days under identical conditions doubled the number of both apoptotic
oligodendrocytes and their precursors. Simultaneous addition of IGF-I
to TNF--treated cultures lowered the number of apoptotic
oligodendrocytes and their precursors toward those in control cultures
(Fig. 4C). Similar results also were observed in cultures treated with
IGF-I and TNF- for 1 day (data not shown).
|
-induced increase in the number of apoptotic
oligodendrocytes is consistent with the decrease in oligodendrocytes
cultured with TNF-, although the number of apoptotic
oligodendrocytes identified was small. The magnitude of apoptosis is
probably not completely quantified by TUNEL and immunostaining in our
culture conditions. We observed a marked increase in cell debris in
TNF--treated culture, and thus, it seems likely that many apoptotic
cells detached from the culture plate. Because TNF- is known to
activate interleukin-1ß converting enzyme (ICE)/caspases and results
in apoptosis in many cell types (see review in Ref. 36), we elected,
therefore, to indirectly evaluate TNF- influence on
oligodendrocyte apoptosis by testing the capacity of BAF, an
inhibitor of ICE/caspases, to prevent oligodendrocyte apoptosis and to
mimic IGF-I’s actions. After 3 days in culture, oligodendrocytes in
control cultures and in the cultures treated with 100 µM
of BAF had similar morphology. Similar to our previous experiments,
TNF- treatment resulted in marked changes in MBP+ oligodendrocyte
morphology and in an approximately 31% decrease in the number of
oligodendrocytes, compared with control cultures (P <
0.05). BAF increased the number of oligodendrocytes by approximately
29% (P < 0.05). Addition of BAF to TNF--treated
cultures ameliorated TNF-’s reduction of oligodendrocyte number
(Fig. 5). In a separated experiment,
cultures treated with TNF- and BAF for 2 days exhibited similar
results. These findings indicate that TNF- induces oligodendrocyte
apoptosis by ICE/caspases-mediated mechanisms, and that IGF-I either
directly inhibits apoptosis or blocks TNF- stimulation of apoptosis,
or both.
|
affects oligodendrocyte function and whether
IGF-I influences any such TNF- effects, Northern blot hybridization
analyses of two mRNAs for myelin-specific proteins, MBP and PLP, were
performed. In these experiments, IFN- (100 U/ml), another cytokine
known to reduce brain myelination (37), also was studied. Compared with
untreated control cultures, addition of IGF-I for 3 days increased MBP
and PLP mRNA abundance by approximately 57% and 43%, respectively
(Fig. 6). In contrast, treatment of
cultures with TNF- reduced MBP and PLP mRNA abundance to
approximately 15% and 32% of control levels, respectively. In an
independent experiment, cultures treated for 1 day with human TNF-,
which only binds to murine type 1 TNF receptor, also exhibited a
decreased MBP mRNA level (data not shown). IFN- (100 U/ml) also
modestly decreased MBP and PLP mRNAs by approximately 30% and 21%,
respectively. Simultaneous addition of IGF-I ameliorated the effects of
TNF- on MBP and PLP mRNA abundance by increasing their levels 2- to
3-fold. IGF-I, however, did not restore the abundance of these
mRNAs to normal. Similarly, IGF-I negated the effects of IFN- on
MBP and PLP mRNA abundance.
|
damage were then characterized. Compared with untreated
cultures, IGF-I by itself stimulated increases in the abundance of MBP
and PLP mRNAs that became significant by 48 h in culture (Fig. 7). In contrast, TNF- markedly reduced
the abundance of MBP and PLP mRNAs after 24 h treatment
(approximately 30% and 50% of control levels, respectively) and
further decreased these levels to less than approximately 20% and 45%
of controls, respectively, by 72 h of treatment. Simultaneous
addition of IGF-I increased the TNF--induced reduction in MBP and
PLP mRNA abundance by 2- to 3-fold at all time studied after 12 h.
In the case of PLP mRNA, IGF-I maintained levels at those of controls.
The effects of IGF-I on TNF--induced reductions in MBP and PLP mRNA
abundance also were directly related to IGF-I’s concentration (Fig. 8). With increasing IGF-I doses from
0–30 ng/ml, the abundance of MBP and PLP mRNAs in TNF--treated
cultures was increased from approximately 20% and 40% to 70–80% of
untreated controls, respectively.
|
|
|
Discussion |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
damages myelination in culture by
reducing the number of mature oligodendrocytes and by blunting their
function, consistent with the data of others that TNF- damages
oligodendrocyte and myelin. The TNF--induced reduction in mature
oligodendrocyte number appears to be predominately due to the promotion
of cell death in oligodendrocytes and their precursors. In contrast,
IGF-I exerts significant growth-promoting actions on oligodendrocytes
and their precursors. As judged by BrdU labeling, IGF-I stimulates the
proliferation of oligodendrocyte precursors. IGF-I’s stimulation of
oligodendrocyte function (judged by an increased myelin protein gene
expression) also suggests that IGF-I promotes the differentiation of
oligodendrocyte precursors. These actions, together with IGF-I’s
antiapoptotic effects, result in an increase in the number of mature
oligodendrocytes. These data are consistent with IGF-I’s actions shown
in transgenic mice and cultured rat oligodendrocytes reported by us and
others (15, 16, 33, 38, 39). Our data also clearly demonstrate that
IGF-I protects oligodendrocytes and their precursors from TNF-’s
deleterious effects. These observations are of potential clinical
importance because TNF-, as well as other cytokines (such as
IFN-), has been implicated in a variety of demyelinating diseases,
such as MS and its animal model, EAE (1).
Our finding that TNF- decreases oligodendrocyte number by apoptotic
mechanisms is consistent with that of others. Louis et al.
(5) demonstrated that TNF- induces an apoptotic-like cell death in
CG4 cells, an oligodendroglial cell line derived from rat brain. These
cells exhibit intense cytoplasmic vacuolation and disruption of the
cell processes within 6 h of TNF- exposure. More recently,
D’Souza et al. (3) also demonstrated that TNF-
induces apoptosis in up to 60% of cultured human oligodendrocytes
after 4 days. Judged by TUNEL method, however, we observed that TNF-
induced apoptosis in only a small fraction of total oligodendrocytes
and precursors (4% and 0.8%, respectively). In our experimental
conditions it is most likely that we underestimated the number of
apoptotic oligodendrocytes because apoptotic cells continually detached
from culture plates, and thus, escaped detection by TUNEL (although
other possibilities exist). Our findings that TNF--treated cultures
exhibit an increase in cell debris and that the ICE/Caspase inhibitor
BAF blocks the TNF--induced reduction in oligodendrocyte number
support this conclusion. They also may explain the discrepancy between
our data and that of others (4, 5). Furthermore, these data indicate
that TNF- apoptotic actions are mediated by ICE/caspases.
The increase in oligodendrocyte number during the 3-day experimental
period, following the initial 8–10 days in culture, indicates
continuous differentiation/maturation of oligodendrocyte precursors
under our culture condition. Addition of IGF-I further increases
oligodendrocyte number. These data, combined with the finding that
IGF-I increases BrdU-labeling of oligodendrocytes, strongly support a
role for IGF-I in the promotion of oligodendrocyte precursor
differentiation. Unlike IGF-I, however, TNF- decreases mature
oligodendrocyte number and their BrdU-labeling, while stimulating small
increases in the number and BrdU-labeling of oligodendrocyte
precursors. These data suggest that TNF- differentially affects
oligodendrocytes and their precursors, and that TNF- might inhibit
oligodendrocyte precursor differentiation. The latter possibility is
supported by the facts that TNF- is capable of decreasing MBP mRNA
abundance in mixed glial cultures (this study) and its immunoactivity
in cultured rat oligodendrocytes (40), and of influencing
differentiation and growth of neuronal precursors (41, 42).
Consistent with the changes in oligodendrocyte number, the abundance of
MBP and PLP mRNAs were increased in IGF-I-treated cultures and reduced
in TNF--treated cultures. The changes in the abundance of MBP and
PLP mRNAs, however, cannot be completely explained by the changes
in the number of oligodendrocytes. For example, treatment with TNF-
for 3 days reduced MBP and PLP mRNA levels to approximately 20% and
30% of controls, respectively, but only decreased the oligodendrocyte
number to approximately 60% of controls. Conversely, IGF-I increased
TNF--reduced-MBP mRNA abundance by approximately 3-fold, but
oligodendrocyte number only by approximately 40%. These data indicate
that TNF- treatment decreases oligodendrocyte function, in addition
to its effects on oligodendrocyte survival. In contrast, IGF-I promotes
oligodendrocyte survival, maturation and the expression of myelin
protein genes.
The mechanism(s) by which IGF-I counteracts TNF-’s effects on
oligodendrocytes involves both the inhibition of apoptosis and the
stimulation of oligodendrocyte precursor proliferation and/or
differentiation. Although treatment with IGF-I alone lowered the number
of apoptotic oligodendrocytes and their precursors, these changes were
small and did not meet tests of statistical significance. IGF-I,
however, significantly reduced TNF--induced apoptosis of
oligodendrocytes, suggesting that IGF-I might directly antagonize the
effects of TNF- on apoptosis. Although it is not clear whether IGF-I
acts directly on oligodendrocytes or indirectly by influencing
astrocytes in our culture condition, this inhibitory action of IGF-I on
TNF--induced oligodendrocyte death may have important consequences
in vivo. Application of BAF mimics IGF-I’s actions on
apoptosis, and raises the possibility that IGF-I blocks TNF-’s
actions by inhibiting the ICE/Caspase pathway. In addition, IGF-I
increases the proliferation and maturation of oligodendrocytes and
their precursors and the expression of myelin protein genes regardless
of TNF- exposure. IGF-I, therefore, appears to promote
oligodendrocyte growth by antagonizing TNF--induced apoptosis, and
by stimulating the proliferation, maturation and function of
oligodendrocyte precursors and oligodendrocytes.
In EAE rats, IGF-I expression is significantly increased in the astrocytes located near lesion areas (17). Furthermore, systemic administration of IGF-I to EAE rats increases myelin-related protein gene expression, whereas reducing lesion severity and clinical deficits (23, 43). These reports are consistent with our data that IGF-I has direct protective effects on oligodendrocytes. This conclusion is supported by the findings that oligodendrocytes express type I IGF receptors in vitro and in vivo (33, 44, 45), and that IGF-I inhibits demyelination induced by anti-white matter antiserum and complement in organotypic nerve tissue culture (22). More recently, IGF-I is shown to reduce the permeability of blood-brain barrier in EAE rats (46), indicating IGF-I might also affect other cell types in EAE rat, in addition to its direct actions on oligodendrocyte lineage. Our data, together with the above-cited data of others, suggest that IGF-I may be useful in the treatment of demyelinating diseases such as MS.
|
Footnotes |
|---|
Received October 1, 1998.
|
References |
|---|
|
TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
|---|
. Proc Natl Acad Sci USA 92:11294–11298 in the central nervous system. Eur J Immunol 27:905–913[Medline]
and lymphotoxin are
not required for induction of acute experimental autoimmune
encephalomyelitis. J Exp Med 185:2177–2182 in the mouse hippocampus
following transient forebrain ischemia. J Cereb Blood Flow Metab 17:491–499[Medline]
on astrocytes, microglia, oligodendrocytes, and
glial precursors in vitro. Dev Neurosci 13:130–137[Medline]
and tumour necrosis factor- on
oligodendroglial lineage cell proliferation and differentiation
in vitro. Eur J Neurosci 8:1106–1116[CrossRef][Medline]
and tumor necrosis factor-: role of receptor
regulation on human neuroblasts. J Natl Cancer Inst 86:1694–1701 (TNF-),
interferon-, and interleukin-6 but not TNF-ß induce
differentiation of neuroblastoma cells: the role of nitric oxide.
J Neurochem 62:1330–1336[Medline]
This article has been cited by other articles:
 |
|
 |
|
  A. Joseph D'Ercole and P. Ye Expanding the Mind: Insulin-Like Growth Factor I and Brain Development Endocrinology, December 1, 2008; 149(12): 5958 - 5962. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 F. G. Mastronardi, D. D. Wood, J. Mei, R. Raijmakers, V. Tseveleki, H.-M. Dosch, L. Probert, P. Casaccia-Bonnefil, and M. A. Moscarello Increased Citrullination of Histone H3 in Multiple Sclerosis Brain and Animal Models of Demyelination: A Role for Tumor Necrosis Factor-Induced Peptidylarginine Deiminase 4 Translocation. J. Neurosci., November 1, 2006; 26(44): 11387 - 11396. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 P. Mana, D. Linares, S. Fordham, M. Staykova, and D. Willenborg Deleterious Role of IFN{gamma} in a Toxic Model of Central Nervous System Demyelination Am. J. Pathol., May 1, 2006; 168(5): 1464 - 1473. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
P. H. Larsen, A. G. DaSilva, K. Conant, and V. W. Yong Myelin formation during development of the CNS is delayed in matrix metalloproteinase-9 and -12 null mice. J. Neurosci., February 22, 2006; 26(8): 2207 - 2214. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 R. G. Smith, L. Betancourt, and Y. Sun Molecular Endocrinology and Physiology of the Aging Central Nervous System Endocr. Rev., April 1, 2005; 26(2): 203 - 250. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 B. Feldhaus, I. D. Dietzel, R. Heumann, and R. Berger Effects of Interferon-{gamma} and Tumor Necrosis Factor-{alpha} on Survival and Differentiation of Oligodendrocyte Progenitors Reproductive Sciences, February 1, 2004; 11(2): 89 - 96. [Abstract] [PDF]
|
|
|
|
|
|
J. L. Mason, S. Xuan, I. Dragatsis, A. Efstratiadis, and J. E. Goldman Insulin-Like Growth Factor (IGF) Signaling through Type 1 IGF Receptor Plays an Important Role in Remyelination J. Neurosci., August 20, 2003; 23(20): 7710 - 7718. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Moralez, W. H. Busby Jr., and D. Clemmons Control of Insulin-Like Growth Factor Binding Protein-5 Protease Synthesis and Secretion by Human Fibroblasts and Porcine Aortic Smooth Muscle Cells Endocrinology, June 1, 2003; 144(6): 2489 - 2495. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 E. Chaum and H. Yang Transgenic Expression of IGF-1 Modifies the Proliferative Potential of Human Retinal Pigment Epithelial Cells Invest. Ophthalmol. Vis. Sci., December 1, 2002; 43(12): 3758 - 3764. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
P. Ye, L. Li, R. G. Richards, R. P. DiAugustine, and A. J. D'Ercole Myelination Is Altered in Insulin-Like Growth Factor-I Null Mutant Mice J. Neurosci., July 15, 2002; 22(14): 6041 - 6051. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. Chrysis, A. S. Calikoglu, P. Ye, and A. J. D'Ercole Insulin-Like Growth Factor-I Overexpression Attenuates Cerebellar Apoptosis by Altering the Expression of Bcl Family Proteins in a Developmentally Specific Manner J. Neurosci., March 1, 2001; 21(5): 1481 - 1489. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. L. Mason, P. Ye, K. Suzuki, A. J. D'Ercole, and G. K. Matsushima Insulin-Like Growth Factor-1 Inhibits Mature Oligodendrocyte Apoptosis during Primary Demyelination J. Neurosci., August 1, 2000; 20(15): 5703 - 5708. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 B. R. Lackey, S. L. Gray, and D. M. Henricks Does the Insulin-Like Growth Factor System Interact with Prostaglandins and Proinflammatory Cytokines During Neurodegeneration? Experimental Biology and Medicine, May 1, 2000; 224(1): 20 - 27. [Abstract] [Full Text]
|
|
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
| Endocrinology | Endocrine Reviews | J. Clin. End. & Metab. |
| Molecular Endocrinology | Recent Prog. Horm. Res. | All Endocrine Journals |
, P
< 0.01; 
