By
From the Department of Neuroimmunology, Max-Planck-Institute for Psychiatry, D-82152 Martinsried, Germany
We explored expression and possible function of interferon- (IFN-
) in cultured fetal (E15)
rat dorsal root ganglion neurons combining whole cell patch-clamp electrophysiology with single cell reverse transcriptase polymerase chain reaction and confocal laser immunocytochemistry. Morphologically, we located IFN-
protein in the cytoplasm of the neurons in culture as
well as in situ during peri- and postnatal development. Transcripts for classic IFN-
and for its
receptor were determined in probes of cytoplasm sampled from individual cultured neurons,
which had been identified by patch clamp electrophysiology. In addition, the cultured neurons expressed both chains of the IFN-
receptor.
Locally produced IFN- acts back on its cellular source. Phosphorylation and nuclear translocation of the IFN-inducible transcriptional factor STAT1 as well as IFN-
-dependent expression of major histocompatibility complex class I molecules on the neuronal membrane
were noted in untreated cultures. However, both processes were substantially blocked in the
presence of antibodies neutralizing IFN-
. Our findings indicate a role of IFN-
in autocrine
regulation of sensory neurons.
Interferon- However, IFN- Thus far, the cellular source of IFN- In this study, we characterized IFN- Cell Culture.
DRG were prepared from Wistar rat fetuses
(E15) obtained from the breeding facility (Max-Planck-Institute
for Psychiatry) as previously described (11). In brief, DRG were
removed from the fetuses and were dissociated by 0.1% trypsin
(Worthington Biochemical Corporation, Freehold, NJ). Cells
were dissociated by trituration and were cultured on poly-L-ornithine (0.1 mg/ml, Sigma Chemical Co., Taufkirchen, Germany)
plus laminin-coated (10 µg/ml, gift from Dr. Ries, Max-Planck-Institute for Biochemistry, Martinsried, Germany) dishes in
DMEM medium (GIBCO BRL, Eggenstein, Germany) supplemented with 10% fetal calf serum (Pansystems GmbH, Nürnberg,
Germany) and 10 ng/ml nerve growth factor (NGF-7S mouse, Sigma Chemical Co.). On the first day cells were treated for 24 h
with 10 µM 5-fluoro-2-deoxyuridine (Sigma Chemical Co.) and 10 µM uridine (Sigma Chemical Co.) to inhibit growth of nonneuronal cells. Antibodies neutralizing IFN- Patch-clamp RT-PCR.
Whole cell recording and reverse transcription of cytoplasmic RNA from single neurons was performed as previously described (14). Cytoplasmic RNA of individual CD4+ myelin basic protein-specific T lymphoblasts (13)
and of satellite cells from the DRG culture was collected with the
patch-clamp pipette and was reverse transcribed using the same
protocol as previously described (14). Oligonucleotides for PCR
amplification were selected with program PRIMER (Whitehead
Institute, Cambridge, MA) and were produced by the Max-Planck-Institute for Biochemistry (Martinsried, Germany). Forward primer
and reverse primer were always chosen from different exons to
detect possible amplification of genomic DNA contamination.
The primer sequences were as follows: for rat IFN- Immunofluorescence Labeling and Confocal Laser Scanning Microscopy.
Cultured rat DRG neurons or frozen sections of rat lumbar DRG
were fixed with 4% paraformaldehyde in PBS. For labeling of MHC class I molecules on the cell membrane, mouse monoclonal antibodies directed against MHC class I (Ox 18, 10 µg/ml;
Serotec Ltd., Oxford, England) were added to the cell culture for
60 min before fixation. Nonspecific binding sites were blocked
with PBS containing 2% goat serum and 2% bovine albumin. For
intracellular labeling, cultured cells were treated with PBS containing 0.1% Triton X-100 for 30 min. In the first step, cells or
tissue sections were labeled with mouse monoclonal antibodies
recognizing rat IFN- Statistical Analyses.
Data were determined by three independent experiments and are indicated as mean ± SEM. Analyses of
STAT1 as well as of MHC class I by confocal laser scanning microscopy revealed significant (P <0.01) difference between the
untreated and IFN- Histological sections
were performed from rat lumbar DRG tissue. IFN-
DRG cells from embryonic rats (E15) were cultured in
the presence of nerve growth factor. Most of the DRG
neurons that differentiated in vitro bound monoclonal antibodies recognizing IFN-
After 7 d
in culture, neurons could be distinguished morphologically
from nonneuronal cells. Their neuronal identity was ascertained by whole cell patch-clamp electrophysiology (15). As
expected, the DRG-derived neurons showed sodium currents activated at different membrane potentials, and responded to current pulses of increasing amplitude with sodium action potentials (Fig. 3).
We then used single-cell RT-PCR analysis to identify the
intraneuronal IFN-
IFN-
Western blot analysis of total cell lysate of DRG
cultures demonstrated phosphorylated STAT1 (data not
shown), but this method does not identify the cell type in
the mixed DRG culture that has the active form of STAT1.
However, confocal laser scanning microscopy clearly located STAT1 in the nuclei of almost all cultured neurofilament-positive DRG neurons (96 ± 4%, Figs. 8 and 9), and also in the glia cells. In contrast, in differentiated hippocampal neuronal cultures, where no IFN-
The possible auto/paracrine action of neuronal IFN- Over the past few years it has become clear that numerous humoral mediators that originally had been thought to
act exclusively either in the immune system or in the nervous system in fact interconnect both organ systems. For
example, neurotrophic factors like nerve growth factor are
not restricted to neural cells, but are produced by and act
on immune cells as well (19). Conversely, a number of cytokines with classical immune functions are also effective in
nervous tissues. Proinflammatory cytokines like TNF- Autocrine gene regulation is a common feature of the
cytokine network. For example, IFN- Most reports on IFN- Autocrine IFN- Demonstration of local production of IFN- (IFN-
), or "immune interferon", is a key
mediator required to correctly orchestrate antimicrobial
and inflammatory tissue responses. It is remarkably pleiotropic, evoking highly diverse effects in many if not all tissues. The cytokine affects proliferation, differentiation, and
the ability to communicate in individual cells. In particular,
IFN-
controls the expression of genes encoding molecules
required in immune reactions, such as MHC products, cell
adhesion molecules, cytokines, and cytocidal proteins. In
contrast to IFN-
's global activities, the cellular sources of
the cytokine are remarkably restricted, with certain sets of
activated T lymphocytes and NK cells as the sole known producers (1, 2).
is not limited to the immune responses. In addition to its proinflammatory function, some
evidence suggests that IFN-
may also affect differentiation
and survival of neuronal cells. For example, in one investigation the cytokine delayed degeneration of sympathetic
neurons caused by withdrawal of nerve growth factor (3).
Furthermore, in the pheochromocytoma cell line PC12,
IFN-
facilitated nerve growth factor-induced neuronal differentiation (4) and induced long-term excitability by
activating transcription of the peripheral nerve type 1 sodium channel (5). Finally, the cytokine promoted cholinergic differentiation of neurons derived from embryonic
septal nuclei (6).
in healthy nervous
tissue has not been determined. Several reports described
IFN-
-like immunoreactivity in dorsal root ganglia (7, 8),
and an "IFN-
-like protein" extracted from sensory trigeminal rat ganglia was shown to share some biological activity with lymphocyte-derived IFN-
(9). However, the
molecular nature of these structures remained elusive. Attempts to identify mRNA for the cytokine were inconclusive (10), and the molecular weight of the "IFN-
-like activity" differed substantially from that of the classic cytokine.
gene transcripts in
cultured fetal rat dorsal root ganglion (DRG)1 neurons
combining patch-clamp electrophysiology with single cell reverse transcriptase (RT)-PCR amplification. We demonstrate that IFN-
immunoreactivity in the cytoplasm of
cultured DRG neurons is indeed associated with the transcription of mRNA for classic IFN-
. We also present
functional evidence of autocrine/paracrine regulatory activity exerted by neuronal IFN-
.
(30 µg/ml, DB1;
Laboserv, Gießen, Germany) were added to the cell culture as
described in the text. Hippocampal cell cultures (12) and T cell
cultures (13) were performed as previously described.
(These sequence data are available from EMBL/GenBank/DDBJ under
accession numbers X02326 and X02327), 5
-AGGATGCATTCATGAGCATCGCC-3
(385-407) and 5
-CACCGACTCCTTTTCCGCTTCCT-3
(223-201); and for rat CD4 (W3/25;
accession number M15768), 5
-GTGCCGAGGCTTCTCTTTCAGG-3
(56-77) and 5
-CCCAGAATTGTCTTTTGGTCAGAGG-3
(247-273). The primer sequences for glyceraldehyde-3-phosphate dehydrogenase (GAPDH), glial fibrillary acidic
protein (GFAP), and IFN-
-receptor have been previously described (14). PCR-amplification was done in a final volume of 50 µl
containing 1 µl of transcribed cDNA probe, the four deoxyribonucleotide triphosphates (final 0.2 mM, Pharmacia), 2.5 U AmpliTaq (Perkin-Elmer/Applied Biosystems GmbH, Weiterstadt, Germany) and 1× PCR-buffer (Perkin-Elmer/Applied Biosystems
GmbH) covered with two drops of mineral oil (Sigma Chemical
Co.). Before PCR amplification on a programmable thermocycler (MultiCycler PTC 200, MJ Research Inc.), denaturated primers (final 100 pmol primers) were added to each tube at 80°C.
The cDNA was denaturated at 95°C for 3 min. PCR was performed on cDNA from single cells with 48-50 cycles (93°C for
60 s; ramp with 0.1°C/s from 93°C to 60°C; 60°C for 60 s; 72°C
for 60 s) and followed by one final cycle at 72°C for 5 min. 10 µl
of the amplified fragments was run along with the molecular
weight marker (
X 174, HaeIII digested; Pharmacia Biotech,
Freiburg, Germany) on a 1.7% agarose gel electrophoresis stained
with ethidium bromide. For Southern blot analysis the DNA
fragments of the gel were transferred onto a nylon membrane (Hybond N+, Amersham Buchler, Braunschweig, Germany).
The resulting blot was hybridized with a rat IFN-
-specific
probe (rat IFN-
; these sequence data are available from EMBL/
GenBank/DDBJ under accession number X02327: 130-153) using the Digoseigenin Southern blot Hybridization System (Boehringer Mannheim, Mannheim, Germany) according to the manufacturer's instructions. For sequencing, the PCR fragments were
gel-purified (QIAGEN, Hilden, Germany) and were sequenced by
MediGene (Martinsried, Germany) with an automated sequence
analyzer.
(DB1; 10 µg/ml), mouse monoclonal antibodies directed against STAT1 (5 µg/ml; PharMingen, Hamburg,
Germany), rabbit polyclonal antibodies recognizing an epitope of
IFN-
-receptor
chain corresponding to amino acids 461-477
(2 µg/ml, K-17; Santa Cruz Biotechnology, Heidelberg, Germany), or rabbit polyclonal antibodies recognizing an epitope of
IFN-
-receptor
chain corresponding to amino acids 312-331
(2 µg/ml, M-20; Santa Cruz Biotechnology) followed by secondary fluorochrome Cy3-conjugated goat antibodies directed against mouse or rabbit immunoglobulin (10 µg/ml; Dianova,
Hamburg, Germany). In a second step, cells were double-labeled
with mouse monoclonal antibodies recognizing neurofilament (2 µg/ml, NN18; Boehringer Mannheim) followed by fluorochrome
(dichlorotriazinyl) aminofluorescein-conjugated antibodies to mouse
immunoglobulin (5 µg/ml; Dianova, Hamburg, Germany). Conventional photographs were taken with an immunofluorescence
microscope (Carl Zeiss, Oberkochen, Germany). Confocal images were acquired with a confocal laser scanning microscope
(Leica, Bensheim, Germany) equipped with ×63 oil objectives.
Baseline labeling for the IFN-
, STAT1, and MHC class I staining procedures was revealed with irrelevant mouse monoclonal
antibodies (10 µg/ml; Dianova) and secondary fluorochrome
Cy3-conjugated goat antibodies to mouse immunoglobulin (10 µg/ml; Dianova). Baseline labeling for the IFN-
-receptor
and
chains was revealed by adding the corresponding (blocking) peptides (2 µg/ml; Santa Cruz Biotechnology) to the polyclonal rabbit antibodies, followed by secondary fluorochrome Cy3-conjugated goat antibodies to rabbit immunoglobulin (10 µg/ml; Dianova).
neutralizing antibody-treated group in the
unpaired two-tail Student's t test.
IFN- Expression in Sensory Neurons.
immunolabeling was detectable in a subpopulation of DRG
cells during peri- and postnatal development (Fig. 1). Histological sections showed strong IFN-
expression at embryonic day 18 and during the first 2 wk after birth. The in
situ IFN-
labeling was very prominent in the neuronal
perikarya, but was also demonstrable in the neuronal processes (Fig. 1).
Fig. 1.
IFN- immunoreactivity in the DRG during postnatal development. (A) Frozen
section from rat postnatal day 9 DRG was immunolabeled with
antibodies directed against IFN-
.
The IFN-
immunoreactivity was detectable in the DRG, but
not in the adjacent muscle tissue. Scale bar: 50 µm. (B) The
perikarya and processes of a subpopulation of neurons were immunolabeled with IFN-
-specific antibodies in a frozen
section of postnatal day 9 DRG.
Scale bar: 20 µm.
[View Larger Version of this Image (192K GIF file)]
. Specific immunofluorescence
was detected after 7 d in culture in 83 ± 8% of DRG neurons identified by neurofilament labeling, but in none of
the nonneuronal cells (Fig. 2). Confocal laser scanning microscopy localized the IFN-
proteins in the cytoplasm,
sparing the nucleus and the cell membrane (Fig. 2), whereas
processes and dendrites showed no or only very weak reactivity.
Fig. 2.
IFN- immunoreactivity in cultured sensory neurons. (A) IFN-
immunofluorescence labeling in neurons.
Embryonic sensory neurons cultured for 7 d were identified by
antibodies recognizing neurofilament and were double-labeled
with antibodies directed against
IFN-
. Scale bar: 10 µm. (B)
Confocal laser scanning microscopy of sensory neurons. Neurofilament labeling is located in
the cytoplasm and the neuronal
processes, whereas IFN-
labeling is only localized in the perinuclear cytoplasm. Scale bar: 10 µm. (C) Incubation with primary control antibodies instead
of IFN-
-specific antibodies showed baseline labeling intensity. Scale bar: 10 µm.
[View Larger Version of this Image (39K GIF file)]
Gene Transcription in Sensory Neurons.
Fig. 3.
Identification of neurons by whole cell patch-clamp electrophysiology. (A) Action potentials were evoked in sensory neurons cultured for 7 d by current pulses of increasing amplitude (200, 300, and 400 pA) and were recorded in the current clamp mode. (B) Whole cell membrane currents were evoked by successive depolarization steps of 10 mV
(from a holding potential of 80 mV to voltages ranging from
50 to
+50 mV) and were recorded in the voltage clamp mode.
[View Larger Version of this Image (23K GIF file)]
-like material. After electrophysiological characterization, we sampled cytoplasmic specimens from
individual neurons through the patch-clamp micropipette,
and assessed current gene transcription by RT-PCR. Oligonucleotides specific for IFN-
were used to amplify gene
transcripts for the cytokine. In addition, as in our previous studies, coamplification of mRNA for the house-keeping
enzyme GAPDH and for cell lineage markers served as internal quality standards (12, 14). The validity of our method
was corroborated by parallel analyses of activated CD4+ T
lymphocytes with a Th1-like phenotype (13) and DRG-derived glia cells (Fig. 4). Each single T lymphoblast analyzed contained mRNA for the lineage marker gene CD4
along with IFN-
. In contrast, satellite cells from DRG cultures, which expressed glia specific GFAP gene transcripts,
and were negative for IFN-
and CD4 gene transcripts
(Fig. 4). In a subsequent cytokine study of 19 individual DRG neurons, mRNA for IFN-
was identified in 13 cells
cultured for 7 d (Fig. 5). Southern blot analysis and sequencing confirmed the identity of the amplified fragments
with classic IFN-
. IFN-
gene expression in cultured DRG
neurons developed steadily over time. Among neurons cultured for 2 h, only a minority (1/11) of the (electrophysiologically immature) cells expressed IFN-
gene transcripts
(Fig. 5). However, the percentage of IFN-
-expressing neurons continuously increased through the first 24 h in
culture. After 6 h, 3/13 neurons, and after 24 h, 7/11 DRG
neurons, expressed IFN-
gene transcripts. In contrast, differentiated neurons derived from hippocampus tissue transcribed IFN-
at no time.
Fig. 4.
Single cell RT-PCR of CD4+ T lymphoblasts and DRG satellite cells. Gene transcripts for GAPDH (A), GFAP (B), CD4 (C), and
IFN- (D) were analyzed by single-cell RT-PCR of activated T cells
(lanes 1-3) and of DRG satellite cells (lanes 4-6). N and M show negative PCR control and molecular weight marker, respectively.
[View Larger Version of this Image (60K GIF file)]
Fig. 5.
Gene transcripts for IFN- detected in sensory neurons by
single-cell RT-PCR. (A) IFN-
gene transcripts amplified from individual neurons. Embryonic sensory neurons were cultured for 7 d and were identified by electrophysiology. Gene transcripts for GAPDH (lanes 1-9)
and IFN-
(lanes 1, 3, 4, 6-9) were detected by single-cell RT-PCR in
individual neurons. N and M show negative PCR control and molecular
weight marker, respectively. (B) Frequency of DRG neurons transcribing
IFN-
in relation to time in culture.
[View Larger Version of this Image (33K GIF file)]
-receptor Expression of Sensory Neurons.
acts
on its target cells by binding to and activating membrane-bound IFN-
receptors, heterodimeric proteins composed by a cytokine-binding
chain and signal-transducing
chain (16). Gene transcripts for the IFN-
receptor
chains were detected in all DRG neurons analyzed (Fig. 6)
Membrane expression of the receptor complex was confirmed by immunofluorescence and confocal laser scanning
microscopy. IFN-
-receptor
chain-specific antibodies labeled 98 ± 4% of neurofilament-positive DRG neurons,
predominantly on the cell membrane (Fig. 7). The signal-transducing
chain was detected on 97 ± 5% sensory neurons (Fig. 7). Simultaneous expression of a cytokine along
with its specific receptor in the same cell provides a necessary, although insufficient, structural basis for autocrine
regulation. This is the proven case in IFN-
production by
activated T lymphocytes (17), and could also hold true for
our cultured DRG neurons. Since the binding of IFN-
to
its receptors results in phosphorylation and translocation of
the transcriptional factor STAT1 from the cytoplasm to the nucleus (16), both features of STAT1 activation should be
demonstrable in the cells of our DRG cultures, and should
be reversed by the addition of neutralizing antibodies
against IFN-
.
Fig. 6.
IFN- receptor gene transcripts in sensory neurons. Gene
transcripts for GAPDH (lanes 1-5) and IFN-
-receptor
chain (lanes 1-5)
were amplified from single neurons by RT-PCR. N and M show negative PCR control and molecular weight marker, respectively.
[View Larger Version of this Image (52K GIF file)]
Fig. 7.
IFN- receptor expression detected on sensory
neurons by confocal laser scanning microscopy. (A) Confocal
localization of IFN-
receptor (
chain) of sensory neurons. Embryonic sensory neurons (cultured for 2 d) were identified by
antibodies recognizing neurofilament and were double labeled
with antibodies directed against
IFN-
-receptor
chain. (B)
Immunolabeling of IFN-
receptor (
chain) on the neuronal
cell membrane. Sensory neurons
(cultured for 2 d) were identified
by double-labeling with neurofilament. Scale bar A and B:
10 µm.
[View Larger Version of this Image (24K GIF file)]
synthesis is demonstrable, all STAT1 immunoreactivity was confined to
the cytoplasm (Fig. 8). IFN-
-neutralizing antibodies to
DRG cultures profoundly interfered with nuclear translocation of STAT1. After neutralization of IFN-
, most neurons displayed STAT1 within their cytoplasm, with nuclear location seen only in a minority of all cells (26 ± 11%,
Figs. 8 and 9).
Fig. 8.
Nuclear localization of STAT1 and cell membrane expression of MHC class I
molecules. (A) Nuclear localization of STAT1 in sensory neurons. Embryonic sensory neurons were cultured for 2 d and
analyzed by confocal laser scanning microscopy for neurofilament and STAT1. (B) Cytoplasmic localization of STAT1 in
neurons of hippocampal cultures.
(C) Cytoplasmic localization of
STAT1 in sensory neurons (cultured for 2 d) in the presence of
neutralizing antibodies directed
against IFN-. (D) MHC class I
molecules detected on the cell
membrane of untreated sensory
neurons. Sensory neurons (cultured for 5 d) were identified by
antibodies recognizing neurofilament and were immunolabeled
with antibodies directed against
MHC class I on the cell surface.
Scale bar A, B, C, and D: 10 µm.
[View Larger Version of this Image (24K GIF file)]
Fig. 9.
Autocrine activation of STAT1 and MHC class I in sensory
neurons. (A) Frequency of sensory neurons (cultured for 2 d with or
without IFN- neutralizing antibodies) demonstrating the nuclear localization of STAT1. (B) MHC class I molecules on the cell membrane of
sensory neurons (cultured for 5 d) untreated or treated with IFN-
neutralizing antibodies.
[View Larger Version of this Image (27K GIF file)]
was supported by an investigation of MHC class I expression on cultured DRG neurons. As previously reported,
IFN-
readily induces MHC class I genes in electrically silent neurons, but much less so in firing neurons (12, 14).
Adult sensory DRG neurons are silent in culture, and are
susceptible to MHC class I induction by IFN-
(18). Thus,
in the case of autocrine activity of neuronal IFN-
, cultured IFN-
-secreting neurons should constitutively express MHC class I on their membranes. That this is the case was documented by confocal microscopy with MHC class I
expression on 67 ± 11% of all sensory neurons (cultured
for 5 d and identified by a neurofilament marker, Figs. 8
and 9). Treatment of the DRG culture with neutralizing
anti-IFN-
monoclonal antibodies reduced the proportion
of MHC class I-positive neurons to 14 ± 3% (Fig. 9).
or
IL-1 influence neuronal function and behavior (20). A proportion of these mediators may be imported through the
blood brain barrier or discharged in the central nervous system by immigrant inflammatory cells. But it has also become clear that many of these cytokines are produced and
released within the nervous system by autochthonous cells
(21, 22). In this study, we have shown that classical IFN-
is produced and used by neurons in the peripheral nervous system.
produced by subsets of activated T cells activates STAT1 factor and thereby
upregulates its own gene expression (17, 23). Likewise, in
the brain, neurotrophils such as brain-derived neurotrophic
factor prevent death of adult sensory neurons acting in autocrine loops (24). Our results now suggest that STAT1 activation in neurons is at least partially regulated by IFN-
produced by the same cells. Although neutralization of IFN-
by monoclonal antibodies suppressed MHC class I membrane expression on neurons almost completely, this procedure inhibited phosphorylation and translocation of STAT1
only partially. This can be explained by the activity of neural mediators ligating the gp130 receptor of the IL-6/CNTF
family of cytokines, which uses besides STAT3 also the
STAT1 activation pathway (25).
activity within the nervous system are related to inflammatory responses, as represented
by experimental autoimmune encephalomyelitis, multiple
sclerosis, or microbial brain infections. In these situations,
the cytokine is mainly produced and released by infiltrating
immune cells (26). This is in contrast to neuronal IFN-
,
whose particular temporal and spatial expression patterns
suggest a developmental function, possibly related to the
differentiation of peripheral nerves.
secretion by DRG neurons may contribute signals controlling neuronal differentiation (4), like
activation of neuron-specific genes encoding the peripheral
nerve type 1 sodium channel (5). Other possible targets of
IFN-
-dependent regulation are the genes controlling neuron transmitter production (6). Alternatively, DRG neuronal
IFN-
could regulate glia cells surrounding the neurons in
situ. In vitro, IFN-
interferes with myelination of central
nervous system (27) and peripheral nerve myelination (28).
In agreement with this, transgenic mice constitutively producing IFN-
in their central nervous system display a
"tremoring" phenotype coinciding with a "dramatic overall decrease in central nervous system myelin" (29).
by small
and medium sized DRG neurons (which are enriched in
our nerve growth factor-dependent culture model) and its
auto/paracrine action on local cells predicts a new and unexpected function of this unusually pleiotropic proinflammatory cytokine.
Address correspondence to Hartmut Wekerle, Neuroimmunology, Max-Planck-Institute for Psychiatry, D-82152 Martinsried, Germany. Phone: 49-89-8578-3550; FAX: 49-89-8578-3790.
Received for publication 27 August 1997 and in revised form 3 October 1997.
The project was supported by grants from the Deutsche Forschungsgemeinschaft (SFB 391) and the European Community (CHRX-CT94-0670).We thank Ms. L. Penner for expert technical assistance; Drs. R. Böhm-Matthaei, E. Hoppe, R. Kiefer, and H.-P. Hartung for technical advice; Dr. A. Ries for supplying us with laminin; Dr. G. Kääb for culturing T lymphoblasts; and Dr. I. Medana for helpful discussion.
1. |
Farrar, M.A., and
R.D. Schreiber.
1993.
The molecular cell
biology of interferon-![]() |
2. |
Boehm, U.,
T. Klamp,
M. Groot, and
J.C. Howard.
1997.
Cellular responses to interferon-![]() |
3. | Chang, J.Y., D.P. Martin, and E.M. Johnson. 1990. Interferon suppresses sympathetic neuronal cell death caused by nerve growth factor deprivation. J. Neurochem. 55: 436-445 [Medline]. |
4. |
Improta, T.,
A.M. Salvatore,
A. DiLuzio,
G. Romeo,
E.M. Coccia, and
P. Calissano.
1988.
IFN-![]() |
5. | Toledo-Aral, J.J., P. Brehm, S. Halegoua, and G. Mandel. 1995. A single pulse of nerve growth factor triggers long-term neuronal excitability through sodium channel gene induction. Neuron. 14: 607-611 [Medline]. |
6. |
Jonakait, G.M.,
R. Wei,
Z.-L. Sheng,
R.P. Hart, and
L. Ni.
1994.
Interferon-![]() |
7. |
Eneroth, A.,
K. Kristensson,
Å. Ljungdahl, and
T. Olsson.
1991.
Interferon-![]() |
8. | Kiefer, R., and G.W. Kreutzberg. 1990. Gamma interferon-like immunoreactivity in the rat nervous system. Neuroscience. 37: 725-734 [Medline]. |
9. |
Olsson, T.,
S. Kelic,
C. Edlund,
M. Bakhiet,
B. Höjeberg,
P. Van der Meide,
Å. Ljungdahl, and
K. Kristensson.
1994.
Neuronal interferon-![]() |
10. | Kiefer, R., C.A. Haas, and G.W. Kreutzberg. 1991. Gamma interferon-like immunoreactive material in rat neurons: evidence against a close relationship to gamma interferon. Neuroscience. 45: 551-560 [Medline]. |
11. | Wood, P.M.. 1976. Separation of functional Schwann cells and neurons from normal peripheral nerve tissue. Brain Res. 115: 361-375 [Medline]. |
12. | Neumann, H., A. Cavalié, D.E. Jenne, and H. Wekerle. 1995. Induction of MHC class I genes in neurons. Science. 269: 549-552 [Medline]. |
13. | Kääb, G., G. Brandl, A. Marx, H. Wekerle, and M. Bradl. 1996. The myelin basic protein specific T cell repertoire in (transgenic) Lewis rat/SCID mouse chimeras: preferential Vb8.2 T cell receptor usage depends on an intact Lewis thymic microenvironment. Eur. J. Immunol. 26: 981-988 [Medline]. |
14. |
Neumann, H.,
H. Schmidt,
A. Cavalié,
D. Jenne, and
H. Wekerle.
1997.
MHC class I gene expression in single neurons of the central nervous system: differential regulation by
interferon-![]() ![]() |
15. | Rohrer, H., S. Henke-Fahle, T. El-Sharkawy, H.D. Lux, and H. Thoenen. 1985. Progenitor cells from embryonic chick dorsal root ganglia differentiate in vitro to neurons: biochemical and neurophysiological evidence. EMBO (Eur. Mol. Biol. Organ.) J. 4: 1709-1714 [Abstract]. |
16. |
Bach, E.A.,
M. Aguet, and
R.D. Schreiber.
1997.
The IFN-![]() |
17. |
Hardy, K.J., and
T. Sawada.
1989.
Human ![]() |
18. |
Fujimaki, H.,
N. Hikawa,
T. Nagoya, and
M. Minami.
1997.
IFN-![]() |
19. | Torcia, M., L. Bracci-Laudiero, M. Lucibello, L. Nencioni, D. Labardi, A. Rubatelli, F. Cozzolino, L. Aloe, and E. Garaci. 1996. Nerve growth factor is an autocrine survival factor for memory B lymphocytes. Cell. 85: 345-356 [Medline]. |
20. | Rothwell, N.J., and S.J. Hopkins. 1995. Cytokines and the nervous system. II. Actions and mechanisms of action. Trends Neurosci. 18: 130-136 [Medline]. |
21. | Murphy, P.G., J. Grondin, M. Altares, and P.M. Richardson. 1995. Induction of interleukin-6 in axotomized sensory neurons. J. Neurosci. 15: 5130-5138 [Abstract]. |
22. |
Tchelingerian, J.-L.,
J. Quinonero,
J. Booss, and
C. Jacque.
1993.
Localization of TNF-![]() ![]() |
23. |
Girdlestone, J., and
M. Wing.
1996.
Autocrine activation by
interferon-![]() |
24. | Acheson, A., J.C. Conover, J.P. Fandl, T.M. DeChiara, M. Russell, A. Thadani, S.P. Squinto, G.D. Yancopoulos, and R.M. Lindsay. 1995. A BDNF autocrine loop in adult sensory neurons prevents cell death. Nature. 374: 450-453 [Medline]. |
25. | Zhong, Z., Z. Wen, and J.E. Darnell. 1994. Stat3: a STAT family member activated by tyrosine phosphorylation in response to epidermal growth factor and interleukin-6. Science. 264: 95-98 [Medline]. |
26. |
Benveniste, E.N., and
D.J. Benos.
1995.
TNF-![]() ![]() |
27. |
Agresti, C.,
D. D'Urso, and
G. Levi.
1996.
Reversible inhibitory effects of interferon-![]() ![]() |
28. |
Schneider-Schaulies, J.,
F. Kirchhoff,
J. Archelos, and
M. Schachner.
1991.
Down-regulation of myelin-associated glycoprotein on Schwann cells by interferon-![]() ![]() |
29. |
Corbin, J.G.,
D. Kelly,
E.M. Rath,
K.D. Baerwald,
K. Suzuki, and
B. Popko.
1996.
Targeted CNS expression of interferon-![]() |