Previous Article | Next Article 
Journal of Virology, December 2001, p. 11700-11708, Vol. 75, No. 23
Institut für Immunologie,
Bundesforschungsanstalt für Viruskrankheiten der Tiere,
Tübingen,1 and Institut
für Neuropathologie, Heinrich-Heine-Universität,
Düsseldorf,2 Germany
Received 7 May 2001/Accepted 9 August 2001
Experimental Borna disease virus (BDV) infection of rats and
natural infection of horses and sheep leads to severe central nervous
system disease based on immunopathological pathways. The virus
replicates slowly, and the cellular immune response results in
immunopathology. CD8+ T cells exert effector cell
functions, and their activity results in the destruction of
virus-infected cells. Previously, Oldach and colleagues (D. Oldach, M. C. Zink, J. M. Pyper, S. Herzog, R. Rott, O. Narayan, and J. E. Clements, Virology 206:426-434, 1995) have reported protection against
Borna disease after inoculation of high-dose cell-adapted BDV. Here we
show that the outcome of the infection, i.e., immunopathology versus
protection, is simply dependent on the amount of virus used for
infection. High-dose BDV (106 FFU) triggers an early
virus-specific reaction of the immune system, as demonstrated by strong
cellular and humoral responses. In particular, the early presence and
function of nucleoprotein-specific CD8+ T cells could be
demonstrated in the brain. We present evidence that in a noncytolytic
and usually persistent virus infection, high-dose input virus mediates
early control of the pathogen due to an efficient induction of an
antiviral immune mechanism. From these data, we conclude that immune
reactivity, in particular the cytotoxic T-cell response, determines
whether the virus is controlled with prevention of the ensuing
immunopathological disease or whether a persistent infection is established.
CD8+ T cells are
important in the control of many intracellular pathogens, where they
function as primary effector cells. Whereas an early and efficient
induction of CD8+ T cells is crucial after infection with
highly cytolytic viruses to eliminate the agent before viral
replication produces viral progeny, the role of CD8+ T
cells in infections with noncytolytic viruses appears to be more
complicated. Noncytolytic viruses are mostly defined as such because
they do not cause overt tissue destruction in vitro. However, in vivo
this situation might change considerably if this type of virus
encounters an intact immune system and induces an antiviral immune
response. Although the virus is mainly innocuous, the induced immune
response produces immunopathological pathways, often resulting in
severe disease. During the initial encounter with a virus, CD8+ T cells bearing T-cell receptors specific for the
given antigen are selected to undergo clonal expansion. In the case of
rapidly replicating viruses, it can be assumed that antigen is produced in an amount that triggers a vigorous immune response that either suffices to eliminate virus-infected cells or is not efficient enough
to control virus infection and results in disease and/or early death.
In viral infections in which only comparably low doses of infectious
virus are passed to a new host or in infections with slowly
replicating, noncytolytic and persistent virus, the concept of early
action of the immune system might not be valid simply due to an
insufficiently strong trigger for the immune system. In this case, only
the increasing number of infected cells over time provides a stimulus
to the immune system; however, this stimulus is too late to eliminate
the virus early after infection and/or to prevent persistence.
Borna disease virus (BDV) is an example of a noncytolytic persistent
virus. In recent years, this viral infection of the central nervous
system (CNS) has been diagnosed in a wide variety of animals including
cattle, cats, dogs, and birds (4, 6, 15, 16, 38).
Furthermore, virus, nucleic acid, and antibodies have been detected in
the blood of patients with psychiatric diseases (1, 5, 13, 18,
26, 31, 37; N. Nowotny and J. Kolodziejek, Letter, Lancet
355:1462-1463, 2000). However, so far no direct correlation
between BDV as the causative agent and any of these human disorders has
been demonstrated. BDV causes a persistent infection of the CNS and
induces Borna disease (BD), an immune-mediated encephalomyelitis
originally described in horses and sheep (14, 19, 30). The
infiltrating immune cells have been characterized as CD4+
CD8+ T cells and macrophages (2, 8, 29).
CD8+ T cells represent the effector cell population,
exhibiting antigen specificity for the nucleoprotein p40, specifically
for the peptide ASYAQMTTY, in the Lewis rat (23-25,
27, 32). No evidence has been presented that antibodies might
contribute to neuropathology, although neutralizing antibodies
apparently control virus tropism and can prevent the spread of virus
from peripheral infection sites to the CNS (9, 11, 34).
After experimental BDV infection of rats, protection against the
immune-mediated brain disease has been achieved by adoptive transfer of
CD4+ T-cell lines, resulting in the loss of virus from the
CNS (20, 24, 28). The underlying mechanisms responsible
for virus elimination have been extensively investigated, and strong
evidence for a role of CD8+ effector cells induced by
virus-specific CD4+ T-cell lines has been provided
(20). In addition to this T-cell-mediated protection,
Oldach et al. have reported protection against disease after infection
with high-dose (HD) cell-attenuated BDV (21); however, the
mechanism of virus control and protection from disease has not been
investigated, and therefore this interesting phenomenon remains to be
elucidated. To determine which effector mechanism might be responsible
for the elimination of BDV from the host after infection with HD virus,
we used HD virus obtained from two different cell types and determined
the immunological nature of this phenomenon by demonstrating enhanced
kinetics of an anti-BDV CD8+ T-cell response.
Viruses and infection. (i) BDV-BR.
The fourth
rat passage of BDV originally obtained from R. Rott and S. Herzog was used for the infection experiments (19).
(ii) BDV-CRL.
CRL1405 cells (guinea pig cell
line) were infected with the fourth rat passage of BDV-BR.
Then the virus was isolated from these cells after two to eight
passages by sonication.
(iii) BDV-MDCK.
Persistently infected
BDV-MDCK cells were lysed by sonication (12).
Five-week-old rats were infected intracerebrally (i.c.) in the left
brain hemisphere with 0.05 ml of the different virus isolates
corresponding to 106, 104, or 102
focus-forming units (FFU). The vaccinia virus (VV)-BDV recombinants (VV-p40, VV-p24, VV-gp18, and VV-gp94) were created by J. C. de la
Torre, Scripps Research Institute, La Jolla, Calif.
Experimental animals and immunosuppression.
Male and female
Lewis rats were bred in the animal-breeding facilities at the
Bundesforschungsanstalt für Viruskrankheiten, Tübingen,
Germany. For immunosuppression, rats thymectomized as adults were
treated with 2 mg of purified mouse monoclonal antibody directed
against rat CD8+ T cells (OX 8) 1 day before and 1 day
after infection. Furthermore, the rats were thymectomized as newborns
within 24 h of birth. Successful T-cell depletion was controlled
by fluorescence-activated cell sorter (FACS) analysis.
Clinical evaluation.
All experimental animals were examined
daily and weighed. Disease symptoms were scored using a scale from 0 to
3 based on the general state of health and the appearance of
neurological signs (score 1, slight incoordination and vigilance; score
2, distinct ataxia or slight paresis; score 3, marked pareses or paralyses).
Infectivity assay and viral antigen detection.
Virus
infectivity of brain homogenates from BDV-infected rats was determined
on CRL1405 cells. Titer determinations were carried out in flat-bottom
96-well microtiter plates. The CRL1405 cells were cultured for 7 days
in the presence of brain homogenates from infected rats. Thereafter the
cells were fixed with 4% formaldehyde-phosphate-buffered saline (PBS)
and treated with 1% Triton X-100-PBS, and viral antigen was
demonstrated in an immunocytochemical reaction using anti-BDV-specific mouse monoclonal antibodies. Nonspecific binding of immunological reagents was blocked by incubation of plates with 10% fetal calf serum-PBS. The reaction of monoclonal antibodies with cells was detected by a secondary anti-mouse biotin-labeled antibody
(Dianova, Hamburg, Germany) and by a streptavidin-peroxidase conjugate
(Dianova). The reaction was visualized with
ortho-phenylendiamine and H2O2 (Sigma, Munich, Germany). Additionally, tissue homogenates were used as
antigens in Western blot analysis, and the presence of virus-specific
antigen was detected by BDV-specific monoclonal antibodies.
Detection of BDV-specific antibodies.
All rat sera were
tested in a solid-phase enzyme-linked immunosorbent assay (ELISA) using
a 1:1,000 dilution of a brain homogenate from BDV-infected rats as
coating antigen and by Western blot analysis with a 10% brain
homogenate from BDV-infected rats. The tests were performed as
described previously (20).
Histology and immunohistochemistry.
At different time points
after infection, brain samples were obtained and immediately either
frozen in isopentane at In situ hybridization.
Digoxigenin-labeled RNA probes
complementary to BDV nucleoprotein p40, phosphoprotein p24, or matrix
protein gp18 mRNAs were used. Brains from experimental animals were
frozen in isopentane to Isolation of effector cells.
Lymphocytes from the brain were
isolated by a method described previously (23). Briefly,
animals were anesthetized and perfused with balanced salt solution. The
brain was homogenized carefully through a stainless steel mesh and
collected in balanced salt solution containing 0.05% collagenase D,
0.1 µg of trypsin inhibitor per ml, 10 µg of DNase I per ml, and 10 mM HEPES. The cell suspension was stirred at room temperature for
1 h and allowed to settle for 25 min. The supernatant was pelleted
at 200 × g for 10 min and resuspended in 10 ml of Ca-
and Mg-free PBS. Then 5 ml of the suspension was layered on top of 10 ml of a modified RPMI-Ficoll gradient and centrifuged at 500 × g for 30 min. The pellet containing the lymphocytes was
resuspended in Iscove modified Dulbecco medium with 2% fetal calf
serum and cultivated overnight at 37°C. Finally, the cells were
counted for further use as effectors in cytotoxicity assays.
51Cr release assay.
Lymphocytes isolated from
the brains of BDV-infected rats were tested at different time points
after the infection in a cytotoxicity assay with BDV-infected or
recombinant VV-infected target cells. The test was performed as
described previously (27). The synthetic peptide
ASYAQMTTY was dissolved in dimethyl sulfoxide, and target cells were labeled as described previously (25).
HD virus from different sources prevents BD.
HD-attenuated BDV
from MDCK cells protects against Borna disease (21). To
define whether this is due to virus properties after passages in tissue
culture or to a protective antiviral immune response to BDV, we
investigated the pathogenic potential of different doses of BDV after
i.c. inoculation in rats. BDV was obtained either from rat brain
(BDV-BR; 104 and 102 FFU) or from
MDCK (BDV-MDCK; 106, 104, and
102 FFU) or CRL1405 (BDV-CRL; 106,
104, and 102 FFU) cells. An infectious dose of
106 BDV-BR was not applicable simply due to
insufficiently high titers in the brain.
0022-538X/01/$04.00+0 DOI: 10.1128/JVI.75.23.11700-11708.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
High-Dose Borna Disease Virus Infection Induces a
Nucleoprotein-Specific Cytotoxic T-Lymphocyte Response and
Prevention of Immunopathology
ABSTRACT
Top
Abstract
Introduction
Materials and Methods
Results
Discussion
References
INTRODUCTION
Top
Abstract
Introduction
Materials and Methods
Results
Discussion
References
MATERIALS AND METHODS
Top
Abstract
Introduction
Materials and Methods
Results
Discussion
References
150°C or fixed in buffered formalin.
Cryostat sections were fixed in isopropanol. All tissue sections were
stained with hematoxylin-eosin. Encephalitic infiltrates were scored on
an arbitrary scale ranging from 0 to 3 based on the number of
infiltrates per section and the number of cell layers in each
infiltrate (score 1, up to 5 small infiltrates per section; score 2, more than 5 small infiltrates per section or more than 3 infiltrates
with multiple layers; score 3, more than 10 small infiltrates per
section or more than 5 infiltrates with multiple layers).
Immunohistochemistry was carried out on cryostat sections for the
presence of BDV-specific antigen, using an anti-nucleoprotein-specific
monoclonal antibody (38/17C1) (36).
150°C. Sections (5 µm) were mounted on
slides and fixed in 4% formaldehyde-PBS. After treatment with 0.1 N
HCl and acetic acid, hybridization was carried out overnight at
65°C with 20 ng of probe per slide. The slides were washed with 5×
SSC (1× SSC is 0.15 M NaCl plus 0.015 M sodium citrate) followed by
2× SSC, both at 56°C. They were then incubated with an alkaline
phosphatase-labeled anti-digoxigenin antibody and then placed overnight
in a 5-bromo-4-chloro-3-indolylphosphate-Nitro Blue Tetrazolium
(BCIP/NBT) solution.
RESULTS
Top
Abstract
Introduction
Materials and Methods
Results
Discussion
References
View larger version (31K):
[in a new window]
FIG. 1.
Appearance of BDV-specific clinical symptoms. The health
status of six rats was monitored daily, and disease symptoms were
scored as described in Materials and Methods. (A and B) The rats were
infected with 102 FFU (A) or 104 FFU (B) of
either BDV-BR ( 
), BDV-CRL (
), or
BDV-MDCK (). (C) For infection with 106 FFU,
only BDV-CRL and BDV-MDCK were used. After
infection with 104 FFU (B) of BDV-MDCK, two
animals developed severe BD () while four animals showed no clinical
symptoms
(
).
Early presence of BDV-specific antibodies after infection with
106 FFU of BDV.
The BDV-specific antibody response was
monitored every third day. As described previously (19),
the first nucleoprotein (p40)-specific antibodies were found around day
15 postinfection (p.i.) and phosphoprotein (p24)-specific antibodies
were found around day 18 p.i. in rats infected with
104 FFU of BDV-BR or BDV-CRL (Fig.
2B). Infection with 102 FFU
of BDV-BR or BDV-CRL resulted in a delayed
production of BDV-specific antibodies (Fig. 2A).
|
|
Detection of antigen, nucleic acid, and infectious virus. After infection with 102 or 104 FFU of BDV-BR or BDV-CRL, Western blot analysis revealed the presence of the nucleoprotein in brain homogenates by day 14 or 9 p.i. and the phosphoprotein by day 16 or 12 p.i., respectively (data not shown). Infectious virus could be isolated from the brains of rats infected with 104 FFU with a titer of 5.7 log10 units around day 10 p.i., reaching maximal titers of 6.3 log10 units on day 21. Similar titers of infectious virus were found when 102 FFU was used for infection; however, there was a delay of 4 to 5 days. BDV-specific nucleic acid and BDV-specific nucleoprotein could be demonstrated in the cortex and hippocampus of infected rats using in situ hybridization and immunohistochemistry (data not shown).
Infection with 104 FFU of BDV-MDCK resulted in the presence of viral antigen and infectious virus on day 18 p.i.; on day 37 p.i., however, little antigen and no infectious virus were found. In situ hybridization revealed the presence of mRNA for the nucleoprotein p40, the phosphoprotein p24, and the matrix protein gp18 in the hippocampus but not in the neocortex. After infection with 102 FFU of BDV-MDCK, no viral antigen or infectious virus was detectable before day 37 p.i. Thereafter, BDV-specific antigen could be detected by Western blotting and an infectious titer of 6.8 log10 units was found in the brain (day 37). In situ hybridization confirmed these results by detecting mRNA for the BDV-specific proteins p40, p24, and gp18 (data not shown). When infection was carried out with HD BDV-MDCK or BDV-CRL, either no or only traces of nucleic acid and antigen and no infectious virus were detectable on day 6 p.i.. By day 12, viral antigen, but no infectious virus, was found in HD BDV-MDCK-infected animals. By day 18, only single BDV-infected cells could be found by immunohistochemistry and in situ hybridization in rats infected with HD BDV-MDCK (Fig. 3A and B), whereas neither viral antigens nor infectious virus, with one exception, could be detected by Western blot analysis and virus titer determination (Table 2). In contrast, significant amounts of nucleic acid, antigen, and infectious BDV were detectable in the brains of HD BDV-CRL-infected rats (Table 2). By day 28, no nucleic acid or antigen was found in HD BDV-MDCK-infected rats, whereas nucleic acid and antigen were found in the cortex in HD BDV-CRL-infected rats, although at reduced levels (Fig. 3C and D). In HD BDV-CRL-infected animals, antigen could be detected in the hippocampus by immunohistochemistry and nucleic acid could be detected by in situ hybridization at levels comparable to those in control animals (Fig. 3E and F). At later time points (day 37), the viral titer was significantly reduced in the brains of HD BDV-CRL-infected rats and no antigen or infectious virus was found in the brains of HD BDV-MDCK-infected rats (Table 2).
|
|
Outcome of HD BDV infection in immunosuppressed rats.
To
investigate whether the immune system is responsible for the absence of
BD symptoms in rats infected with HD BDV-CRL or HD
BDV-MDCK, immunosuppressed animals were infected. For
immunosuppression, animals were thymectomized as adults and then
treated with anti-CD8-specific antibodies to deplete CD8+ T
cells. Other rats were thymectomized as newborns and infected as
adults. Successful thymectomy and T-cell depletion were confirmed by FACS analysis (data not shown). As shown in Table
3, no clinical disease symptoms or
encephalomyelitis could be detected in thymectomized rats depleted of
CD8+ T cells and infected with HD BDV-CRL or HD
BDV-MDCK, but viral antigen and infectious virus were
present in the brain. Comparably, rats thymectomized as newborns and
infected with the same viruses did not show any clinical signs and both
BDV-specific antigen and infectious virus were detectable in the brain.
However, in contrast to animals thymectomized as adults and infected
with HD BDV-MDCK, no antiviral antibodies were found in
those thymectomized as newborns. Interestingly, in those thymectomized
as newborns, infectious titers were higher in the brains after
infection with HD BDV-CRL than in those after infection with
HD BDV-MDCK.
|
Presence of BDV-specific cytotoxic T cells.
In
anti-CD8-treated rats infected with HD BDV, high titers of virus were
found in the brains. Therefore, the cytolytic activity of
CD8+ T cells isolated from the brains of rats infected with
HD BDV-MDCK or HD BDV-CRL was compared to the
activity of cells isolated from the brains of rats infected with
104 FFU of BDV-BR on day 9 p.i. As target
cells, syngeneic fibroblasts from Lewis rats infected with BDV were
used. As shown in Fig. 4A, lymphocytes
isolated from the brains of rats infected with BDV-BR showed
no cytolytic activity, correlating with previous results showing that
the cytolytic activity of CD8+ T cells isolated from the
brains of BDV-BR-infected rats starts around day 18 (20). In contrast, in rats infected with HD
BDV-MDCK or HD BDV-CRL cytolysis of target cells
was found as early as day 9 after infection (Fig. 4A).
|
| |
DISCUSSION |
|---|
|
Top
Abstract
Introduction
Materials and Methods
Results
Discussion
References
|
|---|
In a previous publication, Oldach et al. (21) reported on virus control and protection against disease after infection of rats with HD BDV from tissue culture. In the present study, we have identified the underlying mechanisms as being immunological in nature, demonstrating that a persistent virus infection can be abrogated after infection with a strong trigger for an efficient cellular antiviral immune response by high-input virus titers. As well as an early and strong cell-mediated immune reaction, an early humoral immune response was observed. In this study we used virus obtained from different cell types (BDV-CRL and BDV-MDCK) or from the brains of BDV-infected rats (BDV-BR) to assess whether characteristics of particular virus isolates, e.g., adaptation to tissue culture, contribute to control of the virus from the host or whether this effect is simply triggered by the virus and, in fact, is dependent on the efficiency of the antiviral immune response.
Infection with 102 FFU of BDV-CRL, BDV-BR, or BDV-MDCK in general resulted in late onset of disease. This finding indicates that infection even with low doses of BDV from different preparations causes disease with a comparable clinical picture.
Similarly, using the standard dose (104 FFU) for infection, we were able to demonstrate that BDV-CRL infection results in BD in a comparable time range, with comparable symptoms and severity of disease, as well as onset, quality, and antibody titers in BDV-CRL- and BDV-BR-infected rats. This indicates that BDV obtained from CRL1405 cells (BDV-CRL) and BDV harbored in the brain (BDV-BR) have very similar biological properties as far as the above-mentioned parameters are concerned. In contrast, BDV obtained from persistently infected MDCK cells (BDV-MDCK) and used for infection at 104 FFU apparently differed from BDV-CRL and BDV-BR insofar as only one-third of the animals developed disease within the observation period while the others stayed healthy. These results show that 102 FFU of BDV-MDCK induces disease while infection with 104 FFU apparently represents a breaking point, where some animals develop disease and others control the virus and do not develop BD. This finding also indicates differences among BDV isolates that possibly depend on their passage history in vitro; BDV has been cultured in MDCK cells for more than 20 years (12), whereas it has been passaged in CRL1405 cells only for a short time (9). Infection of rats with HD BDV-CRL or BDV-MDCK (106 FFU) resulted in an early induction of the humoral immune response. The detection of viral antigen and nucleic acid in the hippocampus of BDV-MDCK-infected rats as well as in the cortex and the hippocampus of BDV-CRL-infected rats points to a productive viral infection. Since in both BDV-MDCK- and BDV-CRL-infected rats only traces of virus were detectable, this demonstrates that the infection is controlled, although gradually differently. This is further demonstrated by the detection of infectious virus in BDV-CRL-infected rats but not in BDV-MDCK-infected rats. Furthermore, the quantity of antigen detected in the brains of BDV-MDCK-infected rats was below the detection limit of immunoblotting but antigen was easily detectable in BDV-CRL-infected rats. Encephalitic reactions were seen in the hippocampus of all BDV-CRL-infected rats but only one BDV-MDCK-infected rat. Finally, we injected rats with a low dose of BDV-CRL i.c. and at the same time with HD BDV-CRL subcutaneously and/or intravenously. Unfortunately, rats that were only infected with HD BDV-CRL subcutaneously and/or intravenously developed disease (data not shown). Therefore, this experiment was not suitable to investigate the balance between the local infection in the brain and the induction of a systemic immune response. Control of a virus infection usually is due to the activity of the humoral and/or cellular arm of the immune system. Only recently has it been demonstrated that neutralizing glycoprotein-specific antibodies can have a prophylactic effect in experimental BD (9). However, apart from the early induction of nonneutralizing antibodies against the nucleoprotein and the phosphoprotein, no neutralizing antibodies were found in sera from BDV-CRL- or BDV-MDCK-infected rats. Therefore, and in agreement with previous findings which underline the role of the cellular immune system in both immunopathology and immunoprotection in the experimental model of BD in rats (reviewed in references 3, 33, and 35), we investigated the cellular immune response in more detail.
In BD, T cells represent the major pathway of immunopathology; CD4+ and, particularly, CD8+ T cells are involved in severe encephalitis and the ensuing degenerative encephalopathy (2, 10, 23, 24, 29). On the other hand, immunoprotection is mediated by T cells (28), where CD8+ cytotoxic T cells were again demonstrated as the relevant effector cells (20). Therefore, on the basis of having demonstrated that HD BDV-MDCK-infected rats do not show disease and do not have neutralizing antibodies, we scrutinized the impact of the cellular immune response in the HD phenomenon. Since CD8+ T cells are the most likely effector cells after BDV infection, we concentrated on their role by using different strategies. Rats thymectomized at birth and tested for the absence of T cells in the peripheral blood by flow cytometry did not show disease symptoms or antibody synthesis, whether infected with HD BDV-MDCK or HD BDV-CRL. However, whereas untreated BDV-MDCK-infected rats showed only a very limited presence of viral antigen in the brain and had no detectable amounts of virus, thymectomized rats had high virus titers in the brain. This finding was even more pronounced in immunocompromised HD BDV-CRL-infected rats, which had an extraordinary high virus titer in their brains.
To further define the mechanism of the cellular immune response that is operative in controlling the virus, adult thymectomized rats were infected with virus after they had been treated with an antibody against CD8+ T cells. Neither HD BDV-MDCK- nor HD BDV-CRL-infected rats showed disease or encephalitis, but, again, high virus titers were found in their brains. This result clearly demonstrated that the presence of CD8+ T cells is indispensable and sufficient for virus control in HD BDV-infected rats.
The result of cytotoxicity assays convincingly showed that HD BDV-CRL induces a BDV-specific CD8+ T-cell response as early as day 9 p.i. and that BDV-MDCK triggers a substantial cytotoxic T-cell response, whereas BDV-BR does not cause the lysis of target cells at this early time point. Moreover, we directly demonstrated the presence of specific cytotoxic T cells directed against the nucleoprotein by the use of recombinant VV (27). Finally, a naturally processed Lewis rat major histocompatibility complex class I-associated peptide, ASYAQMTTY, of the BDV nucleoprotein had been identified previously as the relevant target structure (25). These present and earlier results show that a fast and strong immune response directed against the peptide ASYAQMTTY is necessary for the control of BDV and prevention of disease. These data also show that HD BDV does not cause cytotoxic T-lymphocyte exhaustion as has been demonstrated in lymphocytic choriomeningitis virus infection with the consequence of a persistent infection (17). The role of CD8+ T-cell-derived cytokines has not been addressed in this paper in association with protection from disease. These studies are extremely difficult to perform since only very few, if any, of the cytokines produced during an antiviral immune response are uniquely produced by CD8+ T cells.
Our data support previous results observed after infection with other noncytopathic viruses. In lymphocytic choriomeningitis virus infection of mice and hepatitis B virus infection in humans, damage of virus-infected CNS or liver cells is caused by cytotoxic T cells (7, 22, 40). If the host immune response is efficient and quick, the replication and, more important, the spread of the virus can be controlled very early after infection. Therefore, tissue damage is limited, virus is eliminated, and functions are restored within a few weeks after infection (22, 40). If there is no immune response, the virus will spread. Since the virus is noncytopathic, the infection does not cause any damage and the host will become a virus carrier. If there is a low and slow T-cell response, the virus can infect many cells, and as a consequence of the delayed immune response, severe tissue damage occurs due to immunopathology (reviewed in reference 39). This finding is comparable to the situation after BDV infection of rats. If there is no T-cell response, HD BDV-infected rats develop a carrier status. If the T-cell response is excellent, CD8+ T cells prevent a disseminated viral encephalitis and can even control the virus and prevent immunopathology. If the immune response is delayed after BDV infection, the animals develop encephalitis and severe brain cell destruction.
The present work clearly demonstrates that BDV infection does not necessarily result in immunopathology and underlines the role of CD8+ T cells in mediating and preventing disease. Nevertheless, even though it seems unlikely that antibodies, e.g., neutralizing antibodies directed against the glycoprotein (9), play a major role in mediating and preventing disease, they cannot be excluded because they might function at a level that is below detection.
Our data support the hypothesis that CD8+ T-cell-mediated lysis mechanisms are required for immunity to nonlytic viruses. It is not known how deeply gamma interferon, as a product of CD4+ or CD8+ T cells, is involved in the clearance of persistent BDV infection of the brain since cytokine-deficient rats are not available. Therefore, at present the only valid explanation for the control of BDV infection in the brain involves the presence and action of CD8+ T cells, although the question cannot be answered whether these cells exert a noncytolytic mechanism that controls the viral genome with only little tissue damage. On the other hand, a very early and potent cytolytic T-cell response that is capable of controlling virus-infected cells in the brain before the virus disseminates would be suited to limit cellular destruction to only very few cells, resulting in no or only transient and slight disturbances of organ functions.
| |
ACKNOWLEDGMENTS |
|---|
The work was supported in part by Deutsche Forschungsgemeinschaft grants Sti 71/2-2 (to L.S. and O.P.) and Pla 256/1-1 (to O.P. and L.S.) and by the European Union (Pathogenesis of Subacute and Chronic Inflammatory Diseases of the Central Nervous System, grant CHRX-CT94-0670). E.F. is a recipient of a grant from the Schweizer Nationalfonds (SNF) (83EU-048814).
| |
FOOTNOTES |
|---|
* Corresponding author. Mailing address: Institut für Immunologie, Bundesforschungsanstalt für Viruskrankheiten der Tiere, Paul Ehrlich Str. 28, 72076 Tübingen, Germany. Phone: 49 7071 967 254. Fax: 49 7071 967 105. E-mail: oliver.planz{at}tue.bfav.de.
Dedicated to Rudolf Rott on the occasion of his 75th birthday.
| |
REFERENCES |
|---|
|
Top
Abstract
Introduction
Materials and Methods
Results
Discussion
References
|
|---|
| 1. |
Amsterdam, J. D.,
A. Winokur,
W. Dyson,
S. Herzog,
F. Gonzalez,
R. Rott, and H. Koprowski.
1985.
Borna disease virus. A possible etiologic factor in human affective disorders?
Arch. Gen. Psychiatry
42:1093-1096 |
| 2. | Bilzer, T., and L. Stitz. 1994. Immune-mediated brain atrophy. CD8+ T cells contribute to tissue destruction during borna disease. J. Immunol. 153:818-823[Abstract]. |
| 3. | Bilzer, T., and L. Stitz. 1996. Immunopathogenesis of virus diseases affecting the central nervous system. Crit. Rev. Immunol. 16:145-222[Medline]. |
| 4. | Bode, L., R. Durrwald, and H. Ludwig. 1994. Borna virus infections in cattle associated with fatal neurological disease. Vet. Rec. 135:283-284[Medline]. |
| 5. | Bode, L., W. Zimmermann, R. Ferszt, F. Steinbach, and H. Ludwig. 1995. Borna disease virus genome transcribed and expressed in psychiatric patients. Nat. Med. 1:232-236[CrossRef][Medline]. |
| 6. | Caplazi, P., A. Waldvogel, L. Stitz, U. Braun, and F. Ehrensperger. 1994. Borna disease in naturally infected cattle. J. Comp. Pathol. 111:65-72[CrossRef][Medline]. |
| 7. | Cole, G. A., N. Nathanson, and R. A. Prendergast. 1972. Requirement for theta-bearing cells in lymphocytic choriomeningitis virus-induced central nervous system disease. Nature 238:335-337[CrossRef][Medline]. |
| 8. | Deschl, U., L. Stitz, S. Herzog, K. Frese, and R. Rott. 1990. Determination of immune cells and expression of major histocompatibility complex class II antigen in encephalitic lesions of experimental Borna disease. Acta Neuropathol. 81:41-50[CrossRef][Medline]. |
| 9. |
Furrer, E.,
T. Bilzer,
L. Stitz, and O. Planz.
2001.
Neutralizing antibodies in persistent Borna disease virus infection: prophylactic effect of gp94-specific monoclonal antibodies in preventing encephalitis.
J. Virol.
75:943-951 |
| 10. | Hatalski, C. G., W. F. Hickey, and W. I. Lipkin. 1998. Evolution of the immune response in the central nervous system following infection with Borna disease virus. J. Neuroimmunol. 90:137-142[CrossRef][Medline]. |
| 11. | Hatalski, C. G., W. F. Hickey, and W. I. Lipkin. 1998. Humoral immunity in the central nervous system of Lewis rats infected with Borna disease virus. J. Neuroimmunol. 90:128-136[CrossRef][Medline]. |
| 12. | Herzog, S., and R. Rott. 1980. Replication of Borna disease virus in cell cultures. Med. Microbiol. Immunol. (Berlin) 168:153-158[CrossRef][Medline]. |
| 13. | Kishi, M., Y. Arimura, K. Ikuta, Y. Shoya, P. K. Lai, and M. Kakinuma. 1996. Sequence variability of Borna disease virus open reading frame II found in human peripheral blood mononuclear cells. J. Virol. 70:635-640[Abstract]. |
| 14. | Ludwig, H., and P. Thein. 1977. Demonstration of specific antibodies in the central nervous system of horses naturally infected with Borna disease virus. Med. Microbiol. Immunol. 163:215-226[CrossRef][Medline]. |
| 15. | Lundgren, A. L., R. Lindberg, H. Ludwig, and G. Gosztonyi. 1995. Immunoreactivity of the central nervous system in cats with a Borna disease-like meningoencephalomyelitis (staggering disease). Acta Neuropathol. (Berlin) 90:184-193[Medline]. |
| 16. | Malkinson, M., Y. Weisman, S. Perl, and E. Ashash. 1995. A Borna-like disease of ostriches in Israel. Curr. Top. Microbiol. Immunol. 190:31-38[Medline]. |
| 17. | Moskophidis, D., F. Lechner, H. Pircher, and R. M. Zinkernagel. 1993. Virus persistence in acutely infected immunocompetent mice by exhaustion of antiviral cytotoxic effector T cells. Nature 362:758-761[CrossRef][Medline]. |
| 18. |
Nakamura, Y.,
H. Takahashi,
Y. Shoya,
T. Nakaya,
M. Watanabe,
K. Tomonaga,
K. Iwahashi,
K. Ameno,
N. Momiyama,
H. Taniyama,
T. Sata,
T. Kurata,
J. C. de la Torre, and K. Ikuta.
2000.
Isolation of Borna disease virus from human brain tissue.
J. Virol.
74:4601-4611 |
| 19. | Narayan, O., S. Herzog, K. Frese, H. Scheefers, and R. Rott. 1983. Pathogenesis of Borna disease in rats: immune-mediated viral ophthalmoencephalopathy causing blindness and behavioral abnormalities. J. Infect. Dis. 148:305-315[Medline]. |
| 20. |
Nöske, K.,
T. Bilzer,
O. Planz, and L. Stitz.
1998.
Virus-specific CD4+ T cells eliminate Borna disease virus from the brain via induction of cytotoxic CD8+ T cells.
J. Virol.
72:4387-4395 |
| 21. | Oldach, D., M. C. Zink, J. M. Pyper, S. Herzog, R. Rott, O. Narayan, and J. E. Clements. 1995. Induction of protection against Borna disease by inoculation with high-dose-attenuated Borna disease virus. Virology 206:426-434[CrossRef][Medline]. |
| 22. | Peters, M., J. Vierling, M. E. Gershwin, D. Milich, F. V. Chisari, and J. H. Hoofnagle. 1991. Immunology and the liver. Hepatology 13:977-994[CrossRef][Medline]. |
| 23. |
Planz, O.,
T. Bilzer,
M. Sobbe, and L. Stitz.
1993.
Lysis of major histocompatibility complex class I-bearing cells in Borna disease virus-induced degenerative encephalopathy.
J. Exp. Med.
178:163-174 |
| 24. | Planz, O., T. Bilzer, and L. Stitz. 1995. Immunopathogenic role of T-cell subsets in Borna disease virus-induced progressive encephalitis. J. Virol. 69:896-903[Abstract]. |
| 25. |
Planz, O.,
T. Dumrese,
S. Hulpusch,
M. Schirle,
S. Stevanovic, and L. Stitz.
2001.
A naturally processed rat major histocompatibility complex class I- associated viral peptide as target structure of Borna disease virus-specific CD8+ T cells.
J. Biol. Chem.
276:13689-13694 |
| 26. | Planz, O., C. Rentzsch, A. Batra, H.-J. Rziha, and L. Stitz. 1998. Persistence of Borna disease virus-specific nucleic acid in the blood of a psychiatric patient. Lancet 352:623-623[Medline]. |
| 27. |
Planz, O., and L. Stitz.
1999.
Borna disease virus nucleoprotein (p40) is a major target for CD8+-T-cell-mediated immune response.
J. Virol.
73:1715-1718 |
| 28. |
Richt, J. A.,
A. Schmeel,
K. Frese,
K. M. Carbone,
O. Narayan, and R. Rott.
1994.
Borna disease virus-specific T cells protect against or cause immunopathological Borna disease.
J. Exp. Med.
179:1467-1473 |
| 29. |
Richt, J. A.,
L. Stitz,
H. Wekerle, and R. Rott.
1989.
Borna disease, a progressive meningoencephalomyelitis as a model for CD4+ T cell-mediated immunopathology in the brain.
J. Exp. Med.
170:1045-1050 |
| 30. | Rott, R., and H. Becht. 1995. Natural and experimental Borna disease in animals. Curr. Top. Microbiol. Immunol. 190:17-30[Medline]. |
| 31. |
Rott, R.,
S. Herzog,
B. Fleischer,
A. Winokur,
J. Amsterdam,
W. Dyson, and H. Koprowski.
1985.
Detection of serum antibodies to Borna disease virus in patients with psychiatric disorders.
Science
228:755-756 |
| 32. | Sobbe, M., T. Bilzer, S. Gommel, K. Noske, O. Planz, and L. Stitz. 1997. Induction of degenerative brain lesions after adoptive transfer of brain lymphocytes from Borna disease virus-infected rats: presence of CD8+ T cells and perforin mRNA. J. Virol. 71:2400-2407[Abstract]. |
| 33. | Stitz, L., B. Dietzschold, and K. M. Carbone. 1995. Immunopathogenesis of Borna disease. Curr. Top. Microbiol. Immunol. 190:75-92[Medline]. |
| 34. |
Stitz, L.,
K. Noske,
O. Planz,
E. Furrer,
W. I. Lipkin, and T. Bilzer.
1998.
A functional role for neutralizing antibodies in Borna disease: influence on virus tropism outside the central nervous system.
J. Virol.
72:8884-8892 |
| 35. | Stitz, L., and R. Rott. 1999. Borna disease virus (Bornaviridae), p. 167-173. In A. Granoff, and R. G. Webster (ed.), Encyclopedia of virology. Academic Press, Inc., San Diego, Calif. |
| 36. |
Thiedemann, N.,
P. Presek,
R. Rott, and L. Stitz.
1992.
Antigenic relationship and further characterization of two major Borna disease virus-specific proteins.
J. Gen. Virol.
73:1057-1064 |
| 37. | Waltrip, R. W., Jr., R. W. Buchanan, W. T. Carpenter, Jr., B. Kirkpatrick, A. Summerfelt, A. Breier, S. A. Rubin, and K. M. Carbone. 1997. Borna disease virus antibodies and the deficit syndrome of schizophrenia. Schizophr. Res. 23:253-257[CrossRef][Medline]. |
| 38. |
Weissenbock, H.,
N. Nowotny,
P. Caplazi,
J. Kolodziejek, and F. Ehrensperger.
1998.
Borna disease in a dog with lethal meningoencephalitis.
J. Clin. Microbiol.
36:2127-2130 |
| 39. | Zinkernagel, R. M. 1997. Virus-induced immunopathology, p. 163-179. In N. Nathanson (ed.), Viral pathogenesis. Lippincott-Raven, Philadelphia, Pa. |
| 40. |
Zinkernagel, R. M.,
E. Haenseler,
T. Leist,
A. Cerny,
H. Hengartner, and A. Althage.
1986.
T cell-mediated hepatitis in mice infected with lymphocytic choriomeningitis virus. Liver cell destruction by H-2 class I-restricted virus-specific cytotoxic T cells as a physiological correlate of the 51Cr-release assay?
J. Exp. Med.
164:1075-1092 |
Journal of Virology, December 2001, p. 11700-11708, Vol. 75, No. 23
0022-538X/01/$04.00+0 DOI: 10.1128/JVI.75.23.11700-11708.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
This article has been cited by other articles:
-
Perez, M., Clemente, R., Robison, C. S., Jeetendra, E., Jayakar, H. R., Whitt, M. A., de la Torre, J. C.
(2007). Generation and Characterization of a Recombinant Vesicular Stomatitis Virus Expressing the Glycoprotein of Borna Disease Virus. J. Virol.
81: 5527-5536
[Abstract] [Full Text] -
Hausmann, J., Pagenstecher, A., Baur, K., Richter, K., Rziha, H.-J., Staeheli, P.
(2005). CD8 T Cells Require Gamma Interferon To Clear Borna Disease Virus from the Brain and Prevent Immune System-Mediated Neuronal Damage. J. Virol.
79: 13509-13518
[Abstract] [Full Text] -
Bourteele, S., Oesterle, K., Pleschka, S., Unterstab, G., Ehrhardt, C., Wolff, T., Ludwig, S., Planz, O.
(2005). Constitutive Activation of the Transcription Factor NF-{kappa}B Results in Impaired Borna Disease Virus Replication. J. Virol.
79: 6043-6051
[Abstract] [Full Text] -
Hashimoto, Y., Chen, H.-S., Cunningham, C., Malik, T. H., Lai, P. K.
(2003). Two Major Histocompatibility Complex Class I-Restricted Epitopes of the Borna Disease Virus p10 Protein Identified by Cytotoxic T Lymphocytes Induced by DNA-Based Immunization. J. Virol.
77: 6076-6081
[Abstract] [Full Text]
| |||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
Copyright © 2009 by the American Society for Microbiology. For an alternate route to Journals.ASM.org, visit: http://intl-journals.asm.org | More Info»