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1 Protects Mice from Acute Ocular Herpes Simplex Virus Type 1 Infection1
Department of Microbiology, Immunology, and Parasitology, Louisiana State University Medical Center, New Orleans, LA 70112; and Department of Neuropharmacology, The Scripps Research Institute, La Jolla, CA 92037
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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and IFN-ß) play a key role in the
host’s innate defense against viral pathogens. To examine the biologic
relevance of IFN- to a viral pathogen within the confines of the
nervous system, IFN-1 transgenic mice whose transgene is under the
control of the glial fibrillary acidic protein promoter (GFAP-IFN-,
astrocyte specific) were examined for resistance to an ocular herpes
simplex virus type 1 (HSV-1) infection. GFAP-IFN- mice expressed
significantly higher levels of IFN-ß (533 U) in the trigeminal
ganglion compared with nontransgenic mice (70 U) 72 h
postinfection that corresponded with a significant reduction in the
mRNA expression of the HSV-1 immediate early gene infected cell
polypeptide 27 and late gene VP16, as well as the chemokines
monocyte-chemoattractant protein-1 and cytokine response gene-2 in the
eye and trigeminal ganglion. Six days postinfection, the viral load and
the expression of infected cell polypeptide 27, CD8, RANTES, IFN-
,
and IFN- mRNA levels were reduced in the trigeminal ganglion of
GFAP-IFN- mice compared with the wild-type mice. Following the
establishment of HSV-1 latency (i.e., 30 days postinfection), only one
of nine (11%) GFAP-IFN- mice was found to be latent compared with
seven of eight (88%) of the wild-type mice, as determined by the
expression of the latency-associated transcript RNAs. Likewise, only
three of nine GFAP-IFN- mice screened showed seroconversion by day
30 postinfection compared with nine of ten wild-type mice screened.
Collectively, the results show that the IFN-1 transgenic mice are
less susceptible to acute HSV-1 infection and the establishment of
viral latency. |
Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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The mode of transmission, down-regulation of MHC class II expression (13), establishment of latency, and subsequent reactivation are means by which HSV-1 has eluded immune surveillance and achieved remarkable success (14). Most morbidity associated with HSV infection is correlated with the repeated reactivation of latent infection throughout the lifetime of the host. Twenty percent of the U.S. population experience periodic vesicular HSV lesions in the oral pharyngeal region, with the incidence of ocular herpes making up an important subset of HSV-1 disease (15, 16). Reducing or preventing viral reactivation by blocking the establishment of latency or the environmental or biochemical cues that elicit reactivation might facilitate the success of keratoplasties in patients plagued by herpetic eye disease.
Mice receiving HSV-1 Ag 5 days before infection are shown to be
protected from HSV-1-mediated encephalitis, having a reduced viral load
in the TG compared with passively immunized animals (17, 18). However,
the administration of IFN-ß-neutralizing Ab increases the viral
yield in the TG, suggesting that the protective effect of active
immunization is, in part, due to the type I IFNs. To further
investigate the supportive role of type I IFN in ocular HSV-1
infection, a transgenic mouse model expressing IFN-1 under the
control of the glial fibrillary acidic protein (GFAP) promoter was
employed. By constitutive expression of IFN- within the central
nervous system (CNS), it would be possible to determine whether such
expression antagonized HSV-1 replication, retrograde transport, and the
establishment of latency, as well as modify the host’s immune response
to the acute viral infection. The establishment of an antiviral state
mediated by IFN- would indicate IFN- as a possible therapeutic
for ocular/CNS HSV infection.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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The methods for the construction of the GFAP-IFN-1 fusion
gene and the generation and screening of transgenic mice were similar
to those described previously for the generation of GFAP-IL-6
transgenic mice (19). Briefly, a murine IFN- cDNA (0.65 kb)
containing the entire coding region of IFN-1 was inserted between an
upstream SV40 intron and a downstream SV40 poly(A) signal sequence, at
the SalI site within the first exon of the modified GFAP
expression vector. The GFAP-IFN- transgene was microinjected into
the germline of C57BL/6J x BALB/c F1 hybrid mice, and
transgenic offspring were screened for integration by tail DNA dot-blot
hybridization using a 32P-labeled SV40 poly(A) sequence
fragment as probe. Low-expressing GFAP-IFN- transgenic mice were
used as the experimental group compared with the wild-type mice
(C57BL/6J x BALB/c) used as controls.
Virus and cells
L cells and CV-1 African monkey kidney cells (American Type Culture Collection (ATCC), Manassas, VA) were cultured in RPMI 1640 (Mediatech, Washington, D.C.) containing 5% FCS (Life Technologies, Gaithersburg, MD) and an antibiotic/antimycotic solution (Sigma, St. Louis, MO). Cells were incubated at 37°C, 5% CO2, 95% humidity. HSV-1 (McKrae strain) was grown up and harvested as described (10).
Infection of mice
IFN- transgenic and wild-type mice were anesthetized by i.p.
administration of 0.1 ml PBS containing xylazine (6.6 mg/kg) and
ketamine (100 mg/kg). Following corneal scarification, tear film was
blotted from the eyes and the mice were inoculated with 600
plaque-forming units (PFU)/eye of HSV-1 (McKrae strain) in a volume of
3 µl. Infection was verified by swabbing the eyes 2 to 3 days
postinfection (p.i.), placing the swabs in CV-1 monolayer cultures, and
observing the cells for cytopathic effects (CPE). Animals were
subsequently sacrificed at the indicated time p.i.
Measurement of tissue HSV-1 titers
Eyes, TG, and cerebella were removed 6 days p.i. and homogenized in 0.8 ml RMPI 1640 containing 5% FCS in 2-ml microcentrifuge tubes. Homogenates were clarified by centrifugation for 1 min at 13,000 x g. HSV-1 titer in clarified supernatants was determined by plaque assay.
RT-PCR
RT-PCR was conducted as described (10). Briefly, eye and TG RNA
from individual mice were extracted in Ultraspec RNA isolation reagent
(Biotecx, Houston, TX). First strand cDNA was synthesized using AMV
reverse transcriptase (Promega, Madison, WI). PCR was performed in a
thermal cycler (Ericomp 
cycler I; Ericomp, San Diego, CA) with 30
to 35 cycles of 94°C (1 min, 15 s), 57 to 65°C (1 to 1 min,
15 s), and 72°C (30–45 s). PCR primers for
glyceraldehyde-3-phosphate dehydrogenase (GAPDH), infected cell
polypeptide 27 (ICP27), IFN-, LAT, IL-10, and RANTES were as
previously described (10). IFN- (consensus sequence for IFN-1,
-2, and -7), CD4, and CD8 primer sequences were obtained from Clontech
Laboratories (Palo Alto, CA). Primers for IL-6 were
5'-TTCCATCCAGTTGCCTTCTTGG-3' (sense) and 5'-CTTCATGTACTCCAGGTAG-3'
(antisense). Primers for VP16 were 5'-GGACTCGTATTCCAGCTTCAC-3'
(sense) and 5'-CGTCCTCGCCGTCTAAGTG-3' (antisense). Primers for GFAP
were 5'-AAGCTCCAAGATGAAACCAACCTGA-3' (sense) and
5'-GCAAACTTAGACCGATACCACTC-3' (antisense). Primers specific for the
IFN-1 transgene were 5'-ATTCCCGCAGGAGAAGGTGGATGCCCCA-3' (sense)
and 5'-GAGTAGTTACATAGAATAGTACA-3' (antisense), based on the
published sequence for the murine IFN-1 cDNA sequence upstream
primer starting at nucleotide 550 (GenBank accession X01974) and the
downstream primer sequence starting at nucleotide 20 of the SV40 late
region polyadenylation sequence, which was used as a 3' UTR in the
GFAP-IFN-1 fusion gene construct (sequence according to Clontech
Laboratories). Primers for JE/monocyte-chemoattractant protein-1
(MCP-1) and macrophage-inhibitory protein-1ß, and the settings for
the amplification of the specific products were as described (20).
Primers for cytokine response gene (CRG)-2 were
5'-CAGCACCATGAACCCAAGTGC-3' (sense) and
5'-GCTGGTCACCTTTCAGAAGACC-3' (antisense). Following electrophoresis
of the amplified product, ethidium bromide-stained PCR products were
visualized with a Bio-Rad 1000 gel documentation system (Bio-Rad,
Hercules, CA). Densitometric analysis of gel images was performed using
molecular analysis software 3.3 (Bio-Rad).
IFN bioassay
Supernatants from homogenized eye and TG were subjected to
65°C for 20 min to inactivate any HSV-1 in the homogenate, as
previously described (18). The resultant supernatant was passed through
0.22-µm filters (to remove or destroy attenuated virus and virus
particles) and subsequently tested for IFN, as described (21). In the
neutralization experiment, Ab (neutralizing capacity = 2550 U) to
IFN-ß (Access Biomedical, San Diego, CA) or normal rabbit serum
was incubated with the supernatants for 60 min at room temperature
before the addition to L cell monolayers. The supernatant was removed
24 h later, and vesicular stomatitis virus (104
PFU/ml) was added to the monolayers and scored for CPE 48 h after
the addition of virus. One unit of IFN was defined as the reciprocal of
the dilution that blocked 50% CPE.
NK cell assay
Splenocytes from transgenic and wild-type mice infected 3 days previously with HSV-1 were teased from spleens using 20-gauge sterile needles through incisions made in the spleen cuticle in HBSS. RBC were osmotically lysed (0.84% NH4Cl), and the resultant population was placed in RPMI 1640 containing 10% FCS. NK cytolytic activity was determined as previously described (22).
ELISA measurement of anti-HSV-1 Ab titers
HSV-1 virions used as coating Ag were generated as previously described (10). Enzyme immunoassay 96-well plates (EIA; Costar, Cambridge, MA) were coated with HSV-1 virion protein (diluted in carbonate buffer) for 12 h at 4°C. Wells were washed three times with PBS and blocked with 250 µl of 0.5% dry milk dissolved in PBS for 1 h at 37°C. After one rinsing with PBS, duplicate 100-µl samples of diluted mouse serum (1/50 to 1/1600 in PBS) were added to HSV-1 Ag-coated wells and incubated for 1 h at 37°C. After six washings with PBS containing 0.5% Tween 20 (polyoxyethylene-20-sorbitan monolaurate), 100 µl of goat anti-mouse IgG heavy and light chain specific (diluted 1/1500 in PBS; Bio-Rad Laboratories, Richmond, CA), goat anti-mouse IgG1 (diluted 1/2000 in PBS; Caltag Laboratories, Burlingame, CA), or rat anti-mouse IgG2a (diluted 1/1000 in PBS; PharMingen, San Diego, CA) alkaline phosphatase-conjugated Abs were added to their respective well and incubated for 30 to 45 min at 37°C. After six washings in PBS/Tween 20, 100 µl of p-nitrophenyl phosphate (Sigma) was added to each well, and colorimetric development (OD405 nm) was measured in an ELISA plate reader (Bio-tek Instruments, Winooski, VT). To further control for the determination of specific anti-HSV-1 Ab, uninfected CV-1 lysates were used as coating Ag in replicate wells. The OD readings from these wells were subtracted from the HSV-1 Ag-coated wells. Preimmune sera were used to establish background readings to HSV-1 Ag. At a 1/50 dilution, the OD readings ranged from 0.013 to 0.052. A fourfold elevation in titer from preimmune sera was considered positive conversion.
DNA sequencing of mouse TG GFAP
To identify the nucleotide sequence of the GFAP-PCR-amplified
product, DNA template was purified using Qiaex gel extraction kit
(Qiagen, Santa Clarita, CA). Sequencing reactions were performed by end
labeling the DNA template with [-33P]ddNTP using the
Thermo sequence radiolabeled terminator cycle sequencing kit (Amersham
Life Sciences, Cleveland, OH). Twenty-microliter reaction mix was
prepared by combining 100 fmol DNA template, 1 pmol primer, 2 µl
reaction buffer, and 2 µl Thermo sequenase polymerase (4 U/µl).
Then 4.5 µl of the reaction mix was transferred to a termination mix
containing 2 µl dGTP termination mix and 0.5 µl 450 µCi/ml
[-33P]ddNTP. The PCR was performed in a Thermolyne
thermal cycler (Dubuque, IA) with 35 cycles of 95°C (30 s), 60°C
(30 s), and 72°C (60 s). Following the PCR reaction, 4 µl of stop
solution was added and mixed thoroughly to each of the termination
reactions. The samples were heated to 70°C for 2 to 10 min and loaded
(5 µl) immediately on 5% polyacrylamide gels containing 8 M urea.
The amplified DNA sequence was read and compared with known nucleotide
sequence by submission of query sequences to the BLAST at the National
Center for Biotechnology Information.
Statistics
One-way ANOVA and Scheffe multiple comparison test or Tukey’s post hoc t test were used to determine significant (p < 0.05) differences between the indicated groups using the GBSTAT program (Dynamic Microsystems, Silver Springs, MD).
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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transgenic mice
GFAP-IFN- transgenic and wild-type mice were ocularly infected
with HSV-1 (McKrae strain, 600 PFU/eye) and monitored for viral
recovery in the tear film. While all (18/18) of the wild-type and
majority (8/11) of GFAP-IFN- transgenic mice assessed for HSV-1 had
replicating virus in the eye following inoculation, the clearance rate
was rapid in the transgenic mice (Fig. 1
). In addition, the viral load in the
eye and TG was reduced significantly in the transgenic compared with
the wild-type mice at both day 3 and day 6 p.i. (Fig. 2). Ten percent (1/10) of the wild-type
mice had recoverable infectious virus in the cerebellum compared with 0
of 10 transgenic mice 6 days p.i. The poor recovery of HSV-1 in the eye
and TG of the GFAP-IFN- transgenic mice 3 and 6 days p.i. suggests
that even though the virus is capable of replicating in the eye early
in the course of infection (i.e., day 3 p.i.), the environment
quickly becomes antagonistic to the virus-blocking replication and
spread, as evidenced by the recovery of the virus at the latter time
point (i.e., day 6 p.i.) in the TG.
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transgenic mice, indicated by a reduction in cytokine and chemokine
expression
Cytokines such as IFN- and IFN- have been shown to
antagonize HSV-1 replication (23, 24). Since HSV-1 did not appear to
replicate to the same degree in the eye of the transgenic compared with
wild-type mice, cytokine (including chemokines) and viral transcript
expression were investigated over the course of the acute infection to
latency. There was a significant difference in the levels of expression
of CRG-2, but not other chemokine or cytokine transcripts assessed by
RT-PCR in the eye comparing the transgenic to wild-type mice on day
3 p.i. (Fig. 3, Table I). Likewise, there was a significant
reduction in the expression of HSV-1 ICP27 and VP16 in the eyes of the
GFAP-IFN- mice compared with the wild-type controls day 3 p.i.
(Fig. 3, Table I). However, there were no differences in the expression
of CD8 transcripts comparing the two groups. In comparison with the
eye, MCP-1 and CRG-2 mRNA expression were elevated significantly in the
wild-type TG compared with the GFAP-IFN- transgenic mice 3 days p.i.
(Fig. 4, Table II). Likewise, the viral lytic gene
transcript ICP27 and gene transcript VP16 were detected in the TG
of wild-type mice, but absent (ICP27) or significantly reduced (VP16)
in the TG of the transgenic animals. Again, no differences were evident
comparing CD8 transcript expression in the TG 3 days p.i. between the
two groups of animals. CD4, IFN-, and IL-10 transcript expression in
the eye and TG were rarely detected at these time points, with no
differences between the groups (data not shown).
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, and IFN- in the TG of wild-type mice
(Fig. 5, Table II
). However, there were
no significant differences in the expression of IL-6 or IL-10 comparing
the two groups of mice. Likewise, by day 30 p.i., there were no
significant differences in the expression of transcripts in the TG
comparing the GFAP-IFN- to the wild-type mice with the exception of
CD4 and LAT (Fig. 6, Table II). CD4 mRNA
levels were reduced significantly in the transgenic TG, while the
expression of LAT was nearly absent. Specifically, only one of nine
(11%) of the GFAP-IFN- mice had detectable LAT compared with seven
of eight (88%) of the wild-type controls. Consistent with this
finding, only three of nine (33%) of the transgenic mouse sera had
detectable Ab to HSV-1 compared with eight of ten (80%) wild-type mice
30 days p.i. In both transgenic and wild-type mice, the majority of the
anti-HSV-1 Ab was IgG2a, with much less IgG1 detected (Fig. 7).
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transgenic mice produce more IFN- in the TG
following HSV-1 infection
Since the transgenic mice constitutively express IFN- within
the CNS (41) and are resistant to HSV-1 early during the course of
infection, the level of IFN- in the TG and eye was determined.
Although there were no differences in the level of IFN-ß in the
eye comparing the transgenic to wild-type animals, there was a
significant increase in the IFN-ß level in the TG of GFAP-IFN-
mice 3 days p.i. (Table III). Since
IFN- is known to augment NK cytolytic activity (25) and NK cells are
an important first line of defense against HSV-1 (3), the cytolytic
activity of splenocytes was assayed comparing transgenic to wild-type
mice. The results show that while there was a modest elevation in NK
cell activity mediated by splenocytes from the transgenic mice, the
levels did not achieve significance (data not shown).
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1 transgene in the TG of HSV-1-infected
transgenic mice
To determine whether IFN-ß protein levels elevated in the TG
were a result of the localized expression of the transgene, IFN-1
mRNA levels were determined in the eye and TG of transgenic mice. While
the eyes of HSV-1-infected (day 3 p.i.) GFAP-IFN-1 mice
(n = 4) did not express the transgene, one of the four
TG samples analyzed from the transgenic mice expressed the IFN-1
transgene (Fig. 4). In addition, this expression coincided with the
expression of VP16 in the TG of the same animal. No uninfected
transgenic or infected or uninfected wild-type mice (n
= 4) expressed the transgene in the eye or TG, as determined by RT-PCR.
Since the transgene is under the control of the GFAP promoter, an
experiment was conducted to determine whether HSV-1 infection induces
or up-regulates GFAP expression in the TG. RT-PCR analysis revealed
that GFAP mRNA was expressed in uninfected and acutely infected TG
(Fig. 8). However, HSV-1 infection
substantially increased the expression of GFAP mRNA compared with the
uninfected TG. The amplified product was subsequently sequenced and
found to be 97% (226/234 bp) homologous to the mouse GFAP gene (326
bp) (data not shown).
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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and IFN-ß) are potent
immunomodulatory cytokines that elicit antiviral, antitumor, and
antibacterial actions within the target cell (26). Included among the
antiviral actions are the induction of the eIF-2 protein kinase
system that reduces the translation of viral proteins, the 2',5'
oligo-A synthetase system that enzymatically degrades viral RNA, and
the Mx protein that may interfere with viral transcription (26).
Moreover, the rapid expression of type I IFNs (particularly IFN-)
following an infection coupled with the expression of IL-12 are thought
to favor a TH-1 phenotype, resulting in an immune profile that is
likely to antagonize viral infections (27). However, within the central
nervous system (CNS), IFN-ß expression during certain viral
infections such as with lymphocytic choriomeningitis virus is
reportedly low to undetectable (28). By targeting IFN- expression to
the CNS, an assessment of the supportive role this cytokine has against
neurovirulent strains of virus (including HSV-1 McKrae strain), as
measured by virus replication, gene expression, and the immune response
to the infection can be achieved. A recent study has shown that
GFAP-IFN- mice are protected against a lethal inoculum of
lymphocytic choriomeningitis virus, indicating that the expression of
IFN- in the CNS can protect the host from an intracerebral infection
(41). In contrast, the present study was undertaken to determine
whether the expression of IFN- within the CNS modified an infection
in a peripheral sensory ganglion following ocular infection with HSV-1.
Consistent with the protective role of IFN- against HSV-1 infection
(2, 17, 29, 30), the GFAP-IFN- transgenic mice were resistant to
HSV-1. There was a paucity of HSV-1 immediate early, lytic gene
(ICP27), and late gene (VP16) expression in the eye and TG of
HSV-1-infected transgenic mice that correlated with a significant
reduction in the viral load measured in the eye and TG of the infected
GFAP-IFN- animals. Since the basal mRNA levels for the cytokines and
chemokines of the transgenic and wild-type mice did not differ and
were, in many instances, not detected in uninfected animals, there did
not appear to be any preconditioned, enhanced cytokine response in the
TG of the transgenic mice.
Since there was a significant drop in viral titers recovered early
during the acute infection in the transgenic mice and yet, equivalent
amounts of IFN-ß were detected at the site of infection (i.e.,
eye) day 3 p.i., how does resistance develop so quickly? The
protective effect in the eye may be due to an occurrence referred to as
"priming" originally described in 1966 (31). This phenomenon is
defined as an enhanced and rapid response to an IFN-inducing agent
following pretreatment with IFN. It has been found in both in vitro and
in vivo settings, peaking as early as 3 h poststimulation
(32, 33, 34). Consequently, the constitutive expression of IFN-1 in the
transgenic mice may have primed the system to produce IFN faster and in
greater quantity compared with the wild-type controls. The assessment
of IFN-ß levels in the eye 3 days p.i. may have been too late to
detect the priming effect since the duration of the response is
reportedly short-lived (i.e., 8 h) (33). Alternatively, other
immune or nonimmune modifications may have transpired within the
microenvironment of the eye that were not measured or detected that
could explain the protective effect. Changes might have occurred in the
cellular constituency of the eye or TG comparing the GFAP-IFN-
transgenic to wild-type mice, resulting in a cellular environment more
prone to be resistant to viral invasion. However, CD4 and CD8
transcript expression were not altered significantly during the early
(i.e., day 3 p.i.) time point of the infection, suggesting that
these cells were probably not involved in the rapid antiviral state
demonstrated in the transgenic animals. Likewise, splenic NK activity
was not altered comparing the two groups of infected mice, indicating
that the elevation in IFN- is most likely confined to the site of
infection.
Hindrance of the replication and transport of the virus from the site
of initial infection to the innervating sensory ganglion (i.e., TG)
also decreased the establishment of latency, as defined by the reduced
frequency of expression of the LAT gene in the GFAP-IFN- mice. The
reduced expression of LAT coincided with lower levels of CD4
transcripts detected in the GFAP-IFN- mice and fewer transgenic mice
having detectable Ab to HSV-1. The detection of Ab to HSV-1 typically
rises to measurable levels within the first 6 to 8 days p.i., with the
titers continuing to increase throughout latency (10, 17). The lack of
anti-HSV-1 Ig in the majority of the GFAP-IFN- mice implied that
there was a scarcity of Ag to prime the system, even though animals
were inoculated with equal amounts of virus. This assumption is
consistent with the data showing a significant decrease in transcript
expression measured in the TG of the transgenic mice 3 and 6 days p.i.,
including CD8, IFN-, IFN-, the chemokine CRG-2, and the ß
chemokine MCP-1.
The significant enhancement of GFAP mRNA expression in the TG as a
result of HSV-1 may be due to the expansion of Schwann cells in the
peripheral nerve influenced by the inflammatory response to the
infection. GFAP, the major subunit of intermediate filaments in mature
astrocytes, is primarily expressed by astrocytes in the CNS and also
expressed in peripheral nervous system Schwann cells (35). However,
differences between GFAP mRNA in astrocytes and Schwann cells have been
reported in that the GFAP mRNA of Schwann cells contains an extended 5'
untranslated region of 170 bases that is absent in the mRNA of
astrocytes (35). Since extremely low levels of GFAP mRNA were detected
in uninfected TG, it would appear that GFAP expression and, by
inference, the transgene, are nearly quiescent under normal
circumstances. However, HSV-1 replication in the TG (as measured by
VP16 mRNA detection) of the GFAP-IFN- transgenic mice resulted in
the detection of the transgene. Consequently, the increase in
IFN-ß protein levels in the TG of the transgenic mice compared
with the nontransgenic controls day 3 p.i. may reflect the induced
expression of the GFAP-IFN-1. Therefore, the resistance to HSV-1
infection in the transgenic animals may be due to the induced
expression of the IFN-1 transgene in the TG of infected mice as well
as the priming effect of IFN-1 in the CNS.
The constitutive or induced, chronic expression of IFN- is also
known to elicit pathologic effects. Transgenic mice expressing IFN-
under an insulin promoter develop hypoinsulinemic diabetes (36).
Likewise, high-expressing GFAP-IFN- transgenics have been found to
undergo a progressive encephalopathy with marked calcium
mineralization, encephalitis, gliosis, and neurodegeneration (41). This
double-edged sword of IFN- emphasizes the importance of an efficient
regulatory process for this and other cytokines within the immune
system. HSV-1, which periodically may undergo spontaneous reactivation
within the confines of the local microenvironment of the TG in the
mouse model, elicits the chronic expression of cytokines and chemokines
(7, 8, 9, 10, 11, 12) that may mediate unwarranted reactions to self. However, by
regulating the expression or dose of a particular cytokine like
IFN-, it may be possible to use the controlled expression to the
advantage of the host. For example, recent advances in gene expression
(37) have led one group to investigate the efficacy of IFN- gene
vaccinations on a viral infection (38). Using murine CMV as the
prototypic pathogen, Beilharz and colleagues (39) have shown a dramatic
drop in the viral titers of murine CMV in mice vaccinated with IFN-
transgenes (IFN- types 1, 4, and 9, with the IFN-1 transgene
offering the greatest protection) compared with mice vaccinated with
the blank vector. Consequently, the transient expression of endogenous
cytokines in selective target tissues may be a useful alternative to
the current conventional vaccination or other therapeutic protocols for
infectious diseases. Relative to HSV-1, a recent study has shown the
inoculation of plasmid DNA encoding IL-10 on the cornea during acute
HSV-1 infection significantly reduced the lesions associated with the
virus-mediated keratitis (40). The sensitivity of HSV-1 strains as well
as other virus families to IFN- makes this cytokine an appropriate
candidate to further study in DNA expression protocols not only
for the antiviral efficacy, but also for the mode of delivery and
cost effectiveness.
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Footnotes |
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2 Address correspondence and reprint requests to Dr. Daniel J. J. Carr, Department of Microbiology and Immunology, LSU Medical Center Box P6-1, 1901 Perdido Street, New Orleans, LA 70112-1393.
3 Abbreviations used in this paper: HSV-1, herpes simplex virus type 1; CNS, central nervous system; CPE, cytopathic effect; CRG, cytokine response gene; GFAP, glial cell fibrillary protein; ICP27, infected cell polypeptide 27; LAT, latency-associated transcript; MCP-1, monocyte-chemoattractant protein-1; PFU, plaque-forming unit; p.i., postinfection; TG, trigeminal ganglion; VP16, viral protein 16.
Received for publication April 1, 1998. Accepted for publication June 24, 1998.
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References |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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protects mice from herpes simplex virus type 1 corneal disease. J. Gen. Virol. 72:1601.-independent. J. Interferon Cytokine Res. 18:151.[Medline]
interferon and tumor necrosis factor exert a synergistic blockade on the replication of herpes simplex virus. J. Virol. 63:1354.. Proc. Natl. Acad. Sci. USA 91:1198./ß interferons (/ß) and their relationship to IFN-/ß-induced genes in lymphocytic choriomeningitis. J. Virol. 68:7358.1 or interferon-2 enhances the production of both subtypes simultaneously. J. Interferon Cytokine Res. 15:637.[Medline]
in transgenic mice. Science 260:1942. gene by intramuscular injection of naked DNA. J. Interferon Cytokine Res. 17:255.[Medline]
subtypes in vivo: intramuscular injection of interferon expression constructs reduces cytomegalovirus replication. J. Immunol. 160:2932. in the central
nervous system of mice protects against lethal neurotropic viral
infection, but induces inflammation and neurodegeneration. J.
Immunol. In press this issue.
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 A. Gualandris, J. P. Annes, M. Arese, I. Noguera, V. Jurukovski, and D. B. Rifkin The Latent Transforming Growth Factor-beta -binding Protein-1 Promotes In Vitro Differentiation of Embryonic Stem Cells into Endothelium Mol. Biol. Cell, December 1, 2000; 11(12): 4295 - 4308. [Abstract] [Full Text]
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S. Noisakran, I. L. Campbell, and D. J. J. Carr Ectopic Expression of DNA Encoding IFN-{alpha}1 in the Cornea Protects Mice from Herpes Simplex Virus Type 1-Induced Encephalitis J. Immunol., April 1, 1999; 162(7): 4184 - 4190. [Abstract] [Full Text] [PDF]
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