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Department of Microbiology, University of Tennessee, Knoxville, TN 37996
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Abstract |
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 Top Abstract IntroductionMaterials and MethodsResultsDiscussionReferences |
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Introduction |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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Currently, the agonists that drive CD4+ T cell activation in the eye remain ill defined. Peptides derived from viral proteins represent the logical candidate Ags, but this mechanism has been difficult to prove, and the identity of such peptides remains unknown (5). Another idea is that autoantigens, unmasked by HSV infection of the cornea, serve to drive the inflammation (6, 7). The best evidence for this hypothesis was provided by Avery et al. (6), who indicated that a corneal peptide, whose sequence was represented in the IgG2ab Ig isotype, appeared as the inflammatory agonist. Curiously, mice that expressed the IgG2ab Ig isotype, and hence were tolerant to the peptide, were resistant to HSK development (6). More recently, HSV ocular infection was suggested to set off an inflammatory response by expressing a molecular mimic of the Ig peptide (7). These observations await confirmation and have been questioned (8, 9).
An alternative idea to explain HSK lesion development is that the response involves bystander activation of T cells. Support for this mechanism came from studies with TCR-transgenic (Tg) mice backcrossed to SCID or RAG-/- (10, 11, 12). Such animals possessed a limited repertoire of CD4+ T cells and were unable to generate detectable immune responses to HSV Ags. However, unlike SCID or RAG-/- mice, TCR Tg SCID (TgSCID) or TCR Tg RAG-/- (TgRAG-/-) mice could develop HSK lesions upon ocular infection with HSV (10, 11, 12). The mechanism by which HSK occurred in such mice was assumed to involve non-TCR-mediated activation of CD4+ T cells in the cornea. The present study provides more evidence about the mechanism of HSK expression in TgSCID mice. Our results demonstrate that lesion expression requires continuous viral replication most likely necessary to drive production of proinflammatory cytokines. We indicate that activated T cells are involved in HSK expression and that, on the basis of studies with immunosuppressive drugs, CD4+ T cell activation in TgSCID lesions appears as a non-TCR-mediated event. Finally, bystander recruitment and activation are shown to play a significant component in HSK lesions of immunocompetent mice. The results are discussed in terms of the choice of strategies to manage HSK in the clinic.
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Materials and Methods |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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BALB/c mice (4–6 wk) were purchased from Harlan Sprague Dawley
(Indianapolis, IN). DO11.10 (OVA-TCR Tg mice), DO11.10 x SCID
(TgSCID), and CB.17 SCID mice were bred in our special
pathogen-free facility. The DO11.10 OVA-TCR Tg mice were kindly
provided by C. Weaver (University of Alabama, Birmingham, AL).
Generation and immunological profile of the TgSCID mice have been
previously described (10). DO11.10 mice were backcrossed
to CB.17 SCID mice for four generations. Offspring from the first
breeding (F1) were screened for the transgene by
PCR from tail-genomic DNA, and peripheral blood serum was used to
screen for IgM by ELISA. Screened offspring from each generation were
backcrossed to CB.17 SCID mice. After four generations of backcrossing,
brothers and sisters (screened for the transgene and IgM levels) were
mated to develop the TgSCID mice on a homozygous background. Mice have
been brother-sister mated for more than eight generations. Only 70% of
the CD4+ T cells in the DO11.10 mice were
KJ1.26.1 (anti-OVA-TCR Ab) positive, while 
97% of
CD4+ T cells in the TgSCID mice are
KJ+. All food, water, bedding, and instruments
were autoclaved, and all manipulations were done in a laminar flow
hood. To prevent bacterial super infections, all mice were treated
prophylactically with sulfatrim pediatric suspension (Barre-National,
Baltimore, MD) at the rate of 5 ml/200 ml drinking water. All
experimental procedures were in complete agreement with the Association
for Research in Vision and Ophthalmology resolution on the use of
animals in research. The facilities used were accredited by the
American Association for Accreditation of Laboratory Animal
Care.
Virus and reagents
HSV-1 RE and HSV-1 KOS strains were propagated and titrated on monolayers of Vero cells (ATCC CCL81) using standard protocols (13). All virus stocks were aliquoted and stored at -80°C. The thymidine kinase (TK) mutant virus HSV-1 KOS (dlsptk) was kindly provided by D. M. Coen (Harvard Medical School, Cambridge, MA) (14). Cyclosporin A (Sigma, St. Louis, MO) was diluted to 30 mg/ml in castor oil and administered 100 µl i.p. to achieve doses of 100 mg/kg (15, 16, 17). Rapamycin (Sigma) was diluted in ethanol to 1 mg/ml and administered i.p. at a dose of 0.2 mg/kg (16). Acyclovir (ESI Lederle, Philadelphia, PA) was administered in drinking water at a 1 mg/ml concentration (18). CpG sequence 1826 (kindly provided by A. Krieg, University of Iowa, Iowa City, IA) was administered at 30.6 µg in a 100-µl volume in PBS i.p.
Corneal HSV infections and clinical observations
Corneal infections of all mice groups were conducted under deep anesthesia induced by the anesthetic avertin (Pittman Moore, Mondelein, IL). Mice were scarified on their corneas with a 27-gauge needle, and a 4-µl drop containing required viral dose was applied to the eye and gently massaged with the eyelids. The eyes were examined on different days postinfection with a slit lamp biomicroscope (Kowa, Nagoya, Japan), and the clinical severity of keratitis of individually scored mice was recorded. The scoring system was as follows: +1, mild corneal haze; +2, moderate corneal opacity or scarring; +3, severe corneal opacity, but iris invisible; +4, opaque cornea; and +5, necrotizing stromal keratitis.
Virus recovery and titrations
At various time points postinfection, swabs of the corneal surface were taken. The swabs were put into sterile tubes containing 500 µl of DMEM with 10 IU/ml penicillin and 100 µg/ml streptomycin (Life Technologies, Grand Island, NY) and stored at -80°C. For detection and quantification of HSV in the swabs, the samples were thawed and vortexed. Duplicate 200-µl aliquots of each sample of thawed swab medium were plated on Vero cells grown to confluence in 24-well plates at 37°C in 5% CO2 for 1 h and 30 min. Medium was aspirated, and 1 ml of 2x DMEM containing 1% low-melting point agarose was added to each well. Cultures were observed daily for the development of typical cytopathic effect. The titers were calculated as PFU/ml as per standard protocol (13).
Ag-specific lymphoproliferation assay
Individual spleens and cervical and submandibular draining lymph nodes (DLN) were used as responders for lymphoproliferation assays. This method has been described in detail elsewhere (19). Briefly, these responders were restimulated in vitro with irradiated syngeneic splenocytes pulsed with UV HSV-1 KOS (multiplicity of infection (MOI), 1.5) or irradiated naive splenocytes and incubated for 5 days at 37°C. Eighteen hours before harvesting, [3H]thymidine (1 µCi/well) was added to all culture wells, and the plates were read using a beta scintillation counter (Trace 96; Inotech, Lansing, MI). The results were expressed as mean cpm ± SD for six replicates per sample.
Quantification of cytokines by ELISA
Single-cell suspensions of splenic, cervical, and mandibular DLN
cells (2 x 106 cells/ml) were in vitro
restimulated with syngeneic stimulators pulsed with 1.5 MOI UV-HSV-1 RE
or 10 µg/ml OVA323–339 peptide and incubated
for 48–72 h at 37°C. Con A-stimulated (5
µg/106 cells/ml) and unstimulated cells were
used as positive and negative controls, respectively. The supernatants
were analyzed for IFN-
cytokine production by ELISA. Microtiter
plates were coated with 2 µg/ml rat anti-mouse IFN- Ab (BD
PharMingen, San Diego, CA) at 4°C overnight. The plates were then
washed three times with PBS containing 0.5% Tween 20 and blocked with
3% nonfat dry milk for 1 h at 37°C. After washing, serially
diluted samples and standards (rIFN-) were added to the plates and
incubated overnight at 4°C. After washing with PBS, 1 µg/ml
biotinylated anti-IFN- Ab (BD PharMingen) was added to wells and
incubated at 37°C for 2 h. Peroxidase-conjugated streptavidin
(Jackson ImmunoResearch, West Grove, PA) was added and incubated at
37°C for 1 h. The color was developed by adding the substrate
solution (11 mg 2,2'-azino-bis-3-ethylbenzthiazoline-6-sulfonic acid in
25 ml of 0.1 M citric acid, 25 ml of 0.1 M sodium phosphate, and 10
µl of hydrogen peroxide). Quantification was performed using a
Spectramax ELISA reader (softmax version 1.2).
Intracellular cytokine staining
DLN cells from HSV-1 RE-infected DO11.10 or BALB/c mice were
cultured in a 96-well flat-bottom plate and pulsed with
OVA323–339 (10 µg/ml), UV-HSV (MOI 5.0), or
unstimulated naive syngeneic splenocyte for 15 h at 37°C in 5%
CO2. Cell surface staining for FITC
CD4+ (BD PharMingen) was performed, followed by
intracellular staining for PE IFN- (BD PharMingen) using a
Cytofix/Cytoperm kit (BD PharMingen) in accordance with the
manufacturer’s instructions. Golgistop (BD PharMingen) was added for
the last 4 h of the incubation period. All samples were collected
on a FACScan (BD Biosciences, Mountain View, CA), and data were
analyzed using CellQuest 3.1 software (BD Biosciences).
Flow cytometry
Cell preparation. Single-cell suspensions were prepared from eyes, DLN, popliteal lymph nodes, and spleens of mice at different days postinfection. Eyes were digested with collagenase D, as described elsewhere (20), with some modifications. Briefly, enucleated whole eyes were incubated with collagenase for 60 min at 37°C in a humidified atmosphere of 5% CO2. After incubation, eyes were disrupted by grinding with a syringe plunger and passing through a cell scraper. Cells were washed and suspended in RPMI 1640 with 10% FBS. Cells were counted by trypan blue exclusion with high viability.
Staining for flow cytometry. A total of 2 x 106 cells was first blocked with unconjugated anti-CD32/16 for 10 min. Samples were incubated with PE anti-CD4, or PE anti-CD44, or PE anti-CD62L, or PE anti-CD25 (BD PharMingen); PE anti-CCR5 or RPE anti-CD45RB (Calbiochem, La Jolla, CA); and KJ1-26.1 (anti-OVA TCR Ab), followed by biotinylated rat anti-mouse IgG2a and streptavidin PerCP. All samples were collected on a FACScan (BD Biosciences), and data were analyzed using CellQuest 3.1 software (BD Biosciences).
Adoptive transfer
CD4+ T cells were purified to 90% using Cellect Mouse CD4+ T cell columns (Biotex Laboratories, Edmonton, Canada). Cells were stained with CFSE by incubation with 1 µM CFSE in 1x PBS for 10 min at 37°C at 107 cells/ml. Excess CFSE was quenched by adding 10% FBS. Cells were washed three times before i.v. adoptive transfer via tail vein. Cells were stained and analyzed, as described previously.
Histopathology and immunohistochemical staining
Eyes were frozen in optimum cutting temperature (OCT) compound (Miles, Elkart, IN) at different days postinfection. Six-micrometer-thick sections were cut, air dried, and fixed in cold acetone for 5 min. The sections were then blocked with heat-inactivated goat serum and stained for the presence of HSV Ags by the use of rabbit anti-HSV antiserum (DAKO, Carpenteria, CA), which was followed by biotinylated anti-rabbit Ig (1/20 dilution; Biogenex, San Ramon, CA). Sections were then treated with HRP-conjugated streptavidin (1:1000) and 3, 3'-diaminobenzidine (Vector Laboratories, Burlingame, CA), and counterstained with hematoxylin.
Statistical analysis
Wherever specified, data obtained were analyzed for statistical significance by Student’s t test and Wilcoxon rank sum test.
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Results |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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As shown before, whereas immunocompetent mice eliminate virus from
the eye following infection, SCID and TgSCID mice do not, and usually
die of herpetic encephalitis at about day 11–12 postinfection
(10, 11). As shown in Table I
and Fig. 1, if TgSCID mice were treated with the
antiherpes drug acyclovir, continually starting on day 1 or day 4
post-HSV infection, lesions failed to manifest and animals treated at
day 1 survived infection for at least 3 wk (latest time studied). In
experiments with ocularly infected BALB/c or DO11.10 mice (both
competent to respond immunologically to HSV), antiherpes drug acyclovir
treatment begun on day 1 markedly diminished or abrogated HSK lesion
development, but treatment begun on day 4 was without effect (Fig. 1).
These data indicate the necessity for continuous viral replication in
the eyes of TgSCID mice to produce lesions, but only a brief, but
necessary, requirement for replication to induce HSK in immunocompetent
animals. The experiment shown in Fig. 2
demonstrates a failure of replication-competent
TK- virus to induce HSK in TgSCID mice. The
virus mutant fails to replicate in ganglionic tissue (14),
perhaps implying that virus persists in the eye by spreading to the
stroma following replication in the ganglion.
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97% of their CD4+ T cells recognize OVA
peptide (10, 11). Accordingly, we hypothesize that T cell
activation in TgSCID mice occurs other than by TCR recognition. The
data in Fig. 3 add support to this
concept. In these experiments, both BALB/c and TgSCID mice were
ocularly infected with HSV and then treated daily with either
cyclosporin A or rapamycin. As is evident in Fig. 3B, HSK
lesions were markedly diminished in BALB/c mice treated with CsA. In
contrast, the drug had no apparent effect on lesions in TgSCID mice
(Fig. 3A). Rapamycin treatment, however, dramatically
reduced lesions in TgSCID mice. This effect was not the result of
antiviral activity, since virus persisted in ocular washings and
animals succumbed to herpetic encephalitis at the normal time
period (data not shown). In rapamycin-treated BALB/c mice, lesions were
also reduced in severity (Fig. 3, C and D). These
results are consistent with the notion that HSK in TgSCID and BALB/c
mice involve different primary mechanisms of CD4+
T cell activation.
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To measure the role of T cell activation, experiments were done to
compare the severity of HSK lesions in groups of TgSCID mice that were
unmanipulated, with groups immunized previously with OVA peptide in CFA
or exposed daily to immunostimulatory CpG motifs. As is evident in Fig. 4A, animals exposed to either
activation protocol developed more severe lesions than
control-untreated animals. These differences were not striking, but
they were significant (Wilcoxon rank sum test). Interestingly, TgSCID
mice exposed to activation survived for 1–3 days longer than did the
control-untreated animals, most likely the consequence of increased
IFN- production (Fig. 4, B–D). In separate experiments,
both immunized and CpG treatment were shown to result in an increased
percentage of KJ+ splenic T cells that expressed
the activation phenotype (see legend to Fig. 4).
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A). In this model, major
differences were observed. Thus, whereas adoptive transfers of
nonactivated cells resulted in mild HSK and many negative responses,
the transfer of T cells from Ag-stimulated TgSCID resulted in severe
HSK. Moreover, SCID recipients of nonactivated cells also developed
more severe HSK lesions if such mice were additionally immunized with
OVA peptide in CFA.
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B). During the 15 days hiatus, the adoptively transferred
cells presumably underwent homeostatic expansion and activation, as
described by others (21, 22). In support of this, a
decrease in CFSE intensity and an up-regulation of the CD44 marker were
seen in CD4+ KJ+ cells in
mice terminated at day 20 postadoptive transfer (Fig. 5, C
and D). Do bystander recruitment and activation contribute to HSK lesions in immunocompetent mice?
Two types of experiments were performed to evaluate whether or not
bystander recruitment and activation contribute to HSK lesions in
immunocompetent mice. In the first approach, DO11.10 mice were ocularly
infected with HSV, and the phenotype of CD4+ T
cells was measured in HSK lesions, DLN, and spleen. In such mice, about
70% of the CD4+ T cells were
KJ+ and OVA specific, with the remaining cells
most likely reactive with other Ags. As reported previously, DO11.10
mice generate apparently normal immune responses to HSV and recover
normally from infection (10). It was expected that
CD4+ T cells that recognize HSV Ags should be
largely KJ- since the KJ+
cells are known to recognize OVA323–339 peptide
(23). The experiments shown in Fig. 6 support this notion, and show that the
KJ- population from DLN cells of HSV immune mice
contains the predominant HSV reactivity.
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shows corneal sections stained for
CD4 and KJ, and Table II a phenotype
analysis of T cells recovered from the eyes of mice with HSK lesions,
DLN, and spleens of infected and noninfected mice. The results show
only marginal enrichment for KJ- cells in ocular
lesions and DLN of infected mice, indicating that the majority of
CD4+ T cells in DO11.10 ocular lesions are
KJ+ bystanders reactive with OVA peptide. In
fact, an analysis of such cells by flow cytometry revealed that the
majority of intraocular KJ+
CD4+ T cells expressed activation markers (Fig. 8). Accordingly, HSK in DO11.10 mice
contains a major component of bystander recruitment and activation.
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Sample animals showing HSK lesions were killed at intervals
subsequently to isolate cell infiltrates from the inflamed cornea as
well as to determine the phenotype of cells in both DLN, distal lymph
nodes, and spleens (Table III). In corneal samples, on day 16
postinfection, 14.7% of the total recovered cells were
CD4+ (Fig. 9A and Table III). Of these
CD4+ T cells, 10.2% were
KJ+ and CFSE+. Moreover,
97.4% of the recruited KJ+ ocular cells
expressed the
CD44highLsellow activation
phenotype (Fig. 9B), yet were nondividing, as indicated by
the CFSE data (Fig. 9C). In distal LN and spleen samples,
0.9 and 1.6%, respectively, of the CD4+ cells
were KJ+, and such cells did not show the
activation phenotype observed with KJ+ cells
derived from corneal or DLN samples (Table III).
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Discussion |
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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The evolution of HSK lesions following HSV ocular infection involves numerous events (4, 5). Lesion expression appears strictly dependent on HSV replication in the eye. Thus, infection with replication-defective mutants or UV-inactivated virus, even though immunogenic, fails to cause HSK (24). In immunocompetent mice, the period of viral replication is brief, usually no more than 5 days (24, 25), but as shown in this work, it cannot be condensed to 1 day, since mice infected for only that time prior to antiviral drug treatment failed to develop lesions. In the TgSCID model, however, viral replication for longer periods is necessary to induce HSK. Accordingly, antiviral treatment at day 4 prevented the expression of HSK, although not death from herpetic encephalitis. In other studies, virus removal even as late as day 6 had an inhibitory effect on HSK expression (our unpublished observations). Normally, in untreated animals, virus can be demonstrated in the infected cornea until animals die of encephalitis at about day 11–12 postinfection (11). Of interest, whereas the location of viral Ags in immunocompetent mice was the corneal epithelium, after the first few days in TgSCID mice, viral Ag was demonstrable mainly in the stroma in which the inflammatory reaction occurs (11). More than likely, virus arrives at the stromal site by zosteriform spread from the ganglion, an idea supported by the observation that TK- mutant virus, which cannot replicate in ganglionic neurons (14), failed to access the stroma. Furthermore, the mutant virus also failed to cause lesions in TgSCID mice.
Although yet to be formally proven, it is likely that the stromal virus is responsible for inducing the proinflammatory cytokines and chemokines assumed to play a central role in the pathogenesis of TgSCID lesions. This idea was supported by the results of studies on the inhibitory effects of two types of immunosuppressive drugs. Thus, cyclosporin A inhibited HSK in immunocompetent mice, but had no effect on the severity of lesions in TgSCID mice. Lesions in the latter animals were inhibited by rapamycin, a drug that inhibits the response of T cells to cytokine stimulation (15, 16). Since the effect of cyclosporin A on T cells is directed at events triggered by engagement of the TCR, we interpret our result to imply that T cell activation in TgSCID lesions proceeds by a non-TCR activation mechanism. However, this notion requires further evaluation.
The cells involved in TgSCID HSK would seem to be mainly OVA peptide-specific CD4+ T cells, activated by as yet unidentified chemokines and cytokines. Initially, it was suspected that some HSV protein might contain peptides cross-reactive with OVA323–339. A search of the gene bank, however, found no such homology. In addition, other TCR Tg models backcrossed to RAG-/- were also shown to generate HSK (S. Deshpande, manuscript in preparation). It remains unclear why OVA-specific CD4+ T cells should enter the proinflammatory ocular environment. Experiments reported in this work demonstrate that in fact only cells of the effector phenotype appear to participate in the bystander activation process. Curiously, when comparing the efficacy of adoptive transfers of naive with stimulated KJ+ T cells to SCID mice, the stimulated cells supported more severe lesions. In fact, recipients of naive KJ+ cells developed trivial lesions, unless recipient animals were also immunized with OVA peptide in adjuvant. Interestingly, however, KJ+ cells could be induced to express the eye-seeking phenotype in at least two additional ways. One was to expose mice to immunostimulatory CpG motifs. This procedure generated significantly increased numbers of KJ+ cells with the effector phenotype, most likely a consequence of CpG motif-induced cytokine production by APCs.
Perhaps of even more interest was the observation that SCID mice given adoptive transfers of naive KJ+ cells, then rested for 2–3 wk before HSV ocular infection, readily developed HSK. It was recently recognized in such circumstances that naive T cells undergo homeostatic expansion and adopt the phenotype of memory cells (21, 22). They have been referred to as false memory cells (21, 22). Our observations that such cells appear able to mediate immunopathological reactions are, we believe, novel.
The observation that bystander activation can be the principal mechanism of immunopathology is made less interesting by the fact that the model used to show such an effect is unrepresentative of naturally occurring circumstances. However, bystander activation and recruitment were also shown to contribute a significant component of HSK lesions in normal immunocompetent animals. For example, the majority of T cells in HSK lesions of immunocompetent DO11.10 mice were in fact KJ+ cells unreactive with HSV Ags. In such animals, the 30% of CD4+ T cells that are KJ- account principally for the reactivity to HSV Ags. However, such cells were barely more represented in ocular lesions than they were in unstimulated lymphoid tissue. We conclude from such observations that the bulk of the activated CD4+ T cells in the DO11.10 model are in fact bystanders.
Similarly, bystander recruitment and activation were implicated to occur in BALB/c mice that possessed adoptive transfers of KJ+ CFSE-labeled cells. When such animals were infected to induce HSK, at least 10% of cells were bystanders derived from the adoptive transfers. Presumably, many KJ- CD4+ cells from the host itself were also bystanders. Both sets of recruited CD4+ T cells were expected to help orchestrate the inflammatory reaction, most likely by reacting with activating cytokines. However, in the immunocompetent animal model, the source of the activating cytokines is likely to differ, since virus does not continue to replicate in such animals as it does in TgSCID mice. In immunocompetent animals, the T cells themselves, reacting to the so far unidentified Ags, are the likely sources of cytokines.
The ultimate relevance of understanding the inflammatory mechanisms at play during HSK should impact on the choice of treatment. Accordingly, there seems little point in using antiviral drugs since virus is rarely present after the early phase, except in immunocompromised animals. Currently, in humans, corticosteroids are a popular treatment choice (26). However, a combination of other drugs such as cyclosporin and rapamycin would seem to deserve a trial. Indeed, recently, it was reported that cyclosporin A is useful for the control of human HSK (27).
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Acknowledgments |
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Footnotes |
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2 Address correspondence and reprint requests to Dr. Barry T. Rouse, Department of Microbiology, University of Tennessee, M409 Walters Life Sciences Building, Knoxville, TN 37996-0845. E-mail address: btr{at}utk.edu
3 Abbreviations used in this paper: HSK, herpetic stromal keratitis; DLN, draining lymph node; MOI, multiplicity of infection; Tg, transgenic; TK, thymidine kinase.
Received for publication April 27, 2001. Accepted for publication July 3, 2001.
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TopAbstractIntroductionMaterials and MethodsResultsDiscussionReferences |
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 B. Gilbertson, S. Germano, P. Steele, S. Turner, B. F. de St. Groth, and C. Cheers Bystander Activation of CD8+ T Lymphocytes during Experimental Mycobacterial Infection Infect. Immun., December 1, 2004; 72(12): 6884 - 6891. [Abstract] [Full Text] [PDF]
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D. B. McGavern and P. Truong Rebuilding an Immune-Mediated Central Nervous System Disease: Weighing the Pathogenicity of Antigen-Specific versus Bystander T Cells J. Immunol., October 15, 2004; 173(8): 4779 - 4790. [Abstract] [Full Text] [PDF]
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P. S. Biswas, K. Banerjee, M. Zheng, and B. T. Rouse Counteracting corneal immunoinflammatory lesion with interleukin-1 receptor antagonist protein J. Leukoc. Biol., October 1, 2004; 76(4): 868 - 875. [Abstract] [Full Text] [PDF]
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M. Xu, A. J. Lepisto, and R. L. Hendricks CD154 Signaling Regulates the Th1 Response to Herpes Simplex Virus-1 and Inflammation in Infected Corneas J. Immunol., July 15, 2004; 173(2): 1232 - 1239. [Abstract] [Full Text] [PDF]
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R. L. Sparks-Thissen, D. C. Braaten, S. Kreher, S. H. Speck, and H. W. Virgin IV An Optimized CD4 T-Cell Response Can Control Productive and Latent Gammaherpesvirus Infection J. Virol., July 1, 2004; 78(13): 6827 - 6835. [Abstract] [Full Text] [PDF]
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S. K. Lathrop, C. A. Huddleston, P. A. Dullforce, M. J. Montfort, A. D. Weinberg, and D. C. Parker A Signal through OX40 (CD134) Allows Anergic, Autoreactive T Cells to Acquire Effector Cell Functions J. Immunol., June 1, 2004; 172(11): 6735 - 6743. [Abstract] [Full Text] [PDF]
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P. S. Biswas, K. Banerjee, B. Kim, and B. T. Rouse Mice Transgenic for IL-1 Receptor Antagonist Protein Are Resistant to Herpetic Stromal Keratitis: Possible Role for IL-1 in Herpetic Stromal Keratitis Pathogenesis J. Immunol., March 15, 2004; 172(6): 3736 - 3744. [Abstract] [Full Text] [PDF]
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E. Chiffoleau, T. Kobayashi, M. C. Walsh, C. G. King, P. T. Walsh, W. W. Hancock, Y. Choi, and L. A. Turka TNF Receptor-Associated Factor 6 Deficiency during Hemopoiesis Induces Th2-Polarized Inflammatory Disease J. Immunol., December 1, 2003; 171(11): 5751 - 5759. [Abstract] [Full Text] [PDF]
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S. K. Seo, H. Y. Park, J. H. Choi, W. Y. Kim, Y. H. Kim, H. W. Jung, B. Kwon, H. W. Lee, and B. S. Kwon Blocking 4-1BB/4-1BB Ligand Interactions Prevents Herpetic Stromal Keratitis J. Immunol., July 15, 2003; 171(2): 576 - 583. [Abstract] [Full Text] [PDF]
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 D. M. Koelle and L. Corey Recent Progress in Herpes Simplex Virus Immunobiology and Vaccine Research Clin. Microbiol. Rev., January 1, 2003; 16(1): 96 - 113. [Abstract] [Full Text] [PDF]
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J. S. Haring, L. L. Pewe, and S. Perlman Bystander CD8 T Cell-Mediated Demyelination After Viral Infection of the Central Nervous System J. Immunol., August 1, 2002; 169(3): 1550 - 1555. [Abstract] [Full Text] [PDF]
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