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Antibody Prevents Virus Reactivation Within the Central Nervous System

Mark T. Lin, David R. Hinton, Norman W. Marten, Cornelia C. Bergmann and Stephen A. Stohlman
J Immunol June 15, 1999, 162 (12) 7358-7368;
Mark T. Lin
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David R. Hinton
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Norman W. Marten
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Cornelia C. Bergmann
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Stephen A. Stohlman
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Abstract

The neurotropic JHM strain of mouse hepatitis virus (JHMV) produces an acute CNS infection characterized by encephalomyelitis and demyelination. The immune response cannot completely eliminate virus, resulting in persistence associated with chronic ongoing CNS demyelination. The contribution of humoral immunity to viral clearance and persistent infection was investigated in mice homozygous for disruption of the Ig μ gene (IgM−/−). Acute disease developed with equal kinetics and severity in IgM−/− and syngeneic C57BL/6 (wt) mice. However, clinical disease progressed in IgM−/− mice, while wt mice recovered. Viral clearance during acute infection was similar in both groups, supporting a primary role of cell-mediated immunity in viral clearance. In contrast to wt mice, in which infectious virus was reduced to below detection following acute infection, increasing infectious virus was recovered from the CNS of the IgM−/− mice following initial clearance. No evidence was obtained for selection of variant viruses nor was there an apparent loss of cell-mediated immunity in the absence of Ab. Passive transfer of anti-JHMV Ab following initial clearance prevented reactivation of infectious virus within the CNS of IgM−/− mice. These data demonstrate the clearance of infectious virus during acute disease by cell-mediated immunity. However, immunologic control is not maintained in the absence of anti-viral Ab, resulting in recrudescence of infectious virus. These data suggest that humoral immunity plays no role in controlling virus during acute infection, but plays an important role in establishing and maintaining CNS viral persistence.

Viral infections induce vigorous responses in the infected host, including components of both cellular and humoral immunity. The relative importance of each of the individual immune effectors is dependent upon a variety of viral and host factors (1). For example, the ability of a virus to induce cytopathogenicity in vivo has been suggested to correlate with the predominant anti-viral effector mechanisms effective in virus control. Clearance of noncytopathic viruses such as lymphocytic choriomeningitis virus (LCMV)3 is predominantly mediated by a perforin-dependent CTL response (2). By contrast, the clearance of cytopathic viruses is more dependent on noncytolytic responses such as cytokines and anti-viral Ab (3). In addition, the cell type(s) infected influences the predominant effector mechanism responsible for virus clearance. One example is infection of the CNS by the neurotropic JHM strain of mouse hepatitis virus (JHMV), which produces an acute infection characterized by encephalomyelitis and primary demyelination (4, 5). The clearance of infectious JHMV from most CNS cell types occurs before detection of neutralizing Ab (6) and is mediated by a combination of virus-specific CTL and IFN-γ (6, 7, 8). However, the immune response to JHMV infection appears to be only partially effective. Although infectious virus, and the accompanying immunopathology, is eliminated from the CNS, JHMV persists in a noninfectious form associated with chronic ongoing demyelination, which pathologically resembles the chronic human demyelinating disease, multiple sclerosis.

The role(s) of anti-viral effectors during acute self-limiting viral infections is becoming increasingly clear; however, their role(s) in establishing or maintaining persistent viral infections is less well understood. For example, clearance of acute LCMV infection is largely dependent upon the CTL response (2). However, long-term control of LCMV is complex and appears to be dependent upon humoral immunity, CD4+ T cells, and cytokines (9, 10). Similarly, anti-viral Ab responses, influenced by CD4+ T cells, promote recovery from acute influenza virus infection of the murine respiratory tract (11), while virus-specific CTL are the major anti-viral effectors (12). These data suggest that, in addition to a role in controlling some cytopathic viruses, humoral immunity may be a necessary effector mechanism required to control persistent viral infections (13, 14).

A common outcome of viral infection of the CNS is persistence. The presence of the blood-brain barrier, restricted expression of MHC molecules, and the nonrenewable nature of neurons are all unique characteristics that may hinder viral clearance and help establish persistent CNS viral infections (15). For viruses to persist, they must assume a nonlytic phenotype and/or alter gene expression to avoid immune surveillance. For example, noncytopathic viruses may circumvent CTL recognition by infection of MHC class I-negative neurons (16) or by suppressing virus-specific T cell responses during CNS persistence (17). However, Sindbis virus infection of neurons is controlled via an Ab-mediated mechanism distinct from Ab-dependent cell-mediated cytotoxicity or C-dependent lysis (14, 18). In contrast to protection from cytopathic viruses provided by Ab, the virus-specific Ab response modulates viral gene expression and helps maintain persistence following measles virus (19, 20), Sindbis virus (21), and herpes simplex virus infections (22). Viruses with high mutation rates, especially RNA viruses such as JHMV, may persist in the CNS by generating attenuated variants or altering epitopes in response to immune pressure (23, 24).

Infection of IgM−/− mice with JHMV was examined to determine whether the humoral response influenced clearance during acute infection or the ability to establish a persistent CNS infection. The passive transfer of both neutralizing and, in some instances, nonneutralizing mAb before JHMV infection provides protection from lethal disease (25, 26, 27, 28, 29). However, during acute infection JHMV is cleared from the CNS before the detection of serum-neutralizing Ab responses (6). Consistent with data suggesting that cell-mediated immunity is the primary effector of JHMV clearance, the absence of a humoral response did not influence initial clearance of virus from the CNS. However, following initial clearance, reactivation of infectious virus was found in the CNS of IgM−/−, but not control mice. The passive transfer of anti-viral Ab after the majority of infectious virus was cleared from the CNS, but before virus reactivation, prevented the reappearance of infectious virus and diminished immunopathology. These data demonstrate that in spite of an effective cell-mediated immune response initially able to control infectious virus, humoral immunity can maintain control of infectious virus within the CNS.

Materials and Methods

Mice

Breeding pairs of C57BL/6-Igh-6tm1Cgn (IgM−/−) mice (30) and syngeneic C57BL/6 (wt) mice were obtained from The Jackson Laboratory (Bar Harbor, ME). Homozygous IgM−/− mice were bred at the University of Southern California School of Medicine (Los Angeles, CA) under pathogen-free conditions. The absence of Ig in IgM−/− mice was confirmed by testing sera for murine IgG by ELISA, as previously described (31). All mice used were between the ages of 7–8 wk of age.

Viral infection and titer determination

Mice were infected via intracerebral inoculation with 50 plaque-forming units of the 2.2v-1 mAb-derived variant of JHMV in 32 μl of Dulbecco’s PBS, pH 7.4, as previously described (32). The 2.2v-1 strain of JHMV was kindly supplied by J. Fleming (University of Wisconsin, Madison, WI). Tissue levels of infectious virus were determined by plaque assay on monolayers of DBT cells, as previously described (6). Briefly, clarified homogenates from one-half of the brains were serially diluted in serum-free-PBS containing 10% tryptose phosphate broth, and virus was adsorbed for 90 min at room temperature. Plaque numbers were determined following 24–48-h incubation at 37°C. Data represent the average of triplicate samples for groups of four or more mice.

Clinical disease

Clinical disease was graded as previously described (6, 28): 0, healthy; 1, ruffled fur and hunch back; 2, slow mobility and inability to upright; 3, paralysis and wasting; 4, moribund and death. Data represent the average for four or more mice per group.

CTL assays

Spleen cell suspensions were prepared at 7, 11, and 21 days postinfection (p.i.). Splenocytes from three mice per group were pooled, and 1 × 108 cells were cultured for 6 days at 37°C with irradiated (25 Gy) syngeneic JHMV-infected spleen cells in RPMI 1640 medium supplemented with 10% FCS (Gemini Biological Products, Calabasas, CA), 2 mM glutamine, 25 μg/ml gentamicin, 1 mM sodium pyruvate, 5 × 10−5 M 2-ME, and nonessential amino acids. Cytolytic activity was measured as previously described (6, 7). Briefly, syngeneic EL-4 (H-2b) target cells were infected with either a recombinant vaccinia virus (rVV) expressing amino acids 510–518 of the JHMV spike (S) protein (vJS510) or a control rVV expressing the Escherichia coli lacZ gene (vSC8) at a multiplicity of infection of 5 (33). Following incubation for 14 h at 37°C, target cells were labeled with 100 μCi Na51CrO4 (New England Nuclear, Boston, MA) for 1 h at 37°C. Effector cells were added to target cells at various E:T ratios. 51Cr release was measured after 4-h incubation at 37°C. Data are expressed as percent specific release defined as: ((experimental release) − (spontaneous release))/((total release − (spontaneous release)). Maximum spontaneous release values were ≤20% of total release.

RNA expression

Total RNA was prepared from one-half of spinal cords divided longitudinally by homogenization in guanidine isothiocyanate and isolated by centrifugation through 5.4 M cesium chloride at 100,000 × g for 18 h, as previously described (31). The cDNA were prepared using avian myeloblastosis reverse-transcriptase (Promega, Madison, WI) and oligo(dT) primers (Promega) for 1 h at 42°C. PCR was performed using AmpliTaq DNA polymerase (Perkin-Elmer, Branchburg, NJ) and primers specific for IFN-γ (31) and JHMV nucleocapsid protein (N) (viral nucleotides 527-1257): 5′-ATA GGA TCC ATG GCT ACT AGG TTT GCG CCC GGC-3′ and 5′-ACA GTT ACC TAC ATC TGC ACC ACC ATC TTG-3′. Amplification of IFN-γ mRNA was carried out using 35 cycles of one denaturing step for 45 s at 94°C, primer annealing for 45 s at 59°C, and extension for 1.5 min at 72°C. Amplification for JHMV N mRNA was conducted using 25 cycles of one denaturing step at 94°C (45 s), primer annealing at 72°C (45 s), extension step at 72°C (2 min), followed by a final extension step for 7 min. For quantification, PCR products were diluted in denaturing solution (0.4 N NaOH, 25 mM EDTA), neutralized with Tris-HCl (1 M; pH 8), and transferred to 0.45-μm Nytran membranes (Schleicher & Schuell, Keene, NH) using a Minifold I dot-blot apparatus. Membranes were hybridized overnight at 60°C with [32P]ATP-labeled internal oligonucleotide probes for IFN-γ (31) and JHMV N mRNA: 5′-ATA GGA TCC ATG GTT TTG GCT AAG CTC GGT AAA G-3′. Membranes were washed, exposed to Storage Phosphor Screens (Molecular Dynamics, Sunnyvale, CA), and scanned using a PhosphorImaging Scanner (Molecular Dynamics). To adjust for differences in cDNA quantity, levels were normalized to the housekeeping enzyme hypoxanthine phosphoriboxyltransferase, as previously described (31). Mean values for three mice per group are presented.

Sequence analysis

Virus derived from brain samples was propagated once on DBT monolayers, and the cells were lysed by addition of guanidine thiocyanate solution when approximately 80% of the cells exhibited cytopathology. RNA was isolated by phenol/chloroform extraction. RNA from cells infected with parental virus and uninfected cells were used to control for mutations introduced by Taq polymerase and PCR contamination, respectively. RNA (5 μg) was reverse transcribed using avian myeloblastosis virus reverse-transcriptase and random hexanucleotide primers (Promega). A cDNA encompassing the immunodominant CTL epitope within the JHMV S protein (viral nucleotides 1528–1554) plus surrounding 500 nucleotides 1420–1890(1420–1890) was amplified using oligonucleotide primers JS1895 and JS1390, as previously described (8). The cDNAs were sequenced on an ABI Prism automated sequencing apparatus using oligonucleotide JS1390 as primer, as previously described (8).

Ab preparations

Serum-free supernatants from a hybridoma producing JHMV-neutralizing mAb J.2.2 (IgG2b) (34) were concentrated by precipitation with saturated ammonium sulfate. Anti-JHMV polyclonal Ab was obtained from mice hyperimmunized with JHMV (35). Anti-JHMV and control ascites were induced in immunized and naive mice, respectively, by injection of Sarcoma 180 cells (35). Ascites were defibrinated (36) and heat inactivated at 56°C for 30 min before use. Control ascites was tested by ELISA to insure the absence of JHMV Ab, as previously described (31).

Passive transfers

IgM−/− mice received 150 μg of either JHMV-neutralizing mAb J.2.2 or an equal amount of IgG2b myeloma protein (Zymed Laboratories, San Francisco, CA). These Ab and the polyclonal anti-JHMV and control ascites were transferred into the peritoneal cavity of infected IgM−/− mice at 9, 12, and 17 days p.i. Neither the isotype control myeloma protein nor the ascites obtained from naive mice had detectable anti-JHMV Ab, as determined by ELISA. Neutralization titers were determined by the plaque-reduction method (6, 31, 35). Briefly, 50–100 plaque-forming units of either DM (for mAb J.2.2) or the 2.2v-1 (for immune ascites) JHMV strains were mixed with serial 4-fold dilutions. After 1 h at 37°C, residual virus was determined by plaque assay. The dilution that reduced plaque numbers by 50% was designated as titer (35). The neutralizing titers of the J.2.2 mAb and anti-JHMV ascites were 1/1600 and 1/3200, respectively.

Histopathology

Brains, bisected in the mid-coronal plane, and spinal cords were prepared for frozen sections or embedded in paraffin. For paraffin preparations, tissues were fixed for 3 h in Clark’s solution (75% ethanol, 25% glacial acetic acid) before embedding. Sections were stained with either hematoxylin and eosin or luxol fast blue. Distribution of JHMV Ag was determined by immunoperoxidase staining (Vectastain-ABC kit; Vector Laboratories, Burlingame, CA) using the anti-JHMV mAb J.3.3 specific for the carboxyl terminus of the N protein as the primary Ab (34, 37) and horse anti-mouse mAb as secondary Ab (Vector Laboratories). CD4+ and CD8+ T cells were identified in frozen sections fixed with acetone for 2 min at room temperature and stained using anti-CD4 (L3T4; PharMingen, San Diego, CA) or anti-CD8 (Ly-2, PharMingen) mAb. Both primary Ab were detected with a biotinylated rabbit anti-rat serum preabsorbed with mouse serum (Vector Laboratories) and visualized using Vectastain-ABC kits. Apoptotic cells were identified using Oncor ApopTag Kit (Gaithersburg, MD), which utilizes TdT for extension of fragmented DNA, as previously described (6). Tissue processing and staining were performed according to the manufacturer’s instructions. For quantitative comparisons, four serial step sections of individual samples from groups of three to four mice were stained, and all positive cells counted. Student’s t test was used for statistical analysis.

Results

Biphasic pathogenesis in IgM−/− mice

To determine whether humoral immunity contributes to JHMV pathogenesis, clinical disease and mortality were compared in infected IgM−/− and wt mice. Both wt and IgM−/− mice developed clinical signs of disease (mild ruffled fur and hunched back) by 7–8 days p.i. (Fig. 1⇓A), which progressed to hind limb paralysis and wasting in both groups by approximately 13 days p.i. In contrast to the clinical recovery of wt mice, IgM−/− mice showed signs of increasing lethargy and wasting between 14 and 40 days p.i. (Fig. 1⇓A). The increase in clinical signs coincided with an increase in mortality of IgM−/− mice after 25 days p.i. (Fig. 1⇓B). By day 40 p.i., only 8% of JHMV-infected IgM−/− mice survived and the remainder appeared moribund. By contrast, 80% of wt mice had completely recovered from infection and minor gait abnormalities.

FIGURE 1.
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FIGURE 1.

IgM−/− mice exhibit increased morbidity (A) and mortality (B) following JHMV infection. IgM−/− and wt mice were infected with JHMV, and the morbidity and mortality were recorded as follows: 0, healthy; 1, hunched back and ruffled fur; 2, slow mobility and inability to upright; 3, paralysis and wasting; 4, moribund and death. Average scores of 4 or more mice per time point are shown. Error bars indicate the SE means.

The kinetics of viral replication in IgM−/− and wt mice was compared to determine whether increased morbidity and mortality in the JHMV-infected IgM−/− mice resulted from an altered ability to eliminate infectious virus from the CNS. Consistent with the clinical signs of disease, infectious JHMV was cleared from the CNS of both groups with similar kinetics between 3 and 11 days p.i. (Fig. 2⇓). However, by 14 days p.i., when no infectious JHMV could be isolated from the CNS of wt mice, infectious virus had reemerged in the IgM−/− mice after 11 days p.i. Between 14 and 35 days p.i., infectious JHMV continued to increase in the CNS of the surviving IgM−/− mice. By contrast, infectious virus remained undetectable in the CNS of the wt mice. These data confirmed the role of cellular immunity in JHMV clearance during acute CNS infection (6, 7) and suggest that the anti-viral Ab response is critical for maintaining the suppression of infectious virus, resulting in the establishment of a persistent CNS infection.

FIGURE 2.
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FIGURE 2.

JHMV replication in the CNS of IgM−/− and wt mice. Each time point is the mean of four to six samples and the SEMs shown as error bars. Student’s t test was used to test for significance between groups (*, p < 0.05).

CNS inflammation in IgM−/− mice

To determine whether IgM−/− mice exhibited altered inflammatory responses, the extent of mononuclear cell infiltration and frequency of apoptotic cells in the CNS was compared with wt mice. At day 7 p.i., the majority of the mononuclear cell infiltration and apoptotic cells were localized to the perivascular areas, with only a few CD4+ or CD8+ T cells detected within the parenchyma in both groups (data not shown). At day 21 p.i., the CNS of the IgM−/− mice showed increased numbers of perivascular mononuclear cells (Fig. 3⇓). This distribution of inflammatory cells is reminiscent of the perivascular inflammation observed during acute JHMV infection (38). Indeed, a 10-fold increase in perivascular CD4+ T cells was observed in the IgM−/− mice compared with the wt mice at day 21 p.i. (Fig. 3⇓). In both groups, the CD8+ T cells were present in a diffuse pattern within the parenchyma (Fig. 3⇓). Although the numbers of CD8+ T cells varied dramatically in individual microscopic fields, in contrast to the CD4+ T cells, the total number of CD8+ T cells in the IgM−/− mice was not statistically different from wt mice. Fewer apoptotic cells were detected at 21 days p.i. compared with 7 days p.i.; however, no difference in the numbers or distribution of apoptotic cells was observed comparing the two groups at 21 days p.i. (Fig. 3⇓). These data suggest, in contrast to the absence of CD4+ T cells that dramatically increased the number of apoptotic lymphocytes (38), the absence of humoral immunity did not result in an increase in apoptosis of effector cells within the CNS. Both wt and IgM−/− mice had prominent demyelination within the spinal cord (Fig. 4⇓). In contrast to wt mice, demyelinating lesions were detected in the brains of IgM−/− mice in areas in which virus Ag was found (data not shown). These data suggest that the cell-mediated immune response is not compromised in the IgM−/− mice, although the increase in infectious virus is associated with increased demyelination.

FIGURE 3.
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FIGURE 3.

Immunopathology in the CNS of JHMV-infected IgM−/− and wt mice at 21 days p.i. IgM−/− mice (A) show increased cellular infiltrates localized to the perivascular areas (small arrows) compared with wt mice (B). IgM−/− mice have increased numbers of perivascular CD4+ T cells (small arrows) (C) compared with wt mice (D), although both groups have similar total numbers of CD4+ T cells within the CNS. Although focal areas of the wt mice showed increased numbers of parenchymal CD8+ T cells (arrowheads), the total numbers present were similar in both groups (E and F). Few apoptotic cells (large arrows) were detected in either IgM−/− (G) or wt mice (H). Sections in A and B were stained with hematoxylin and eosin. CD4+ and CD8+ T cells were detected by immunoperoxidase stain using the avidin-biotin-peroxidase complex method on acetone-fixed frozen sections (C–F). Apoptotic cells were detected using TUNEL stain on frozen sections. Magnification for A–F is ×125. For G and H, magnification is ×400.

FIGURE 4.
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FIGURE 4.

Demyelination in the CNS of IgM−/− and wt mice infected with JHMV. Demyelination in the spinal cords of IgM−/− (A) and wt mice (B) at 21 days p.i. Sections were stained with luxol fast blue, and areas of demyelination are outlined by arrows.

Cellular tropism

IgM−/− mice and wt controls were infected with the JHMV 2.2v-1 variant derived by selection from a highly virulent JHMV with neutralizing mAb J.2.2 (32). A possible explanation for the recrudescence of JHMV within the CNS of the IgM−/− mice is reversion of the 2.2v-1 phenotype to a more virulent phenotype (39). To examine this possibility, mAb J.2.2 was passively administered to infected IgM−/− mice at 9, 12, and 17 days p.i. and compared with infected IgM−/− mice receiving an isotype-matched Ab with no detectable ability to recognize JHMV. These time points were chosen to parallel the initial detection of neutralizing Ab in JHMV-infected mice (6). Neither passive transfer of mAb J.2.2 nor the control Ab altered the clinical course of infection or prevented recrudescence of JHMV at 21 days p.i. (Table I⇓). Furthermore, no difference in the extent of inflammation or distribution of viral Ag was detected comparing the mAb J.2.2 recipients, the control Ab recipients, or untreated IgM−/− mice (data not shown). These data suggest that reversion to the parental phenotype was not responsible for the reactivation of JHMV in the CNS of the IgM−/− mice following initial clearance.

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Table I.

Passive transfer of mAb J.2.2 into IgM−/− mice

Altered cellular tropism of JHMV within the CNS as a contributing factor to the phenotype observed in IgM−/− mice was examined by determining the distribution of viral Ag. At day 7 p.i., the majority of viral Ag was present in oligodendrocytes, microglia, and astrocytes in both groups (Fig. 5⇓). Only rarely were neurons positive for viral Ag in either group. At 11 days p.i., few Ag-positive cells were detected in the brains of either group (Fig. 5⇓), consistent with the reduction in infectious virus (Fig. 2⇑). A similar cellular distribution of viral Ag in both groups during the acute phase of disease (7–11 days p.i.) suggests that altered cellular tropism did not contribute to the recrudescence of infectious JHMV in the absence of Ab. Similarly, only focal areas containing viral Ag-positive cells were detected within the spinal cords of wt mice at 21 days p.i. (Fig. 5⇓), consistent with the inability to recover infectious virus at this time point (Fig. 2⇑). By contrast, the brains of IgM−/− mice showed numerous Ag-positive cells (Fig. 5⇓), consistent with the presence of infectious virus (Fig. 2⇑), and the spinal cords showed a more diffuse distribution of Ag-positive cells. In both sites, virus Ag was predominantly found in the white matter; however, focal involvement of gray matter was also seen. The cellular distribution of virus Ag at 21 days p.i. in the IgM−/− mice was similar to that seen at earlier time points with predominant localization to astrocytes, oligodendrocytes, and microglia. A small number of Ag-positive neurons was also present. With the exception of viral spread to a small number of neurons, the distribution of viral Ag was comparable in the two groups, suggesting no change in cell tropism in the absence of humoral immunity.

FIGURE 5.
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FIGURE 5.

Distribution of JHMV Ag within the CNS. JHMV Ag (arrowheads) was detected in the brains of both IgM−/− (A and C) and wt mice (B and D) at days 7 (A and B) and 11 (C and D) p.i. without apparent differences in the extent or distribution of virus-infected cells. Note the decrease in Ag-positive cells at day 11 p.i. At day 21 p.i., viral Ag was localized to cells within the white matter of the spinal cord (E), but not the brain (F) of wt mice. In IgM−/− mice, viral Ag was present in the both the spinal cord (G) and brain (H). Immunoperoxidase stain used the avidin-biotin-peroxidase complex method for viral Ag (mAb J.3.3) with hematoxylin and eosin counterstain (magnification, ×125).

CTL responses in IgM−/− mice

CTL responses in the spleens of IgM−/− and wt mice were comparable at day 7 p.i. (Fig. 6⇓), consistent with the predominant role of these effectors in JHMV clearance from the CNS (6, 7). CTL activity in the spleens of wt mice was equivalent to the day 7 p.i. level at 21 days p.i. By contrast, peripheral CTL activity in IgM−/− mice was reduced compared with the levels detected in wt mice (Fig. 6⇓), suggesting the retention of CTL within the CNS due to the presence of infectious virus and viral Ag within the CNS (Figs. 2⇑ and 5⇑). Accumulation of IFN-γ mRNA and the mRNA encoding the N protein of JHMV within the CNS of infected IgM−/− mice was examined to demonstrate that the decreased peripheral CTL activity in IgM−/− mice represented continuous recruitment and/or sequestration in the CNS due to the presence of infectious virus (40). The IFN-γ mRNA levels in the CNS of IgM−/− and wt mice were approximately equivalent during acute infection and viral clearance (7, 9, and 11 days p.i.; Fig. 7⇓). The kinetics of accumulation of JHMV N mRNA paralleled the kinetics of virus replication within the CNS of both groups (Fig. 7⇓). However, while the levels of IFN-γ mRNA in wt mice decreased to below detection limits as infectious virus was eliminated (Fig. 5⇑), IFN-γ mRNA levels in the CNS of the IgM−/− mice remained elevated (Fig. 7⇓). Although both CD4+ and CD8+ T cells could contribute to IFN-γ mRNA within the CNS, CD8+ T cells are a major source of this cytokine (41, 42), suggesting that reduced splenic CTL in the IgM−/− mice is due to the presence of activated CTL within the CNS (Fig. 3⇑).

FIGURE 6.
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FIGURE 6.

CTL activity following JHMV infection. CTL activity at 7 (A) and 21 (B) days p.i. Splenocytes from four infected mice/group were pooled and restimulated for 6 days. JHMV-specific CTL activity was tested in a 4-h 51Cr release assay. EL4 (H-2b) target cells were infected with either a rVV expressing the Escherichia coli lacZ gene (vSC8) or an rVV expressing amino acids 510–518 of the JHMV S protein (vJS510).

FIGURE 7.
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FIGURE 7.

Kinetics of IFN-γ and JHMV N mRNA accumulation in the CNS of mice during JHMV infection of IgM−/− and wt mice. RNA was extracted from the spinal cords of JHMV-infected IgM−/− and wt mice at various times p.i. Levels of IFN-γ (A) and N (B) mRNA are expressed as a relative amount value for comparison. Each point represents the mean value of at least three mice per group, and error bars represent the SEM.

The possibility that CTL escape variants contributed to viral reactivation in IgM−/− mice was examined by sequence analysis of the region of the gene encoding the JHMV S protein, which contains the immunodominant H-2Db-restricted CTL epitope (amino acids 510–518) (33, 43). The 500 nucleotides of the gene (nucleotides 1420–1890) surrounding the CTL epitope (nucleotides 1528–1554) were sequenced from viruses isolated from infected IgM−/− mice. None of the viruses isolated at 7, 11, or 21 days p.i. had dominant base substitutions or deletions within the S gene region encoding the immunodominant CTL epitope. Two of three mice analyzed at 30 days p.i. had wt sequences. In a third mouse, however, sequence analysis suggested that approximately 50% of the S genes contained a mutation within the CTL epitope. Consistent with this estimate, five of nine cloned PCR products contained the identical mutation (G→T at nucleotide 1539), resulting in a W→C change at amino acid 513. None of the six cloned PCR products from another 30 day p.i. mouse had any detectable mutations within the S gene. To determine whether the W→C mutation altered CTL recognition, viruses with both the wt and mutant sequences were plaque purified from mice 30 days p.i., and the presence of wt and mutation was confirmed by sequence analysis. Target cells (IC-21 (H-2b)) infected with both viruses were recognized by JHMV-specific CTL (data not shown). These data suggest that mutations within the immunodominant CTL epitope can indeed be obtained in the absence of humoral immunity. However, the paucity of mutants obtained only well after virus reactivation supports a functional cell-mediated immune response within the CNS of infected IgM−/− mice.

Passive transfer eliminates recrudescence of infectious JHMV

Serum JHMV-neutralizing Ab is detected approximately 9 days p.i. in JHMV-infected wt mice (6). These data suggest that the Ab response contributes to preventing JHMV recrudescence. To demonstrate that the absence of an Ab response is responsible for the reactivation of infectious virus, anti-JHMV Ab was passively transferred to infected IgM−/− mice at 9, 12, and 17 days p.i., and its ability to inhibit CNS virus reactivation was determined at 21 days p.i. Control IgM−/− mice received an equal volume of ascites obtained from naive mice. IgM−/− mice that received the passive transfer of anti-JHMV Ab had reduced clinical signs compared with IgM−/− mice that received control Ab (Table II⇓). Consistent with the reduction in clinical signs, IgM−/− mice treated with anti-JHMV Ab also had either undetectable or significantly (p < 0.01) reduced levels of infectious JHMV within the CNS (Table II⇓). By contrast, high titers of infectious virus were detected in IgM−/− mice that received nonimmune ascites (Table II⇓). Anti-JHMV Ab-protected IgM−/− mice showed little or no viral Ag in the brain at 21 days p.i. (Fig. 8⇓), while extensive demyelination and viral Ag in cells morphologically consistent with oligodendrocytes were found in spinal cords (Fig. 8⇓). By contrast, both the brain and spinal cord of IgM−/− mice treated with nonimmune ascites had numerous Ag-positive cells and prominent demyelination (Fig. 8⇓) similar to untreated JHMV-infected IgM−/− mice (Figs. 3⇑ and 4⇑). These data support a direct role of the anti-viral Ab response in controlling the reactivation of JHMV within the CNS and establishing persistent JHMV infection.

FIGURE 8.
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FIGURE 8.

Immunopathology in anti-JHMV Ab-treated IgM−/− mice. Brain and spinal cords from anti-JHMV Ab-treated and control IgM−/− mice at 21 days p.i. were stained for JHMV Ag with J.3.3 mAb and counterstained with hematoxylin and eosin. Sections from IgM−/− mice treated with control Ab show extensive cellular infiltrates and numerous Ag-positive cells (arrowheads) (A and B). By contrast, fewer viral Ag-positive cells (arrowheads) were detected in the brains (C) and the spinal cords (D) of IgM−/− mice, which received the passive transfer of anti-JHMV Ab (magnification, ×125).

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Table II.

Passive transfer of anti-JHMV Ab provides protection

Discussion

The immune response to viral infection of the CNS can play dual roles in pathogenesis. Effective responses protect by eliminating the infectious agent and thereby limit the immunopathology associated with both virally induced and immune-mediated pathology (1, 6, 13, 14, 15, 44). By contrast, immune responses may either promote or maintain persistent viral infections, either as a direct response to the virus or as a consequence of the desire to limit CNS immunopathology (1, 15, 20). CNS infections of mice with JHMV result in acute encephalomyelitis accompanied by primary demyelination. In immunocompetent mice, infectious virus is generally cleared within 2 wk of infection (6, 31, 32). However, following resolution of the acute infection, survivors develop chronic demyelination associated with persistent viral Ag (45, 46, 47), but the absence of detectable infectious virus. These data suggest the anti-viral effectors are inhibited before successfully completing virus clearance, and this directly contributes to establishing persistence. Alternatively, infection of a specific CNS cell type(s) refractory to immune recognition such as oligodendroglia or neurons may contribute to viral persistence. The adoptive transfer of JHMV-specific CTL mediates protection via suppression of virus replication in astrocytes and microglia (7). However, CTL have little effect on virus replication in oligodendrocytes or within the rarely infected neuron (7, 48). IFN-γ, most likely secreted by CTL (38, 41, 42), was demonstrated to control JHMV replication in oligodendrocytes (8), but not neurons (48). Therefore, CTL, either directly via a perforin-dependent cytolytic mechanism (6) or indirectly via IFN-γ release (8), appear to be the predominant effector mechanism for virus clearance from the majority of infected cells during acute infection. Similar to infection of IgM−/− mice with CMV (49), LCMV (9, 50), and influenza B virus (51), the kinetics of viral clearance in JHMV-infected IgM−/− mice demonstrates that cellular immunity alone is sufficient to eliminate infectious virus during acute infection. However, during JHMV infection it is insufficient to control persistence.

The potential role(s) of humoral immunity has been implicated in various models of JHMV infection of the CNS. For example, resistant Brown Norway rats develop a more rapid neutralizing Ab response compared with susceptible Lewis rats (52). Suckling mice weaned on immunized dams are protected from acute encephalitis; however, approximately 40% subsequently develop chronic disease associated with the evolution of CTL escape variants (24). These data suggest that partial Ab-mediated protection could, in some instances, predispose the host to the evolution of viruses able to escape distinct effector mechanisms. However, during JHMV infection of adult mice, no evidence for CTL escape mutants was obtained even after reactivation of infectious virus due to the absence of an Ab response or in the absence of IFN-γ (8). The ability of virus-specific mAb to alter JHMV infection has also been extensively studied. Passive transfer of some, but not all, neutralizing anti-S protein mAb protect from lethal challenge and limit immunopathology (25). However, the passive transfer of nonneutralizing mAb specific for other viral structural proteins is also protective (26, 27, 28, 29). Limiting infection of neurons and increased severity of chronic disease were often observed following passive protection (25, 28). These experiments were generally conducted by mAb transfer before or concomitant with infection. However, they suggested that even in those cases in which infectious virus was not reduced, Ab may play a role in limiting immunopathology. The present study using IgM−/− mice was unable to confirm a role of humoral immunity in limiting immunopathology during resolution of acute infection. Until virus reactivation, there was no difference in the extent or distribution of mononuclear cell infiltrates or in the number of apoptotic cells comparing IgM−/− and wt mice. Analysis of JHMV infection in IgM−/− mice demonstrates an essential role for humoral immunity in preventing virus reactivation within the CNS. However, the mechanism(s) via which Ab prevents JHMV reactivation is not clear. Ab induces a variety of changes within virus-infected cells that could help establish persistence. Anti-measles virus and anti-herpes simplex virus Ab not only reduces surface glycoprotein expression, but also alters viral gene expression (19, 20). Anti-JHMV Ab prevents both cell death and cytopathology while selecting for JHMV variants in vitro (53, 54). Reduced expression of viral glycoproteins has also been observed during persistent infections with paramyxovirus, retroviruses, rhabdoviruses, and arenaviruses (55, 56, 57, 58). The immunodominant CTL epitope in H-2b mice is encoded within the JHMV S glycoprotein (33, 43); therefore, it is possible that interaction between Ab and the S protein reduces expression of the immunodominant epitope, leading to viral persistence.

The accumulation of CD8+ T cells within the CNS coincides with declining infectious virus, and depletion of CD8+ T cells prevents viral clearance (59, 60). Therefore, CTL appear to be the major effectors of protective JHMV immunity (6, 7, 38), similar to many other viral infections (1, 3). There appears to be no relative reduction of CD8+ T cells or increase in apoptotic cells within the CNS of the IgM−/− mice during reactivation of JHMV. Similar to LCMV infection of IgM−/− mice (9, 50), decreasing peripheral CTL activity was observed in IgM−/− mice infected with JHMV. Recent data demonstrated a decrease in Ag-specific IFN-γ secretion by splenocytes from IgM−/− mice chronically infected with LCMV (61). Interestingly, the number of LCMV-specific CD8+ T cells capable of IFN-γ secretion was not significantly altered. The defect in IFN-γ secretion appears more pronounced in the CD4+ T cell population during LCMV persistence (61). Increased apoptosis of CD8+ T cells within the CNS parenchyma is indicative of a defect in CD4+ T cell function during JHMV infection (38). Similar numbers of apoptotic cells were found within the CNS parenchyma of IgM−/− and wt mice. In addition, the increased IFN-γ mRNA in the CNS during JHMV reactivation suggests that the CD4+ T cell compartment is not compromised in JHMV-infected IgM−/− mice. Alternatively, increased IFN-γ mRNA in the absence of IFN-γ secretion could reflect an abortive attempt to control JHMV reactivation. Although infection with high doses of LCMV or with a rapidly replicating strain may result in a phenomena termed CTL exhaustion (9, 50, 62, 63), this mechanism appears unlikely to account for persistence during an organ-specific infection such as JHMV infection of the CNS. In contrast to LCMV infection, CD8+ and CD4+ T cells, along with high IFN-γ mRNA levels, were detected in the CNS of IgM−/− mice during virus reactivation. CD8+ T cells exhibiting virus-specific cytolysis can be activated by in vitro culture of peripheral lymphoid organs (40, 64). Sequestration, local activation, or expansion during JHMV infection was previously suggested to account for the absence of ex vivo CTL activity detected in peripheral lymphoid organs compared with cells isolated from the CNS (40). Indeed, low level direct JHMV-specific cytotoxic activity was detected ex vivo using cells isolated from the CNS of IgM−/− mice at day 35 p.i. (data not shown). These data suggest that the decrease in splenic CTL activity may result from sequestration of CTL within the CNS, and argue against Ag-driven CTL exhaustion as a cause for JHMV reactivation in the CNS of infected IgM−/− mice.

In the apparent absence of exhaustion, the inability of the CTL to affect clearance of JHMV in the absence of Ab may reflect an inability to overcome a normal homeostatic mechanism designed to prevent excessive immunopathology within the CNS or decreased expression of the S protein. Indeed, the ability of passively transferred S protein-specific mAb to protect from JHMV (25) and the ability of anti-JHMV Ab to facilitate virus persistence in vitro without S protein-mediated cell-cell fusion (53) support the possibility that decreased S protein expression facilitates JHMV persistence in the CNS. However, since CTL effector function requires MHC class I expression, clearance of virus from neurons and oligodendroglia require additional effector mechanisms. Indeed, recent data have implicated IFN-γ as the major anti-viral effector mechanism controlling JHMV infection of oligodendroglia (8). The minimal increase in infected neurons in the IgM−/− mice suggests the possibility that the Ab response may play an important role in preventing the relatively late spread of JHMV to neurons, consistent with the analysis of passively transferred virus-specific mAb before infection (25). Together, these observations suggest that, in addition to viral cytopathology (3) properties, the host cells and their phenotypes (15, 16) are also important in determining the relative importance of the Ab and cell-mediated immune responses.

In summary, results from the analysis of JHMV infection of the CNS of IgM−/− mice clearly demonstrate that cell-mediated immunity, in the absence of Ab or other related factors, effectively controls virus replication during acute infection. However, in the absence of Ab, infectious virus cannot be effectively cleared from the CNS nor is a persistent CNS infection established. This results in an apparent reactivation of infectious virus and increased mortality despite initial virus clearance from the CNS. These data raise a number of important issues relative to the regulation of immunopathology within the CNS and the mechanism via which Ab suppresses expression of infectious virus within the CNS. These data demonstrate that both cellular and humoral effector mechanisms of the anti-viral immune response play critical, yet apparently distinct roles in the resolution of CNS viral infection.

Footnotes

  • ↵1 This work was supported by Grant NS18146 from the National Institutes of Health.

  • ↵2 Address correspondence and reprint requests to Dr. Stephen Stohlman, University of Southern California School of Medicine, 1333 San Pablo Street, MCH 142, Los Angeles, CA 90033. E-mail address: stohlman{at}hsc.usc.edu

  • ↵3 Abbreviations used in this paper: LCMV, lymphocytic choriomeningitis virus; JHMV, JHM strain of mouse hepatitis virus; N, JHMV nucleocapsid protein; p.i., postinfection; rVV, recombinant vaccinia virus; S, JHMV spike envelope glycoprotein; wt, wild type.

  • Received October 29, 1998.
  • Accepted March 30, 1999.
  • Copyright © 1999 by The American Association of Immunologists

References

  1. ↵
    Zinkernagel, R. M.. 1996. Immunology taught by viruses. Science 271: 173
    OpenUrlAbstract
  2. ↵
    Kagi, D., B. Ledermann, K. Burki, P. Seiler, B. Odermatt, K. J. Olsen, E. R. Podack, R. M. Zinkernagel, H. Hengartner. 1994. Cytotoxicity mediated by T cells and natural killer cells is greatly impaired in perforin-deficient mice. Nature 369: 31
    OpenUrlCrossRefPubMed
  3. ↵
    Kagi, D., P. Seiler, J. Pavlovic, B. Ledermann, K. Burki, R. M. Zinkernagel, H. Hengartner. 1995. The roles of perforin- and Fas-dependent cytotoxicity in protection against cytopathic and noncytopathic viruses. Eur. J. Immunol. 25: 3256
    OpenUrlCrossRefPubMed
  4. ↵
    Weiner, L. P.. 1973. Pathogenesis of demyelination induced by a mouse hepatitis. Arch. Neurol. 28: 298
    OpenUrlCrossRefPubMed
  5. ↵
    Lampert, P. W., J. K. Sims, A. J. Kniazeff. 1973. Mechanism of demyelination in JHM virus encephalomyelitis. Acta Neuropathol. 24: 76
    OpenUrlCrossRefPubMed
  6. ↵
    Lin, M. T., S. A. Stohlman, D. R. Hinton. 1997. Mouse hepatitis virus is cleared from the central nervous systems of mice lacking perforin-mediated cytolysis. J. Virol. 71: 383
    OpenUrlAbstract/FREE Full Text
  7. ↵
    Stohlman, S. A., C. C. Bergmann, R. C. van der Veen, D. R. Hinton. 1995. Mouse hepatitis virus-specific cytotoxic T lymphocytes protect from lethal infection without eliminating virus from the central nervous system. J. Virol. 69: 684
    OpenUrlAbstract/FREE Full Text
  8. ↵
    Parra, B., D. R. Hinton, N. W. Marten, C. C. Bergmann, M. T. Lin, C. S. Yang, and S. A. Stohlman. 1998. γ Interferon is required for viral clearance from central nervous system oligodendroglia. J. Immunol. In press.
  9. ↵
    Thomsen, A. R., J. Johansen, O. Marker, J. P. Christensen. 1996. Exhaustion of CTL memory and recrudescence of viremia in lymphocytic choriomeningitis virus-infected MHC class II-deficient mice and B cell-deficient mice. J. Immunol. 157: 3074
    OpenUrlAbstract
  10. ↵
    Planz, O., S. Ehl, E. Furrer, E. Horvath, M. A. Brundler, H. Hengartner, R. M. Zinkernagel. 1997. A critical role for neutralizing-antibody-producing B cells, CD4+ T cells, and interferons in persistent and acute infections of mice with lymphocytic choriomeningitis virus: implications for adoptive immunotherapy of virus carriers. Proc. Natl. Acad. Sci. USA 94: 6874
    OpenUrlAbstract/FREE Full Text
  11. ↵
    Mozdzanowska, K., M. Furchner, G. Washko, J. Mozdzanowski, W. Gerhard. 1997. A pulmonary influenza virus infection in SCID mice can be cured by treatment with hemagglutinin-specific antibodies that display very low virus-neutralizing activity in vitro. J. Virol. 71: 4347
    OpenUrlAbstract/FREE Full Text
  12. ↵
    Doherty, P. C., D. J. Topham, R. A. Tripp, R. D. Cardin, J. W. Brooks, P. G. Stevenson. 1997. Effector CD4+ and CD8+ T-cell mechanisms in the control of respiratory virus infections. Immunol. Rev. 159: 105
    OpenUrlCrossRefPubMed
  13. ↵
    Amor, S., M. F. Scallan, M. M. Morris, H. Dyson, J. K. Fazakerley. 1996. Role of immune responses in protection and pathogenesis during Semliki Forest virus encephalitis. J. Gen. Virol. 77: 281
    OpenUrlAbstract/FREE Full Text
  14. ↵
    Griffin, D., B. Levine, W. Tyor, S. Ubol, P. Despres. 1997. The role of antibody in recovery from alphavirus encephalitis. Immunol. Rev. 159: 155
    OpenUrlCrossRefPubMed
  15. ↵
    Sedgwick, J. D., R. Dorries. 1991. The immune system response to viral infection of the CNS. Neuroscience 3: 93
  16. ↵
    Joly, E., L. Mucke, M. B. Oldstone. 1991. Viral persistence in neurons explained by lack of major histocompatibility class I expression. Science 253: 1283
    OpenUrlAbstract/FREE Full Text
  17. ↵
    Moskophidis, D., M. Battegay, M. van den Broek, E. Laine, U. Hoffmann-Rohrer, R. M. Zinkernagel. 1995. Role of virus and host variables in virus persistence or immunopathological disease caused by a non-cytolytic virus. J. Gen. Virol. 76: 381
    OpenUrlAbstract/FREE Full Text
  18. ↵
    Levine, B., J. M. Hardwick, B. D. Trapp, T. O. Crawford, R. C. Bollinger, D. E. Griffin. 1991. Antibody-mediated clearance of alphavirus infection from neurons. Science 254: 856
    OpenUrlAbstract/FREE Full Text
  19. ↵
    Fujinami, R. S., M. B. Oldstone. 1979. Antiviral antibody reacting on the plasma membrane alters measles virus expression inside the cell. Nature 279: 529
    OpenUrlCrossRefPubMed
  20. ↵
    Liebert, U. G., S. Schneider-Schaulies, K. Baczko, V. ter Meulen. 1990. Antibody-induced restriction of viral gene expression in measles encephalitis in rats. J. Virol. 64: 706
    OpenUrlAbstract/FREE Full Text
  21. ↵
    Levine, B., D. E. Griffin. 1993. Molecular analysis of neurovirulent strains of Sindbis virus that evolve during persistent infection of scid mice. J. Virol. 67: 6872
    OpenUrlAbstract/FREE Full Text
  22. ↵
    Garcia-Blanco, M. A., B. R. Cullen. 1991. Molecular basis of latency in pathogenic human viruses. Science 254: 815
    OpenUrlAbstract/FREE Full Text
  23. ↵
    Narayan, O., M. C. Zink, D. Huso, D. Sheffer, S. Crane, S. Kennedy-Stoskopf, P. E. Jolly, J. E. Clements. 1988. Lentiviruses of animals are biological models of the human immunodeficiency viruses. Microb. Pathog. 5: 149
    OpenUrlCrossRefPubMed
  24. ↵
    Pewe, L., G. F. Wu, E. M. Barnett, R. F. Castro, S. Perlman. 1996. Cytotoxic T cell-resistant variants are selected in a virus-induced demyelinating disease. Immunity 5: 253
    OpenUrlCrossRefPubMed
  25. ↵
    Buchmeier, M. J., H. A. Lewicki, P. J. Talbot, R. L. Knobler. 1984. Murine hepatitis virus-4 (strain JHM)-induced neurologic disease is modulated in vivo by monoclonal antibody. Virology 132: 261
    OpenUrlCrossRefPubMed
  26. ↵
    Nakanaga, K., K. Yamanouchi, K. Fujiwara. 1986. Protective effect of monoclonal antibodies on lethal mouse hepatitis virus infection in mice. J. Virol. 59: 168
    OpenUrlAbstract/FREE Full Text
  27. ↵
    Lecomte, J., V. Cainelli-Gewbara, G. Mercier, S. Mansour, P. J. Talbot, G. Lussier, D. Oth. 1987. Protection from mouse hepatitis virus type 3-induced acute disease by an anti-nucleoprotein monoclonal antibody. Arch. Virol. 97: 123
    OpenUrlCrossRefPubMed
  28. ↵
    Fleming, J. O., R. A. Shubin, M. A. Sussman, N. Casteel, S. A. Stohlman. 1989. Monoclonal antibodies to the matrix (E1) glycoprotein of mouse hepatitis virus protect mice from encephalitis. Virology 168: 162
    OpenUrlCrossRefPubMed
  29. ↵
    Yokomori, K., S. C. Baker, S. A. Stohlman, M. M. C. Lai. 1992. Hemagglutinin-esterase-specific monoclonal antibodies alter the neuropathogenicity of mouse hepatitis virus. J. Virol. 66: 2865
    OpenUrlAbstract/FREE Full Text
  30. ↵
    Kitamura, D., K. Rajewsky. 1992. Targeted disruption of μ chain membrane exon causes loss of heavy-chain allelic exclusion. Nature 356: 154
    OpenUrlCrossRefPubMed
  31. ↵
    Parra, B., D. R. Hinton, M. T. Lin, D. J. Cua, S. A. Stohlman. 1997. Kinetics of cytokine mRNA expression in the central nervous system following lethal and nonlethal coronavirus-induced acute encephalomyelitis. Virology 233: 260
    OpenUrlCrossRefPubMed
  32. ↵
    Fleming, J. O., M. Trousdale, F. El-Zaatari, S. A. Stohlman, L. P. Weiner. 1986. Pathogenicity of antigenic variants of murine coronavirus JHM selected with monoclonal antibodies. J. Virol. 58: 869
    OpenUrlAbstract/FREE Full Text
  33. ↵
    Bergmann, C. C., Q. Yao, M. Lin, S. A. Stohlman. 1996. The JHM strain of mouse hepatitis virus induces a spike protein-specific Db-restricted cytotoxic T cell response. J. Gen. Virol. 77: 315
    OpenUrlAbstract/FREE Full Text
  34. ↵
    Fleming, J. O., S. A. Stohlman, R. C. Harmon, M. M. C. Lai, J. A. Frelinger, L. P. Weiner. 1983. Antigenic relationships of murine coronaviruses: analysis using monoclonal antibodies to JHM (MHV-4) virus. Virology 131: 296
    OpenUrlCrossRefPubMed
  35. ↵
    Childs, J. C., S. A. Stohlman, L. Kingsford, R. Russell. 1983. Antigenic relationships of murine coronaviruses. Arch. Virol. 78: 81
    OpenUrlCrossRefPubMed
  36. ↵
    Chiewsilp, D., J. M. McCown. 1972. Elimination of repeated clot formation in mouse ascitic fluid containing arbovirus antibodies. Appl. Microbiol. 24: 288
    OpenUrlPubMed
  37. ↵
    Stohlman, S. A., C. C. Bergmann, D. Cua, H. Wege, R. van der Veen. 1994. Location of antibody epitopes within the mouse hepatitis virus nucleocapsid protein. Virology 202: 146
    OpenUrlCrossRefPubMed
  38. ↵
    Stohlman, S. A., C. C. Bergmann, M. T. Lin, D. J. Cua, D. R. Hinton. 1998. CTL effector function within the central nervous system requires CD4+ T cells. J. Immunol. 160: 2896
    OpenUrlAbstract/FREE Full Text
  39. ↵
    Wang, F. I., J. O. Fleming, M. M. C. Lai. 1992. Sequence analysis of the spike protein gene of murine coronavirus variants: study of genetic sites affecting neuropathogenicity. Virology 186: 742
    OpenUrlCrossRefPubMed
  40. ↵
    Stohlman, S. A., S. Kyuwa, J. M. Polo, D. Brady, M. M. C. Lai, C. C. Bergmann. 1993. Characterization of mouse hepatitis virus-specific cytotoxic T cells derived from the central nervous system of mice infected with the JHM strain. J. Virol. 67: 7050
    OpenUrlAbstract/FREE Full Text
  41. ↵
    Ruby, J., I. Ramshaw. 1991. The antiviral activity of immune CD8+ T cells is dependent on interferon-γ. Lymphokine Cytokine Res. 10: 353
    OpenUrlPubMed
  42. ↵
    Young, H. A., K. J. Hardy. 1995. Role of interferon-γ in immune cell regulation. J. Leukocyte Biol. 58: 373
    OpenUrlAbstract
  43. ↵
    Castro, R. F., S. Perlman. 1995. CD8+ T-cell epitopes within the surface glycoprotein of a neurotropic coronavirus and correlation with pathogenicity. J. Virol. 69: 8127
    OpenUrlAbstract/FREE Full Text
  44. ↵
    Dethlefs, S., M. Brahic, E. L. Larsson-Sciard. 1997. An early, abundant cytotoxic T-lymphocyte response against Theiler’s virus is critical for preventing viral persistence. J. Virol. 71: 8875
    OpenUrlAbstract/FREE Full Text
  45. ↵
    Wang, F., D. Hinton, W. Gilmore, M. Trousdale, J. Fleming. 1992. Sequential infection of glial cells by the murine hepatitis virus JHM strain (MHV-4) leads to a characteristic distribution of demyelination. Lab. Invest. 66: 744
    OpenUrlPubMed
  46. ↵
    Perlman, S., D. Ries. 1987. The astrocyte is a target cell in mice persistently infected with mouse hepatitis virus, strain JHM. Microb. Pathog. 3: 309
    OpenUrlCrossRefPubMed
  47. ↵
    Stohlman, S. A., L. P. Weiner. 1981. Chronic central nervous system demyelination in mice after JHM virus infection. Neurology 31: 38
    OpenUrlAbstract/FREE Full Text
  48. ↵
    Lane, T. E., A. D. Paoletti, M. J. Buchmeier. 1997. Disassociation between the in vitro and in vivo effects of nitric oxide on a neurotropic murine coronavirus. J. Virol. 71: 2202
    OpenUrlAbstract/FREE Full Text
  49. ↵
    Jonjic, S., I. Pavic, B. Polic, I. Crnkovic, P. Lucin, U. H. Koszinowski. 1994. Antibodies are not essential for the resolution of primary cytomegalovirus infection but limit dissemination of recurrent virus. J. Exp. Med. 179: 1713
    OpenUrlAbstract/FREE Full Text
  50. ↵
    Brundler, M. A., P. Aichele, M. Bachmann, D. Kitamura, K. Rajewsky, R. M. Zinkernagel. 1996. Immunity to viruses in B cell-deficient mice: influence of antibodies on virus persistence and on T cell memory. Eur. J. Immunol. 26: 2257
    OpenUrlCrossRefPubMed
  51. ↵
    Epstein, S. L., C.-Y. Lo, J. A. Misplon, J. R. Bennink. 1998. Mechanism of protective immunity against influenza virus infection in mice without antibodies. J. Immunol. 160: 322
    OpenUrlAbstract/FREE Full Text
  52. ↵
    Schwender, S., H. Imrich, R. Dorries. 1991. The pathogenic role of virus-specific antibody-secreting cells in the central nervous system of rats with different susceptibility to coronavirus-induced demyelinating encephalitis. Immunology 74: 533
    OpenUrlPubMed
  53. ↵
    Stohlman, S. A., L. P. Weiner. 1978. Stability of neurotropic mouse hepatitis virus (JHM strain) during chronic infection of neuroblastoma cells. Arch. Virol. 57: 53
    OpenUrlCrossRefPubMed
  54. ↵
    Stohlman, S., A. Sakaguchi, L. Weiner. 1979. Characterization of the cold-sensitive murine hepatitis virus mutants rescued from latently infected cells by cell fusion. Virology 98: 448
    OpenUrlCrossRefPubMed
  55. ↵
    O’Rourke, E. J., W. H. Guo, A. S. Huang. 1983. Antibody-induced modulation of proteins in vesicular stomatitis virus-infected fibroblasts. Mol. Cell. Biol. 3: 1580
    OpenUrlAbstract/FREE Full Text
  56. ↵
    Roux, L., P. Beffy, A. Portner. 1984. Restriction of cell surface expression of Sendai virus hemagglutinin-neuraminidase glycoprotein correlates with its higher instability in persistently and standard plus defective interfering virus infected BHK-21 cells. Virology 138: 118
    OpenUrlCrossRefPubMed
  57. ↵
    Stevenson, M., C. Meier, A. M. Mann, N. Chapman, A. Wasiak. 1988. Envelope glycoprotein of HIV induces interference and cytolysis resistance in CD4+ cells: mechanism for persistence in AIDS. Cell 53: 483
    OpenUrlCrossRefPubMed
  58. ↵
    Oldstone, M. B., M. J. Buchmeier. 1982. Restricted expression of viral glycoprotein in cells of persistently infected mice. Nature 300: 360
    OpenUrlCrossRefPubMed
  59. ↵
    Williamson, J. S., K. C. Sykes, S. A. Stohlman. 1991. Characterization of brain-infiltrating mononuclear cells during infection with mouse hepatitis virus strain JHM. J. Neuroimmunol. 32: 199
    OpenUrlCrossRefPubMed
  60. ↵
    Dorries, R., S. Schwender, H. Imrich, H. Harms. 1991. Population dynamics of lymphocyte subsets in the central nervous system of rats with different susceptibility to coronavirus-induced demyelinating encephalitis. Immunology 74: 539
    OpenUrlPubMed
  61. ↵
    Homann, D., A. Tishon, D. P. Berger, W. O. Weigle, M. G. von Herrath, M. B. A. Oldstone. 1998. Evidence of an underlying CD4 helper and CD8 T-cell defect in B-cell-deficient mice: failure to clear persistent virus infection after adoptive immunotherapy with virus-specific memory cells from μMT/μMT mice. J. Virol. 72: 9208
    OpenUrlAbstract/FREE Full Text
  62. ↵
    Moskophidis, D., F. Lechner, H. Pircher, R. M. Zinkernagel. 1993. Virus persistence in acutely infected immunocompetent mice by exhaustion of antiviral cytotoxic effector T cells. Nature 362: 758
    OpenUrlCrossRefPubMed
  63. ↵
    Moskophidis, D., F. Lechner, H. Hengartner, R. M. Zinkernagel. 1994. MHC class I and non-MHC-linked capacity for generating an anti-viral CTL response determines susceptibility to CTL exhaustion and establishment of virus persistence in mice. J. Immunol. 152: 4976
    OpenUrlAbstract
  64. ↵
    Castro, R. F., S. Perlman. 1996. Differential antigen recognition by T cells from the spleen and central nervous system of coronavirus-infected mice. Virology 222: 247
    OpenUrlCrossRefPubMed
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The Journal of Immunology: 162 (12)
The Journal of Immunology
Vol. 162, Issue 12
15 Jun 1999
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Antibody Prevents Virus Reactivation Within the Central Nervous System
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Antibody Prevents Virus Reactivation Within the Central Nervous System
Mark T. Lin, David R. Hinton, Norman W. Marten, Cornelia C. Bergmann, Stephen A. Stohlman
The Journal of Immunology June 15, 1999, 162 (12) 7358-7368;

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Antibody Prevents Virus Reactivation Within the Central Nervous System
Mark T. Lin, David R. Hinton, Norman W. Marten, Cornelia C. Bergmann, Stephen A. Stohlman
The Journal of Immunology June 15, 1999, 162 (12) 7358-7368;
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Print ISSN 0022-1767        Online ISSN 1550-6606