Impaired T cell immunity in B cell-deficient mice following viral central nervous system infection

CD8(+) T cells are required to control acute viral replication in the CNS following infection with neurotropic coronavirus. By contrast, studies in B cell-deficient (muMT) mice revealed Abs as key effectors in suppressing virus recrudescence. The apparent loss of initial T cell-mediated immune control in the absence of B cells was investigated by comparing T cell populations in CNS mononuclear cells from infected muMT and wild-type mice. Following viral recrudescence in muMT mice, total CD8(+) T cell numbers were similar to those of wild-type mice that had cleared infectious virus; however, virus-specific T cells were reduced at least 3-fold by class I tetramer and IFN-gamma ELISPOT analysis. Although overall T cell recruitment into the CNS of muMT mice was not impaired, discrepancies in frequencies of virus-specific CD8(+) T cells were most severe during acute infection. Impaired ex vivo cytolytic activity of muMT CNS mononuclear cells, concomitant with reduced frequencies, implicated IFN-gamma as the primary anti viral factor early in infection. Reduced virus-specific CD8(+) T cell responses in the CNS coincided with poor peripheral expansion and diminished CD4(+) T cell help. Thus, in addition to the lack of Ab, limited CD8(+) and CD4(+) T cell responses in muMT mice contribute to the ultimate loss of control of CNS infection. Using a model of virus infection restricted to the CNS, the results provide novel evidence for a role of B cells in regulating T cell expansion and differentiation into effector cells.     

Mechanisms of central nervous system viral persistence: the critical role of antibody and B cells.

Contributions of humoral and cellular immunity in controlling neurotropic mouse hepatitis virus persistence within the CNS were determined in B cell-deficient J(H)D and syngeneic H-2(d) B cell+ Ab-deficient mice. Virus clearance followed similar kinetics in all mice, confirming initial control of virus replication by cellular immunity. Nevertheless, virus reemerged within the CNS of all Ab-deficient mice. In contrast to diminished T cell responses in H-2(b) B cell-deficient muMT mice, the absence of B cells or Ab in the H-2(d) mice did not compromise expansion, recruitment into the CNS, or function of virus-specific CD4+ and CD8+ T cells. The lack of B cells and lymphoid architecture thus appears to manifest itself on T cell responses in a genetically biased manner. Increasing viral load did not enhance frequencies or effector function of virus-specific T cells within the CNS, indicating down-regulation of T cell responses. Although an Ab-independent antiviral function of B cells was not evident during acute infection, the presence of B cells altered CNS cellular tropism during viral recrudescence. Reemerging virus localized almost exclusively to oligodendroglia in B cell+ Ab-deficient mice, whereas it also replicated in astrocytes in B cell-deficient mice. Altered tropism coincided with distinct regulation of CNS virus-specific CD4+ T cells. These data conclusively demonstrate that the Ab component of humoral immunity is critical in preventing virus reactivation within CNS glial cells. B cells themselves may also play a subtle role in modulating pathogenesis by influencing tropism.     

Antibody-independent antiviral function of memory CD4+ T cells in vivo requires regulatory signals from CD8+ effector T cells

Previous studies have shown that vaccine-primed CD4(+) T cells can mediate accelerated clearance of respiratory virus infection. However, the relative contributions of Ab and CD8(+) T cells, and the mechanism of viral clearance, are poorly understood. Here we show that control of a Sendai virus infection by primed CD4(+) T cells is mediated through the production of IFN-gamma and does not depend on Ab. This effect is critically dependent on CD8(+) cells for the expansion of CD4(+) T cells in the lymph nodes and the recruitment of memory CD4(+) T cells to the lungs. Passive transfer of a CD8(+) T cell supernatant into CD8(+) T cell-depleted, hemagglutinin-neuraminidase (HN)(421-436)-immune muMT mice substantially restored the virus-specific memory CD4(+) response and enhanced viral control in the lung. Together, the data demonstrate for the first time that in vivo primed CD4(+) T cells have the capacity to control a respiratory virus infection in the lung by an Ab-independent mechanism, provided that CD8(+) T cell "help" in the form of soluble factor(s) is available during the virus infection. These studies highlight the importance of synergistic interactions between CD4(+) and CD8(+) T cell subsets in the generation of optimal antiviral immunity.     

Perforin and gamma interferon-mediated control of coronavirus central nervous system infection by CD8 T cells in the absence of CD4 T cells

Infection of the central nervous system (CNS) with the neurotropic JHM strain of mouse hepatitis virus produces acute and chronic demyelination. The contributions of perforin-mediated cytolysis and gamma interferon (IFN-gamma) secretion by CD8(+) T cells to the control of infection and the induction of demyelination were examined by adoptive transfer into infected SCID recipients. Untreated SCID mice exhibited uncontrolled virus replication in all CNS cell types but had little or no demyelination. Memory CD8(+) T cells from syngeneic wild-type (wt), perforin-deficient, or IFN-gamma-deficient (GKO) donors all trafficked into the infected CNS in the absence of CD4(+) T cells and localized to similar areas. Although CD8(+) T cells from all three donors suppressed virus replication in the CNS, GKO CD8(+) T cells expressed the least antiviral activity. A distinct viral antigen distribution in specific CNS cell types revealed different mechanisms of viral control. While wt CD8(+) T cells inhibited virus replication in all CNS cell types, cytolytic activity in the absence of IFN-gamma suppressed the infection of astrocytes, but not oligodendroglia. In contrast, cells that secreted IFN-gamma but lacked cytolytic activity inhibited replication in oligodendroglia, but not astrocytes. Demyelination was most severe following viral control by wt CD8(+) T cells but was independent of macrophage infiltration. These data demonstrate the effective control of virus replication by CD8(+) T cells in the absence of CD4(+) T cells and support the necessity for the expression of distinct effector mechanisms in the control of viral replication in distinct CNS glial cell types.     

Roles of CD4+ T-cell-independent and -dependent antibody responses in the control of influenza virus infection: evidence for noncognate CD4+ T-cell activities that enhance the therapeutic activity of antiviral antibodies

Previous studies have indicated that B cells make a significant contribution to the resolution of influenza virus infection. To determine how B cells participate in the control of the infection, we transferred intact, major histocompatibility complex class II (MHC-II)-negative or B-cell receptor (BCR)-transgenic spleen cells into B-cell-deficient and CD8(+) T-cell-depleted muMT mice, termed muMT(-8), and tested them for ability to recover from infection. muMT(-8) mice that received no spleen cells invariably succumbed to the infection within 20 days, indicating that CD4(+) T-cell activities had no significant therapeutic activity on their own; in fact, they were harmful and decreased survival time. Interestingly, however, they became beneficial in the presence of antiviral antibody (Ab). Injection of MHC-II((-/-)) spleen cells, which can provide CD4(+) T-cell-independent (TI) but not T-cell-dependent (TD) activities, delayed mortality but only rarely resulted in clearance of the infection. By contrast, 80% of muMT(-8) mice injected with normal spleen cells survived and resolved the infection. Transfer of BCR-transgenic spleen cells, which contained approximately 10 times fewer virus-specific precursor B cells than normal spleen cells, had no significant impact on the course of the infection. Taken together, the results suggest that B cells contribute to the control of the infection mainly through production of virus-specific Abs and that the TD Ab response is therapeutically more effective than the TI response. In addition, CD4(+) T cells appear to contribute, apart from promoting the TD Ab response, by improving the therapeutic activity of Ab-mediated effector mechanisms.     

Contributions of CD8+ T cells and viral spread to demyelinating disease

Acute and chronic demyelination are hallmarks of CNS infection by the neurotropic JHM strain of mouse hepatitis virus. Although infectious virus is cleared by CD8+ T cells, both viral RNA and activated CD8+ T cells remain in the CNS during persistence potentially contributing to pathology. To dissociate immune from virus-mediated determinants initiating and maintaining demyelinating disease, mice were infected with two attenuated viral variants differing in a hypervariable region of the spike protein. Despite similar viral replication and tropism, one infection was marked by extensive demyelination and paralysis, whereas the other resulted in no clinical symptoms and minimal neuropathology. Mononuclear cells from either infected brain exhibited virus specific ex vivo cytolytic activity, which was rapidly lost during viral clearance. As revealed by class I tetramer technology the paralytic variant was superior in inducing specific CD8+ T cells during the acute disease. However, after infectious virus was cleared, twice as many virus-specific IFN-gamma-secreting CD8+ T cells were recovered from the brains of asymptomatic mice compared with mice undergoing demyelination, suggesting that IFN-gamma ameliorates rather than perpetuates JHM strain of mouse hepatitis virus-induced demyelination. The present data thus indicate that in immunocompetent mice, effector CD8+ T cells control infection without mediating either clinical disease or demyelination. In contrast, demyelination correlated with early and sustained infection of the spinal cord. Rapid viral spread, attributed to determinants within the spike protein and possibly perpetuated by suboptimal CD8+ T cell effector function, thus ultimately leads to the process of immune-mediated demyelination.     

Recruitment kinetics and composition of antibody-secreting cells within the central nervous system following viral encephalomyelitis

Infection by the neurotropic JHM strain of mouse hepatitis virus produces an acute demyelinating encephalomyelitis. While cellular immunity initially eliminates infectious virus, CNS viral persistence is predominantly controlled by humoral immunity. To better understand the distinct phases of immune control within the CNS, the kinetics of humoral immune responses were determined in infected mice. Early during clearance of the JHM strain of mouse hepatitis virus, only few virus-specific Ab-secreting cells (ASC) were detected in the periphery or CNS, although mature B cells and ASC without viral specificity were recruited into the CNS concomitant with T cells. Serum antiviral Ab and CNS virus-specific ASC became prominent only during final elimination of infectious virus. Virus-specific ASC peaked in lymphoid organs before the CNS, suggesting peripheral B cell priming and maturation. Following elimination of infectious virus, virus-specific ASC continued to increase within the CNS and then remained stable during persistence, in contrast to declining T cell numbers. These data comprise three novel findings. Rapid recruitment of B cells in the absence of specific Ab secretion supports a potential Ab-independent effector function involving lysis of virus-infected cells. Delayed recruitment relative to viral clearance and subsequent maintenance of a stable CNS ASC population demonstrate differential regulation of T and B lymphocytes within the infected CNS. This supports a critical role of humoral immunity in regulating viral CNS persistence. Lastly, altered antiviral ASC specificities following clearance of infectious virus suggest ongoing recruitment of peripheral memory cells and/or local B cell differentiation.     

Perforin-mediated effector function within the central nervous system requires IFN-gamma-mediated MHC up-regulation

CD8(+) T cells infiltrating the CNS control infection by the neurotropic JHM strain of mouse hepatitis virus. Differential susceptibility of infected cell types to clearance by perforin or IFN-gamma uncovered distinct, nonredundant roles for these antiviral mechanisms. To separately evaluate each effector function specifically in the context of CD8(+) T cells, pathogenesis was analyzed in mice deficient in both perforin and IFN-gamma (PKO/GKO) or selectively reconstituted for each function by transfer of CD8(+) T cells. Untreated PKO/GKO mice were unable to control the infection and died of lethal encephalomyelitis within 16 days, despite substantially higher CD8(+) T cell accumulation in the CNS compared with controls. Uncontrolled infection was associated with limited MHC class I up-regulation and an absence of class II expression on microglia, coinciding with decreased CD4(+) T cells in CNS infiltrates. CD8(+) T cells from perforin-deficient and wild-type donors reduced virus replication in PKO/GKO recipients. By contrast, IFN-gamma-deficient donor CD8(+) T cells did not affect virus replication. The inability of perforin-mediated mechanisms to control virus in the absence of IFN-gamma coincided with reduced class I expression. These data not only confirm direct antiviral activity of IFN-gamma within the CNS but also demonstrate IFN-gamma-dependent MHC surface expression to guarantee local T cell effector function in tissues inherently low in MHC expression. The data further imply that IFN-gamma plays a crucial role in pathogenesis by regulating the balance between virus replication in oligodendrocytes, CD8(+) T cell effector function, and demyelination.     

CD4+ T-cell responses are required for clearance of West Nile virus from the central nervous system

Although studies have established that innate and adaptive immune responses are important in controlling West Nile virus (WNV) infection, the function of CD4(+) T lymphocytes in modulating viral pathogenesis is less well characterized. Using a mouse model, we examined the role of CD4(+) T cells in coordinating protection against WNV infection. A genetic or acquired deficiency of CD4(+) T cells resulted in a protracted WNV infection in the central nervous system (CNS) that culminated in uniform lethality by 50 days after infection. Mice surviving past day 10 had high-level persistent WNV infection in the CNS compared to wild-type mice, even 45 days following infection. The absence of CD4(+) T-cell help did not affect the kinetics of WNV infection in the spleen and serum, suggesting a role for CD4-independent clearance mechanisms in peripheral tissues. WNV-specific immunoglobulin M (IgM) levels were similar to those of wild-type mice in CD4-deficient mice early during infection but dropped approximately 20-fold at day 15 postinfection, whereas IgG levels in CD4-deficient mice were approximately 100- to 1,000-fold lower than in wild-type mice throughout the course of infection. WNV-specific CD8(+) T-cell activation and trafficking to the CNS were unaffected by the absence of CD4(+) T cells at day 9 postinfection but were markedly compromised at day 15. Our experiments suggest that the dominant protective role of CD4(+) T cells during primary WNV infection is to provide help for antibody responses and sustain WNV-specific CD8(+) T-cell responses in the CNS that enable viral clearance.     

Role of viral persistence in retaining CD8(+) T cells within the central nervous system

The continued presence of virus-specific CD8(+) T cells within the central nervous system (CNS) following resolution of acute viral encephalomyelitis implicates organ-specific retention. The role of viral persistence in locally maintaining T cells was investigated by infecting mice with either a demyelinating, paralytic (V-1) or nonpathogenic (V-2) variant of a neurotropic mouse hepatitis virus, which differ in the ability to persist within the CNS. Class I tetramer technology revealed more infiltrating virus-specific CD8(+) T cells during acute V-1 compared to V-2 infection. However, both total and virus-specific CD8(+) T cells accumulated at similar peak levels in spinal cords by day 10 postinfection (p.i.). Decreasing viral RNA levels in both brains and spinal cords following initial virus clearance coincided with an overall progressive loss of both total and virus-specific CD8(+) T cells. By 9 weeks p.i., T cells had largely disappeared from brains of both infected groups, consistent with the decline of viral RNA. T cells also completely disappeared from V-2-infected spinal cords coincident with the absence of viral RNA. By contrast, a significant number of CD8(+) T cells which contained detectable viral RNA were recovered from spinal cords of V-1-infected mice. The data indicate that residual virus from a primary CNS infection is a vital component in mediating local retention of both CD8(+) and CD4(+) T cells and that once minimal thresholds of stimuli are lost, T cells within the CNS cannot survive in an autonomous fashion.     

CD4 T cells contribute to virus control and pathology following central nervous system infection with neurotropic mouse hepatitis virus

Replication of the neurotropic mouse hepatitis virus strain JHM (JHMV) is controlled primarily by CD8(+) T-cell effectors utilizing gamma interferon (IFN-gamma) and perforin-mediated cytotoxicity. CD4(+) T cells provide an auxiliary function(s) for CD8(+) T-cell survival; however, their direct contribution to control of virus replication and pathology is unclear. To examine a direct role of CD4(+) T cells in viral clearance and pathology, pathogenesis was compared in mice deficient in both perforin and IFN-gamma that were selectively reconstituted for these functions via transfer of virus-specific memory CD4(+) T cells. CD4(+) T cells from immunized wild-type, perforin-deficient, and IFN-gamma-deficient donors all initially reduced virus replication. However, prolonged viral control by IFN-gamma-competent donors suggested that IFN-gamma is important for sustained virus control. Local release of IFN-gamma was evident by up-regulation of class II molecules on microglia in recipients of IFN-gamma producing CD4(+) T cells. CD4(+) T-cell-mediated antiviral activity correlated with diminished clinical symptoms, pathology, and demyelination. Both wild-type donor CD90.1 and recipient CD90.2 CD4(+) T cells trafficked into the central nervous system (CNS) parenchyma and localized to infected white matter, correlating with decreased numbers of virus-infected oligodendrocytes in the CNS. These data support a direct, if limited, antiviral role for CD4(+) T cells early during acute JHMV encephalomyelitis. Although the antiviral effector mechanism is initially independent of IFN-gamma secretion, sustained control of CNS virus replication by CD4(+) T cells requires IFN-gamma.     

Regulatory T cells selectively preserve immune privilege of self-antigens during viral central nervous system infection

Regulatory T cells (Tregs) are important for the attenuation of immune reactions. During viral CNS infections, however, an indiscriminate maintenance of CNS immune privilege through Treg-mediated negative regulation could prevent autoimmune sequelae but impair the control of viral replication. We analyzed in this study the impact of Tregs on the development of acute viral encephalomyelitis, T cell-mediated antiviral protection, and prevention of CNS autoimmunity following intranasal infection with the gliatropic mouse hepatitis virus strain A59. To assess the contribution of Tregs in vivo, we specifically depleted CD4(+)Foxp3(+) T cells in a diphtheria toxin-dependent manner. We found that depletion of Tregs had no impact on viral distribution and clearance and did not significantly alter virus-specific CD4(+) and CD8(+) T cell responses. However, Treg depletion led to a more severe CNS inflammation associated with neuronal damage. Dissection of the underlying immunopathological mechanisms revealed the elaborate Treg-dependent regulation of self-reactive CD4(+) T cell proliferation within the CNS-draining lymph node and downtuning of CXCR3 expression on T cells. Taken together, these results suggest that Tregs preserve CNS immune privilege through selective control of CNS-specific Th cells while keeping protective antiviral immunity fully operative.     

Synergistic roles of antibody and interferon in noncytolytic clearance of Sindbis virus from different regions of the central nervous system

Sindbis virus (SINV) is an alphavirus that causes infection of neurons and encephalomyelitis in adult immunocompetent mice. Recovery can occur without apparent neurological damage. To better define the factors facilitating noncytolytic clearance of SINV in different regions of the central nervous system (CNS) and the roles of innate and adaptive immune responses at different times during infection, we have characterized SINV infection and clearance in the brain, brain stem, and spinal cords of severe combined immunodeficiency (SCID) and C57BL/6 (wild-type [WT]) mice and mice deficient in beta interferon (IFN-beta) (BKO), antibody (muMT), IFN-gamma (GKO), IFN-gamma receptor (GRKO), and both antibody and IFN-gamma (muMT/GKO). WT mice cleared infectious virus by day 8, while SCID mice had persistent virus replication at all sites. For 3 days after infection, BKO mice had higher titers at all sites than WT mice, despite similar IFN-alpha production, but cleared virus similarly. GKO and GRKO mice cleared infectious virus from all sites by days 8 to 10 and, like WT mice, displayed transient reactivation at 12 to 22 days. muMT mice did not clear virus from the brain, and clearance from the brain stem and lumbar spinal cord was delayed, followed by reactivation. Eighty-one days after infection, muMT/GKO mice had not cleared virus from any site, but titers were lower than for SCID mice. These studies show that IFN-beta is independently important for early control of CNS virus replication, that antiviral antibody is critical for clearance from the brain, and that both antibody and IFN-gamma contribute to prevention of reactivation after initial clearance.     

CD8(+) T lymphocytes mediate Borna disease virus-induced immunopathology independently of perforin

Perforin-mediated lysis of target cells is the major antiviral effector mechanism of CD8(+) T lymphocytes. We have analyzed the role of perforin in a mouse model for CD8(+) T-cell-mediated central nervous system (CNS) immunopathology induced by Borna disease virus. When a defective perforin gene was introduced into the genetic background of the Borna disease-susceptible mouse strain MRL, the resulting perforin-deficient mice developed strong neurological disease in response to infection indistinguishable from that of their perforin-expressing littermates. The onset of disease was slightly delayed. Brains of diseased perforin-deficient mice showed similar amounts and a similar distribution of CD8(+) T cells as wild-type animals. Perforin deficiency had no impact on the kinetics of viral spread through the CNS. Unlike brain lymphocytes from diseased wild-type mice, lymphocytes from perforin-deficient MRL mice showed no in vitro cytolytic activity towards target cells expressing the nucleoprotein of Borna disease virus. Taken together, these results demonstrate that CD8(+) T cells mediate Borna disease independent of perforin. They further suggest that the pathogenic potential of CNS-infiltrating CD8(+) T cells does not primarily reside in their lytic activity but rather in other functions.     

Effects of an epitope-specific CD8+ T-cell response on murine coronavirus central nervous system disease: protection from virus replication and antigen spread and selection of epitope escape mutants

Both CD4(+) and CD8(+) T cells are required for clearance of the murine coronavirus mouse hepatitis virus (MHV) during acute infection. We investigated the effects of an epitope-specific CD8(+) T-cell response on acute infection of MHV, strain A59, in the murine CNS. Mice with CD8(+) T cells specific for gp33-41 (an H-2D(b)-restricted CD8(+) T-cell epitope derived from lymphocytic choriomeningitis glycoprotein) were infected with a recombinant MHV-A59, also expressing gp33-41, as a fusion protein with enhanced green fluorescent protein (EGFP). By 5 days postinfection, these mice showed significantly (approximately 20-fold) lower titers of infectious virus in the brain compared to control mice. Furthermore mice with gp33-41-specific CD8(+) cells exhibited much reduced levels of viral antigen in the brain as measured by immunohistochemistry using an antibody directed against viral nucleocapsid. More than 90% of the viruses recovered from brain lysates of such protected mice, at 5 days postinfection, had lost the ability to express EGFP and had deletions in their genomes encompassing EGFP and gp33-41. In addition, genomes of viruses from about half the plaques that retained the EGFP gene had mutations within the gp33-41 epitope. On the other hand, gp33-41-specific cells failed to protect perforin-deficient mice from infection by the recombinant MHV expressing gp33, indicating that perforin-mediated mechanisms were needed. Virus recovered from perforin-deficient mice did not exhibit loss of EGFP expression and the gp33-41 epitope. These observations suggest that the cytotoxic T-cell response to gp33-41 exerts a strong immune pressure that quickly selects epitope escape mutants to gp33-41.     

Blockade of PD-1/B7-H1 interaction restores effector CD8+ T cell responses in a hepatitis C virus core murine model

The impaired function of CD8(+) T cells is characteristic of hepatitis C virus (HCV) persistent infection. HCV core protein has been reported to inhibit CD8(+) T cell responses. To determine the mechanism of the HCV core in suppressing Ag-specific CD8(+) T cell responses, we generated a transgenic mouse, core(+) mice, where the expression of core protein is directed to the liver using the albumin promoter. Using a recombinant adenovirus to deliver Ag, we demonstrated that core(+) mice failed to clear adenovirus-LacZ (Ad-LacZ) infection in the liver. The effector function of LacZ-specific CD8(+) T cells was particularly impaired in the livers of core(+) mice, with suppression of IFN-gamma, TNF-alpha, and granzyme B production by CD8(+) T cells. In addition, the impaired CD8(+) T cell responses in core(+) mice were accompanied by the enhanced expression of the inhibitory receptor programmed death-1 (PD-1) by LacZ-specific CD8(+) T cells and its ligand B7-H1 on liver dendritic cells following Ad-LacZ infection. Importantly, blockade of the PD-1/B7-H1 inhibitory pathway (using a B7-H1 blocking antibody) in core(+) mice enhanced effector function of CD8(+) T cells and cleared Ad-LacZ-infection as compared with that in mice treated with control Ab. This suggests that the regulation of the PD-1/B7-H1 inhibitory pathway is crucial for HCV core-mediated impaired T cell responses and viral persistence in the liver. This also suggests that manipulation of the PD-1/B7-H1 pathway may be a potential immunotherapy to enhance effector T cell responses during persistent HCV infection.     

Functional expression of chemokine receptor CCR5 on CD4(+) T cells during virus-induced central nervous system disease

Intracranial infection of C57BL/6 mice with mouse hepatitis virus (MHV) results in an acute encephalomyelitis followed by a demyelinating disease similar in pathology to the human disease multiple sclerosis (MS). CD4(+) T cells are important in amplifying demyelination by attracting macrophages into the central nervous system (CNS) following viral infection; however, the mechanisms governing the entry of these cells into the CNS are poorly understood. The role of chemokine receptor CCR5 in trafficking of virus-specific CD4(+) T cells into the CNS of MHV-infected mice was investigated. CD4(+) T cells from immunized CCR5(+/+) and CCR5(-/-) mice were expanded in the presence of the immunodominant epitope present in the MHV transmembrane (M) protein encompassing amino acids 133 to 147 (M133-147). Adoptive transfer of CCR5(+/+)-derived CD4(+) T cells to MHV-infected RAG1(-/-) mice resulted in CD4(+)-T-cell entry into the CNS and clearance of virus from the brain. These mice also displayed robust demyelination correlating with macrophage accumulation within the CNS. Conversely, CD4(+) T cells from CCR5(-/-) mice displayed an impaired ability to traffic into the CNS of MHV-infected RAG1(-/-) recipients, which correlated with increased viral titers, diminished macrophage accumulation, and limited demyelination. Analysis of chemokine receptor mRNA expression by M133-147-expanded CCR5(-/-)-derived CD4(+) T cells revealed reduced expression of CCR1, CCR2, and CXCR3, indicating that CCR5 signaling is important in increased expression of these receptors, which aid in trafficking of CD4(+) T cells into the CNS. Collectively these results demonstrate that CCR5 signaling is important to migration of CD4(+) T cells to the CNS following MHV infection.     

Alphavirus-induced encephalomyelitis: antibody-secreting cells and viral clearance from the nervous system

Sindbis virus (SINV) infection of the central nervous system (CNS) provides a model for understanding the role of the immune response in recovery from alphavirus infection of neurons. Virus clearance occurred in three phases: clearance of infectious virus (days 3 to 7), clearance of viral RNA (days 8 to 60), and maintenance of low levels of viral RNA (>day 60). The antiviral immune response was initiated in the cervical lymph nodes with rapid extrafollicular production of plasmablasts secreting IgM, followed by germinal center production of IgG-secreting and memory B cells. The earliest inflammatory cells to enter the brain were CD8(+) T cells, followed by CD4(+) T cells and CD19(+) B cells. During the clearance of infectious virus, effector lymphocytes in the CNS were primarily CD8(+) T cells and IgM antibody-secreting cells (ASCs). During the clearance of viral RNA, there were more CD4(+) than CD8(+) T cells, and B cells included IgG and IgA ASCs. At late times after infection, ASCs in the CNS were primarily CD19(+) CD38(+) CD138(-) Blimp-1(+) plasmablasts, with few fully differentiated CD38(-) CD138(+) Blimp-1(+) plasma cells. CD19(+) CD38(+) surface Ig(+) memory B cells were also present. The level of antibody to SINV increased in the brain over time, and the proportion of SINV-specific ASCs increased from 15% of total ASCs at day 14 to 90% at 4 to 6 months, suggesting specific retention in the CNS during viral RNA persistence. B cells in the CNS continued to differentiate, as evidenced by accumulation of IgA ASCs not present in peripheral lymphoid tissue and downregulation of major histocompatibility complex (MHC) class II expression on plasmablasts. However, there was no evidence of germinal center activity or IgG avidity maturation within the CNS.     

B cell-deficient mice are highly resistant to Leishmania donovani infection, but develop neutrophil-mediated tissue pathology

Resolution of Leishmania infection is T cell-dependent, and B lymphocytes have been considered to play a minimal role in host defense. In this study, the contribution of B lymphocytes to the response against Leishmania donovani was investigated using genetically modified IgM transmembrane domain (muMT) mutant mice, which lack mature B lymphocytes. When compared with wild-type mice, muMT mice cleared parasites more rapidly from the liver, and infection failed to establish in the spleen. The rapid clearance of parasites in muMT mice was associated with accelerated and more extensive hepatic granuloma formation compared with wild-type mice. However, the liver of infected muMT mice also showed signs of destructive pathology, associated with the presence of increased numbers of neutrophils. The role of neutrophils in controlling parasite growth in the viscera was determined by depletion with the mAb RB6-8C5. This treatment led to a dramatic enhancement of parasite growth in both the liver and spleen of muMT and wild-type mice. As assessed by transfer of both normal and chronic-infection serum, Ig protects microMT mice from destructive hepatic pathology, but minimally alters their resistance compared with wild-type mice. However, adoptive transfer of CD4+ and CD8+ T cells into recombinase activating gene 1 (RAG1-/-) recipients, suggested that T cell function was not altered by maturation in a B cell-deficient environment. Taken together, these data suggest an inhibitory role for B lymphocytes in resistance to L. donovani unrelated to the presence or absence of Ig. However, Ig protects muMT mice from the exaggerated pathology that occurs during infection.     

Antibody prevents virus reactivation within the central nervous system.

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 mu 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.