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

This study demonstrates that neutralizing-antibody-producing B cells, CD4(+) T cells, and interferons (IFNs) are of key importance in virus control both in adoptive immunotherapy of persistent infection and in the late phase of acute infection with the WE strain of lymphocytic choriomeningitis virus (LCMV). We report the following results. (i) Clearance of LCMV-WE from C57BL/6 carrier mice by adoptive transfer of memory spleen cells requires B cells and CD4(+) T cells but not necessarily CD8(+) T cells. (ii) At the doses examined, CD8(+) T cells contribute to the initial reduction of viral titers but are alone not sufficient to clear the virus because they are exhausted. (iii) In the presence of functional IFN-gamma, virus clearance correlates well with the generation of neutralizing antibodies in the treated carrier mice. (iv) In the absence of receptors for IFN-gamma, virus clearance is not achieved. (v) Adoptive immunotherapy of mice persistently infected with a distinct virus isolate, LCMV-Armstrong, revealed only low levels of neutralizing antibodies; in this case, CD8(+) T cells were needed for virus clearance in addition to B and CD4(+) T cells. (vi) After low dose infection of C57BL/6 mice with LCMV-WE, virus is eliminated below detectable levels by CD8(+) T cells, but long-term (>2 months) virus control is usually not achieved in the absence of B cells or CD4(+) T cells; reappearance of the virus is paralleled either by exhaustion of virus-specific cytotoxic T lymphocytes or lethal immunopathology. These findings are of importance for adoptive immunotherapy strategies against persistent virus infections in humans.     

Disassociation between the in vitro and in vivo effects of nitric oxide on a neurotropic murine coronavirus

Intranasal inoculation of the neuroattenuated OBLV60 strain of mouse hepatitis virus results in infection of mitral neurons in the olfactory bulb, followed by spread along olfactory and limbic pathways to the brain. Immunocompetent BALB/c mice were able to clear virus by 11 days postinfection (p.i.). Gamma interferon (IFN-gamma) may play a role in clearance of OBLV60 from infected immunocompetent BALB/c mice through a nonlytic mechanism. Among the variety of immunomodulatory activities of IFN-gamma is the induction of expression of inducible nitric oxide synthase (iNOS), an enzyme responsible for the production of nitric oxide (NO). Studies were undertaken to investigate the role of IFN-gamma and NO in host defense and clearance of OBLV60 from the central nervous system (CNS). Exposure of OBLV60-infected OBL21a cells, a mouse neuronal cell line, to the NO-generating compound S-nitroso-L-acetyl penicillamine resulted in a significant decrease in viral replication, indicating that NO interfered with viral replication. Furthermore, infection of IFN-gamma knockout (GKO) mice and athymic nude mice with OBLV60 resulted in low-level expression of iNOS mRNA and protein in the brains compared to that of OBLV60-infected BALB/c mice. Nude mice were unable to clear virus and eventually died between days 11 and 14 p.i. (B. D. Pearce, M. V. Hobbs, T. S. McGraw, and M. J. Buchmeier, J. Virol. 68:5483-5495, 1994); however, GKO mice survived infection and cleared virus by day 18 p.i. These data suggest that IFN-gamma production in the olfactory bulb contributed to but may not be essential for clearance of OBLV60 from the brain. In addition, treatment of OBLV60-infected BALB/c mice with aminoguanidine, a selective inhibitor of iNOS activity, did not result in any increase in mortality, and the mice cleared the virus by 11 days p.i. These data suggest that although NO was able to block replication of virus in vitro, expression of iNOS with NO release in vivo did not appear to be the determinant factor in clearance of OBLV60 from CNS neurons.     

Human diploid cell culture rabies vaccine (HDCV) and purified chick embryo cell culture rabies vaccine (PCECV) both confer protective immunity against infection with the silver-haired bat rabies virus strain (SHBRV)

The demonstration of extensive differences in the antigenic makeups of the silver-haired bat rabies virus (SHBRV) and canine rabies virus (COSRV) strains raised concerns as to whether current licensed rabies vaccines are sufficiently protective against SHBRV. NIH mouse protection test results show that both the human diploid cell culture rabies vaccine (HDCV) and the purified chicken embryo cell rabies vaccine (PCECV) protected against lethal infection with SHBRV as well as the canine rabies strain COSRV. However, in this investigation, the potencies of both vaccines in mice were found to be significantly higher for COSRV than for SHBRV. The in vivo protection data are confirmed by in vitro virus neutralizing antibody (VNA) test results which demonstrate that mice immunized with HDCV or PCECV develop significantly higher VNA titres against COSRV than against SHBRV. In contrast, VNA tests of sera from individuals immunized with HDCV or PCECV showed that humans, as opposed to mice, develop significantly higher VNA titres against SHBRV than against COSRV. These data suggest that HDCV and PCECV will protect humans against infection with the silver-haired but rabies virus strain in addition to canine rabies virus strains.     

The range and distribution of murine central nervous system cells infected with the gamma(1)34.5- mutant of herpes simplex virus 1

Wild-type herpes simplex virus 1 (HSV-1) multiplies, spreads, and rapidly destroys cells of the murine central nervous system (CNS). In contrast, mutants lacking both copies of the gamma(1)34.5- gene have been shown to be virtually lacking in virulence even after direct inoculation of high-titered virus into the CNS of susceptible mice (J. Chou, E. R. Kern, R. J. Whitley, and B. Roizman, Science 250:1262-1266, 1990). To investigate the host range and distribution of infected cells in the CNS of mice, 4- to 5-week-old mice were inoculated stereotaxically into the caudate/putamen with 3 x 10(5) PFU of the gamma(1)34.5- virus R3616. Four-micrometer-thick sections of mouse brains removed on day 3, 5, or 7 after infection were reacted with a polyclonal antibody directed primarily to structural proteins of the virus and with antibodies specific for neurons, astrocytes, or oligodendrocytes. This report shows the following: (i) most of the tissue damage caused by R3616 was at the site of injection, (ii) the virus spread by retrograde transport from the site of infection to neuronal cell nuclei at distant sites and to ependymal cells by cerebrospinal fluid, (iii) the virus infected neurons, astrocytes, oligodendrocytes, and ependymal cells and hence did not discriminate among CNS cells, (iv) viral replication in some neurons could be deduced from the observation of infected astrocytes and oligodendrocytes at distant sites, and (v) infected cells were being efficiently cleared from the nervous system by day 7 after infection. We conclude that the gamma(1)34.5- attenuation phenotype is reflected in a gross reduction in the ability of the virus to replicate and spread from cell to cell and is not due to a restricted host range. The block in viral replication appears to be a late event in viral replication.     

Failure of protection induced by a Brazilian vaccine against Brazilian wild rabies viruses

This report shows that the SMB vaccine currently used in Brazil for human immunisation provides different degrees of protection in mice, depending on the rabies virus strain used as challenge. Using the NIH and Habel potency tests to evaluate the protective activity of rabies vaccine, we observed that vaccinated mice showed a higher resistance to a challenge with a fixed rabies virus (CVS-Challenge Virus Strain). The vaccine potency using the Habel or NIH tests was respectively > 6.4 (log 10) and 1.0 (Relative Potency-RP) when the fixed rabies virus was used for challenge, and from 2.9 to 4.3 (log 10) or 0.13 to 0.8 (RP) when different wild rabies viruses were used for challenge. The presence of virus neutralising antibodies (VNA) could not explain the differences of susceptibility after vaccination, since sera of vaccinated animals had similar VNA levels against both fixed and wild strains before virus challenge (respectively, 5.6 +/- 0.24 and 5.0 +/- 0.25 IU/ml of VNA against the fixed rabies virus and the 566-M strain of wild rabies virus in sera of mice vaccinated with 0.2 units of vaccine). Only cell-mediated immunity parameters correlated with the protection induced by vaccination. The IFN gamma titers found in sera and brain tissues of animals challenged with CVS strain were higher (from 36.7 +/- 5.7 to 293.3 +/- 46.2 IU/ml) than those found in mice challenged with 566-M virus strain (from 16.7 +/- 5.8 to 36.7 +/- 5.8). The proliferation index of spleen cells obtained with CVS stimulation reached a maximal value of 15.1 +/- 0.7 while spleen cells from vaccinated mice stimulated with 566-M virus failed to proliferate. The implications of these data in human protection by vaccination are discussed.     

B-cell activation in the mesenteric lymph nodes of resistant BALB/c mice infected with the murine nematode parasite Trichuris muris

Immune responses in resistant BALB/c mice infected with the murine nematode parasite Trichuris muris were examined. Following the establishment of infection, worm burdens of T. muris were expelled by BALB/c mice by day 21 postinfection (p.i.). Specific immunoglobulin G1 (IgG1) antibodies to T. muris excretory/secretory (E/S) antigens were detected in sera from infected mice, though specific IgG2a antibodies were not observed during infection. Ig-producing cells increased in the mesenteric lymph nodes (MLN) of infected mice on days 7, 14, and 21 p.i., with the greatest increase in numbers of IgG- and IgA-producing cells occurring on day 14. Marked increases in the relative percentages of B220+ and surface Ig+ (sIg+) cells were observed in the MLN of infected mice on days 14 and 21 p.i. Furthermore, cellular expansion of the MLN in infected mice resulted in an increase in the absolute numbers of B220+ and sIg+ cells. The levels of interleukin 2 (IL-2), IL-4, and interferon-gamma (IFN-gamma) detected in the supernatants from concanavalin A-stimulated MLN cells of infected mice were higher than those found in normal mice. Consequently, the expulsion of T. muris in resistant BALB/c mice was concomitant with cytokine production and B-cell activation in the MLN of infected mice. These results suggest the involvement of B-cell responses in protective immunity to T. muris infection.     

The role of the clinical psychologist in gynecological cancer

To assess the gamma delta TCR T cells in the control of the timing of the mucosal response to enteric parasitic infections, we used C57BL mice, orally infected with 200 viable T. spiralis larvae. The small intestine, spleens and Peyer's patches (PP) were excised on 1, 4, 7, 14, 21 and 29 postinfection days (p.i.) for immunophenotyping and histological studies. Uninfected mice served as control. Characterization of isolated lymphocytes of C57BL control mice, confirmed that T cell immunophenotype differs in spleen, PP and i-IEL. Practically all i-IEL were CD3+ cells (83%). In addition, most of the i-IEL expressed Ly-2 (65%). Among the i-IEL, the level of gamma delta TCR+ cells was significantly higher (29%) than that found in spleen (3%) and PP (3%). The expression was high on CD3+ and Ly-2+ (26 and 21%, respectively) and low on L3T4+ i-IEL (< 1%). During T. spiralis infection alpha beta TCR+ CD3+, gamma delta TCR+ CD3+ and gamma delta TCR+ Ly-2+ i-IEL increased on day 4 and 7. However, infected mice displayed a reduction in i-IEL number from 14 to 29 p.i. day. At the same time the proportion of gamma delta TCR on spleen Ly-2+ and on PP CD3+ and Ly-2+ cells increased on 14 and 21 p.i. day. Adult worms were expelled from the gut by day 14. Thus, the kinetics of gamma delta TCR+ i-IEL, but not spleen and PP gamma delta TCR, corresponded to the kinetics of worm expulsion in C57BL mice. Most murine i-IEL of the gamma delta T cell lineage tend to be cytolytic when activated. We speculated that gamma delta T cells of i-IEL during the early stages of infection recognize and eliminate damaged epithelial cells generated by parasite antigens, simultaneously accelerating the worm expulsion.     

Association of CD4+ T cell-dependent keratitis with genetic susceptibility to Pseudomonas aeruginosa ocular infection

The role of T lymphocytes in susceptibility to Pseudomonas aeruginosa corneal infection was studied in inbred C57Bl/6 (B6) beta2-microglobulin+/+ (beta2m+/+) and beta2m-/- knockout (KO) mice on a B6 genetic background. The corneas of both B6 and KO mice perforated by 7 days postinfection (p.i.). Histopathology revealed a similar inflammatory response characterized by an infiltration of polymorphonuclear neutrophilic leukocytes by 24 h p.i. in both groups of mice. CD4+ and CD8+ (latter absent in KO) T cells were present in cornea by 3 days p.i., and by 5 days, IL-2R-positive cells were positively immunostained. Corneas of B6 beta2m+/+ mice depleted of CD4+ T cells and infected with P. aeruginosa did not perforate at 7 days p.i. vs mice depleted of CD8+ T cells or treated with an irrelevant mAb. Neutralization of IFN-gamma before infecting B6 mice prevented corneal perforation and was associated with a lower delayed-type hypersensitivity than in B6 mice similarly treated with an irrelevant mAb. These data provide evidence that a CD4+ T cell (Th1)-dominated response following P. aeruginosa corneal infection is associated with genetic susceptibility and corneal perforation in inbred B6 mice.     

Host immune response to vesicular stomatitis virus infection of the central nervous system in C57BL/6 mice

In this report, the kinetics of cellular inflammatory changes in the brain of vesicular stomatitis virus (VSV)-infected C57BL/6 (B6) mice was determined. The behavior and survival rate of infected B6 were carefully monitored each day. Infectious viral titers and VSV antigen distribution were determined at several time points during the course of infection. Strong activation of both astrocytes and microglia was observed after VSV infection. Induction of type II nitric oxide synthase (iNOS) was detected in activated microglia in the olfactory bulb (OB) starting at day 4 postinfection. Induced expression of major histocompatibility complex (MHC) molecules and rapid infiltration of both T cells and natural killer (NK) cells were detected in the VSV-infected CNS. Collectively, these data indicate that the response to CNS infection in B6 mice, which is often primarily Th1 in characteristics, is comparable to BALB/c mice, a strain that often shows a Th2-dominated immune response.     

Resistance to and recovery from lethal influenza virus infection in B lymphocyte-deficient mice

In the adaptive immune response to most viruses, both the cellular and humoral arms of the immune system play complementary roles in eliminating virus and virus-infected cells and in promoting recovery. To evaluate the relative contribution of CD4+ and CD8+ effector T lymphocytes in virus clearance and recovery, we have examined the host response to lethal type A influenza virus infection in B lymphocyte-deficient mice with a targeted disruption in the immunoglobulin mu heavy chain. Our results indicate that naive B cell-deficient mice have a 50- 100-fold greater susceptibility to lethal type A influenza virus infection than do wild type mice. However, after priming with sublethal doses of influenza, immune B cell-deficient animals show an enhanced resistance to lethal virus infection. This finding indicates that an antibody-independent immune-mediated antiviral mechanism accounts for the increased resistance to lethal virus challenge. To assess the contribution of influenza-specific CD4+ and CD8+ effector T cells in this process, defined clonal populations of influenza-specific CD4+ and CD8+ effector T cells were adoptively transferred into lethally infected B cell-deficient mice. Cloned CD8+ effectors efficiently promoted recovery from lethal infection, whereas cloned CD4+ T cells conferred only partial protection. These results suggest that memory T lymphocytes can act independently of a humoral immune response in order to confer resistance to influenza infection in immune individuals. The potential implications of these results for vaccination against human influenza infection are discussed.     

Vaccination protects beta 2 microglobulin deficient mice from immune mediated mortality but not from persisting viral infection

Intracranial (i.c.) infection of immunocompetent mice with lymphocytic choriomeningitis virus (LCMV) results in immunopathological lethal meningitis mediated by CD8+ cytotoxic T lymphocytes (CTL). Vaccination of immunocompetent mice elicits a CD8+ CTL response that can protect the mice from lethal meningitis. beta 2 microglobulin-deficient (beta 2m-/-) mice are deficient in CD8+ CTL, exhibit CD4+ CTL, and, after i.c. LCMV infection, undergo a less severe meningitis with decreased mortality and additionally develop a wasting disease. Both wasting disease and mortality in beta 2m-/- mice are mediated by CD4+ T cells. We studied the effects of vaccination and challenge dose on weight loss, mortality and viral clearance after i.c. LCMV infection in beta 2m-/- mice. Unvaccinated beta 2m-/- mice had significant weight loss and mortality at doses of 200 and 10(3) p.f.u. LCMV, while a dose of 10(6) p.f.u. LCMV elicited significant mortality but less weight loss. Vaccination with u.v.-inactivated LCMV in complete Freund's adjuvant or with vaccinia virus expressing the LCMV glycoprotein or nucleoprotein genes protected beta 2m-/- mice from mortality but not weight loss after 200 p.f.u. LCMV challenge. Although protected from mortality, beta 2m-/- mice were unable to clear LCMV from their brains or spleens. Therefore, we show that vaccination can protect against lethal immune-meningitis in the face of persistent infection.     

Role of T lymphocyte subsets in immunity to spotted fever group Rickettsiae

To evaluate the roles of CD4 and CD8 T lymphocytes in immunity to disseminated endothelial infection with Rickettsia conorii (Malish 7 strain), these T cell subsets were depleted or adoptively transferred into subsequently infected C3H/HeN mice. CD4 T lymphocyte-depleted and sham-depleted mice underwent a similar course of illness with a sublethal rickettsial dose, cleared the infection by day 10, and recovered on days 10 to 11. In contrast, mice depleted of CD8 lymphocytes or CD4 and CD8 lymphocytes died or remained persistently infected through day 15 with the ordinarily sublethal dose. Endothelium was the major site of rickettsial persistence, including sites in the vital organs, brain, and lungs of CD8 lymphocyte-depleted mice. In nondepleted animals, CD8 T lymphocytes were observed in apposition to endothelial cells on day 10 at the time of rickettsial clearance. Adoptive transfer of immune CD4 or CD8 T lymphocytes protected mice against a lethal dose of R. conorii in the disseminated endothelial target model. Nonimmune CD4 or CD8 lymphocytes and immune lymphocytes that had passed through columns that depleted both CD4 and CD8 lymphocytes failed to protect mice against R. conorii. These studies represent the first analysis of the role of T lymphocyte subsets in immunity to spotted fever group rickettsiae and the first demonstration that clearance of spotted fever group rickettsiae from endothelial cells requires immune CD8 T lymphocytes.     

Cooperation of B cells and T cells is required for survival of mice infected with vesicular stomatitis virus

To define the role of T cells and B cells in resistance to vesicular stomatitis virus (VSV) infection, knockout mice with different specific immune defects on an identical background were infected i.v. and the outcome of infection was compared; in this way a more complete picture of the relative importance of various host defence mechanisms could be obtained. Compared to T and B cell-deficient SCID mice which all succumbed from encephalitis within 5-9 days of infection, T cell-deficient nude mice generally lived longer, but within a period of approximately 1 month after challenge all died. In contrast, B cell-deficient mice were highly susceptible even to low doses of virus and mortality could be prevented by transfer of naive B cells prior to challenge as well as by immune serum given after challenge. Analysis of MHC class I- and class II-deficient mice revealed that CD8+ T cells could exert some antiviral activity, but CD4+ T cells sufficed for survival and were required for optimal resistance. Consistent with this it was found that in nude mice a lethal outcome could be prevented by transfer of CD8-depleted cells from B cell-deficient mice. Thus our results clearly demonstrate that while antibodies are pivotal for survival in the early phase of VSV infection, T cells are required for long-term survival, with CD4+ T cells being more effective in controlling this infection than CD8+ T cells.     

Apoptosis mediated by Fas but not tumor necrosis factor receptor 1 prevents chronic disease in mice infected with murine cytomegalovirus

The role of Fas- and TNF-receptor 1 (TNF-R1)-mediated apoptosis in the clearance of virally infected cells and in the regulation of the immune response was analyzed after murine cytomegalovirus (MCMV) infection of C57BL/6 (B6)-+/+ mice, Fas-mutant B6-lpr/lpr mice, TNF-R1 knockout B6-tnfr0/0 mice, and double-deficient B6-tnfr0/0 lpr/lpr mice. There was approximately equivalent clearance of MCMV in B6-+/+, B6-tnfr0/0, and B6-lpr/lpr mice, and by day 28 no infectious virus could be detected in the liver, kidney, lung, or peritoneal exudate. However, delayed virus clearance was observed in B6-tnfr0/0 lpr/lpr mice. An acute inflammatory response occurred in the liver, lung, and kidney of all mice, which was most severe 7 d after MCMV infection, but resolved by day 28 in B6-+/+ and B6-tnfr0/0 mice, but not in B6-lpr/lpr or B6-tnfr0/0 lpr/lpr mice. These results indicate that apoptosis mediated by either Fas or TNF-R1 is sufficient for rapid clearance of the virus. However, apoptosis induced by Fas, but not TNF-R1, is required for the downmodulation of the immune response to the virus and prevention of a chronic inflammatory reaction.     

Administration-time-dependent effects of diltiazem on the 24-hour blood pressure profile of essential hypertension patients

Cells of the central nervous system (CNS) normally do not express detectable levels of major histocompatibility complex (MHC) Class I antigens. However, MHC Class I expression can be induced after virus infection. We tested the hypothesis that virus-induced Class I expression is mediated by lymphocytes or cytokines using lymphocyte- and cytokine-deficient mice. We used Theiler's murine encephalomyelitis virus (TMEV), which induces CNS demyelination that maps genetically to the D region of MHC Class I and is associated with high levels of Class I products. TMEV infection of severe combined immunodeficiency (SCID) and recombination activation gene-1-deficient mice, which lack B and T lymphocytes, resulted in equivalent H-2D and H-2K expression in brain and spinal cord, according to analysis of the area and intensity of immunoperoxidase staining. Class I antigens were demonstrated as early as 6 hours after infection, and they were more widely distributed than viral RNA, indicating that expression was induced indirectly via a soluble factor. To determine whether cytokines induced the expression, we infected mice lacking receptors for interferon-alpha/beta (IFN-alpha/beta R (-/-)), interferon-gamma (IFN-gamma R(-/-)), and tumor necrosis factor-alpha (TNFRp55(-/-)). TMEV-infected IFN-gamma R(-/-) and TN-FRp55(-/-) mice expressed Class I antigens in the CNS, whereas IFN-alpha/beta R(-/-) mice did not, establishing that IFN-alpha/beta mediated the expression. In contrast to the equivalent expression in SCID mice, we observed greater area and higher intensity of H-2D versus H-2K antigens in infected SCID mice reconstituted with normal spleen cells. Collectively, the data indicate that after TMEV infection, early generalized MHC Class I expression is mediated by IFN-alpha/beta independently of lymphocytes, but the differential regulation of H-2D over H-2K may be controlled by B and/or T lymphocytes.     

The inflammatory response to nonfatal Sindbis virus infection of the nervous system is more severe in SJL than in BALB/c mice and is associated with low levels of IL-4 mRNA and high levels of IL-10-producing CD4+ T cells

SJL mice are susceptible to inflammatory autoimmune diseases of the central nervous system (CNS), while BALB/c mice are relatively resistant. To understand differences in immune responses that may contribute to autoimmune neurologic disease, we compared the responses of SJL and BALB/c mice to infection with Sindbis virus, a virus that causes acute nonfatal encephalomyelitis in both strains of mice. Clearance of virus was similar, but SJL mice developed a more intense inflammatory response in the brain and spinal cord and inflammation persisted for several weeks. Analysis of lymphocytes isolated from brains early after infection showed an absence of NK cells in SJL mice, while both strains of mice showed CD4+ and CD8+ T cells. During the second week after infection, CD4+ T cells increased in SJL mice and the proportion of CD8+ T cells decreased, while the opposite pattern was seen in BALB/c mice. Expression of IL-10 mRNA was higher and IL-4 mRNA was lower in the brains of infected SJL than in BALB/c mice, while expression of the mRNAs of IL-6, IL-1beta, TNFalpha, and the Th1 cytokines IL-2, IL-12, and IFN-gamma was similar. Lymphocytes isolated from the CNS of SJL mice produced large amounts of IL-10. CNS lymphocytes from both strains of mice produced IFN-gamma in response to stimulation with Sindbis virus, but not in response to myelin basic protein. These data suggest that IL-10-producing CD4+ T cells are differentially recruited to or regulated within the CNS of SJL mice compared with BALB/c mice infected with Sindbis virus, a characteristic that may be related to low levels of IL-4, and is likely to be involved in susceptibility of SJL mice to CNS inflammatory diseases.     

Cellular mechanisms involved in protection against influenza virus infection in transgenic mice expressing a TCR receptor specific for class II hemagglutinin peptide in CD4+ and CD8+ T cells

Mice transgenic for a TCR that recognizes peptide110-120 of hemagglutinin of PR8 influenza virus in the context of MHC class II I-Ed molecules express the transgenes in both CD4+ and CD8+ T cells. We have found that these TCR-hemagglutinin (TCR-HA) transgenic mice display a significantly increased resistance to the primary infection with PR8 virus compared with the wild-type mice. The TCR-HA transgenic mice mounted significant MHC type II and enhanced MHC type I-restricted cytotoxicity as well as increased cytokine responses in both spleen and lungs after infection with PR8 virus. In contrast, the primary humoral response against PR8 virus was not significantly different from that of the wild-type mice. In vivo depletion and adoptive cell transfer experiments demonstrated that both CD4+ and CD8+ TCR-HA+ T cell subsets were required for the complete clearance of pulmonary virus following infection with a dose that is 100% lethal in wild-type mice. Whereas CD4+ TCR-HA+ T cells were necessary for effective activation and local recruitment of CD8+ T cells, CD8+ TCR-HA+ T cells showed a Th1-biased pattern and MHC type II-restricted cytotoxicity. However, in the absence of in vivo expression of MHC type I molecules on the infected cells, the protection conferred by the TCR-HA+ T cells was impaired, indicating that the enhanced MHC class I-restricted cytotoxicity due to TCR-HA+ CD4+ Th cells was a critical element for clearance of the pulmonary virus by the transgenic mice.     

Theiler's virus infection of perforin-deficient mice

Theiler's virus, a murine picornavirus, infects the central nervous systems of C57BL/6 mice and is cleared after approximately 10 days by a process which requires CD8+ cytotoxic T cells. We used perforin-deficient C57BL/6 mice to test the role of this protein in viral clearance. Perforin-deficient mice died from viral encephalomyelitis between days 12 and 18 postinoculation. They had high levels of viral RNA in their central nervous systems until the time of death. In contrast, viral RNA had disappeared by day 11 postinoculation in wild-type C57BL/6 mice. Cytotoxic T cells can kill infected cells by two main mechanisms: the secretion of the pore-forming protein perforin or the interaction of the Fas ligand with the apoptosis-inducing Fas molecule on the target cell. Our results demonstrate that clearance of Theiler's virus from the central nervous system in C57BL/6 mice is perforin dependent.