Interleukin-1 receptor type 1 is essential for control of cerebral but not systemic listeriosis.

Listeria monocytogenes may infect the central nervous system and several peripheral organs. To explore the function of IL-1 receptor type 1 (IL-1R1) in cerebral versus systemic listeriosis, IL-1R1(-/-) and wild-type mice were infected either intracerebrally or intraperitoneally with L. monocytogenes. After intracerebral infection with various numbers of attenuated Listeria, IL-1R1(-/-) mice succumbed due to an insufficient control of intracerebral Listeria, whereas all wild-type mice survived, efficiently restricting growth of Listeria. IL-1R1(-/-) mice recruited increased numbers of leukocytes, especially granulocytes, to the brain compared with wild-type mice. In contrast, both IL-1R1(-/-) and wild-type mice survived a primary and secondary intraperitoneal infection with Listeria without differences in the hepatic bacterial load. In addition, both strains developed similar frequencies of Listeria-specific CD4 and CD8 T cells after primary and secondary intraperitoneal infection. However, an intraperitoneal immunization before intracerebral challenge infection neither protected IL-1R1(-/-) mice from death nor reduced the intracerebral bacterial load, although numbers of intracerebral Listeria-specific CD4 and CD8 T cells and levels of inducible nitric oxide synthase, tumor necrosis factor, and interferon-gamma mRNA were identical in IL-1R1(-/-) and wild-type mice. Collectively, these findings illustrate a crucial role of IL-1R1 in cerebral but not systemic listeriosis.

TNF is important for pathogen control and limits brain damage in murine cerebral listeriosis.

Cerebral listeriosis is a life-threatening disease. However, little is known about the bacterial virulence factors responsible for the severe course of disease and the factors of the immune system contributing to the control of Listeria monocytogenes (LM) or even to the damage of the brain. To analyze the importance of the actA gene of LM, which mediates cell-to-cell spread of intracellular LM, the function of TNF in murine cerebral listeriosis was studied. C57BL/6 mice survived an intracerebral (i.c.) infection with actA-deficient LM, but succumbed to infection with wild-type (WT) LM. Upon infection with actA-deficient LM, macrophages and microglial cells rapidly, and later LM-specific CD4 and CD8 T cells, produced TNF. In contrast to WT mice, TNF-deficient animals succumbed to the infection within 4 days due to failure of control of LM. Histology identified a more severe meningoencephalitis, brain edema, and neuronal damage, but a reduced inducible NO synthase expression in TNF-deficient mice. Reciprocal bone marrow chimeras between WT and TNF-deficient mice revealed that hematogenously derived TNF was essential for survival, whereas TNF produced by brain-resident cells was less important. Death of TNF-deficient mice could be prevented by LM-specific T cells induced by an active immunization before i.c. infection. However, brain pathology and inflammation of immunized TNF-deficient mice were still more severe. In conclusion, these findings identify a crucial role of TNF for the i.c. control of LM and survival of cerebral listeriosis, whereas TNF was not responsible for the destruction of brain tissue.

Organ- and disease-stage-specific regulation of Toxoplasma gondii-specific CD8-T-cell responses by CD4 T cells.

Toxoplasma gondii induces a persistent central nervous system infection, which may be lethally reactivated in AIDS patients with low CD4 T-cell numbers. To analyze the role of CD4 T cells for the regulation of parasite-specific CD8 T cells, mice were infected with transgenic T. gondii expressing the CD8 T-cell antigen beta-galactosidase (beta-Gal). Depletion of CD4 T cells prior to infection did not affect frequencies of beta-Gal(876-884)-specific (consisting of residues 876 to 884 of beta-Gal) CD8 T cells but resulted in a pronounced reduction of intracerebral beta-Gal-specific gamma interferon (IFN-gamma)-producing and cytolytic CD8 T cells. After cessation of anti-CD4 treatment a normal T. gondii-specific CD4 T-cell response developed, but IFN-gamma production of intracerebral beta-Gal-specific CD8 T cells remained impaired. The important supportive role of CD4 T cells for the optimal functional activity of intracerebral CD8 T cells was also observed in mice that had been depleted of CD4 T cells during chronic toxoplasmosis. Reinfection of chronically infected mice that had been depleted of CD4 T cells during either the acute or chronic stage of infection resulted in an enhanced proliferation of beta-Gal-specific IFN-gamma-producing splenic CD8 T cells. However, reinfection of chronically infected mice that had been depleted of CD4 T cells in the acute stage of infection did not reverse the impaired IFN-gamma production of intracerebral CD8 T cells. Collectively, these findings illustrate that CD4 T cells are not required for the induction and maintenance of parasite-specific CD8 T cells but, depending on the stage of infection, the infected organ and parasite challenge infection regulate the functional activity of intracerebral CD8 T cells.

Both lymphotoxin-alpha and TNF are crucial for control of Toxoplasma gondii in the central nervous system.

Immunity to Toxoplasma gondii critically depends on TNFR type I-mediated immune reactions, but the precise role of the individual ligands of TNFR1, TNF and lymphotoxin-alpha (LTalpha), is still unknown. Upon oral infection with T. gondii, TNF(-/-), LTalpha(-/-), and TNF/LTalpha(-/-) mice failed to control intracerebral T. gondii and succumbed to an acute necrotizing Toxoplasma encephalitis, whereas wild-type (WT) mice survived. Intracerebral inducible NO synthase expression and-early after infection-splenic NO levels were reduced. Additionally, peritoneal macrophages produced reduced levels of NO upon infection with T. gondii and had significantly reduced toxoplasmastatic activity in TNF(-/-), LTalpha(-/-), and TNF/LTalpha(-/-) mice as compared with WT animals. Frequencies of parasite-specific IFN-gamma-producing T cells, intracerebral and splenic IFN-gamma production, and T. gondii-specific IgM and IgG titers in LTalpha(-/-) and TNF/LTalpha(-/-) mice were reduced only early after infection. In contrast, intracerebral IL-10 and IL-12p40 mRNA expression and splenic IL-2, IL-4, and IL-12 production were identical in all genotypes. In addition, TNF(-/-), LTalpha(-/-), and TNF/LTalpha(-/-), but not WT, mice succumbed to infection with the highly attenuated ts-4 strain of T. gondii or to a subsequent challenge infection with virulent RH toxoplasms, although they had identical frequencies of IFN-gamma-producing T cells as compared with WT mice. Generation and infection of bone marrow reconstitution chimeras demonstrated an exclusive role of hematogeneously produced TNF and LTalpha for survival of toxoplasmosis. These findings demonstrate the crucial role of both LTalpha and TNF for control of intracerebral toxoplasms.

Mice with neonatally induced inactivation of the vascular cell adhesion molecule-1 fail to control the parasite in Toxoplasma encephalitis.

Under various inflammatory conditions, cell adhesion molecules are up-regulated in the central nervous system (CNS) and may contribute to the recruitment of leukocytes to the brain. In the present study, the functional role of vascular cell adhesion molecule (VCAM)-1 in Toxoplasma encephalitis (TE) was addressed using VCAM(flox/flox MxCre) mice. Neonatal inactivation of the VCAM-1 gene resulted in a lack of induction of VCAM-1 on cerebral blood vessel endothelial cells, whereas the constitutive expression of VCAM-1 on choroid plexus epithelial cells and the ependyma was unaffected; in these animals, resistance to T. gondii was abolished, and VCAM(flox/flox MxCre) mice died of chronic TE caused by a failure to control parasites in the CNS. Although leukocyte recruitment to the CNS was unimpaired, the B cell response was significantly reduced as evidenced by reduced serum levels of anti-T. gondii-specific IgM and IgG antibodies. Furthermore, the frequency and activation state of intracerebral T. gondii-specific T cells were decreased, and microglial activation was markedly reduced. Taken together, these data demonstrate the crucial requirement of VCAM-1-mediated immune reactions for the control of an intracerebral infectious pathogen, whereas other cell adhesion molecules can efficiently compensate for VCAM-1-mediated homing across cerebral blood vessels.

The induction and kinetics of antigen-specific CD8 T cells are defined by the stage specificity and compartmentalization of the antigen in murine toxoplasmosis.

Toxoplasma gondii forms different life stages, fast-replicating tachyzoites and slow-growing bradyzoites, in mammalian hosts. CD8 T cells are of crucial importance in toxoplasmosis, but it is unknown which parasite stage is recognized by CD8 T cells. To analyze stage-specific CD8 T cell responses, we generated various recombinant Toxoplasma gondii expressing the heterologous Ag beta-galactosidase (beta-gal) and studied whether 1) secreted or cytoplasmic Ags and 2) tachyzoites or bradyzoites, which persist intracerebrally, induce CD8 T cells. We monitored the frequencies and kinetics of beta-gal-specific CD8 T cells in infected mice by MHC class I tetramer staining. Upon oral infection of B6C (H-2(bxd)) mice, only beta-gal-secreting tachyzoites induced beta-gal-specific CD8 T cells. However, upon secondary infection of mice that had received a primary infection with tachyzoites secreting beta-gal, beta-gal-secreting tachyzoites and bradyzoites transiently increased the frequency of intracerebral beta-gal-specific CD8 T cells. Frequencies of splenic and cerebral beta-gal-specific CD8 T cells peaked at day 23 after infection, thereafter persisting at high levels in the brain but declining in the spleen. Splenic and cerebral beta-gal-specific CD8 T cells produced IFN-gamma and were cytolytic upon specific restimulation. Thus, compartmentalization and stage specificity of an Ag determine the induction of CD8 T cells in toxoplasmosis.

Protective immunosurveillance of the central nervous system by Listeria-specific CD4 and CD8 T cells in systemic listeriosis in the absence of intracerebral Listeria.

The invasion of the CNS by pathogens poses a major risk for damage of the highly vulnerable brain. The aim of the present study was to analyze immunological mechanisms that may prevent spread of infections to the CNS. Intraperitoneal application of Listeria monocytogenes to mice induced infection of the spleen, whereas pathogens remained absent from the brain. Interestingly, Listeria-specific CD4 and CD8 T cells homed to the brain and persisted intracerebrally for at least 50 days after both primary and secondary infection. CD4 and CD8 T cells resided in the leptomeninges, in the choroid plexus, and, in low numbers, in the brain parenchyma. CD4 and CD8 T cells isolated from the brain early after infection (day 7) were characterized by an activated phenotype with spontaneous IFN-gamma production, whereas at a later stage of infection (day 28) restimulation with Listeria-specific peptides was required for the induction of IFN-gamma production by CD4 and CD8 T cells. In contrast to splenic T cells, T cells in the brain did not exhibit cytotoxic activity. Adoptively transferred T cells isolated from the brains of Listeria-infected mice reduced the bacterial load in cerebral listeriosis. The frequency of intracerebral Listeria-specific T cells was partially regulated by the time of exposure to Listeria and cross-regulated by CD4 and CD8 T cells. Collectively, these data reveal a novel T cell-mediated pathway of active immunosurveillance of the CNS during bacterial infections.

Phenotype and regulation of persistent intracerebral T cells in murine Toxoplasma encephalitis.

Toxoplasma gondii is a parasite causing asymptomatic, persistent encephalitis. Protective CD4 and CD8 T cells are recruited to and accumulate in the brain in acute Toxoplasma encephalitis (TE), with slowly decreasing numbers in chronic TE. It is unclear how the size of the intracerebral T cell pool is regulated. Conceivably, permanent recruitment, proliferation, and apoptosis may be involved. We observed that in murine TE recruitment of T cells to the brain was terminated in chronic TE. In vivo 5-bromo-2'-deoxyuridine incorporation and in vitro T cell proliferation experiments revealed that intracerebral T cells did not proliferate, which was explained by the expression of the cell cycle inhibitors p21(Waf/cip1) and p27(Kip1) and the inhibitory activity of intracerebral F4/80(+) cells. TUNEL staining detected apoptotic T cells at low frequency corresponding to an increased expression of the anti-apoptotic molecules Bcl-2 and Bcl-x(L) and a reduced expression of the pro-apoptotic molecules Bad, Bax, and Fas ligand in CD4 and CD8 T cells. During progression from acute to chronic TE, both CD4 and CD8 T cells down-regulated CD45RB expression and expressed a differential pattern of cytokines. From these experiments it is concluded that the number of intracerebral T cells increases by recruitment of T cells during acute infection, whereas proliferation of intracerebral T cells does not play a role. In chronic TE, T cell recruitment is terminated, the phenotype of intracerebral T cells changes, and their number is gradually downsized by low level apoptosis, which, however, does not completely resolve the T cell infiltrates.