Elsevier

Journal of Neuroimmunology

Volume 113, Issue 1, 1 February 2001, Pages 95-108
Journal of Neuroimmunology

Selective role for the p55 Kd TNF-α receptor in immune unresponsiveness induced by an acute viral encephalitis

https://doi.org/10.1016/S0165-5728(00)00427-6Get rights and content

Abstract

Brain infection by the laboratory strain challenge virus standard (CVS), a highly neurotropic strain of rabies virus, causes splenocytes to become less responsive to in vitro stimulation with ConA. CVS-induced immune unresponsiveness is less severe in mice lacking the p55 Kd TNF-α receptor (p55TNFR−/−) than in C57BL/6 mice, despite a similar invasion of the brain. Comparison of CVS infection in these two strains of mice indicated that decreased immune responsiveness is associated with: (1) an in vivo reduction of the percentages of Th1 (IL-2, IFN-γ and TNF-α) but not of Th2 (IL-4) cytokine-secreting T cells; and (2) an in vivo increase of the percentages of CD25 and CD69-expressing splenocytes. In contrast, CVS-induced immune unresponsiveness is not associated with abnormal percentage of T, B, NK cells or monocytes in vivo. The reductions of the CD4/CD8 ratio and of splenocyte expression of I-Ab during CVS infection are similar in p55TNFR−/− and C57BL/6 mice indicating that these two parameters are not linked to the decreased responsiveness of splenocytes. These data suggest that CVS-induced immune unresponsiveness is under the control of the p55 Kd TNF-α receptor. We propose that T cell activation through this receptor, in an environment of poor antigen presentation, results in a state of T cells characterized by the reduced production of IL-2, TNF-α and IFN-γ in vivo, the decreased responsiveness of splenocytes to ConA stimulation in vitro and the expression of the activation markers CD25 and CD69.

Introduction

LCMV, MCMV, HIV, Measles virus (MV) and other viruses that infect several types of tissues including the central nervous system (CNS) induce defects in immune function such that they evade the immune response. These defects include the reduction of the proliferative capacity of immune cells in response to mitogens, IL-2 or viral antigens (Rouse and Horohov, 1986, Shearer, 1998). Immune dysfunction induced by viral infections may be direct and results from: (1) apoptosis of immune cells both in vivo and in vitro (Ameisen and Capron, 1991, Fugier-Vivier et al., 1997, Lohman and Welsh, 1998); (2) decreased expression of MHC class II and B7 co-stimulatory molecules resulting in the loss of antigenic presentation and immune activation (Dudhane et al., 1996, Ennen et al., 1990, Grosjean et al., 1997, Jacobs and Cole, 1976, Meyaard et al., 1993, Polyak et al., 1997, Redpath et al., 1999); (3) altered cytokine production, usually an increased IL-10 production and decreased IL-2 and IL-12 secretion (Blackett and Mims, 1988, Chougnet et al., 1996, Clerici and Shearer, 1993, Redpath et al., 1999); or (4) activation of suppresser cells such as CD8+ T lymphocytes or macrophages (Badley et al., 1996, Badley et al., 1997, Herbein et al., 1998).

However, perturbations of immune functions in vivo induced by viral infections may also result from indirect effects. For instance homeostasis and functions of the immune system are regulated by the CNS. Neuroptropic viruses may alter immune function through the local delivery of neurotransmitters by nerves in the peripheral lymphoid organs (Felten et al., 1985, Felten et al., 1988) or by the release of immuno-modulator hormone such as corticotropin (ACTH) or corticotropin releasing hormone (CRH) through the stimulation of the hypothalamic–pituitary–adrenal (HPA) axis. Since viruses such as LCMV, MCMV, HIV and MV infect cells of both the immune and the nervous system, it is difficult to determine whether the immune alterations are the result from infection of cells in the periphery, the CNS or both. In order to discriminate between perturbations of the immune function resulting from direct alteration of the immune system or from infection of the nervous system we studied the decreased immune responsiveness induced by a CNS infection with a strictly neurotropic strain (CVS) of rabies virus. This strain provokes a fatal encephalitis associated with a loss of cellular mediated immunity (Wiktor et al., 1977) and an unresponsiveness of spleen cells to in vitro stimulation with ConA. It has been proposed that unresponsiveness of spleen cells from CVS-infected mice to ConA stimulation may be associated with the inability of these cells to produce and to respond to IL-2 in vitro (Hirai et al., 1992). Involvement of suppresser CD8+ T cells has also been proposed (Hirai et al., 1992). Independently, others (Perry et al., 1990) have shown that there is a decrease of the MHC class II I-Ab expression on spleen lymphocytes following rabies infection, suggesting a lower capacity of APC to present foreign antigen to T cells. However, the mechanisms underlying this decreased immune responsiveness have not been clearly elucidated.

TNF-α produced in the brain after rabies virus infection (Camelo et al., 2000, Marquette et al., 1996, Theerasurakarn and Ubol, 1998) is present in larger amount after infection with CVS a highly neurotropic strain which induces decreased immune responsiveness than with the virus strain PV4 of lower pathogenicity which does not induce decreased immune responsiveness (Baloul et al., in preparation). As most of the perturbations of the immune system could be due to a disregulation of the cytokine network and since TNF-α has been proposed to be a candidate in several models of immunosuppression (Nokta et al., 1996, Tomioka et al., 1995, Tomioka et al., 1996) we used mice deficient for the p55 Kd TNF-α receptor (p55TNFR−/−) infected with CVS to study the role of TNF-α in rabies virus-induced immune unresponsiveness.

To assess whether the reduced proliferative capacity of spleen cells in vitro was linked to alterations of the immune system in vivo, the percentages of IL-2, IL-4, TNF-α and IFN-γ-producing cells among both T and non T lymphocytes, were determined by intracellular cytokine detection. Finally, we compared C57BL/6 mice and p55TNFR−/− mice. P55TNFR−/− mice are less susceptible to decreased immune responsiveness and can thus be use to identify factors involved in the decreased immune responsiveness induced by CVS. The percentage of CD2, CD3, CD4, CD8, CD11b, CD25, CD45 (B220), CD69 and I-Ab expressing cells in p55TNFR−/− and C57BL/6 mice were compared by flow cytometry.

We found that acute rabies encephalitis induces a state of unresponsiveness of spleen cells characterized by a lack of proliferation to mitogen stimulation, a decrease in cytokine production, and an increase in the percentage of activated cells expressing CD25 and CD69. Altogether, these characteristics suggest that CVS-induced immune unresponsiveness is not related to modifications in the percentages of splenocytes subpopulations and that abnormal TNF-α production activates splenocytes through the p55 Kd TNF-α receptor and induces decreased immune responsiveness.

Section snippets

Viruses

The rabies laboratory strain Challenge Virus Standard, CVS, obtained from the American Type Cell Collection, Rockville, MD (Vr959) and the Pasteur Virus strain 4, PV4, (Lafon et al., 1988) were propagated on BSR cells, a baby hamster kidney (BHK-21)-derived cell line. Cell culture supernatants were used as viral inocula.

Mice, infection and assessment of clinical symptoms

Experiments were performed with 6-week-old female mice. BALB/c (H-2d, I-E+) and C57BL/6 (H-2b, I-E) mice were purchased from Janvier (St. Berthevin, France). P55 Kd

Immune unresponsiveness is induced following infection with CVS but not PV4 strain of rabies virus

To characterize rabies virus-induced immune unresponsiveness, we studied the kinetics of proliferation of spleen cells from mice infected with the CVS strain of rabies virus, in response to an in vitro ConA stimulation in C57BL/6 and BALB/c mice, respectively. As shown in Fig. 1A (result shown only for C57BL/6), spleen cells from CVS-infected mice responded equally or better than spleen cell from uninfected mice (158% of reference spleen cell proliferation at day 4), until day 5 p.i. From day 6

Discussion

We report evidence that the rabies virus-induced immune unresponsiveness is characterized by a decrease of the proliferative capacity of splenocytes to ConA stimulation and a decrease in the percentage of Th1 but not Th2 cytokine-secreting splenocytes predominantly among CD4-positive T cells. This suggests, as previouly reported for other viruses such as HIV, CMV and MV (Blackett and Mims, 1988, Chougnet et al., 1996, Clerici and Shearer, 1993, Redpath et al., 1999) that the CVS-induced immune

Acknowledgements

This work was supported by an institutional grant from the Institut Pasteur. We would like to thank Dr Werner Lesslauer of Hoffmann Laroche, Ltd for kindly providing p55 Kd TNF-α receptor-deficient mice.

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