Mary K. Kennedy

CD40 Ligand Is Required for Protective Cell-Mediated Immunity to Leishmania major

The CD40-CD40 ligand (CD40L) signaling process is a pivotal component of multiple immunoregulatory pathways. Although the role that CD40L plays in humoral immune responses is fairly well defined, its function(s) in cell-mediated responses in vivo has not been established. We investigated this issue by assessing the course of Leishmania major infection in CD40L knockout (CD40LKO) mice that were generated on a resistant background. In response to parasite challenge, CD40LKO mice developed ulcerating cutaneous lesions and failed to mount a vigorous Th1-like response. The impaired Th1-like response appears to be related to a defect in the ability of CD40LKO T cells to induce the production of IL-12 from macrophages. Treatment with exogenous IL-12 prevented disease progression in CD40LKO mice, and administration of recombinant CD40L provided partial protection against infection. Thus, a protective cell-mediated immune response to L. major appears to be dependent upon CD40L-induced IL-12 secretion by antigen-presenting cells.

Induction of active and adoptive relapsing experimental autoimmune encephalomyelitis (EAE) using an encephalitogenic epitope of proteolipid protein

Proteolipid protein (PLP) is a major component of the central nervous system (CNS) myelin membrane and has been shown to induce acute experimental autoimmune encephalomyelitis (EAE) in genetically susceptible animals. Here we describe conditions by which a relapsing-remitting form of EAE can be reliably induced in SJL/J mice either actively immunized with the major encephalitogenic PLP peptide, PLP13-151(S), or following adoptive transfer of PLP139-151(S)-specific T cells. The disease follows a reliable relapsing-remitting course with acute clinical signs first appearing 6-20 days after priming or transfer and relapses first appearing at 30-45 days. The initial onset of disease correlates with delayed-type hypersensitivity (DTH) reactivity specific for PLP139-151(S), in the apparent absence of T cell reactivity to the major myelin basic protein (MBP) peptide. Histologically, both the active and adoptive forms of the disease are characterized by extensive mononuclear cell infiltration and severe demyelination of the CNS. These results suggest that T cell responses specific for PLP139-151(S) are sufficient to induce clinical and histological R-EAE in SJL/J mice. This model should prove useful for examination of the cellular and molecular events involved in clinical relapses and perhaps in determining the role of PLP-specific T cell responses in multiple sclerosis (MS).

Class II-restricted T cell responses in Theiler's murine encephalomyelitis virus (TMEV)-induced demyelinating disease. III. Failure of neuroantigen-specific immune tolerance to affect the clinical course of demyelination

Abstract Intracerebral inoculation of Theilers murine encephalomyelitis virus (TMEV) into susceptible mouse strains produces a chronic demyelinating disease in which mononuclear cell-rich infiltrates in the central nervous system (CNS) are prominent. Current evidence strongly supports an immune-mediated basis for myelin breakdown, with an effector role proposed for TMEV-specific, major histocompatibility complex (MHC) class II-restricted delayed-type hypersensitivity (DTH) responses in which lymphokine-activated macrophages mediate bystander demyelination. The present study examined the possibility that concomitant or later-appearing neuroantigen-specific autoimmune T cell responses, such as those demonstrated in chronic-relapsing experimental allergic encephalomyelitis (R-EAE), may contribute to the demyelinating process following TMEV infection. T cell responses against intact, purified major myelin proteins (myelin basic protein (MBP) and proteolipid protein (PLP), and against altered myelin constituents were readily demonstrable in SJL/J mice with R-EAE, but were not detectable in SJL/J mice with TMEV-induced demyelinating disease. TMEV-infected mice also did not display T cell responses against the peptide fragments of MBP(91–104) and PLP(139–151) recently shown to be encephalitogenic in SJL/J mice. In addition, induction of neuroantigen-specific tolerance to a heterogeneous mixture of CNS antigens, via the i.v. injection of syngeneic SJL/J splenocytes covalently coupled with mouse spinal cord homogenate, resulted in significant suppression of clinical and histologic signs of R-EAE and the accompanying MBP- and PLP-specific DTH responses. In contrast, neuroantigen-specific tolerance failed to alter the development of clinical and histologic signs of TMEV-induced demyelinating disease or the accompanying virus-specific DTH and humoral immune responses. These findings demonstrate that TMEV-induced demyelinating disease can occur in the apparent absence of neuroantigen-specific autoimmune responses. The relationship of the present results to the immunopathology of multiple sclerosis is discussed.

Monoclonal antibody-induced inhibition of relapsing EAE in SJL/J mice correlates with inhibition of neuroantigen-specific cell-mediated immune responses☆

Relapsing experimental allergic encephalomyelitis (R-EAE) in SJL/J mice was examined in relation to the development of neuroantigen-specific T cell proliferative (Tprlf) and delayed-type hypersensitivity (DTH) responses. R-EAE was induced by injecting syngeneic mouse spinal cord homogenate in CFA on days 0 and 7 over the shaved flanks of female SJL/J mice. Mice primed in this manner exhibited significant Tprlf and DTH responses specific for both major myelin proteins, myelin basic protein (MBP) and proteolipid protein (PLP). A time course comparison between the induction of R-EAE and the development of neuroantigen-specific cell-mediated immune (CMI) responses (Tprlf and DTH) revealed that the MBP- and PLP-specific Tprlf and DTH responses peaked prior to the onset of initial clinical symptoms and the DTH responses remained at significant levels throughout the relapsing course of the disease. Monoclonal antibodies were used to determine whether in vivo inhibition of class II-restricted Tprlf and DTH responses correlated with inhibition of R-EAE. In vivo administration of a total of 100 micrograms anti-L3T4 antibody, but not anti-Lyt-2 antibody, resulted in delayed onset and reduced severity of clinical signs of R-EAE concomitant with significantly reduced levels of MBP- and PLP-specific Tprlf and DTH responses. Treatment with a total of 300 micrograms of purified anti-L3T4 resulted in total abrogation of R-EAE induction and neuroantigen-specific CMI. Thus, clinical signs of R-EAE were found to correlate with the activity of neuroantigen-specific, class II-restricted T cells.

Specific Immunoregulation of the Induction and Effector Stages of Relapsing EAE via Neuroantigen-Specific Tolerance Induction

The effects of neuroantigen-specific tolerance on the induction and effector stages of relapsing experimental autoimmune encephalomyelitis (R-EAE) were examined. The incidence of clinical and histologic signs of active MSCH-induced R-EAE, and accompanying neuroantigen-specific DTH responses, were dramatically reduced in SJL/J mice tolerized via the i.v. injection of syngeneic splenocytes coupled with MSCH, PLP, or encephalitogenic PLP peptides 7-14 days before priming. MBP-specific tolerance was not effective in preventing active R-EAE. In contrast to MSCH-induced active R-EAE, treatment of recipient mice with splenocytes coupled with MBP and the encephalitogenic MBP 84-104 peptide, but not with PLP, suppressed of clinical signs of adoptive R-EAE mediated by MBP-specific effector T cells in a dose-dependent manner. Neuroantigen-coupled splenocytes were also efficient in treating established disease as tolerization of SJL/J mice after the first incidence of clinical disease significantly reduced the incidence and severity of subsequent paralytic relapses. Antigen-specific tolerance thus provides a powerful approach for the prevention and/or treatment of autoimmune disease.

Mechanisms of genetic control of immune responses: II. Nonresponsiveness in BALB/c GT-specific cell-mediated immune responses does not correlate with the absence of functional T cells or the induction of suppressor T cells

The mechanisms underlying Ir gene control of CMI were addressed by examining the DTH and Tprlf responses specific for the synthetic polymers GT, GAT, and GA. We show that BALB/c mice (GAT/GA responders, GT nonresponders) primed with GT fail to develop DTH and Tprlf responses specific for GT, GAT, or GA. GAT immunization resulted in DTH responses that could be elicited not only with GAT and GA but also with GT, demonstrating that GT-specific TDH are present in nonresponder mice. GT-specific DTH was transferred with Thy-1+ Lyt-1+2−, H-2 Irestricted, nylon wool nonadherent cells. GA-primed BALB/c mice developed GAT- and GA-, but not GT-apecific DTH responses, indicating that GA and GT do not cross-react at the T-cell level. The ability of GAT [but not a mixture of GA plus GT, or GT electrostatically complexed to the immunogenic carrier MBSA (GT-MBSA)] to induce GT-specific DTH suggested a requirement for covalent linkage of stimulatory ‘GA’ and nonstimulatory ‘GT’ determinants present on the GAT molecule. Similarly, GT-specific in vitro Tprlf responses could be demonstrated in GAT-primed mice exhibiting significant levels of GT-specific DTH but not in GT- or GT-MBSA-primed mice. Tolerization experiments also suggested that GT-specific Th were involved in the development of GT-specific DTH in GAT-primed mice. The GT nonresponsiveness of BALB/c mice for DTH and Tprlf responses could not be reversed by treatments designed to abrogate Ts activity (priming with GT-MBSA and CY injection), nor could GT-primed cells be shown to inhibit the development or elicitation of GT-specific CMI in GAT-primed mice during the afferent and/or efferent stages of DTH. Our results suggest that GT nonresponsiveness does not result from the absence of GT-specific T cells or preferential induction of Ts. The results are discussed in the context of hole-in-the-repertoire and antigen presentation (determinant selection) models of Ir gene control.

CD40 Ligand Knockout Mice

CD40 is a 50-kDa glycoprotein expressed on B-cells, monocytes, dendritic cells, follicular dendritic cells, thymic epithelial cells, and certain carcinomas (for review, see ref. 1). Its counterstructure, CD40 ligand (CD40L), is a 33-kDa glycoprotein expressed primarily on activated CD4+ T-cells, but also found on CD8+ T-cells, mast cells, basophils (1), and eosinophils (2). CD40L belongs to the tumor necrosis factor (TNF) superfamily (3), members of which stimulate a broad range of biological functions including proliferation, induction of cytokine secretion, and apoptosis (For review, see ref. 4).

Successful treatment of adoptive relapsing EAE in SJL/J mice via neuroantigen-specific tolerance induction

REGULATION OF ENCEPHALITOGENIC T CELLS WITH T CELL RECEPTOR PEPTIDES Arthur A. Vandenbark, George Hashim, and Halina Offner, Veterans Affairs Medical Center, Portland, OR. Immunization of Lewis rats with basic protein in adjuvant induces eneephalitogenic T cells specific for the immunodominant 72-89 epitop6, :rid for a secondary epitope, residues 8799. Evaluation of T coil receptor genes utilized by these T cells revealed that the 72-89 specific clones isolated from lymph nodes and spinal cord predominantly utilized V[38.2, whereas the 87-99 specific clones from both organs predominae.tly utilized VI56. These data indicate that the same T cell specificities recovered from the circulation actually penetrate the CNS compartment during induction of EAE. The expression of common V genes allowed the use of TCR peptides for the induction of autoregulatory T cells and antibodies. Injection of the CDR2 peptide corresponding to V~8.2 residues 39-59 or 44-54 not only prevented EAE, but also was eff~ctive as treatment, halting disease progression and speeding recovery from EAE. The rapid offect of TCR peptide therapy suggested triggering of a regulatory recall response to the TCR peptide, resulting in decreased activity of encephalitoganic specificities. In direct support of this idea, TCR peptide treated rats had increased frequencies of TCR peptide specific T cells and decreased frequencies of encephalitogenic T cells in the lymph node, blood, and spinal cords compared to untreated paralyzed rats. These data demonstrate that TCR peptides can induce selective autoregulation of pathogenic T cells, with potential therapeutic application to human autoimmune diseases.