Leishmania donovani infection of a susceptible host results in apoptosis of Th1-like cells: rescue of anti-leishmanial CMI by providing Th1-specific bystander costimulation

A protective immune response against Leishmania donovani infection is mediated by T-helper type 1 (Th1) cells. Th1 induced cell-mediated immunity (CMI), as assessed by anti-leishmanial DTH response, is lost in a susceptible host such as BALB/c mice. Although the impaired Th1 function eventuates in unhindered parasite growth and in manifestation of the susceptible phenotype, the mechanism of down-regulation of the Th1 function is yet to be elucidated. Here, we provide evidence that the parasite down-regulates the expression of a Th1-specific costimulatory molecule, M150, on the surface of infected BALB/c mice-derived macrophages. Th cells are rendered unresponsive to anti-CD3 Ab-mediated stimulation after interaction with infected macrophages. The anergized T cells produce much less IL-2, IL-4 and IFN-gamma compared to those T cells which were costimulated using normal macrophages. The defect in proliferation, anti-CD3 Ab induced unresponsiveness and IFN-gamma but not IL-4 production can be restored by providing bystander costimulation through M150. These results not only unfold a novel immune evasion strategy used by the parasite but also clarify the mechanism of Th1 cell debilitation during the disease. Recovery of Th1 cytokine production by bystander costimulation through M150 may help in formulating a new strategy for the elimination of intracellular parasites.     

Anticryptococcal activity by alveolar macrophages from rats treated with cortisone acetate during different periods of time

Recently mouse models have shown that expression of costimulatory molecules such as B7-1 on tumor cells can induce tumor-specific immunity, suggesting that tumor cells modified to express costimulatory molecules can be a potential tumor vaccine. To investigate the importance of B7-1 co-stimulation in induction of autologous tumor immunity in humans, we established a renal carcinoma cell line, RCC-1, from a tumor resection and studied the patient's antitumor immune responses in vitro. The RCC-1 cell line constitutively expressed major histocompatibility complex (MHC) class I, intercellular adhesion molecule (ICAM)-1, and leukocyte function-associated antigen (LFA)-3 molecules, and MHC class II molecules were induced by interferon-gamma (IFN-gamma) treatment in vitro. However, neither RCC-1- nor IFN-gamma-treated RCC-1 cells expressed B7-1, and both failed to induce T-cell proliferative responses in mixed lymphocyte and tumor cell reaction (MLTR) assays, suggesting that the costimulatory signals provided by cell adhesion molecules such as ICAM-1 and LFA-3 were not sufficient to elicit an antitumor immune response. However, on transfection of the human B7-1 into RCC-1, these cells were able to induce a significant T-cell proliferation in MLTR assays. This T-cell response could be blocked by anti-B7 mAb treatment of the tumor cells. RCC-1B7 cells also induced the generation of tumor-specific cytolytic T lymphocytes to the parent RCC-1 cells in vitro, with little nonspecific cytolysis of an unrelated RCC line, A498, or autologous phytohemagglutinin (PHA) blasts. This specific cytotoxicity could be abrogated by anti-CD8 mAb and complement treatment. In summary, our study indicates that B7-1-CD28 interaction plays a critical role in induction of autologous tumor-specific cytotoxic T lymphocytes (CTLs) in humans, suggesting that the costimulatory molecule transfected tumor cells could be useful in expanding tumor-specific autologous CTL in vitro for adoptive tumor immunotherapy.     

The mucosal adjuvanticity of cholera toxin involves enhancement of costimulatory activity by selective up-regulation of B7.2 expression

Cholera toxin (CT) is a potent mucosal immunogen and adjuvant that can strongly prime mucosal T cells. The present study was undertaken to investigate the effects of CT on the expression and functional activity of the costimulatory molecules B7.1 and B7.2 on macrophages and the relationship of these effects to the mucosal adjuvanticity of CT. Bone marrow macrophages (BMM) were generated by culturing bone marrow with macrophage CSF or granulocyte-macrophage CSF. After treatment with either CT alone or IFN-gamma alone, B7.2 expression on BMM was moderately up-regulated and was further increased when BMM were treated with both CT and IFN-gamma together. Interestingly, CT had no effect on B7.1 expression despite the close relationship between these two molecules. Up-regulation of B7.2 expression by CT was mediated by intracellular cAMP production, in that CT-B subunit had no effect and dibutyryl cAMP could mimic the effect. CT increased functional costimulatory activity of macrophages for both anti-CD3-stimulated and allostimulated T cells, an increase that was blocked by anti-B7.2, but not anti-B7.1, Ab. B7.2 expression by Mac1+ Peyer's patch cells was increased after intraluminal exposure to CT in vivo. Treatment of mice with anti-B7.2 Ab in vivo inhibited both the mucosal adjuvanticity and the immunogenicity of CT. We conclude that CT enhances the costimulatory activity of mucosal APC by differentially up-regulating B7.2 expression, an effect that appears to be important for its mucosal adjuvanticity and immunogenicity.     

Regulation of T helper cell differentiation in vivo by soluble and membrane proteins provided by antigen-presenting cells

The aim of this study was to test whether the nature of the antigen-presenting cell (APC) can influence the Th1/Th2 balance in vivo. Our data show that dendritic cells (DC), pulsed extracorporeally with antigen, induced the development of cells secreting IL-2, IFN-gamma and IL-4 upon antigen rechallenge in vitro. Priming with peritoneal macrophages sensitized cells that produced IL-4 but not IFN-gamma. To identify the factors involved in T helper development, mice were primed with APC with or without treatment with neutralizing antibodies to costimulatory molecules or cytokines. Our results indicate that priming with DC or macrophages is strictly dependent on the CD28-CTLA4/B7 interaction. Of note, CD86 provides the initial signal to induce naive T cells to become IL-4 producers, whereas CD80 is a more neutral differentiation signal. IL-12, released by the DC, appears as a potent and obligatory inducer of differentiation for IFN-gamma-producing cells. IL-6, although produced by both APC populations, is necessary to direct activation of the Th2-type response by macrophages but not by DC.     

Impaired categorical perception of synthetic speech sounds in schizophrenia

BACKGROUND: The rationale for the study was based on the hypothesis that decreased or absent expression on tumor cells of adhesion molecules, the class I or class II major histocompatibility complex (MHC) molecules, or costimulatory molecules might be responsible, in part, for the poor ability of squamous cell carcinoma of the head and neck (SCCHN) to induce generation of antitumor effector cells in vitro and in vivo. OBJECTIVE: To investigate expression of intercellular adhesion molecule-1 (ICAM-1) and lymphocyte function associated antigen-3 (LFA-3) and distribution of the costimulatory molecules, B7.1, B7.2, and CD40, and of class I and class II MHC molecules on SCCHN cells in situ and on SCCHN cell lines. SETTING: University medical centers. DESIGN: Expression of ICAM-1, LFA-3, MHC molecules, B7.1, B7.2, and CD40 was evaluated in human SCCHN biopsy specimens by immunohistochemistry and on SCCHN cell lines by flow cytometry. To confirm our hypothesis that impaired T-cell activation observed in patients with SCCHN is caused by the absence of costimulatory B7 molecules, a B7-negative SCCHN cell line was transduced with the B7.1 gene, using a retroviral vector, and tested in mixed lymphocyte tumor cocultures. RESULTS: In contrast to abundant expression of ICAM-1, LFA-3, class I MHC molecules, and CD40, the absence of B7.1, B7.2, and class II MHC molecules on tumor cells was observed in situ and in vitro. Lymphocytes and antigen-presenting cells in inflammatory infiltrates surrounding tumor cell clusters expressed both costimulatory and adhesion molecules. The SCCHN lines negative for B7.1 and class II MHC antigens failed to induce proliferation of T cells in mixed lymphocyte tumor cocultures. However, when these cell lines were transduced with the B7.1 gene, their ability to induce T-cell proliferation in mixed lymphocyte tumor cocultures was restored. CONCLUSIONS: The absence of B7 protein or class II MHC antigen expression on human SCCHN cells is responsible for the failure of these tumors to induce proliferation of T cells in vitro. Transduction of the B7.1 gene into SCCHN restores the ability of the tumor to induce T-cell proliferation in vitro.     

Dependence of mesenchymal cell responses on duration of exposure to bone morphogenetic protein-2 in vitro

Murine Leydig (TM3) and Sertoli (TM4) cell lines were studied as nonprofessional antigen-presenting cells using the antigen model of human choriogonadotropin (hCG alpha/beta) and specific T-cell hybridomas. Both cell lines were treated with IFN-gamma to induce I-A(d) and I-E(d) molecules expression. Only the TM3 cell line, which expressed MHC-class II molecules upon IFN-gamma stimulation, was able to uptake, process, and present the human choriogonadotropin beta subunit to related T-cell hybridomas. Interestingly, the TM3 cell line was incapable of presenting the human choriogonadotropin alpha subunit, the presentation of which, by classical APC, is highly efficient. Using T-cell hybridomas directed against the immunogenic regions of hCG alpha/beta previously described in BALB/c mice, we showed that the TM3 cell line generated a narrower peptide repertoire than classical APC (i.e., B cells, macrophages, and dendritic cells). This experimental system suggests that Leydig cells could initiate, in vivo, an autoimmune process directed against gonadal tissues. In particular, such a mechanism has been evoked in experimental autoimmune orchitis.     

Vector-host-parasite inter-relationships in leishmaniasis. I. The effect of Leishmania parasites on rate of digestion of blood proteins from various vertebrate hosts by the sand fly Phlebotomus langeroni (Diptera: Psychodidae)

Phlebotomus langeroni collected from a leishmaniasis endemic focus at Et Agamy, Alexandria, Egypt, were found to have fed on blood from man, dogs (Canis familiaris) and rats (Rattus rattus). The effect of the kind of blood meal on the development and the life-cycle of L. infantum and L. major in laboratory reared P. langeroni was therefore investigated. A membrane feeding technique was used to infect sand flies. Gut smears of infected females were examined immediately after feeding and daily for 16 days. Nectomonads and short promastigote forms of L. infantum or L. major were detected in females fed on human, dog and rat bloods at all intervals. Paramastigotes (infective stage) were present only in females fed on dog blood containing L. infantum or L. major and in those fed on rat blood containing L. major. It is concluded that among the factors influencing the Leishmania-phlebotomus relationship is the natural medium in which the parasite is present in vivo. The blood of the natural reservoir host(s) is the key factor for the development of the infective parasite form in the sand fly and P. langeroni could be considered a potential vector for transmitting L. infantum from dogs and L. major from rats and dogs but not from man. This investigation offers a new concept for the study of interactions among vector, host and parasites in Leishmania transmission.     

Activation of human dendritic cells following infection with Mycobacterium tuberculosis

Dendritic cells (DC) play an essential role in the initiation of primary T cell responses to foreign Ag. It is likely that these potent APC are critical in the initiation of immune responses to pathogens, such as bacteria or parasites. However, little is known about the interaction of these important APC with pathogens. To address this issue, the interaction of the bacterium Mycobacterium tuberculosis with human DC was studied. DC generated from human peripheral blood by short term culture in medium containing recombinant human cytokines granulocyte-macrophage-CSF and IL-4 were capable of phagocytosing M. tuberculosis. Infection of DC with live M. tuberculosis bacilli resulted in increased APC surface expression of the costimulatory molecules CD54, CD40, and B7.1, as well as MHC class I molecules. In addition, infected DC secreted elevated levels of inflammatory cytokines, including TNF-alpha, IL-1, and IL-12. M. tuberculosis-infected human monocytes also secreted inflammatory cytokines, but exhibited no enhancement of costimulatory or MHC class I molecule expression. These data indicate that infection with M. tuberculosis results in the direct activation and maturation of these DC. In vivo, such activation may facilitate migration to the lymph nodes, and enhance presentation of Ag to T cells, thereby facilitating the induction of the immune response against this pathogen.     

Prolonged administration of dexamethasone induces limited reactivation of visceral leishmaniasis in chronically infected BALB/c mice

Leishmania parasites persist in their vertebrate host after the treatment-induced clinical cure and in the asymptomatic infection. They confer resistance to reinfection but represent a risk of occurrence of acute leishmaniosis in immunosuppressed conditions. We examined the effects of prolonged dexamethasone administration on a chronic Leishmania infantum infection. Splenic T cell populations from the long-term-infected BALB/c mice were reduced by 55%, whereas those from uninfected controls were depleted by 85%. The ability of the remaining spleen cells to produce IL-2, IFN-gamma, IL-4 and TNF-alpha after in vitro specific stimulation decreased twofold, and the specific anti-leishmanial antibodies declined 3- to 5-fold. Liver, spleen and bone marrow are the main L. infantum targets in natural and experimental infections. Three-fold increase of amastigote burden was evidenced in the spleen, after dexamethasone administration was prolonged for over 2 months. No reactivation of Leishmania proliferation was disclosed in the liver and bone marrow. These results show a decreased sensitivity of splenic T cells to dexamethasone in a chronic Leishmania infection and a distinct response of the Leishmania-infected target organs to the dexamethasone-induced immunosuppression.     

Morphology and structure of photosensitive dye J-aggregates adsorbed on AgBr microcrystals grown in gelatin

Our purpose was to determine the expression of the major histocompatibility complex (MHC) class I and class II gene products as well as the costimulatory molecules B7-1 and B7-2 on cervical epithelial cells, and to determine to what extent inflammatory cytokines regulate their expression. Immunohistology and flow cytometry techniques were used to identify and quantify MHC class I and class II molecules, and the costimulatory molecules B7.1 and B7.2, on sections and primary epithelial cell cultures of human endo- and ectocervix. MHC class I but not class II molecules were constitutively expressed on tissue sections and primary epithelial cell cultures derived from endo- and ectocervix. Expression of MHC class I and class II was upregulated in vitro by IFN-gamma in a time and dose dependent fashion. The induction of class II expression was more pronounced on ectocervical cells than on endocervical cells. MHC class I but not class II expression was also enhanced by IFN-alpha as well as TNF-alpha. TNF-alpha and TGF-beta1 inhibited the IFN-gamma induced MHC class II expression. Expression of the costimulatory molecules B7-1 and B7-2 were not detected in tissue sections or on resting or cytokine-treated cervical epithelial cells in vitro. The present results support the concept that endo- and ectocervical epithelial cells, like their counterparts at other mucosal sites. constitutively express MHC class I molecules and can express MHC class II upon cytokine stimulation, indicating that they are capable of presenting antigens to T-cells.     

Total hip arthroplasty: imaging evaluation

Crosslinking of CD28 receptors on resting T lymphocytes by B7 costimulatory molecules expressed by antigen-presenting cells (APCs) plays a critical role in T-cell activation. Human melanomas express major histocompatibility complex (MHC)-restricted tumor-associated antigens that can be recognized by cytotoxic T lymphocytes (CTL), yet they remain poorly immunogenic. One mechanism for the failure of T-cell response is the lack of expression of costimulatory molecules by human melanoma cells. We have transfected the B7-1 gene into three HLA-A2-expressing human melanoma cell lines, and studied their capacity to stimulate primary human T cells. B7-expressing melanoma cells were excellent inducers of T-cell proliferation, cytokine production, and cytolytic activity in allogeneic mixed lymphocyte cultures through a process dependent on the function of the T-cell receptor as well as interactions between B7:CD28, CD2:LFA-3, and LFA-1:ICAM-1. Subset analysis demonstrated that CD4+ T cells or addition of exogenous interleukin-2 was required for the induction of CD8+ CTL. Untransfected parental melanoma cells were inert as APCs in these cultures. Rotating stimulation of T cells with the three B7-expressing cell lines led to the generation of T-cell lines that were cytolytic for HLA-A2+ melanoma cells and other HLA-A2+ targets that were pulsed with HLA-A2-restricted MART-1 peptides. These data demonstrate that expression of B7-1 by human melanoma cells converts them into effective APCs for the in vitro induction of MHC-restricted, melanoma-specific CTL.     

The role of chemokines and accessory cells in the immunoregulation of cutaneous leishmaniasis

The course of infection with Leishmania parasites is determined by the type of the developing CD4+ T cell immune response. Macrophages and Langerhans cells/dendritic cells play a decisive role in the interaction between the parasites and the host's immune system because they serve as host cells, as accessory cells that present parasite antigen, deliver costimulatory signals and secrete cytokines modulating the T cell activity and as effector cells eliminating the microorganisms. Therefore, we put particular emphasis on characterizing the role of these cells in cutaneous leishmaniasis and the factors regulating their activities. Our results show that (1) expression of the chemokine monocyte chemoattractant protein 1 (MCP-1) is associated with macrophage infiltration into the lesion and stimulation of leishmanicidal activity, (2) Langerhans cells are required for the transport of Leishmania from the infected skin to the draining lymph node and initiation of the specific T cell immune response in the early phase of infection, (3) lymph node dendritic cells containing persistent parasites may be involved in the maintenance of specific immunity, (4) Langerhans cells are able to present L. major LPG to T cells and (5) treatment of mice with antigen-pulsed Langerhans cells induces protective immunity against cutaneous leishmaniasis.     

CD4-mediated and CD8-mediated cytotoxic and proliferative immune responses to Toxoplasma gondii in seropositive humans

Both CD4+ and CD8+ cytotoxic T lymphocytes (CTL) are part of the human immune response to Toxoplasma gondii infection. To further our understanding of Toxoplasma immunity, we investigated factors influencing stimulation of CD4+ or CD8+ human T. gondii-specific immune cells. Both antigen-pulsed and Toxoplasma-infected antigen-presenting cells (APC) induced cell proliferation. Toxoplasma-infected APC elicited strong proliferation of CD4+ cells, but little or no proliferation of CD8+ cells, unless high antigen loads were used. Toxoplasma-infected APC stimulated specific cytotoxicity poorly or not at all, owing to death of stimulated cultures, whereas antigen-pulsed APC strongly elicited specific cytotoxicity. Cytotoxicity elicited by either type of APC resided exclusively in CD4+ T cells in polyclonal cultures. Thus, Toxoplasma-infected APC elicited stronger CD4-mediated than CD8-mediated cell proliferation and generated CD4+ CTL more readily than CD8+ CTL. Nonetheless, specific CD8+ memory cells were demonstrated, and rare CD8+ Toxoplasma-specific CTL were subcloned. Fixed Toxoplasma-infected APC (which induce CD8+ CTL) also elicited cell proliferation, but polyclonal cultures stimulated with these infected APC did not die. Unfixed Toxoplasma-infected APC strongly inhibited phytohemagglutinin-induced cell proliferation, whereas fixed APC did not. These data suggested that infected APC were inhibitory or lethal to some immune cells. Further investigations into interactions between immune cells and Toxoplasma-infected cells likely will help elucidate factors involved in the immunopathogenesis of Toxoplasma infection. As other intracellular parasites, including Plasmodium spp. and Leishmania spp., also elicit CD4+ CTL, such work may help establish paradigms governing immunity to intracellular parasites.     

IL-12 up-regulates CD40 ligand (CD154) expression on human T cells

IL-12 is a heterodimeric cytokine produced by APC that promotes the development of CD4+ Th1 cells and their IFN-gamma production after TCR/CD3 triggering. We here investigated the capacity of IL-12 to modify the expression on T cells of CD40 ligand (CD40L or CD154), a molecule transiently expressed on activated T cells and known to be of utmost importance for cognate interaction with B cells and for activation of dendritic cells and macrophages. Our data demonstrate that IL-12 up-regulates CD40L expression on anti-CD3-activated human peripheral blood T cells. For optimal induction of CD40L, IL-12 synergizes with IL-2 as well as with other costimulatory interactions, such as B7/CD28. The effect of IL-12 was observed at both the protein and the mRNA level. T cells costimulated by IL-12 provided more efficient help for IL-4-dependent B cell proliferation and for IgG production than when activated in the absence of IL-12. This helper activity was blocked by an mAb against CD40L, indicating that the effect of IL-12 on B cells is mediated indirectly through CD40L. The data thus suggest that the effects of IL-12 on cellular and humoral immune responses are partly mediated through CD40L induction.     

B7-1 and B7-2 monoclonal antibodies modulate the severity of murine Lyme arthritis

We assessed the role of B7-1 and B7-2 costimulatory molecules on the course of murine Lyme borreliosis because experimental Lyme arthritis is dependent, at least partially, upon the development of the host immune response and these costimulatory molecules have been implicated in CD4+ T-cell differentiation. We demonstrated that Borrelia burgdorferi infection upregulated the surface expression of B7-1 and B7-2 in macrophages and B7-2 expression in B cells. Anti-B7-2 monoclonal antibody (MAb) or both anti-B7-2 and anti-B7-1 MAbs produced a dose-dependent increase in the severity of Lyme arthritis in C3H/HeN mice. In contrast, the administration of an anti-B7-1 MAb reduced the degree of arthritis. These effects occurred independently of significant alteration in B. burgdorferi-specific immune responses, including splenocyte proliferative responses to B. burgdorferi, B. burgdorferi antibody levels and specificity, and mRNA levels of gamma interferon, interleukin-4 (IL-4), IL-10, and IL-12 in the spleen. These results demonstrate that signaling delivered by B7-1 and B7-2 plays a role in determining the severity of acute murine Lyme arthritis.     

The effects of a community-based pulmonary rehabilitation programme on exercise tolerance and quality of life: a randomized controlled trial

The molecules B7.1 and B7.2 deliver costimulatory signals of critical importance to naive T cells, and may thus be involved in abrogation of oral tolerance in IBD. Functional disparity apparently exists among antigen-presenting cells in vivo. We wanted to examine if differential B7 expression occurs on mucosal macrophage subsets. Cryosections of bowel specimens from patients with IBD and normal controls were subjected to immunofluorescence and immunoperoxidase staining. In normal mucosa, selective subepithelial accumulation of B7.2+ cells was found. In inflamed IBD mucosa, however, subsets appeared consisting of both B7.2(hi) and B7.1(hi) cells as well as CD14(hi) macrophages. Notably, outside lymphoid aggregates the prominent fraction of recently recruited CD14(hi) macrophages comprised most (approximately 80%) of the B7.1(hi) cells, whereas most (approximately 70%) B7.2(hi) cells were identified as resident mucosal macrophages (CD14(lo) or CD14-). Differential expression of B7.1 and B7.2 on two functionally different subsets of intestinal macrophages implies separate immunoregulatory roles for the two molecules. This finding is in keeping with recent experimental data demonstrating that monocyte-derived cells are crucial for immune responses at mucosal surfaces. Preferential B7.1 up-regulation might be critical in breaking the immunological tolerance to luminal antigens in IBD, but it cannot be excluded that it is a secondary pathogenic event.     

Increased expression of costimulatory molecules on peripheral blood monocytes in patients with Crohn's disease

BACKGROUND: Activation of T lymphocytes and monocytes/macrophages has been implicated in the pathogenesis of Crohn's disease (CD). Costimulatory molecules play important roles in optimal T-cell activation. METHODS: With flow cytometric analysis we have investigated the expression of the costimulatory molecules B7-1 (CD80), B7-2 (CD86), and CD18 and the intercellular adhesion molecule-1 (ICAM-1) on peripheral blood monocytes and the expression of the activation markers HLA-DR and IL-2R (CD25) on peripheral blood T lymphocytes from 31 CD patients, 17 ulcerative colitis (UC) patients, and 10 healthy controls. RESULTS: In CD patients the percentage of activated T cells (CD3+ HLA-DR+ and CD3+ IL-2R+) was significantly increased compared with those of controls and UC patients (P < 0.05). Most monocytes from all three groups expressed B7-2, CD18, and ICAM-1 molecules (all > 79%), but only a few positive cells expressed B7-1 molecules (< 5%). No significant differences were detected in the percentage positivity of all costimulatory molecules tested among CD, UC, and controls. The mean fluorescence intensity (MFI) of B7-1 in all three groups was very weak and not significantly different. However, in CD patients there was a significantly increased MFI of B7-2, CD18, and ICAM-1 molecules compared with UC and controls (P < 0.05). On the other hand, both the percentage positivity and MFI of HLA-DR molecules on monocytes of UC patients were significantly lower than those of CD patients and controls (P < 0.05). CONCLUSIONS: Expression of the costimulatory molecules B7-2, CD18, and ICAM-1 on peripheral blood monocytes of CD patients is increased. In CD patients activation of peripheral T lymphocytes may correlate with increased expression of these costimulatory molecules on peripheral blood monocytes.     

Characterization of and functional antigen presentation by central nervous system mononuclear cells from mice infected with Theiler's murine encephalomyelitis virus

We examined the phenotype and function of cells infiltrating the central nervous system (CNS) of mice persistently infected with Theiler's murine encephalomyelitis virus (TMEV) for evidence that viral antigens are presented to T cells within the CNS. Expression of major histocompatibility complex (MHC) class II in the spinal cords of mice infected with TMEV was found predominantly on macrophages in demyelinating lesions. The distribution of I-As staining overlapped that of the macrophage marker sialoadhesin in frozen sections and coincided with that of another macrophage/microglial cell marker, F4/80, by flow cytometry. In contrast, astrocytes, identified by staining with glial fibrillary acidic protein, rarely expressed detectable MHC class II, although fibrillary gliosis associated with the CNS damage was clearly seen. The costimulatory molecules B7-1 and B7-2 were expressed on the surface of most MHC class II-positive cells in the CNS, at levels exceeding those found in the spleens of the infected mice. Immunohistochemistry revealed that B7-1 and B7-2 colocalized on large F4/80(+) macrophages/microglia in the spinal cord lesions. In contrast, CD4(+) T cells in the lesions expressed mainly B7-2, which was found primarily on blastoid CD4(+) T cells located toward the periphery of the lesions. Most interestingly, plastic-adherent cells freshly isolated from the spinal cords of TMEV-infected mice were able to process and present TMEV and horse myoglobin to antigen-specific T-cell lines. Furthermore, these cells were able to activate a TMEV epitope-specific T-cell line in the absence of added antigen, providing conclusive evidence for the endogenous processing and presentation of virus epitopes within the CNS of persistently infected SJL/J mice.