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IL-10-producing B cells have a regulatory effect in various mouse models for immune-mediated disorders via secretion of IL-10, a potent immunoregulatory cytokine. However, currently, the signaling pathways that regulate IL-10 production in B cells are not well understood. Here, we show that TLR signaling, but not BCR activation or CD40 ligation, induces potent production of IL-10 in human B cells. We demonstrate that the activation of STAT3 and ERK is required for TLR-induced IL-10 production by B cells, since inhibition of STAT3 or ERK activation abrogates TLR-induced IL-10 production. We also uncover a novel function of the TLR-MyD88-STAT3 pathway in B cells, namely controlling IL-10 production, in addition to the known role for this pathway in antibody production. Furthermore, IFN-α, a member of the type I IFN family, differentially modulates TLR7/8- and TLR9-activated STAT3 and ERK in B cells, which provides an explanation for our findings that IFN-α enhances TLR7/8-induced, but not TLR9-induced IL-10 production. These results yield insights into the mechanisms by which TLR signaling regulates IL-10 production in B cells and how type I IFN modulates TLR-mediated IL-10 production by B cells, therefore providing potential targets to modulate the function of IL-10-producing B cells.

The recognition of microbial patterns by Toll-like receptors (TLRs) is critical for activation of the innate immune system. Although TLRs are expressed by human CD4(+) T cells, their function is not well understood. Here we found that engagement of TLR7 in CD4(+) T cells induced intracellular calcium flux with activation of an anergic gene-expression program dependent on the transcription factor NFATc2, as well as unresponsiveness of T cells. As chronic infection with RNA viruses such as human immunodeficiency virus type 1 (HIV-1) induces profound dysfunction of CD4(+) T cells, we investigated the role of TLR7-induced anergy in HIV-1 infection. Silencing of TLR7 markedly decreased the frequency of HIV-1-infected CD4(+) T cells and restored the responsiveness of those HIV-1(+) CD4(+) T cells. Our results elucidate a previously unknown function for microbial pattern-recognition receptors in the downregulation of immune responses.

Immunogenetic Variation Many genetic variants have been implicated in disease but their effects in function across tissues and cell-types remain to be resolved. Raj et al. (p. 519 ) present an analysis of expression quantative trait loci (eQTL) measuring messenger RNA levels and examined correlations between genotypes and gene expression in purified monocytes and T cells in healthy individuals of European, African, and Asian descent. Most, but not all, of the eQTLs and their effects on expression were shared between the populations, as well as a substantial proportion between the cell types. Links were found with disease-associated variants and loci that previous genome-wide analyses have implicated in neurodegenerative and autoimmune diseases.

We identified three retinoid-related orphan receptor gamma t (RORγt)-specific inhibitors that suppress T helper 17 (Th17) cell responses, including Th17-cell-mediated autoimmune disease. We systemically characterized RORγt binding in the presence and absence of drugs with corresponding whole-genome transcriptome sequencing. RORγt acts as a direct activator of Th17 cell signature genes and a direct repressor of signature genes from other T cell lineages; its strongest transcriptional effects are on cis-regulatory sites containing the RORα binding motif. RORγt is central in a densely interconnected regulatory network that shapes the balance of T cell differentiation. Here, the three inhibitors modulated the RORγt-dependent transcriptional network to varying extents and through distinct mechanisms. Whereas one inhibitor displaced RORγt from its target loci, the other two inhibitors affected transcription predominantly without removing DNA binding. Our work illustrates the power of a system-scale analysis of transcriptional regulation to characterize potential therapeutic compounds that inhibit pathogenic Th17 cells and suppress autoimmunity., United States. National Institutes of Health (DP1OD003958-01)

Multiple sclerosis (MS) is a common neurological disease among young adults that typically presents with relapsing, remitting neurological symptoms. Historically it has been described as a demyelinating disease; however, research has shown that axonal degeneration, in addition to diffuse inflammation in normal-appearing white matter, contribute significantly to its pathology. This article will address MS genetics and immunology, differences between white and gray matter pathology, and the role of advanced imaging techniques. The clinical and pathological hallmarks of acute disseminated encephalomyelitis are also reviewed.

Multiple sclerosis (MS) is a multifocal inflammatory autoimmune disease that grossly affects the CNS white matter resulting in progressive neurodegeneration in genetically susceptible hosts. In the past half-century great strides have been made in understanding the genetic risk, immunomodulation, and immune dysregulation that is responsible for establishing MS, as well as in the development of various disease modifying therapies that have succeeded in controlling disease evolution and survival. While MS immunopathology has long been attributed to the Th1 subset of T cells, recent work has identified novel genetic susceptibility loci in addition to establishing a putative role for the regulatory T cell and Th17 T cell subsets, among a potential role for CD8+ T cell and B cell subsets. Nevertheless the identification of an antigen or infectious agent responsible for driving the ongoing inflammatory and humoral response in MS remains to be elucidated. This chapter examines the paradigms currently being utilized for deciphering MS, highlights current disease modifying therapies, and defines compelling future directions for MS-based research.

In biomedical applications, an experimenter encounters different potential sources of variation in data such as individual samples, multiple experimental conditions, and multivariate responses of a panel of markers such as from a signaling network. In multiparametric cytometry, which is often used for analyzing patient samples, such issues are critical. While computational methods can identify cell populations in individual samples, without the ability to automatically match them across samples, it is difficult to compare and characterize the populations in typical experiments, such as those responding to various stimulations or distinctive of particular patients or time-points, especially when there are many samples. Joint Clustering and Matching (JCM) is a multi-level framework for simultaneous modeling and registration of populations across a cohort. JCM models every population with a robust multivariate probability distribution. Simultaneously, JCM fits a random-effects model to construct an overall batch template – used for registering populations across samples, and classifying new samples. By tackling systems-level variation, JCM supports practical biomedical applications involving large cohorts. Software for fitting the JCM models have been implemented in an R package EMMIX-JCM, available from http://www.maths.uq.edu.au/~gjm/mix_soft/EMMIX-JCM/.

Multiple sclerosis (MS) is an inflammatory disease of the central nervous system (CNS) characterized by autoimmune-mediated demyelination and neurodegeneration. The CNS of patients with MS harbors expanded clones of antigen-experienced B cells that reside in distinct compartments including the meninges, cerebrospinal fluid (CSF), and parenchyma. It is not understood whether this immune infiltrate initiates its development in the CNS or in peripheral tissues. B cells in the CSF can exchange with those in peripheral blood, implying that CNS B cells may have access to lymphoid tissue that may be the specific compartment(s) in which CNS-resident B cells encounter antigen and experience affinity maturation. Paired tissues were used to determine whether the B cells that populate the CNS mature in the draining cervical lymph nodes (CLNs). High-throughput sequencing of the antibody repertoire demonstrated that clonally expanded B cells were present in both compartments. Founding members of clones were more often found in the draining CLNs. More mature clonal members derived from these founders were observed in the draining CLNs and also in the CNS, including lesions. These data provide new evidence that B cells traffic freely across the tissue barrier, with the majority of B cell maturation occurring outside of the CNS in the secondary lymphoid tissue. Our study may aid in further defining the mechanisms of immunomodulatory therapies that either deplete circulating B cells or affect the intrathecal B cell compartment by inhibiting lymphocyte transmigration into the CNS.

Thymectomy in mice on neonatal day 3 leads to the development of multiorgan autoimmune disease due to loss of a CD+CD25+ T cell regulatory population in their peripheral lymphoid tissues. Here, we report the identification of a CD4+ population of regulatory T cells in the circulation of humans expressing high levels of CD25 that exhibit in vitro characteristics identical with those of the CD4+CD25+ regulatory cells isolated in mice. With TCR cross-linking, CD4+CD25high cells did not proliferate but instead totally inhibited proliferation and cytokine secretion by activated CD4+CD25− responder T cells in a contact-dependent manner. The CD4+CD25high regulatory T cells expressed high levels of CD45RO but not CD45RA, akin to the expression of CD45RBlow on murine CD4+CD25+ regulatory cells. Increasing the strength of signal by providing either costimulation with CD28 cross-linking or the addition of IL-2 to a maximal anti-CD3 stimulus resulted in a modest induction of proliferation and the loss of observable suppression in cocultures of CD4+CD25high regulatory cells and CD4+CD25− responder cells. Whereas higher ratios of CD4+CD25high T cells are required to suppress proliferation if the PD-L1 receptor is blocked, regulatory cell function is shown to persist in the absence of the PD-1/PD-L1 or CTLA-4/B7 pathway. Thus, regulatory CD4 T cells expressing high levels of the IL-2 receptor are present in humans, providing the opportunity to determine whether alterations of these populations of T cells are involved in the induction of human autoimmune disorders.

Antibiotic treatment-resistant Lyme arthritis is a chronic inflammatory joint disease that follows infection with Borrelia burgdorferi (Bb). A marked Ab and T cell response to Bb outer surface protein A (OspA) often develops during prolonged episodes of arthritis. Furthermore, cross-reaction between the bacterial OspA and human LFA-1αL at the T cell level and the inability to detect Bb in the joint implicate an autoimmune mechanism. To analyze the nature of response to OspA and LFA-1αL, we used OspA-specific T cell hybrids from DR4 transgenic mice, as well as cloned human cells specific for OspA165–184, the immunodominant epitope, from five DRB1*0401+ patients, using OspA-MHC class II tetramers. Although OspA165–184 stimulated nearly all OspA-specific human T cell clones tested to proliferate and secrete IFN-γ and IL-13, LFA-1αL326–345 stimulated ∼10% of these clones to proliferate and a greater percentage to secrete IL-13. Assays with LFA- or OspA-DR4 monomers revealed that higher concentrations of LFA-DR4 were needed to stimulate dual-reactive T cell hybrids. Our analysis at the clonal level demonstrates that human LFA-1αL326–345 behaves as a partial agonist, perhaps playing a role in perpetuating symptoms of arthritis.

Rapamycin is a fungal macrolide that inhibits the proliferation of T cells. Studies in both animals and humans have found that rapamycin significantly reduces graft rejection. However, though CD8+ T cells are involved in graft infiltration and rejection, little is known regarding the effects of rapamycin on CD8+ human T cell responses. In this study, we examined the mechanism of rapamycin-induced inhibition of Ag-driven activation of CD8+ T cells. Surprisingly, a heterogeneous proliferative response in the presence of rapamycin was observed among different Ag-specific CD8+ T cell clones; this was also observed in CD8+ peripheral blood T cells activated with TCR cross-linking ex vivo. Inhibition of T cell proliferation by rapamycin was controlled by both the strength of signal delivered through the Ag receptor as well as the specific costimulatory signals received by the T cell. Rapamycin-resistant proliferation occurred despite inhibition of p70s6 kinase activity. Moreover, rapamycin-resistant proliferation of the CD8+ T cell clones was blocked by anti-IL-2 Abs, suggesting that while some of the parallel pathways triggered by IL-2R signaling are sensitive to the effects of rapamycin, others account for the Ag-driven rapamycin resistance. These data provide a new framework for examining the specific mechanism of action of rapamycin in human disease.

The cytokine profile of invariant CD4(-)CD8(-)V alpha 24J alpha Q T cells from patients with multiple sclerosis (MS) was compared with that of healthy controls. CD4(-)CD8(-)V alpha 24(+) T cells from the peripheral blood of 12 patients with relapsing-remitting MS (RR-MS), 5 patients with progressive MS (CP-MS), and 9 control individuals were directly sorted into single wells and expanded in vitro for analysis of IL-4 and IFN-gamma secretion; 315 V alpha 24J alpha Q T cell clones were generated and their T cell receptor (TCR) sequenced. T cell functionality was determined by examining cytokine secretion upon TCR cross-linking. RR-MS patients exhibited lower frequencies of IL-4 secreting CD4(-)CD8(-)V alpha 24J alpha Q T cell clones than patients with CP-MS and controls. No differences in IFN-gamma secretion were observed between the groups. An IL-4 positive cytokine profile could be correlated to the cloning efficiency of the V alpha 24J alpha Q T cells. We conclude that alterations in cytokine secretion patterns of CD4(-)CD8(-)V alpha 24J alpha Q T cells may influence the immune system and thus contribute to relapsing-remitting MS.

Multiple sclerosis (MS) is an inflammatory disease of the central nervous system white matter. The association of the disease with MHC genes, the inflammatory white matter infiltrates, similarities with animal models, and the observation that MS can be treated with immunomodulatory and immunosuppressive therapies support the hypothesis that autoimmunity plays a major role in the disease pathology. Evidence supports activated CD4+ myelin-reactive T cells as major mediators of the disease. In addition, a renewed interest in the possible contribution of B cells to MS immunopathology has been sparked by nonhuman primate and MS pathological studies. This review focuses on the immunopathology of MS, outlining the hypothetical steps of tolerance breakdown and the molecules that play a role in the migration of autoreactive cells to the CNS. Particular focus is given to autoreactive T cells and cytokines as well as B cells and autoantibodies and their role in CNS pathogenesis in MS.

The application of self-antigens as therapeutic tools is validated in inbred animal models of autoimmune disease. Mechanisms of antigen-induced tolerance (apoptosis, anergy, regulatory T cells and immune deviation) are being clarified in relation to the properties of antigens and the modes and routes of their delivery. Mucosa-mediated tolerance remains the predominant mode of antigen-specific therapy but awaits demonstration of clinical efficacy in human autoimmune disease.

Following infection by human T cell lymphotrophic virus-I (HTLV-I), high frequencies of polyclonal Tax11–19-reactive CD8+ T cells can be detected in the peripheral blood. To investigate whether there are differences in the effector functions of these cells, we generated a panel of Tax11–19-reactive T cell clones by single cell sorting of HLA-A2/Tax11–19 tetramer binding CD8+ T cells followed by repeated stimulation with PHA and IL-2. Examination of the TCRs revealed 17 different T cell clones with unique clonal origins. Nine representative CD8+ T cell clones showed a similar cytotoxic dose-response activity against Ag-pulsed target cells, even though they express different TCRs. This cytotoxic effector function was not influenced by the engagement of either CD28 or CD2 costimulatory molecules. In contrast to the cytotoxic activity, qualitatively different degrees of proliferative response and cytokine secretion were observed among T cell clones of different clonal origin. The induction of proliferation and cytokine secretion required the engagement of costimulatory molecules, particularly CD2-LFA-3 interaction. These results indicate that functionally diverse, polyclonal CTL populations can be activated specific to a single immunodominant viral epitope; they can manifest virtually identical cytotoxic effector function but have marked differences in proliferation and cytokine secretion.

We identified a novel population of human T cells, studied directly ex vivo, that co-express surface B7-1 and intracellular IL-4. These peripheral blood B7-1+/CD4+ T cells expressed cell surface molecules associated with differentiation including CD45RO and MHC class II, yet were CD69(-) and CD25(-). In short-term cultures, T cell receptor (TCR) cross-linking induced further IL-4 production with little IFN-gamma or TNF-alpha. In marked contrast, CD4+ T cells negative for B7-1 expressed intracellular IFN-gamma and high amounts of TNF-alpha but little IL-4 upon TCR cross-linking. The CD4+/B7-1+/IL-4-expressing T cells were of polyclonal origin based on their diverse TCR repertoire. To explore the biological significance of this B7-1+/IL-4+ T cell population and to assess its potential regulatory role in autoimmune disease, we examined whether these T cells isolated ex vivo were altered in subjects with multiple sclerosis (MS). While the frequency of B7-1+ T cells was enhanced in patients with MS as compared to normal subjects, there was a significant diminution of B7-1+/IL-4+ T cells in the patients. The decrease in these IL-4-producing T cells in patients with autoimmune disease is consistent with a possible role as immunoregulatory T cells.

Quantitative and qualitative defects in CD1d-restricted T cells have been demonstrated in human and murine autoimmune diseases. To investigate the transcriptional consequences of T cell receptor activation in human Vα24JαQ T cell clones, DNA microarrays were used to quantitate changes in mRNA levels after anti-CD3 stimulation of clones derived from identical twins discordant for type 1 diabetes and IL-4 secretion. Activation resulted in significant modulation of 226 transcripts in the IL-4 secreting clone and 86 in the IL-4-null clone. Only 28 of these genes were in common. The differences observed suggest both ineffective differentiation of diabetic Vα24JαQ T cells and a role for invariant T cells in the recruitment and activation of cells from the myeloid lineage.

We examined the effect of glatiramer acetate, a random copolymer of alanine, lysine, glutamic acid, and tyrosine, on antigen-specific T-cell responses in patients with multiple sclerosis (MS). Glatiramer acetate (Copaxone) functioned as a universal antigen, inducing proliferation, independent of any prior exposure to the polymer, in T-cell lines prepared from MS or healthy subjects. However, for most patients, daily injections of glatiramer acetate abolished this T-cell response and promoted the secretion of IL-5 and IL-13, which are characteristic of Th2 cells. The surviving glatiramer acetate-reactive T cells exhibited a greater degree of degeneracy as measured by cross-reactive responses to combinatorial peptide libraries. Thus, it appears that, in some individuals, in vivo administration of glatiramer acetate induces highly cross-reactive T cells that secrete Th2 cytokines. To our knowledge, glatiramer acetate is the first agent that suppresses human autoimmune disease and alters immune function by engaging the T-cell receptor. This compound may be useful in a variety of autoimmune disorders in which immune deviation to a Th2 type of response is desirable.

T-cell co-stimulation delivered by the molecules B7-1 or B7-2 through CD28 has a positive effect on T-cell activation, whereas engagement of cytotoxic T-lymphocyte antigen 4 (CTLA-4) by these molecules inhibits activation. In vivo administration to mice of blocking monoclonal antibodies or Fab fragments against CTLA-4 can augment antigen-specific T-cell responses and, thus, therapy with monoclonal antibody against CTLA-4 has potential applications for tumor therapy and enhancement of vaccine immunization. The effects of B7-1 and B7-2 co-stimulation through CD28 depend on the strength of the signal delivered through the T-cell receptor (TCR) and the activation state of T cells during activation. Thus, we sought to determine whether these factors similarly influence the effect of B7-mediated signals delivered through CTLA-4 during T-cell activation. Using freshly isolated human T cells and Fab fragments of a monoclonal antibody against CTLA-4, we demonstrate here that CTLA-4 blockade can enhance or inhibit the clonal expansion of different T cells that respond to the same antigen, depending on both the T-cell activation state and the strength of the T-cell receptor signal delivered during T-cell stimulation. Thus, for whole T-cell populations, blocking a negative signal may paradoxically inhibit immune responses. These results provide a theoretical framework for clinical trials in which co-stimulatory signals are manipulated in an attempt to modulate the immune response in human disease.

Objective: To determine if there are variables linked to responsiveness to pulse cyclophosphamide/methylprednisolone therapy in progressive Multiple Scerosis (MS). Background: MS is a presumed autoimmune disease of the CNS in which immunosuppressive and immunomodulatory treatments are being used. We have treated patient with the progressive form of MS using a regimen consisting of pulse cyclophosphamide/methylprednisolone that is given as an outpatient at 4-8 week intervals similar to lupus nephritis protocols. Design/Methods: We investigated a series of 95 consecutive progressive MS patient treated in an open label fashion in an effort to identify factors linked to response to treatment. Clinical outcome measures included status at 12 months and time to failure determined by EDSS change and global physician impression. For each endpoint associations were examined between outcome and patient characteristics including gender age at onset of disease and treatment, EDSS 1 year previously and at start of treatment, duration of MS, previous treatment, age at onset and duration of progression, and primary vs secondary progressive MS. Result: Of the variables studied, age, gender, age at onset, and age at treatment did not correlate with response to therapy. The most significant variable that correlated with response was length of time the patient was in the progressive phase (P=0.048, 12 month change in EDSS; P=0.017, risk for time to failure). Patient that improved on therapy at 12 months had progressive disease for an average of 2.1 years prior to treatment, whereas those stable or worse had progressive disease for 5.0 and 4.1 years respectively. There was a trend (P=0.08) favoring positive clinical responses in secondary progressive as opposed to primary progressive patients. Conclusions: Our data suggest that progressive MS may become refractory to immunosuppressive therapy with time and early intervention when patient enter the progressive stage should be considered. Furthermore, in trials of immunosuppressive agent for progressive MS, duration of progression should be considered as a randomization and analysis variable.

NK T cells are a T cell subset in the human that express an invariant α-chain (Vα24invt T cells). Because of the well-described immunomodulation by glucocorticoids on activation-induced cell death (AICD), the effects of dexamethasone and anti-CD3 stimulation on Vα24invt T cell clones and CD4+ T cell clones were investigated. Dexamethasone significantly enhanced anti-CD3-mediated proliferation of Vα24invt T cells, whereas CD4+ T cells were inhibited. Addition of neutralizing IL-2 Ab partially abrogated dexamethasone-induced potentiation of Vα24invt T cell proliferation, indicating a role for autocrine IL-2 production in corticosteroid-mediated proliferative augmentation. Dexamethasone treatment of anti-CD3-stimulated Vα24invt T cells did not synergize with anti-Fas blockade in enhancing proliferation or preventing AICD. The Vα24invt T cell response to dexamethasone was dependent on the TCR signal strength. In the presence of dexamethasone, lower doses of anti-CD3 inhibited proliferation of Vα24invt T cells and CD4+ T cells; at higher doses of anti-CD3, which caused inhibition of CD4+ T cells, the Vα24invt T cell clones proliferated and were rescued from AICD. These results demonstrate significant differences in TCR signal strength required between Vα24invt T cells and CD4+ cells, and suggest important immunomodulatory consequences for endogenous and exogenous corticosteroids in immune responses.

Markedly elevated levels of serum IL-4 were reported previously in 50% of a small group of type 1 diabetes nonprogessors. To determine the patterns of expression for this phenotype, a larger cohort of 58 families containing type 1 diabetic patients was examined. Analysis of the two-site ELISA assay used to measure serum IL-4 revealed evidence for heterophile antibodies, i.e., nonanalyte substances in serum capable of binding antibodies mutivalently and providing erroneous analyte (e.g., IL-4) quantification. Interestingly, relatives without type 1 diabetes were significantly more likely to have this phenotype than were patients with the disease ( P = 0.003). In addition, the trait appears to have clustered within certain families and was associated with the protective MHC allele DQB1*0602 ( P = 0.008). These results suggest that heterophile antibodies represent an in vivo trait associated with self-tolerance and nonprogression to diabetes.

CD4+T helper 1 (TH1) cells are important mediators of inflammation and are regulated by numerous pathways, including the negative immune receptor Tim-3. We found that Tim-3 is constitutively expressed on cells of the innate immune system in both mice and humans, and that it can synergize with Toll-like receptors. Moreover, an antibody agonist of Tim-3 acted as an adjuvant during induced immune responses, and Tim-3 ligation induced distinct signaling events in T cells and dendritic cells; the latter finding could explain the apparent divergent functions of Tim-3 in these cell types. Thus, by virtue of differential expression on innate versus adaptive immune cells, Tim-3 can either promote or terminate TH1 immunity and may be able to influence a range of inflammatory conditions.

Acute disseminated encephalomyelitis (ADEM), a postinfectious illness of the central nervous system (CNS), is thought to be an autoimmune disease. Here, we characterized the cytokines secreted by myelin-reactive T cells generated from patients with ADEM. The frequency of MBP-reactive T cell lines was ten-fold higher in patients with ADEM compared to patients with encephalitis and normal subjects. Whereas there was no significant IFN-gamma secretion, the predominant cytokine secreted by MBP-reactive T cell lines was IL-4 in patients with ADEM. In contrast, IL-4 secretion was only rarely detected in the controls. The presence of high frequencies of MBP-reactive IL-4 secreting T cells in subjects with ADEM during their recovery phase may be similar to myelin reactive IL-4 secreting T cells observed during the spontaneous recovery of animals with EAE.

Researchers from the fields of basic immunology and autoimmune disease, largely but not exclusively focused on multiple sclerosis, came together at a recent workshop1The International Workshop on Th1/Th2: T-Cell Differentiation in MS and Autoimmunity was held at Savannah, GA, USA, on 1–5 April 1998 1 to develop a better understanding of T helper cell responses in autoimmune disease.

A novel population of both IFN-γ- and IL-10-secreting human T cells that differentiate in the presence of exogenous IL-12 in vitro has recently been described. Whether this T cell population exists in vivo is unknown. Borrelia burgdorferi, the etiologic agent of Lyme disease, can induce a chronic infection in the presence of a vigorous humoral immune response. We established T cell lines specific for B. burgdorferi and tetanus toxoid from subjects with chronic B. burgdorferi infection and healthy controls in limiting dilution experiments and assessed proliferation and cytokine secretion. As expected, higher frequencies of B. burgdorferi-specific precursor T cells were observed in Lyme patients compared with controls. In both groups of subjects, T cell lines specific for B. burgdorferi secreted high amounts of IFN-γ. However, in patients with Lyme disease, 27% of T cell lines secreted not only IFN-γ but also IL-10, which was only observed in 0.6% of B. burgdorferi-reactive T cell lines generated from controls and in none of the tetanus toxoid-reactive T cell lines generated from either Lyme patients and controls. Single cell PHA cloning confirmed that both cytokines were secreted from one clonally expanded precursor cell. Whole mononuclear cells from B. burgdorferi-infected individuals, but not from controls, secreted IL-12. Moreover, neutralizing anti-IL-12 mAbs inhibited the generation of the IFN-γ/IL-10 population. These data demonstrate that this novel population of IL-12-induced IFN-γ/IL-10-secreting T cells is generated in response to chronic B. burgdorferi infection.

Multiple sclerosis (MS) is an inflammatory disease of the myelinated central nervous system that is postulated to be induced by myelin-reactive CD4 T cells. T cell activation requires an antigen-specific signal through the TCR and a costimulatory signal, which can be mediated by B7–1 or B7–2 engagement of CD28. To directly examine the activation state of myelin-reactive T cells in MS, the costimulation requirements necessary to activate myelin basic protein (MBP) or tetanus toxoid (TT)-reactive CD4 T cells were compared between normal controls and MS patients. Peripheral blood T cells were stimulated with Chinese hamster ovary (CHO) cells transfected either with DRB1*1501/DRA0101 chains (t-DR2) alone, or in combination with, B7–1 or B7–2. In the absence of costimulation, T cells from normal subjects stimulated with the recall antigen TT p830–843 were induced to expand and proliferate, but stimulation with MBP p85–99 did not have this effect. In marked contrast, T cells from patients with MS stimulated with MBP p85–99 in the absence of B7–1 or B7–2 signals expanded and proliferated. Thus, MBP-reactive CD4 T cells in patients with MS are costimulation independent and have been previously activated in vivo. These experiments provide further direct evidence for a role of activated MBP-specific CD4 T cells in the pathogenesis of MS.

Type 1 diabetes (insulin-dependent diabetes mellitus, IDDM) is a disease controlled by the major histocompatibility complex (MHC) which results from T-cell-mediated destruction of pancreatic beta-cells. The incomplete concordance in identical twins and the presence of autoreactive T cells and autoantibodies in individuals who do not develop diabetes suggest that other abnormalities must occur in the immune system for disease to result. We therefore investigated a series of at-risk non-progressors and type 1 diabetic patients (including five identical twin/triplet sets discordant for disease). The diabetic siblings had lower frequencies of CD4-CD8- Valpha24JalphaQ+ T cells compared with their non-diabetic sibling. All 56 Valpha24JalphaQ+ clones isolated from the diabetic twins/triplets secreted only interferon (IFN)-gamma upon stimulation; in contrast, 76 of 79 clones from the at-risk non-progressors and normals secreted both interleukin (IL)-4 and IFN-gamma. Half of the at-risk non-progressors had high serum levels of IL-4 and IFN-gamma. These results support a model for IDDM in which Thl-cell-mediated tissue damage is initially regulated by Valpha24JalphaQ+ T cells producing both cytokines; the loss of their capacity to secrete IL-4 is correlated with IDDM.

Multiple sclerosis (MS) is postulated to be a Th1-type cell-mediated autoimmune disease. Thus therapies that decrease T cell interferon (IFN)-gamma production or increase interleukin (IL)-4 production would be expected to have an ameliorating effect on MS. Some progressive MS patients receiving pulse cyclophosphamide therapy developed peripheral blood eosinophilia. We investigated whether cyclophosphamide-treated patients had immune deviation toward Th2 responses. We measured cytokine production in patients receiving either monthly intravenous methylprednisolone (MP), intravenous cyclophosphamide plus methylprednisolone (CY/MP), methotrexate, IFN-beta1b, in untreated MS patients, and in healthy controls. Minimal IL-4 was secreted in untreated patients (129 +/- 62 pg/ml), methotrexate-treated patients (99 +/- 79 pg/ml), and healthy controls (50 +/- 13 pg/ml). A marked increase in IL-4 was observed in CY/MP patients (1,503 +/- 291 pg/ml). Patients treated with MP (418 +/- 160 pg/ml) or IFN-beta1b (425 +/- 167 pg/ml) showed small increases. Eosinophilia in CY/MP-treated patients (6.0 +/- 0.7%) correlated with increased IL-4. IL-10 production was also increased in CY/MP-treated patients. Both CY/MP- and MP-treated groups had decreased production of IFN-gamma compared with untreated MS. These findings demonstrate pronounced immune deviation favoring Th2-type responses after pulse cyclophosphamide therapy.

The frequency of clonally expanded and persistent T cells recognizing the immunodominant autoantigenic peptide of myelin basic protein (MBP)p85-99 was directly measured ex vivo in subjects with typical relapsing remitting multiple sclerosis (MS). T cells expressing mRNA transcripts encoding T cell receptor (TCR)-α and -β chains found in T cell clones previously isolated from these subjects recognizing the MBPp85-99 epitope were examined. In contrast to frequencies of 1 in 105–106 as measured by limiting dilution analysis, estimates of the T cell frequencies expressing MBPp85-99–associated TCR chain transcripts were as high as 1 in 300. These high frequencies were confirmed by performing PCR on single T cells isolated by flow cytometry. MBPp85-99 TCR transcripts were present in IL-2 receptor α–positive T cells which were induced to undergo Fas-mediated cell death upon antigen stimulation. These data demonstrate that at least a subpopulation of patients with MS can have a very high frequency of activated autoreactive T cells.

Human T-cell leukemia virus type I (HTLV-I)-infected T cells expanded in vitro by single-cell cloning provide a unique system for investigating virus-cell interactions in nonimmortalized T cells. By analysis of clones generated randomly from the blood of virus carriers, we confirm that CD4 T cells are the major reservoir of HTLV-I in vivo and show that most infected cells contain a single integrated provirus. Contrary to the situation in HTLV-I immortalized cell lines, the HTLV-I provirus was found to be transcriptionally silent in a high proportion of randomly generated T-cell clones and could not be reactivated by mitogenic stimulation. The spontaneous proliferation previously documented in HTLV-I–infected T-cell clones was not observed in silently infected cells, and therefore correlates directly with the expression of tax and other viral genes. The only cytokine mRNA found to be significantly elevated in the virus-producing clones was interleukin-6; however, receptor-blocking experiments argue against a role for IL-6 in the virus-induced cell proliferation. We observed a striking variation in the ability of individual HTLV-I–producing clones to immortalize fresh peripheral blood lymphocytes. This ability did not correlate with the levels of viral mRNA expression, gag p24 production, spontaneous proliferation, or tax-transactivation, possibly suggesting a role for host cell factors as determinants of viral infectivity or immortalization. Studies to elucidate the basis of this phenotypic heterogeneity should enhance our understanding of viral spread and pathogenesis.

Developed societies, although having successfully reduced the burden of infectious disease, constitute an environment where metabolic, cardiovascular, and autoimmune diseases thrive. Living in westernized countries has not fundamentally changed the genetic basis on which these diseases emerge, but has strong impact on lifestyle and pathogen exposure. In particular, nutritional patterns collectively termed the "Western diet", including high-fat and cholesterol, high-protein, high-sugar, and excess salt intake, as well as frequent consumption of processed and 'fast foods', promote obesity, metabolic syndrome, and cardiovascular disease. These factors have also gained high interest as possible promoters of autoimmune diseases. Underlying metabolic and immunologic mechanisms are currently being intensively explored. This review discusses the current knowledge relative to the association of "Western diet" with autoimmunity, and highlights the role of T cells as central players linking dietary influences to autoimmune pathology.

T-cell immunoglobulin and mucin domain 3 (TIM-3) is an Ig-superfamily member expressed on IFN-γ-secreting Th1 and Tc1 cells and was identified as a negative regulator of immune tolerance. TIM-3 is expressed by a subset of activated CD4(+) T cells, and anti-CD3/anti-CD28 stimulation increases both the level of expression and the number of TIM-3(+) T cells. In mice, TIM-3 is constitutively expressed on natural regulatory T (Treg) cells and has been identified as a regulatory molecule of alloimmunity through its ability to modulate CD4(+) T-cell differentiation. Here, we examined TIM-3 expression on human Treg cells to determine its role in T-cell suppression. In contrast to mice, TIM-3 is not expressed on Treg cells ex vivo but is upregulated after activation. While TIM-3(+) Treg cells with increased gene expression of LAG3, CTLA4, and FOXP3 are highly efficient suppressors of effector T (Teff) cells, TIM-3(-) Treg cells poorly suppressed Th17 cells as compared with their suppression of Th1 cells; this decreased suppression ability was associated with decreased STAT-3 expression and phosphorylation and reduced gene expression of IL10, EBI3, GZMB, PRF1, IL1Rα, and CCR6. Thus, our results suggest that TIM-3 expression on Treg cells identifies a population highly effective in inhibiting pathogenic Th1- and Th17-cell responses.

CD4(+) T helper cells are a central element of the adaptive immune system. They protect the organism against a wide range of pathogens and are able to initiate and control many immune reactions in combination with other cells of the adaptive and the innate immune system. Starting from a naive cell, CD4(+) T cells can differentiate into various effector cell populations with specialized function. This subset specific differentiation depends on numerous signals and the strength of stimulation. However, recent data have shown that differentiated CD4(+) T cell subpopulations display a high grade of plasticity and that their initial differentiation is not an endpoint of T cell development. In particular, FoxP3(+) regulatory T cells (Treg) and Th17 effector T cells demonstrate a high grade of plasticity, which allow a functional adaptation to various physiological situations during an immune response. However, the plasticity of Treg and Th17 cells might also be a critical factor for autoimmune disease. Here we discuss the recent developments in CD4(+) T cell plasticity with a focus on Treg and Th17 cells and its role in human autoimmune disease, in particular multiple sclerosis (MS).

The major histocompatibility complex (MHC) region is strongly associated with multiple sclerosis (MS) susceptibility. HLA-DRB1*15:01 has the strongest effect, and several other alleles have been reported at different levels of validation. Using SNP data from genome-wide studies, we imputed and tested classical alleles and amino acid polymorphisms in 8 classical human leukocyte antigen (HLA) genes in 5,091 cases and 9,595 controls. We identified 11 statistically independent effects overall: 6 HLA-DRB1 and one DPB1 alleles in class II, one HLA-A and two B alleles in class I, and one signal in a region spanning from MICB to LST1. This genomic segment does not contain any HLA class I or II genes and provides robust evidence for the involvement of a non-HLA risk allele within the MHC. Interestingly, this region contains the TNF gene, the cognate ligand of the well-validated TNFRSF1A MS susceptibility gene. The classical HLA effects can be explained to some extent by polymorphic amino acid positions in the peptide-binding grooves. This study dissects the independent effects in the MHC, a critical region for MS susceptibility that harbors multiple risk alleles., R01NS049477, R01NS026799, NIH/NINDS R01NS049510, R01NS0495103, NIH/NIAID R01AI076544

Analyses of somatic hypermutation (SHM) patterns in B cell immunoglobulin (Ig) sequences contribute to our basic understanding of adaptive immunity, and have broad applications not only for understanding the immune response to pathogens, but also to determining the role of SHM in autoimmunity and B cell cancers. Although stochastic, SHM displays intrinsic biases that can confound statistical analysis, especially when combined with the particular codon usage and base composition in Ig sequences. Analysis of B cell clonal expansion, diversification, and selection processes thus critically depends on an accurate background model for SHM micro-sequence targeting (i.e., hot/cold-spots) and nucleotide substitution. Existing models are based on small numbers of sequences/mutations, in part because they depend on data from non-coding regions or non-functional sequences to remove the confounding influences of selection. Here, we combine high-throughput Ig sequencing with new computational analysis methods to produce improved models of SHM targeting and substitution that are based only on synonymous mutations, and are thus independent of selection. The resulting "S5F" models are based on 806,860 Synonymous mutations in 5-mer motifs from 1,145,182 Functional sequences and account for dependencies on the adjacent four nucleotides (two bases upstream and downstream of the mutation). The estimated profiles can explain almost half of the variance in observed mutation patterns, and clearly show that both mutation targeting and substitution are significantly influenced by neighboring bases. While mutability and substitution profiles were highly conserved across individuals, the variability across motifs was found to be much larger than previously estimated. The model and method source code are made available at http://clip.med.yale.edu/SHM.

Little is known about how human genetic variation affects the responses to environmental stimuli in the context of complex diseases. Experimental and computational approaches were applied to determine the effects of genetic variation on the induction of pathogen-responsive genes in human dendritic cells. We identified 121 common genetic variants associated in cis with variation in expression responses to Escherichia coli lipopolysaccharide, influenza, or interferon-β (IFN-β). We localized and validated causal variants to binding sites of pathogen-activated STAT (signal transducer and activator of transcription) and IRF (IFN-regulatory factor) transcription factors. We also identified a common variant in IRF7 that is associated in trans with type I IFN induction in response to influenza infection. Our results reveal common alleles that explain interindividual variation in pathogen sensing and provide functional annotation for genetic variants that alter susceptibility to inflammatory diseases., National Human Genome Research Institute (U.S.) (Grant P50 HG006193), National Institutes of Health (U.S.). Pioneer Award (DP1 CA174427), Howard Hughes Medical Institute, National Institutes of Health (U.S.) (Grant HG004037), National Institutes of Health (U.S.). Pioneer Award (DP1 MH100706), National Institutes of Health (U.S.) (Transformative R01 Grant R01 DK097768), W. M. Keck Foundation, McKnight Foundation, Merkin, Richard N., Damon Runyon Cancer Research Foundation, Searle Scholars Program, Simons Foundation

Acute disseminated encephalomyelitis (ADEM) and relapsing remitting multiple sclerosis (MS) are distinct forms of autoimmune-mediated demyelination, yet they may share polysymptomatic neurological deficits, multifocal white matter lesions and laboratory findings. This can render initial prediction of monophasic outcome (typical of ADEM) versus prediction of relapsing-remitting MS, difficult and observation over time is essential to define outcome. An improved understanding of the biological factors that distinguish transient immune targeting of the CNS from chronic CNS-directed immunity has both diagnostic and potential therapeutic implications. Molecular biomarkers that can distinguish between MS and ADEM are therefore highly sought after.1–3 The etiology of MS is multi-factorial, with a role for both genetic and environmental factors. The natural history of MS involves episodic exacerbations overlying a chronic and progressive decline, though life-long immunomodulatory therapy can moderately attenuate the disease. The current diagnosis of MS is based on clinical and radiologic confirmation of relapsing inflammatory demyelination that occurs in multiple CNS regions separated in both space and time.4 In contrast, ADEM is a monophasic disorder in more than 90% of cases and affects primarily children. Many patients report recent upper respiratory illness, prompting consideration of an infectious trigger for the immune response. The criteria for diagnosing ADEM remain a subject of debate, though many clinicians apply the consensus criteria proposed by a 2007 international panel of experts who describe the disorder as a first episode of inflammatory demyelination with multi-focal CNS involvement and encephalopathy.5 In both ADEM and MS, autoantibodies to specific myelin proteins and lipids may contribute to disease pathogenesis6–9 and activate complement in some patients.10–12 We have developed antigen arrays for profiling autoantibodies in a variety of autoimmune diseases, including demyelinating disorders.13–15 Preliminary studies suggest that the two diseases have distinct autoantibody profiles,2, 16 including differences in immunoglobulin (Ig) subtype.1, 8, 9, 12, 17–19 Based on the available literature, we hypothesized that IgG and IgM autoantibodies in ADEM and MS patients might offer disease-specific profiles. In this study, we use antigen arrays to identify serum anti-myelin autoantibody profiles that can distinguish between ADEM and MS.

A recent epidemiological study showed that eating ‘fast food’ items such as potato chips increased likelihood of obesity, whereas eating yogurt prevented age-associated weight gain in humans. It was demonstrated previously in animal models of obesity that the immune system plays a critical role in this process. Here we examined human subjects and mouse models consuming Westernized ‘fast food’ diet, and found CD4[superscript +] T helper (Th)17-biased immunity and changes in microbial communities and abdominal fat with obesity after eating the Western chow. In striking contrast, eating probiotic yogurt together with Western chow inhibited age-associated weight gain. We went on to test whether a bacteria found in yogurt may serve to lessen fat pathology by using purified Lactobacillus reuteri ATCC 6475 in drinking water. Surprisingly, we discovered that oral L. reuteri therapy alone was sufficient to change the pro-inflammatory immune cell profile and prevent abdominal fat pathology and age-associated weight gain in mice regardless of their baseline diet. These beneficial microbe effects were transferable into naïve recipient animals by purified CD4[superscript +] T cells alone. Specifically, bacterial effects depended upon active immune tolerance by induction of Foxp3[superscript +] regulatory T cells (Treg) and interleukin (Il)-10, without significantly changing the gut microbial ecology or reducing ad libitum caloric intake. Our finding that microbial targeting restored CD4[superscript +] T cell balance and yielded significantly leaner animals regardless of their dietary ‘fast food’ indiscretions suggests population-based approaches for weight management and enhancing public health in industrialized societies., National Institutes of Health (U.S.) (Grant P30-ES002109), National Institutes of Health (U.S.) (Grant RO1CA108854), National Institutes of Health (U.S.) (Grant P01 AI045757), National Institutes of Health (U.S.) (Grant U19 AI046130), National Institutes of Health (U.S.) (Grant U19 AI070352), National Institutes of Health (U.S.) (Grant P01 AI039671), National Institute of Neurological Disorders and Stroke (U.S.) (Jacob Javits Merit Award NS2427), The Penates Foundation, Nancy Taylor Foundation for Chronic Diseases, Inc.

Multiple sclerosis (MS) is an inflammatory CNS disease with a substantial genetic component, originally mapped to only the human leukocyte antigen (HLA) region. In the last 5 years, a total of seven genome-wide association studies and one meta-analysis successfully identified 57 non-HLA susceptibility loci. Here, we merged nominal statistical evidence of association and physical evidence of interaction to conduct a protein-interaction-network-based pathway analysis (PINBPA) on two large genetic MS studies comprising a total of 15,317 cases and 29,529 controls. The distribution of nominally significant loci at the gene level matched the patterns of extended linkage disequilibrium in regions of interest. We found that products of genome-wide significantly associated genes are more likely to interact physically and belong to the same or related pathways. We next searched for subnetworks (modules) of genes (and their encoded proteins) enriched with nominally associated loci within each study and identified those modules in common between the two studies. We demonstrate that these modules are more likely to contain genes with bona fide susceptibility variants and, in addition, identify several high-confidence candidates (including BCL10, CD48, REL, TRAF3, and TEC). PINBPA is a powerful approach to gaining further insights into the biology of associated genes and to prioritizing candidates for subsequent genetic studies of complex traits.

Multiple sclerosis (MS) is a genetically mediated autoimmune disease of the central nervous system. B cells have recently emerged as major contributors to disease pathogenesis, but the mechanisms responsible for the loss of B cell tolerance in patients with MS are largely unknown. In healthy individuals, developing autoreactive B cells are removed from the repertoire at 2 tolerance checkpoints during early B cell development. Both of these central and peripheral B cell tolerance checkpoints are defective in patients with rheumatoid arthritis (RA) and type 1 diabetes (T1D). Here, we found that only the peripheral, but not the central, B cell tolerance checkpoint is defective in patients with MS. We show that this specific defect is accompanied by increased activation and homeostatic proliferation of mature naive B cells. Interestingly, all of these MS features parallel defects observed in FOXP3-deficient IPEX patients, who harbor nonfunctional Tregs. We demonstrate that in contrast to patients with RA or T1D, bone marrow central B cell selection in MS appears normal in most patients. In contrast, patients with MS suffer from a specific peripheral B cell tolerance defect that is potentially attributable to impaired Treg function and that leads to the accumulation of autoreactive B cell clones in their blood.

Multiple sclerosis (MS) is a multifocal demyelinating disease of the central nervous system (CNS), caused by an autoimmune response to self-antigens in a genetically susceptible individual. It is characterized by progressive neurodegeneration and by CNS lesions containing a high number of infiltrating autoreactive B and T cells. In healthy individuals, regulatory T (Treg) cells can control potentially pathogenic autoreactive T cells, while Treg cells in MS patients show insufficient regulatory abilities. The fact that autoreactive T cells can be found in the peripheral blood of healthy individuals without causing any autoimmune diseases underlies the importance of an efficient control of immune responses. Treg cells are key regulators of immune homeostasis and self-tolerance.

Objective: To profile the reactivity of CSF-derived immunoglobulin from patients with multiple sclerosis (MS) against a large panel of antigens, to identify disease-specific reactivities. Methods: CSF from subjects with MS with elevated immunoglobulin G and CSF from control subjects presenting with other inflammatory neurologic disease were screened against a protein array consisting of 9,393 proteins. Reactivity to a candidate protein identified using these arrays was confirmed with ELISA and immunocytochemistry. Results: Autoantibodies against one protein on the array, recombination signal binding protein for immunoglobulin kappa J region (RBPJ), discriminated between patients with MS and controls (p = 0.0052). Using a large validation cohort, we found a higher prevalence of autoantibodies against RBPJ in the CSF of patients with MS (12.5%) compared with the CSF of patients with other neurologic diseases (1.6%; p = 0.02) by ELISA. This difference in reactivity was restricted to the CSF as serum reactivity against RBPJ did not differ between patients and controls. The presence of CSF autoantibodies against RBPJ was further confirmed by immunocytochemistry. Conclusions: These data indicate that RBPJ, a ubiquitous protein of the Notch signaling pathway that plays an important role in Epstein-Barr virus infection, is a novel MS autoantigen candidate that is recognized by CSF-derived immunoglobulin G in a subset of patients with MS.

T cells recognize antigen by formation of a trimolecular complex in which the T-cell receptor (TCR) recognizes a specific peptide antigen within the groove of a major histocompatibility complex (MHC) molecule. It has generally been assumed that T-cell recognition of two distinct MHC–antigen complexes is due to similarities in the three-dimensional structure of the complexes. Here we report results of experiments examining the crossreactivity of TCRs recognizing the myelin basic protein peptide MBPp85–99 and several of its analogs in the context of MHC. We demonstrate that single conservative amino acid substitutions of the antigenic peptide at the predominant TCR contact residues at positions 91 and 93 totally abrogate reactivity of specific T-cell clones. Yet, when a conservative substitution is made at position 91 concomitant with a substitution at position 93, the T-cell clones regain reactivity equivalent with that of the original stimulating peptide. Thus, the exact nature of the amino acid side chains engaging one TCR functional pocket may change the apparent selectivity of the other predominant TCR functional pocket, thus suggesting a remarkable degree of receptor plasticity. This ability of the TCR–MHC–peptide complex to undergo conformational changes provides a conceptual framework for reconciling the apparent paradox of the extreme selectivity of the TCR and its remarkable crossreactivity with different MHC–peptide complexes.

Multiple sclerosis is a presumed autoimmune disease, associated with inflammation in the CNS white matter, mediated by autoreactive T cells. We previously reported that oral myelin tolerization of relapsing-remitting MS patients resulted in fewer attacks, as compared to a placebo-fed group. Here, we examined whether oral tolerization with bovine myelin resulted in altered autoreactive T-cell populations or altered T-cell fraction. We generated 4,620 T-cell lines from 34 relapsing-remitting MS patients (17 were fed bovine myelin daily), and each line was examined for proliferation to MBP, PLP, and TT and for secretion of IL-4, IFN-gamma, and TGF-beta1. The frequency of TGF-beta1-secreting T-cell lines after MBP and PLP stimulation in fed patients was greater than that of nonfed patients. These experiments demonstrate that oral tolerization with autoantigen results in altered cytokine secretion in a human autoimmune disease with the generation of TGF-beta1-secreting T cells that may regulate the inflammatory response at the site of the demyelinating lesions in multiple sclerosis. These data provide the first evidence of antigen-specific modification of cytokine secretion in a human autoimmune disease.

Understanding the immune response to myelin antigens in regard to the peptide/MHC/TCR complex is important in defining pathogenesis of demyelinating autoimmune diseases and in developing antigen-specific therapies. We previously reported that individual multiple sclerosis patients may use certain dominant TCR V beta chains to recognize immunodominant MBP peptides. In examining the TCR beta chain usage, we observed repeated TCR VDJ sequences among different T-cell lines isolated from the same patient. This suggested that a few expanded T-cell clones may dominate the immune response to immunodominant MBP peptides. Here, we report experiments where TCR rearrangements were used as a probe for the clonal origin of MBP specific T-cells cultured from blood lymphocytes of MS patients and normal subjects. In two patients with the DR2 haplotype that were analyzed in detail, the T-cell response to MBP was focused on the MBP (84-102) peptide and in vivo expanded population(s) dominated the response to the MBP (84-102) peptide. Two MBP (84-102) specific T-cell clones from a normal subject with the DR2 haplotype were also found to have identical TCR sequences. Clonality was proven by demonstrating that independent clones had identical TCR alpha and beta chain sequences as well as identical sequences of a TCR gamma chain or of a second TCR alpha chain rearrangement. These data suggest that the response to human MBP is dominated in at least some subjects by expanded clones that may persist in vivo for relatively long periods of time.

The autologous mixed lymphocyte reaction (AMLR) involves the activation of T cells by autologous antigen presenting cells. Cells are generated during the course of the AMLR that have suppressive properties in vitro. In the present study we investigated the induction of CD8+ T cells in the AMLR with suppressive properties and the mechanism by which these cells downregulate in vitro proliferative responses. Purified CD8+ but not CD4+ T cells activated in the AMLR in conditioned medium inhibited proliferation of autologous T cells by anti-CD3 or PPD. Nonactivated CD8+ T cells did not suppress. The CD8+ T cells activated in the AMLR in the presence of conditioned medium (CD8+ Tact) were CD11b negative and were noncytotoxic. The inhibitory effect of CD8+ Tact cells was completely abrogated by anti-IFN-gamma antibody, but not by anti-IL-4, anti-IL-10, or anti-TGF-beta antibody. The induction of CD8+ Tact cells in the AMLR was blocked by anti-IL-2 or by anti-GM-CSF antibody and the combination of these two recombinant cytokines could support the induction of suppressive CD8+ Tact cells. CD8+ Tact cells were defective in patients with chronic progressive multiple sclerosis (MS) as compared to patients with relapsing-remitting MS or normal controls. Our studies provide a basis for understanding the mechanism of suppression by human CD8+ T cells in terms of specific cytokines, and demonstrate the potential importance of these cells in a human autoimmune disease as their function is defective in patients with progressive MS.

Tropical spastic paraparesis (TSP) is a retroviral disease characterized predominantly by a chronic myelopathy and progressive leg weakness. Four patients from the northern coast of Colombia with chronic spastic paraparesis and serum positivity for antibodies to human T cell lymphotropic virus type 1 by enzyme-linked immunosorbent assay and Western blot are reported. All patients had mixed ethnic origins (white, black, and amerindian). This is the first report of TSP on the Caribbean coast of Colombia. This study extends the geographic boundaries of TSP in the Caribbean basin.