Characterization of the antigen specificity and TCR repertoire,and TCR-based DNA vaccine therapy in myelin basic protein-induced autoimmune encephalomyelitis in DA rats

Like Lewis rats, DA rats are an experimental autoimmune encephalomyelitis (EAE)-susceptible strain and develop severe EAE upon immunization with myelin basic protein (MBP). However, there are several differences between the two strains. In the present study we induced acute EAE in DA rats by immunization with MBP and MBP peptides and examined the Ag specificity and TCR repertoire of encephalitogenic T cells. It was found that although immunization with MBP and a peptide corresponding to its 62-75 sequence (MBP(62-75)) induced clinical EAE, the responses of lymph node T cells isolated from MBP-immunized rats to MBP(62-75) was marginal, indicating that this peptide contains major encephalitogenic, but not immunodominant, epitopes. The TCR analysis by CDR3 spectratyping of spinal cord T cells revealed that Vbeta10 and Vbeta15 spectratype expansion was always found in MBP(62-75)-immunized symptomatic rats. On the basis of these findings, we examined the encephalitogenicity of Vbeta10- and Vbeta15-positive T cells. First, the adoptive transfer experiments revealed that Vbeta10-positive T line cells derived from MBP(62-75)-immunized rats induced clinical EAE in recipients. Second, administration of DNA vaccines encoding Vbeta10 and Vbeta15, alone or in combination, ameliorated MBP(62-75)-induced EAE. Collectively, it was strongly suggested that Vbeta10- and Vbeta15-positive T cells are encephalitogenic. Analyses of the Ag specificity and T cell repertoire of pathogenic T cells performed in this study provide useful information for designing specific immunotherapies against autoimmune diseases.     

TGF-beta 1 regulates antigen-specific CD4+ T cell responses in the periphery

T cell expansion typically is due to cognate interactions with specific Ag, although T cells can be experimentally activated through bystander mechanisms not involving specific Ag. TGF-beta1 knockout mice exhibit a striking expansion of CD4+ T cells in the liver by 11 days of age, accompanied by CD4+T cell-dependent necroinflammatory liver disease. To examine whether hepatic CD4+T cell expansion in TGF-beta1(-/-) mice is due to cognate TCR-peptide interactions, we used spectratype analysis to examine the diversity in TCR Vbeta repertoires in peripheral CD4+T cells. We reasoned that Ag-nonspecific T cell responses would yield spectratype profiles similar to those derived from control polyclonal T cell populations, whereas Ag-specific T cell responses would yield perturbed spectratype profiles. Spleen and liver CD4+T cells from 11-day-old TGF-beta1(-/-) mice characteristically exhibited highly perturbed nonpolyclonal distributions of TCR Vbeta CDR3 lengths, indicative of Ag-driven T cell responses. We quantitatively assessed spectratype perturbation to derive a spectratype complexity score. Spectratype complexity scores were considerably higher for TGF-beta1(-/-) CD4+ T cells than for TGF-beta1(+/-) CD4+T cells. TCR repertoire perturbations were apparent as early as postnatal day 3 and preceded both hepatic T cell expansion and liver damage. By contrast, TGF-beta1(-/-) CD4+ single-positive thymocytes from 11-day-old mice exhibited normal unbiased spectratype profiles. These results indicate that CD4+ T cells in TGF-beta1(-/-) mice are activated by and respond to self-Ags present in the periphery, and define a key role for TGF-beta1 in the peripheral regulation of Ag-specific CD4+ T cell responses.     

Administration of PLP139-151 primes T cells distinct from those spontaneously responsive in vitro to this antigen

We examined the TCR repertoire used by naive SJL mice in their in vitro spontaneous response to proteolipid protein (PLP) 139-151 by Vbeta-Jbeta spectratyping and compared it to that used after immunization with the peptide. T cells from immunized mice use the public rearrangement Vbeta10-Jbeta1.1, but naive mice do not; in contrast, TCR CDR3-beta rearrangements of Vbeta18-Jbeta1.2 and Vbeta19-Jbeta1.2 consistently are associated with the spontaneous response. T cells involved in spontaneous and induced responses can each recognize PLP(139-151) presented in vivo, but its s.c. administration has different consequences for the two repertoires. Four days after immunization, T cells associated with spontaneous responsiveness appear in the draining lymph nodes but disappear by day 10 and never appear elsewhere. Simultaneously, Vbeta10-Jbeta1.1 T cells are likewise activated in the lymph nodes by day 4 and spread to the spleen by day 10. Eight- to 10-wk-old naive mice use a narrower repertoire of TCRs than do immunized age-matched mice. Induced Vbeta10-Jbeta1.1 T cells home to the CNS during experimental autoimmune encephalomyelitis, whereas we failed to detect Vbeta18-Jbeta1.2 and Vbeta19-Jbeta1.2 TCR rearrangements in the CNS. Thus, we observe that administration of PLP(139-151) primes a T cell repertoire distinct from the one responsible for spontaneous responsiveness. This "immunized" repertoire substitutes for the naive one and becomes dominant at the time of disease onset.     

Successful TCR-based immunotherapy for autoimmune myocarditis with DNA vaccines after rapid identification of pathogenic TCR

The identification of TCRs of autoimmune disease-inducing T cells within a short period of time is a key factor for designing TCR-based immunotherapy during the course of the disease. In this study, we show that experimental autoimmune carditis-associated TCRs, Vbeta8.2 and Vbeta10, were determined by complementarity-determining region 3 (CDR3)-spectratyping analysis and subsequent sequencing of the CDR3 region of spectratype-derived TCR clones. Immunotherapy targeting both Vbeta8.2 and Vbeta10 TCRs using mAbs and DNA vaccines significantly reduced the histological severity of experimental autoimmune carditis and completely suppressed the inflammation in some animals. Since depletion or suppression of one of two types of effector cells does not improve the severity of the disease significantly, combined TCR-based immunotherapy should be considered as a primary therapy for T cell-mediated autoimmune diseases. TCR-based immunotherapy after rapid identification of autoimmune disease-associated TCRs by CDR3 spectratyping can be applicable, not only to animal, but also to human autoimmune diseases whose pathomechanism is poorly understood.     

Complementarity-determining region 3 spectratyping analysis of the TCR repertoire in multiple sclerosis

Multiple sclerosis (MS) is considered to be an autoimmune disease mediated by T cells reactive with Ags in the CNS. Therefore, it has been postulated that neuroantigen-reactive T cells bearing particular types of TCRs are expanded clonally during the course of the disease. However, there is a controversy with regard to the TCR usage by T cells associated with the development of MS. By the use of complementarity-determining region 3 spectratyping analysis that is shown to be a useful tool for identification of pathogenic TCR in autoimmune disease models, we tried to demonstrate that spectratype was T cells bearing particular types of TCR are activated in MS patients. Consequently, it was found that Vbeta5.2 were often oligoclonally expanded in peripheral blood of MS patients, but not of healthy subjects. Sequence analysis of the complementarity-determining region 3 region of spectratype-derived TCR clones revealed that the predominant TCR clone was different from patient to patient, but that similar results were obtained in a patient examined at different time points. More importantly, examination of cerebrospinal fluid T cells and longitudinal studies of PBLs from selected patients revealed that Vbeta5.2 expansion was detectable in the majority of patients examined. These findings suggest that Vbeta5.2 spectratype expansion is associated with the development of MS and that TCR-based immunotherapy can be applicable to MS patients if the TCR activation pattern of each patient is determined at different stages of the disease.     

Public T cell receptor beta-chains are not advantaged during positive selection

Studies of human and murine T cells have shown that public TCR beta-chain rearrangements can dominate the Ag-specific and naive repertoires of distinct individuals. We show that mouse T cells responding to the minor histocompatibility Ag HYDbSmcy share an invariant Vbeta8.2-Jbeta2.3 TCR gene rearrangement. The dominance of this rearrangement shows that it successfully negotiated thymic selection and was highly favored during clonal expansion in all animals examined. We hypothesized that such beta-chains are advantaged during thymic and/or peripheral selection and, as a result, may be over-represented in the naive repertoire. A sequencing study was undertaken to examine the diversity of Vbeta8.2-Jbeta2.3 CDR3 loops from naive T cell repertoires of multiple mice. Public TCR beta-chain sequences were identified across different repertoires and MHC haplotypes. To determine whether such public beta-chains are advantaged during thymic selection, individual chains were followed through T cell development in a series of novel bone marrow competition chimeras. We demonstrate that beta-chains were positively selected with similar efficiency regardless of CDR3 loop sequence. Therefore, the establishment and maintenance of public beta-chains in the periphery is predominantly controlled by post-thymic events through modification of the primary, thymus-derived TCR repertoire.     

Identification of encephalitogenic V beta-4-bearing T cells in SJL mice. Further evidence for the V region disease hypothesis?

Experimental allergic encephalomyelitis (EAE) is an autoimmune disease of the central nervous system mediated by T cells bearing TCR of restricted heterogeneity. Thus, in the murine PL strain, V beta-8.2 is used by 80% of the encephalitogenic T cells. This observation has led to the successful prevention and reversal of EAE by the in vivo use of mAb directed to these restricted gene products. In SJL mice, the V beta-17a gene product has been shown to be used by approximately 50% of encephalitogenic T cells subsequent to immunization with a myelin basic protein (MBP)-derived peptide. However, the other V beta genes used by encephalitogenic T cells in SJL EAE have remained uncharacterized. We now report, for the first time, the beta-chain-encoding DNA sequence of two encephalitogenic, MBP-reactive, SJL-derived T cell clones. These clones which are specific for H-2s and the carboxyl-terminus (amino acid 92-103) of MBP, use TCR encoded by V beta-4. In addition, we demonstrate that the transfer of EAE by a heterogenous SJL-derived encephalitogenic T cell line can be prevented using an anti-V beta-4 antibody in vivo. V beta-4 usage has been previously described in a H-2u/MBP amino-terminus-reactive encephalitogenic T cell. The present findings may thus further support the "V region-disease" hypothesis.     

CDR3 spectratyping analysis of the TCR repertoire in myasthenia gravis

Because myasthenia gravis (MG) is an autoimmune disease mediated by Abs specific for the acetylcholine receptor, helper T cells play a role in Ab production. In this study, we have performed large-scale cross-sectional and longitudinal TCR studies by CDR3 spectratyping using PBL and thymus tissues from MG patients. We found that there was no preferential usage of any particular TCR beta-chains that was identical among MG patients. However, the longitudinal study clearly demonstrated that one or more TCR Vbeta expansions persisted frequently in MG patients. Importantly, persistent TCR expansions correlated with clinical severity and high anti-acetylcholine receptor Ab titer. Finally, examinations of T cells expressing CXCR5, i.e., follicular B-helper T cells, revealed that spectratype expansions in MG patients were detected mainly in the CD4+ CXCR5+ T cell populations, whereas CD8+ T cells were the major source of clonal expansion in healthy subjects. These findings suggest that persistent clonal expansions of T cells in MG patients are associated with the development and maintenance of MG. Close examination of pathogenic T cells in MG provides useful information to elucidate the pathogenesis and to estimate the disease status.     

Analysis of T-cell receptor Vbeta gene from infiltrating T cells in insulitis and myocarditis in encephalomyocarditis virus-infected BALB/C mice

Encephalomyocarditis (EMC) virus induces insulin-dependent diabetes and myocarditis in several strains of mice. The T-cell receptor (TCR) Vbeta genes of infiltrating T cells in the pancreas and myocardium of BALB/C mice infected with EMC virus D-variant (EMC-D virus) were analyzed. Using a nested two-step polymerase chain reaction (PCR), TCR Vbeta cDNAs were cloned and sequenced. Two and four kinds of TCR Vbeta clones were obtained from T cells infiltrating into the pancreas and myocardium of BALB/C mice infected with EMC-D virus, respectively. The infiltrating lymphocytes in the diabetic mice expressed Vbeta 8.1, 8.2, and 8.3 genes predominantly. Previously, the use of Vbeta 8.2 has been reported in autoimmune diseases such as murine experimental allergic encephalomyelitis (EAE) and non-obese diabetic (NOD) mouse. This study suggests that mice infected with EMC virus are a useful animal model for autoimmune diseases such as insulin-dependent diabetes.     

Pertussis toxin prevents the induction of peripheral T cell anergy and enhances the T cell response to an encephalitogenic peptide of myelin basic protein.

In a murine model of T cell-mediated autoimmune disease, experimental autoimmune encephalitis (EAE), 80% of all encephalitogenic T cell clones in H-2u mice use the V beta 8.2 TCR element. To induce EAE in susceptible strains of mice either heat-killed Bordetella pertussis organisms or Bordetella pertussis toxin (PT) must be injected in addition to Ag in CFA. We investigated the mechanisms by which PT facilitates the induction of EAE. Our data show, that PT interferes with the induction of Ag-induced peripheral T cell anergy. Furthermore it has a specific adjuvanticity for the autoantigen pAc1-11 in vivo and acts as a selective mitogen in vitro. We also tested the hypothesis that PT is a bacterial superantigen that specifically expands the V beta 8.2+ subset of T cells, thereby expanding the encephalitogenic T cell clones that are contained in this subset, so that the number of autoreactive T cells is brought over a critical threshold, necessary to induce autoimmune disease. Our data show that PT is not a superantigen. Staphylococcal enterotoxin B, a V beta 8.2-specific superantigen, does not enhance the immune response to the encephalitogenic peptide.     

Gamma delta T cell regulation of IFN-gamma production by central nervous system-infiltrating encephalitogenic T cells: correlation with recovery from experimental autoimmune encephalomyelitis

Interferon-gamma has been shown to be important for the resolution of inflammation associated with CNS autoimmunity. Because one of the roles of gamma delta T cells is the regulation of inflammation, we asked whether gamma delta T cells were able to regulate CNS inflammation using the autoimmune disease mouse model experimental autoimmune encephalomyelitis (EAE). We show that the presence of gamma delta T cells was needed to promote the production of IFN-gamma by both CD4 and CD8 T cells in the CNS before the onset of EAE. This regulation was shown to be independent of the ability of gamma delta T cells to produce IFN-gamma, and was specific to T cells in the CNS, as no alterations in IFN-gamma production were detectable in gamma delta T cell-deficient mice in the spleen and lymph nodes of mice with EAE or following immunization. Analysis of TCR gamma delta gene usage in the CNS showed that the only TCR delta V gene families present in the CNS before EAE onset are from the DV7s6 and DV105s1 gene families. We also show that the primary IFN-gamma-producing cells in the CNS are the encephalitogenic T cells, and that gamma delta T cell-deficient mice are unable to resolve EAE disease symptoms like control mice, thus exhibiting a long-term chronic disease course similar to that observed in IFN-gamma-deficient mice. These data suggest that CNS resident gamma delta T cells promote the production of IFN-gamma by encephalitogenic T cells in the CNS, which is ultimately required for the recovery from EAE.     

Role of the complementarity-determining region 3 (CDR3) of the TCR-beta chains associated with the V alpha 14 semi-invariant TCR alpha-chain in the selection of CD4+ NK T Cells

The NK1.1(+)TCRalphabeta(int) CD4(+), or double negative T cells (NK T cells) consist of a mixture of CD1d-restricted and CD1d-unrestricted cells. The relationships between CD4(+)NK1.1(+) T cells and conventional T cells are not understood. To compare their respective TCR repertoires, NK1.1(+)TCRalphabeta(int), CD4(+) T cells have been sorted out of the thymus, liver, spleen, and bone marrow of C57BL/6 mice. Molecular analysis showed that thymus and liver used predominantly the Valpha14-Jalpha281 and Vbeta 2, 7, and 8 segments. These cells are CD1d restricted and obey the original definition of NK T cells. The complementarity-determining region 3 (CDR3) sequences of the TCR Vbeta8.2-Jbeta2.5 chain of liver and thymus CD4(+) NK T cells were determined and compared with those of the same rearrangements of conventional CD4(+) T cells. No amino acid sequence or usage characteristic of NK T cells could be evidenced: the Vbeta8.2-Jbeta2.5 diversity regions being primarily the same in NK T and in T cells. No clonal expansion of the beta-chains was observed in thymus and liver CD1d-restricted CD4(+)NK T cells, suggesting the absence of acute or chronic Ag-driven stimulation. Molecular analysis of the TCR used by Valpha14-Jalpha281 transgenic mice on a Calpha(-/-) background showed that the alpha-chain can associate with beta-chains using any Vbeta segment, except in NK T cells in which it paired predominately with Vbeta 2, 7, and 8(+) beta-chains. The structure of the TCR of NK T cells thus reflects the affinity for the CD1d molecule rather than a structural constraint leading to the association of the invariant alpha-chain with a distinctive subset of Vbeta segment.     

neonatally primed lymph node, but not splenic T cells, display a Gly-Gly motif within the TCR beta-chain complementarity-determining region 3 that controls affinity and may affect lymphoid organ retention

Ig-proteolipid protein 1 (Ig-PLP1) is an Ig chimera expressing the encephalitogenic PLP1 peptide corresponding to amino acid residues 139-151 of PLP. Newborn mice given Ig-PLP1 in saline on the day of birth and challenged 7 wk later with PLP1 peptide in CFA develop an organ-specific neonatal immunity that confers resistance against experimental allergic encephalomyelitis. The T cell responses in these animals are comprised of Th2 cells in the lymph node and anergic Th1 lymphocytes in the spleen. Intriguingly, the anergic splenic T cells, although nonproliferative and unable to produce IFN-gamma or IL-4, secrete significant amounts of IL-2. Studies were performed to determine whether the two populations display any structural differences in the TCR H chain variable region that could contribute to the differential affinity and retention in different organs. Responsive Th2 lymph node T cells and anergic splenic lymphocytes were immortalized, and the structures of their TCR Vbeta were determined. The results show that Vbeta and Jbeta usage was random, but the CDR3 regions of the lymph node cells had a conserved Gly-Gly motif. Analysis of TCR affinity/avidity correlated the Gly-Gly motif with lower affinity and retention of the Th2 cells in the lymph node. Also, it is suggested that a higher TCR affinity may be a contributing factor for the development of the neonatal Th1 response in the spleen.     

Characterization of the immune response to a secondary encephalitogenic epitope of basic protein in Lewis rats. I. T cell receptor peptide regulation of T cell clones expressing cross-reactive V beta genes.

In Lewis rats, immunization with myelin basic protein induces two distinct encephalitogenic T cell populations, those responding to the immunodominant 72-89 epitope and those specific for a secondary epitope including residues 87-99. The 72-89 specific T cells were I-A restricted and preferentially expressed V beta 8.2 in their TCR. To determine the fine specificity, MHC restriction, and TCR V beta gene use in T cells reactive to the secondary epitope, we characterized 23 T cell clones from the lymph nodes (LN) and spinal cords (SC) of rats immunized with either whole basic protein or synthetic peptides S85-99 and S87-99 that were found to be functionally similar. The S85-99/S87-99 specific clones from LN and SC were all encephalitogenic despite differences in recognition of intact basic protein and class II MHC restriction. Unlike LN clones that overexpressed V beta 8 (46%+) and V beta 6 (31%+), however, SC clones were strongly biased (86%+) in their expression of V beta 6. This V gene bias raised the possibility of TCR peptide therapy using V beta 6 peptides. The V beta 6 sequence was similar to V beta 8.2 in the CDR2 region, and the corresponding peptides from this region were found to be cross-reactive in vivo. Moreover, both peptides were effective in the treatment of EAE induced with either S85-99, biased in V beta 6+ and V beta 8+ T cells, or guinea pig basic protein, biased only in V beta 8+ T cells. These data demonstrate the presence of common immunogenic epitopes among subsets of TCR V region gene families that possess important regulatory activity on effector T cell function.     

Oligoclonality of CD8+ T cells in breast cancer patients.

Substantial evidence has suggested that T cells play an important role in antitumor immunity. T cells with cytotoxic activity against tumors have been isolated from in vitro culture of tumor-infiltrated lymphocytes of cancer patients. In addition, clonal expansions of T cells have been identified in lesions of tumors by using a PCR-based CDR3 analysis of T cell receptors (TCR). Since the CDR3 region of the T cell receptor directly interacts with the antigen-MHC complex and is thus highly polymorphic, a dominant CDR3 length in a particular TCR V beta population will indicate the clonal expansion of a specific T cell clone. Utilizing this technique, we have analyzed the T cell repertoire in lymph nodes (LNs) and peripheral blood of 20 breast cancer patients. Our results show that in most cases, peripheral blood mononuclear cells (PB-MCs) and LN express dominant CD8+ T cell clones in different V beta gene families, and the number of dominant clones is higher in PBMC than in the LN. Furthermore, in 7 out of 16 patients' lymph nodes, there is a dominant V beta 18 T cell clonal expansion in the CD8+ T cell subset. The frequency of an oligoclonal expansion of V beta 18 CD8+ T cells in non-breast cancer lymph nodes is 1 out of 9, but no obvious motif in the CDR3 region of V beta 18 TCR can be identified. The prevalence of the clonal dominance found in breast cancer is discussed in the context of a possible tumor-related antigen stimulation.     

Anti-TCR antibody treatment activates a novel population of nonintestinal CD8 alpha alpha+ TCR alpha beta+ regulatory T cells and prevents experimental autoimmune encephalomyelitis

CD8alphaalpha+CD4-TCRalphabeta+ T cells are a special lineage of T cells found predominantly within the intestine as intraepithelial lymphocytes and have been shown to be involved in the maintenance of immune homeostasis. Although these cells are independent of classical MHC class I (class Ia) molecules, their origin and function in peripheral lymphoid tissues are unknown. We have recently identified a novel subset of nonintestinal CD8alphaalpha+CD4-TCRalphabeta+ regulatory T cells (CD8alphaalpha Tregs) that recognize a TCR peptide from the conserved CDR2 region of the TCR Vbeta8.2-chain in the context of a class Ib molecule, Qa-1a, and control- activated Vbeta8.2+ T cells mediating experimental autoimmune encephalomyelitis. Using flow cytometry, spectratyping, and real-time PCR analysis of T cell clones and short-term lines, we have determined the TCR repertoire of the CD8alphaalpha regulatory T cells (Tregs) and found that they predominantly use the TCR Vbeta6 gene segment. In vivo injection of anti-TCR Vbeta6 mAb results in activation of the CD8alphaalpha Tregs, inhibition of the Th1-like pathogenic response to the immunizing Ag, and protection from experimental autoimmune encephalomyelitis. These data suggest that activation of the CD8alphaalpha Tregs present in peripheral lymphoid organs other than the gut can be exploited for the control of T cell-mediated autoimmune diseases.     

Synthetic peptides representing sequence 39 to 59 of rat V beta 8 TCR fail to elicit regulatory T cells reactive with V beta 8 TCR on rat encephalitogenic T cells.

Subpathogenic doses of syngeneic autoreactive T cells protect experimental animals against associated autoimmune disease. Preferential use of the TCR of encephalitogenic T cells suggests that this molecule serves as the target for immunoregulation in experimental autoimmune encephalomyelitis (EAE). Whether peptides derived from the V beta 8 of the rat TCR elicit regulatory T cells and produce the same vaccinating effect against EAE as do whole T cells remains unknown. Here we show that immunization of Lewis rats with V beta 8(39-59), a peptide representing residues 39 to 59 of the rat V beta 8 TCR, does not induce the production of regulatory T cells reactive to the intact TCR V beta 8 containing this sequence. Moreover, animals that had recovered from both actively induced EAE and transferred EAE did not generate regulatory T cells that recognized the V beta 8(39-59) peptide. Further, transfusion of large doses of peptide-specific T cells did not protect the animals from EAE. Our results suggest that the V beta 8(39-59) peptide may comprise so-called cryptic epitopes, which function as immunogens only when dissociated from large protein complexes.     

Nonencephalitogenic CD4-CD8- V alpha 2V beta 8.2+ anti-myelin basic protein rat T lymphocytes inhibit disease induction.

Recently there has been a number of reports suggesting that CD4-CD8- T cells participate in the processes of inflammatory reaction. In an attempt to delineate the distinctive functions of double negative (DN) T lymphocytes in an autoimmune-induced disease, we isolated and cloned such T cells, along with control CD4+ cells, from Lewis rats immunized with guinea-pig myelin basic protein in CFA. Both clones proliferated in response to the guinea-pig myelin basic protein and its synthetic encephalitogenic peptide, and expressed the same TCR V genes homologous to the mouse V alpha 2 and V beta 8.2 families that appear to be the defining entity of experimental autoimmune encephalomyeltis (EAE). Moreover, the TCR D and J region gene products of the DN cell were found to be similar to another encephalitogenic rat T cell clone. The two T clones did not differ markedly in their ability to produce TNF and IL-2 and to adhere to vascular wall-derived extracellular matrix- and laminin-coated plates. Surprising, therefore, was the finding that, although the CD4+ T lymphocytes were capable of inducing EAE, the DN cells did not elicit disease but rather inhibited subsequent EAE induction. Thus, TCR V alpha 2V beta 8.2 and its junctional region gene products are not the only prerequisite segment for a T cell to become encephalitogenic. We suggest that the important determinants of the T cell ability to induce disease are features of the T cell, other than or in addition to, the T cell receptor.     

Influence of the route of infection on development of T-cell receptor beta-chain repertoires of reovirus-specific cytotoxic T lymphocytes

It is well established that the route of infection affects the nature of the adaptive immune response. However, little is known about the effects of the route of exposure on development of cytotoxic T-lymphocyte (CTL) responses. Alternative antigen-presenting cell populations, tissue-restricted expression of class I major histocompatibility complex-encoded molecules, and unique T-cell receptor (TCR)-bearing cells in mucosal tissues could influence the selection and expansion of responder T cells. This study addresses the question of whether the route of virus infection affects the selection and expansion of subpopulations of virus-specific CTLs. Mice were infected orally or in the hind footpads with reovirus, and the repertoires of TCR beta-chains expressed on virus-specific CD8(+) T cells in Peyer's patches or lymph nodes and spleens were examined. CD8(+) cells expressing the variable gene segment of the TCR beta-chain 6 (Vbeta6) expanded in the spleens of mice infected by either route and in CTL lines established from the spleens and draining lymphoid tissues. Adoptively transferred Vbeta6(+) CD8(+) T cells from orally or parenterally infected donors expanded in reovirus-infected severe combined immunodeficient recipient mice and mediated cytotoxicity ex vivo. Furthermore, recovered Vbeta6(+) cells were enriched for clones utilizing uniform complementarity-determining region 3 (CDR3) lengths. However, sequencing of CDR3beta regions from Vbeta6(+) CD8(+) cells indicated that Jbeta gene segment usage is significantly more restricted in CTLs from orally infected mice, suggesting that the route of infection affects selection and/or subsequent expansion of virus-specific CTLs.