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Mucosal-associated invariant T (MAIT) cells are a major subset of innate-like T cells mediating protection against bacterial infection through recognition of microbial metabolites derived from riboflavin biosynthesis. Mouse MAIT cells egress from the thymus as two main subpopulations with distinct functions, namely, T-bet-expressing MAIT1 and RORγt-expressing MAIT17 cells. Previously, we reported that inducible T-cell costimulator and interleukin (IL)-23 provide essential signals for optimal MHC-related protein 1 (MR1)-dependent activation and expansion of MAIT17 cells in vivo. Here, in a model of tularemia, in which MAIT1 responses predominate, we demonstrate that IL-12 and IL-23 promote MAIT1 cell expansion during acute infection and that IL-12 is indispensable for MAIT1 phenotype and function. Furthermore, we showed that the bias toward MAIT1 or MAIT17 responses we observed during different bacterial infections was determined and modulated by the balance between IL-12 and IL-23 and that these responses could be recapitulated by cytokine coadministration with antigen. Our results indicate a potential for tailored immunotherapeutic interventions via MAIT cell manipulation.

MHC-related protein 1 (MR1) presents microbial riboflavin metabolites to mucosal-associated invariant T (MAIT) cells for surveillance of microbial presence. MAIT cells express a semi-invariant T-cell receptor (TCR), which recognizes MR1-antigen complexes in a pattern-recognition-like manner. Recently, diverse populations of MR1-restricted T cells have been described that exhibit broad recognition of tumor cells and appear to recognize MR1 in association with tumor-derived self-antigens, though the identity of these antigens remains unclear. Here, we have used TCR gene transfer and engineered MR1-expressing antigen-presenting cells to probe the MR1 restriction and antigen reactivity of a range of MR1-restricted TCRs, including model tumor-reactive TCRs. We confirm MR1 reactivity by these TCRs, show differential dependence on lysine at position 43 of MR1 (K43) and demonstrate competitive inhibition by the MR1 ligand 6-formylpterin. TCR-expressing reporter lines, however, failed to recapitulate the robust tumor specificity previously reported, suggesting an importance of accessory molecules for MR1-dependent tumor reactivity. Finally, MR1-mutant cell lines showed that distinct residues on the α1/α2 helices were required for TCR binding by different MR1-restricted T cells and suggested central but distinct docking modes by the broad family of MR1-restricted αβ TCRs. Collectively, these data are consistent with recognition of distinct antigens by diverse MR1-restricted T cells.

Conventional T cells recognise protein-derived antigens in the context of major histocompatibility complex (MHC) class Ia and class II molecules and provide anti-microbial and anti-tumour immunity. Conventional T cells have also been implicated in type IV (also termed delayed-type or T cell-mediated) hypersensitivity reactions in response to protein-derived allergen antigens. In addition to conventional T cells, subsets of unconventional T cells exist, which recognise non-protein antigens in the context of monomorphic MHC class I-like molecules. These include T cells that are restricted to the cluster of differentiation 1 (CD1) family members, known as CD1-restricted T cells, and mucosal-associated invariant T cells (MAIT cells) that are restricted to the MHC-related protein 1 (MR1). Compared with conventional T cells, much less is known about the immune functions of unconventional T cells and their role in hypersensitivities. Here, we review allergen antigen presentation by MHC-I-like molecules, their recognition by unconventional T cells, and the potential role of unconventional T cells in hypersensitivities. We also speculate on possible scenarios of allergen antigen presentation by MHC-I-like molecules to unconventional T cells, the hallmarks of such responses, and the expected frequencies of hypersensitivities within the human population.

© 2021 The Authors., The third edition of Flow Cytometry Guidelines provides the key aspects to consider when performing flow cytometry experiments and includes comprehensive sections describing phenotypes and functional assays of all major human and murine immune cell subsets. Notably, the Guidelines contain helpful tables highlighting phenotypes and key differences between human and murine cells. Another useful feature of this edition is the flow cytometry analysis of clinical samples with examples of flow cytometry applications in the context of autoimmune diseases, cancers as well as acute and chronic infectious diseases. Furthermore, there are sections detailing tips, tricks and pitfalls to avoid. All sections are written and peer-reviewed by leading flow cytometry experts and immunologists, making this edition an essential and state-of-the-art handbook for basic and clinical researchers., Hyun-Dong Chang is supported by the Dr. Rolf M. Schwiete Foundation. Susanne Melzer and Attila Tarnok thank De Novo Software for providing FCS Express. Enrico Lugli is supported by a grant from the Associazione Italiana per la Ricerca sul Cancro (AIRC IG20676). Gabriele De Simone and Giovanni Galletti were supported by Fellowships from the Fondazione Italiana per la Ricerca sul Cancro-Associazione Italiana per la Ricerca sul Cancro (FIRC-AIRC). Jun Dong is supported by Deutsche Forschungsgemeinschft (DFG, German Research Foundation) Projektnummer 389687267 and Chinesisch-Deutsches Zentrum für Wissenschaftsförderung [Sino-German Center for Research Promotion (SGC)] grant C-0072. Nicola Gagliani, Samuel Huber and Franziska Muscate are supported by DFG fundings: SFB841,GA 2441/3-1, HU 1714/10-1. The tetramer APC-conjugated H-2K (d) HIV-1 gag197-205 AMQMLKETI used in TDS assay for mouse blood T cells was obtained through the NIH Tetramer Facility. Larissa Nogueira Almeida was supported by DFG research grant MA 2273/14-1. Supported by the following grants: AIRC 5X1000 2018 id. 21147 (Lorenzo Moretta); AIRC IG 2017 id. 19920 (Lorenzo Moretta); RC-2020 OPBG (Lorenzo Moretta); AIRC and European Union's Horizon 2020 research and innovation program under the Marie Skłodowska-Curie grant agreement No 800924 (Linda Quatrini). Dirk Baumjohann was supported by Deutsche Forschungsgemeinschaft (DFG, German Research Foundation) under Emmy Noether Programme BA 5132/1-2 (252623821) and Germany's Excellence Strategy EXC2151 (390873048).

Mucosal‐associated invariant T (MAIT) cells represent up to 10% of circulating human T cells. They are usually defined using combinations of non‐lineage‐specific (surrogate) markers such as anti‐TRAV1‐2, CD161, IL‐18Rα and CD26. The development of MR1‐Ag tetramers now permits the specific identification of MAIT cells based on T‐cell receptor specificity. Here, we compare these approaches for identifying MAIT cells and show that surrogate markers are not always accurate in identifying these cells, particularly the CD4+ fraction. Moreover, while all MAIT cell subsets produced comparable levels of IFNγ, TNF and IL‐17A, the CD4+ population produced more IL‐2 than the other subsets. In a human ontogeny study, we show that the frequencies of most MR1 tetramer+ MAIT cells, with the exception of CD4+ MAIT cells, increased from birth to about 25 years of age and declined thereafter. We also demonstrate a positive association between the frequency of MAIT cells and other unconventional T cells including Natural Killer T (NKT) cells and Vδ2+ γδ T cells. Accordingly, this study demonstrates that MAIT cells are phenotypically and functionally diverse, that surrogate markers may not reliably identify all of these cells, and that their numbers are regulated in an age‐dependent manner and correlate with NKT and Vδ2+ γδ T cells.

The major histocompatibility complex (MHC) class-I related molecule MR1 is a monomorphic and evolutionary conserved antigen (Ag)-presenting molecule that shares the overall architecture of MHC-I and CD1 proteins. However, in contrast to MHC-I and the CD1 family that present peptides and lipids, respectively, MR1 specifically presents small organic molecules. During microbial infection of mammalian cells, MR1 captures and presents vitamin B precursors, derived from the microbial biosynthesis of riboflavin, on the surface of antigen-presenting cells. These MR1-Ag complexes are recognized by the mucosal-associated invariant T cell receptor (MAIT TCR), which subsequently leads to MAIT cell activation. Recently, MR1 was shown to trap chemical scaffolds including drug and drug-like molecules. Here, we review this metabolite Ag-presenting molecule and further define the key molecular interactions underlying the recognition and reactivity of MAIT TCRs to MR1 in an Ag-dependent manner.

TCR-αβ(+) double negative (DN) T cells (CD3(+) TCR-αβ(+) CD4(-) CD8(-) NK1.1(-) CD49b(-) ) represent a minor heterogeneous population in healthy humans and mice. These cells have been ascribed pro-inflammatory and regulatory capacities and are known to expand during the course of several autoimmune diseases. Importantly, previous studies have shown that self-reactive CD8(+) T cells become DN after activation by self-antigens, suggesting that self-reactive T cells may exist within the DN T-cell population. Here, we demonstrate that programmed cell death 1 (PD-1) expression in unmanipulated mice identifies a subset of DN T cells with expression of activation-associated markers and a phenotype that strongly suggests they are derived from self-reactive CD8(+) cells. We also found that, within DN T cells, the PD-1(+) subset generates the majority of pro-inflammatory cytokines. Finally, using a TCR-activation reporter mouse (Nur77-GFP), we confirmed that in the steady-state PD-1(+) DN T cells engage endogenous antigens in healthy mice. In conclusion, we provide evidence that indicates that the PD-1(+) fraction of DN T cells represents self-reactive cells.

This overview presents nomenclature and serology information on human leucocyte antigens, or HLA molecules, which are encoded by a cluster of genes linked on the short arm of chromosome 6. This region is known as the major histocompatibility complex and codes for class I and class II molecules, which are distinguished from each other based upon their structure, tissue distribution, and source of peptide antigen, as well as upon their interactions with T cell subsets. © 2017 by John WileySons, Inc.

The T-cell antigen receptor is a heterodimeric αβ protein (TCR) expressed on the surface of T-lymphocytes, with each chain of the TCR comprising three complementarity-determining regions (CDRs) that collectively form the antigen-binding site. Unlike antibodies, which are closely related proteins that recognize intact protein antigens, TCRs classically bind, via their CDR loops, to peptides (p) that are presented by molecules of the major histocompatibility complex (MHC). This TCR-pMHC interaction is crucially important in cell-mediated immunity, with the specificity in the cellular immune response being attributable to MHC polymorphism, an extensive TCR repertoire and a variable peptide cargo. The ensuing structural and biophysical studies within the TCR-pMHC axis have been highly informative in understanding the fundamental events that underpin protective immunity and dysfunctional T-cell responses that occur during autoimmunity. In addition, TCRs can recognize the CD1 family, a family of MHC-related molecules that instead of presenting peptides are ideally suited to bind lipid-based antigens. Structural studies within the CD1-lipid antigen system are beginning to inform us how lipid antigens are specifically presented by CD1, and how such CD1-lipid antigen complexes are recognized by the TCR. Moreover, it has recently been shown that certain TCRs can bind to vitamin B based metabolites that are bound to an MHC-like molecule termed MR1. Thus, TCRs can recognize peptides, lipids, and small molecule metabolites, and here we review the basic principles underpinning this versatile and fascinating receptor recognition system that is vital to a host's survival.

T cells that express clonally distributed αβ T-cell receptors (TCRs) corecognize antigenic peptides (p) bound to major histocompatibility complex class I (MHC-I) and class II molecules (MHC-II). Extensive human leukocyte antigen (HLA) polymorphism enables HLA molecules from different haplotypes to capture an array of self- and microbe-derived peptide antigens that is fundamental to adaptive immunity. T cells developing in the thymus are selected for weak binding to self-peptide-HLA complexes generating a vast repertoire of clonally distinct T cells in the periphery. Indeed, diversity within germline loci and the finally assembled TCR genes, coupled with inherent TCR cross-reactivity, enables CD8(+) T cells to survey the multitude of pHLA-I landscapes. Precisely how does the TCR ligate to pHLA-I, and how does knowledge of the detailed structural interactions inform immunobiology? A recent number of our structural studies concerning the TCR-pMHC-I axis, alongside others in the field, have provided insight into HLA-I polymorphism, pMHC-I flexibility, TCR bias, TCR polymorphism, maintenance of self-tolerance, T-cell cross-reactivity, and alloreactivity. Collectively, the data also provide an opportunity to address the structural correlates of MHC-I restriction. Here, we provide our perspective concerning these advances in the field. Although much key information has been gleaned, the structural data show that some of the key concepts surrounding the TCR-pMHC-I interaction remain controversial and unresolved.

Objective This study was undertaken to determine the molecular characteristics of clonotypic autoantibodies in the sera of patients with primary Sjogren's syndrome (SS). This characterization is hampered by the presence of mixed anti-Ro/La specificities that may conceal clonotypic species. In order to narrow clonotypic diversity, a positive selection step was performed on a peg-like determinant of Ro 60 (termed Ro 60–peg) prior to analysis of the autoantibody proteome. Methods Monospecific anti–Ro 60–peg IgG were isolated by affinity purification from the sera of 7 patients with primary SS and anti-Ro/La and subjected to 2-dimensional gel electrophoresis and high-resolution orbitrap mass spectrometric sequencing. V regions of heavy and light chains were analyzed by combined database and de novo amino acid sequencing. Results Proteomic analysis revealed a Ro 60–peg–specific IgG1κ-restricted monoclonal autoantibody that was present in the sera of all patients and specified by a VH3–23 heavy chain paired with a Vκ3–20 light chain. The public anti–Ro 60–peg clonotype was specified further by common mutations in the heavy-chain and light-chain complementarity-determining regions. Titers and relative affinities of clonotypic IgG did not vary over the course of the disease. Conclusion The expression of a Ro 60–reactive public B cell clonotype in a subset of patients with primary SS as a long-lived, class-switched circulating autoantibody implies a common breach of B cell tolerance checkpoints in these patients. The unique heavy chain/light chain signature opens the possibility of tracking the development of a “forbidden” clone against a bona fide systemic autoantigen in human disease.

The association between the immune system and malignancy is at the forefront of research aimed at identifying and developing effective immunotherapies to treat and suppress cancer. Understanding the mechanisms underlying the biology, differentiation and effector functions of key immune cells in the tumor microenvironment is central to this process. The stimulation of T cells results in their activation, triggering the release of cellular mediators, including cytokines and chemokines. The interplay between these components determines the polarization of T cells into phenotypically-, functionally- and transcriptionally-distinct subsets (e.g. T-helper [Th] 1, Th2, Th17 and T-regulatory cells). Given that antigen recognition, costimulation and the cytokine milieu are critical signals in T-cell activation and differentiation, they represent potential and important therapeutic targets for modulating immune responses in disease. The aim of this article is to review the role of T cells in immune protection and pathogenesis, and to discuss the key pathways involved in these processes.

αβ T-cell receptors (TCRs), which can engage a broad array of foreign peptide-laden major histocompatibility complex (pMHC) landscapes, have an essential role in protective immunity. TCRs are selected by pMHC molecules in the thymus and in the periphery, and so are restricted to recognizing 'self'-major histocompatibility complex (MHC) molecules. Accordingly, T cells are inherently cross-reactive, and although the versatility and specificity of this MHC-restricted response are the hallmarks of adaptive immunity, 'unwanted' TCR interactions, such as those observed in T-cell alloreactivity, often occur. Recent data have shown that direct T-cell alloreactivity can arise from peptide-dependent molecular mimicry, as well as distinct pMHC-binding modes. Here we review recent advances in the field, focusing on structural data pertaining to alloreactivity, and discuss the implications for T-cell-mediated transplant rejection.

Controversy still surrounds the importance of cross-presentation versus endogenous or direct presentation of MHC-I restricted Ag in CD8(+) T-cell (T(CD8+)) immunity. It is even less clear what relative role these pathways play in shaping the T-cell repertoire specific for ubiquitous self-antigens, especially in cases where both Ag presentation pathways could potentially be involved. Here we provide evidence that a T(CD8+) repertoire specific for a determinant from the nuclear autoantigen La-SSB is largely shaped by direct presentation. In this system, mouse T(CD8+) reactive to a xenogeneic human La (hLa(51-58)) K(b) peptide did not recognize directly presented peptide on either spleen cells from hLa-Tg mice or hLa transfected syngeneic cells. Interestingly, the same T(CD8+) were activated by in vivo challenge with allogeneic APC expressing either the Tg hLa or loaded with intact recombinant hLa protein, indicating functional cross-presentation of the hLa(51-58). However, in irradiated bone marrow chimeric mice, DC expressing Tg hLa, but not WT DC that matured in hLa-Tg mice, constitutively presented the hLa(51-58) to T(CD8+). These data demonstrate that although both the direct- and cross-presentation pathways are potentially operative in revealing hLa(51-58) to T(CD8+), the T(CD8+) repertoire to this determinant is shaped quantitatively according to the efficiency of Ag presentation.

To analyse the immune correlates in a setting of recurrent exposure to hepatitis C virus (HCV), we studied T(CD8) responses in injecting drug users (IDUs) with different disease outcomes. Ex vivo HCV-specific T(CD8) responses assessed by interferon-gamma (IFNgamma) enzyme-linked immunospot (ELISPOT) were comparable in human lymphocyte antigen (HLA)-matched IDUs with spontaneous HCV clearance or persistent infection. A detailed characterization of these T(CD8) cells in age and HLA-matched IDUs demonstrated that HCV clearance and protection from reinfection correlated with HCV-specific T(CD8) cells that could proliferate in vitro, possessed cytotoxic potential and produced IFNgamma and tumour-necrosis factor-alpha, rather than with the circulating frequency of responding T(CD8) cells determined ex vivo. While validating the importance of multifunctional T(CD8) in mediating protection in IDUs with recurrent exposure to HCV our findings highlight that the magnitude and/or breadth of HCV-specific T(CD8) determined in ex vivo ELISPOT may not be the sole determinant of protection especially in a setting of recurrent exposure.

Patients with Sjögren's syndrome (SS) in association with either Rheumatoid Arthritis (RA), Systemic Lupus Erythematosus (SLE) or Sclerodactyly were tested for HLA-A, B and Dr antigens and the phenotypes of the 4th component of complement (CA). Significant associations were identified between HLA-Bw62, Dr4, C4B2.9, C4A8 and females with SS-RA. The MHC antigen frequencies noted for secondary Sjögren's syndrome differed from those reported for their respective primary diseases and the implications of these observations are discussed.

Objective T cells are implicated in the production of anti-La/SSB and anti-Ro/SSA autoantibodies commonly associated with the DR3/DQ2 haplotype in systemic lupus erythematosus and Sjogren's syndrome. This study was undertaken to investigate the DR3/DQ2-restricted T cell response to wild-type human La (hLa) and a truncated form of mutant La. Methods Humanized transgenic mice expressing HLA–DRB1*0301/DQB1*0201 (DR3/DQ2) were immunized with recombinant antigen and examined for development of autoantibodies and T cell proliferation against overlapping peptides spanning the La autoantigen. HLA restriction and peptide binding of identified T cell epitopes to DR3 or DQ2 were determined using blocking monoclonal antibodies and a direct binding assay. Results DR3/DQ2-transgenic mice generated an unusually rapid class-switched humoral response to hLa with characteristic spreading to Ro 52 and Ro 60 proteins following hLa protein immunization. Seven T cell determinants in hLa were restricted to the HLA–DR3/DQ2 haplotype. Six epitopes tested were restricted to HLA–DR and bound DR3 with semiconserved DR3 binding motifs. No DQ restriction of these epitopes was demonstrable despite efficient DQ binding activity in some cases. No neo–T cell epitopes were identified in mutant La; however, T cells primed with mutant La exhibited a striking increase in proliferation to the epitope hLa151–168 compared with T cells primed with hLa. Conclusion Multiple DR3-restricted epitopes of hLa have been identified. These findings suggest that truncation of La produced by somatic mutation or possibly granzyme B–mediated cleavage alters the immunodominance hierarchy of T cell responsiveness to hLa and may be a factor in the initiation or maintenance of anti-La autoimmunity.

Polyphenol oxidases include tyrosinase and catechol oxidase. There have been multiple reports in the literature describing the apparent synergistic action when different polyphenol oxidase substrat...

Many autoimmune conditions have close genetic linkages to particular human histocompatibility leukocyte antigen (HLA) class II genes. With the aim of establishing a murine model of autoimmune disease, we have generated an HLA DR4-DQ3 haplotype transgenic (Tg) mouse that expresses a 440-kb yeast artificial chromosome harbouring DRA, DRB1*040101, DRB4*010301, DQA1*030101, DQB1*0302 and all the internal regulatory segments. This Tg mouse line was crossed to human CD4 (hCD4) Tg mice and endogenous class II knockout mice (I-A(o/o) and I-E(o/o)) lines to generate a DR4-DQ3.hCD4.IAE(o/o) Tg line. The Tg DR and DQ molecules are expressed on the physiological cell types in these animals, i.e. on most B cells (>85%), dendritic cells (DCs) and macrophages but not on T cells, with levels of expression comparable with those of human B cells (where DR > DQ expression). The DR4/DQ3 transgenes fully reconstituted the CD4 T-cell compartment, in both the thymus and the periphery, and the analysis of the T-cell receptor repertoire in the Tg mice confirmed that these class II molecules were able to mediate thymic selection of a broad range of Vbeta families. HLA DR4- and DQ3-restricted T-cell responses were elicited following immunization with known T-cell determinants presented by these molecules. Furthermore, the DR4-DQ3-restricted CD4(+) T cells conferred protective antibody-mediated immunity against an otherwise lethal infection with Salmonella enterica var. typhimurium. These new DR4-DQ3 Tg mice should prove to be valuable tools for dissecting the importance of this class II haplotype in autoimmune disorders like rheumatoid arthritis.

Alphabeta T-cell receptor (TcR) recognition of antigenic peptides bound to the major histocompatibility complex (pMHC), is integral to the cellular immune system. Crystallographic studies over the last decade have provided significant insight into this unique trimolecular recognition event. The TcR-pMHC structural information has been paralleled by biophysical studies that have further explored the emerging binding models in an attempt to answer fundamental immunological questions regarding MHC restriction, T-cell immunodominance and TcR cross-reactivity. However, despite the important data that has been generated regarding TcR-pMHC interactions, the scope of this information is still incomplete due to the limited range of TcRs that have been studied. These limitations are primarily due to difficulties in obtaining high yields of recombinant alphabeta TcR for crystallographic and biophysical analysis; here we will discuss some of the protein engineering strategies that have been employed to expand the pool of recombinant TcRs suitable for crystallographic studies and the subsequent studies that have utilized these proteins.

We describe a strategy for identifying ligands of human leukocyte antigen (HLA) class I molecules based on a peptide library-mediated in vitro assembly of recombinant class I molecules. We established a microscale class I assembly assay and used a capture ELISA to quantify the assembled HLA-peptide complexes. The identity of the bound ligands was then deduced by mass spectrometry. In this method, HLA complexes assembled in vitro in the presence of components of a mixture of peptides were immunoprecipitated and the bound peptide(s) identified by matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass spectrometry. This process of epitope extraction is robust and can be used with complex mixtures containing in excess of 300 candidate ligands. A library of overlapping peptides representing all potential octamers, nonamers and decamers from human preproinsulin was synthesized using unique library chemistry. Peptides from the library were used to initiate assembly of recombinant HLA-B8, HLA-B15 and HLA-A2, facilitating the identification of candidate T-cell epitopes from preproinsulin.

In 1998, the first International Conference on human leukocyte antigen-G (HLA-G) was held in Paris. At that time, HLA-G was still a new HLA class I molecule, few aspects of its immunological functions were known, and its expression by tumors was just being described. In 1998, tools to properly study HLA-G were lacking, especially monoclonal antibodies, and three conclusions were drawn after the congress: (i) animal models were needed, (ii) the biology of HLA-G isoforms had to be confirmed, and (iii) HLA-G expression by tumors required clarification. Five years later, these three issues have been addressed. HLA-G is now gaining pace and is investigated for its immuno-inhibitory functions in the context of multiple pathologies. Eighty five oral presentations were given this year for more than 200 investigators working on HLA-G by speakers from over 20 countries. The success of the 3rd International Conference on HLA-G reflects the interest and tremendous work of the many research teams which, over the years, contributed to the publication of more than 500 peer-review articles. We summarize the key points that were presented and discussed during this meeting.

Background Hypersensitivity reactions to carbamazepine (CBZ) have been shown to be strongly associated with specific human leukocyte antigen (HLA) alleles, with carriers of the HLA alleles presenting an increased risk of developing hypersensitivity. HLA-B*15:02 was detected in almost all cases of CBZ-induced Stevens-Johnson syndrome (SJS) in patients of Han Chinese or South-East Asian ancestry, and its functional role in CBZ-induced SJS has been well characterised. HLA-A*3101 is associated with all clinical phenotypes of CBZ-induced hypersensitivity in Caucasian and Japanese patients. However, functional studies investigating the role of HLA-A*31:01 in CBZ-specific T-cell responses have not been performed. Furthermore, CBZ-specific T-cells of CD4+ and CD8+ phenotype are readily detectable in Caucasian patients, which is in stark contrast to the dominant CD8+ T-cell response in Han Chinese. In this study we therefore investigated the HLA restriction of CBZ-reactive T-cells from a HLA-A*31:01 positive CBZ hypersensitive patient, focusing on both the CD4+ and CD8+ cells.

We report the distribution of genes encoding 11 killer cell immunoglobulin-like receptors (KIR) and 2 CD94:NKG2 receptors, in 32 Caucasians, 67 Australian Aborigines and 59 Vietnamese. The inhibitory and the activating KIR genes were found at different frequency in the three populations. No correlation was found between the polymorphism of the KIR genes and the HLA specificities of the tested samples. The most significant KIR associations were 2DL2 with 2DS2; 2DL2 with 2DS3 and 3DL1 with 2DS4 in all three study groups. In Caucasians and Vietnamese 2DS2 was associated with 2DS3 and 2DS1with 3DS1. KIR 2DL1 was strongly associated with three other KIRs: 2DL3, 3DL1 and 2DS4 in Aborigines. The distribution of the KIR phenotypes was different in the three populations. The AA1 phenotype was frequent in Vietnamese (42.4%) and Caucasians (31.2%), but very rare in Aborigines (1.5%). In contrast, the BB7 phenotype was very common for Aborigines (22.4%) and was absent in the two other groups. Our data demonstrate that different associations and putative KIR haplotypes could be distinguished in different populations.

Background: Reactive arthritis (ReA) is an inflammatory arthritis triggered by certain gastrointestinal and genitourinary infections. Single source outbreaks of triggering infections provide an opportunity to elucidate host susceptibility factors in this disease. Aim: To determine the role of Major Histocompatibility Complex (MHC) Class I alleles in ReA susceptibility after two large single source outbreaks of Salmonella Typhimurium gastroenteritis. Methods: A questionnaire screening for features of ReA and a request for HLA class I typing were sent to all patients affected by two single source outbreaks of S. Typhimurium gastroenteritis. Individuals with arthritis of recent onset were interviewed, examined and diagnostic criteria for ReA applied. Results: Nineteen cases of reactive arthritis, 11 female, were diagnosed in the 424 respondents with S. Typhimurium gastroenteritis from both outbreaks. Clinical features of the arthritis were similar to those described after other large single source outbreaks of Salmonella infection. HLA-B27 was expressed by only two of the 19 ReA patients and therefore did not predict susceptibility to this form of arthritis. Caucasians were, however, more likely to develop reactive arthritis than Asians. Conclusions: In this study, susceptibility to ReA was not increased in HLA-B27 positive individuals or males but was greater in those of Caucasian descent.

Three previously unknown A24-related alleles were identified by PCR-SSO typing and confirmed by DNA sequencing in Australian Aboriginal populations (A*2406, 2413) and in individuals of South American descent (A*2414). A*2406 and A*2413 both have two adjacent (but different) nucleotide substitutions in codon 156 in exon 3 compared to A*2402, resulting in a single amino acid replacement in each allele. The South American A*2414 is apparently a hybrid between A2 and A24 with a segment of the A*24 sequence between codons 95 and 107 in exon 3 replaced with the A*02 sequence. Interallelic sequence exchange is the most likely mechanism in the generation of all three novel alleles. Compared to A*2402, the four amino acid substitutions in the A*2414 molecule would be expected to significantly change the shape of the peptide binding cleft, leading to selection of different peptide ligands. The single amino acid replacements in position 156 of the two Australian Aboriginal A*24 alleles may also have significant functional effects. In particular, Trp replacing Gln in position 156 (A*2406) is predicted to markedly reduce the volume of the peptide binding cleft, influence the interaction of HLA pockets with peptide side chains, and therefore, cause major changes in peptide presentation. These newly defined alleles may reflect the adaptive process of HLA genes to local environments.

The memory response to the immunodominant Epstein-Barr virus (EBV) epitope FLRGRAYGL, which associates with HLA B8, is exceptionally restricted, being dominated by cytotoxic T lymphocytes (CTL) with a single, public T cell receptor (TCR). CTL clones that express this receptor fortuitously cross-react with the alloantigen HLA B44. However, of the two major subtypes of this HLA, B*4402 and B*4403, that differ by a single amino acid, only the former is recognized by these mature CTL clones. Individuals heterozygous for HLA B8 and B*4402 use alternative TCR for the EBV determinant since the dominant TCR is potentially self-reactive. We now demonstrate that this clonotype is also essentially absent from the repertoire of CTL directed against the viral epitope in seven from seven unrelated individuals heterozygous for HLA B8 and B*4403. Thus immune tolerance of these CTL recognizing HLA B*4402 is associated with expression of either B*4402 or B*4403. This suggests that tolerance in the human T cell compartment requires a lower threshold of recognition than for effector function, thus providing a buffer zone minimizing the risk of autoimmunity. These data also illustrate the potential for non-restricting HLA molecules to bias dramatically the T cell repertoire used for specific immune responses. Such influences may be the basis of the "protective" effects of certain HLA alleles in susceptibility to autoimmune disorders.

A serological family study identified an HLA-A "blank" segregating through three generations of apparently healthy individuals. The HLA-A*0301 allele was assigned by DNA genotyping in each of the three individuals. Complete absence of cellular expression of the HLA-A3 antigen was associated with a 6 nucleotide deletion in exon 3 of the A*0301 gene. The in-frame deletion of nucleotides 373-378 results in the absence of residues C101 and D102 from the mature HLA-A heavy chain. Cysteine 101 is involved in the formation of the highly conserved disulfide bridge in the alpha 2 domain of the class I molecule, and deletion of this residue is believed to be highly disruptive to proper folding and function of the class I molecule.

T cell proliferative responses to rye and Bermuda grass pollen allergens have been studied in a series of 51 atopic and 18 non-atopic subjects. Mean T cell responses were higher in the atopic group than in the non-atopic group (P < 0.001), and there was a strong correlation between the magnitude of reaction in the T cell assay and in the skin test (rye P < 0.01, Bermuda P < 0.05). A similar association was shown between T cell reactivity and serum levels of allergen-specific IgE (rye P < 0.05, Bermuda P < 0.05), but no relationship was found between serum allergen-specific IgG levels and any other parameter studied. T cell reactivity was not found in three cord blood samples tested. Discordance between positivity for T cell responses and skin test reactions in some cases might reflect reactivity by T cell subsets that promote IgG antibody or cell-mediated responses without IgE antibody production. A precise knowledge of T cell recognition of grass pollen allergens will provide exciting new prospects for more effective and safer immunotherapy strategies for allergic diseases including asthma.

Traditional methods of serological typing have largely used antisera of Caucasoid origin, which can overlook HLA heterogeneity in non-Caucasoid populations. Therefore, we have used molecular techniques to evaluate potential polymorphism in HLA class I molecules of Aborigines from the central desert and northern coast of Australia. The DNA sequence of common Aboriginal HLA-A and B antigens were compared with serological reaction patterns which suggested new polymorphisms. Although serological data indicated that long and short variants of A34 may exist, regardless of the serological pattern, all individuals carried the A*3401 allele. Therefore, the variation in A34 reaction pattern observed serologically was not attributable to primary sequence variation in the HLA A*3401 allele. Similarly, there was no detectable polymorphism in the sequences of selected HLA-B alleles, even though some of these alleles showed unusual serological reaction patterns. However, a new allele of B15 (B*1521) was detected in two individuals carrying this serotype. The cells from both of these individuals showed ambiguous reaction patterns with monospecific B62 and B75 sera. cDNA sequencing of the HLA B15 gene from these cells revealed a B15 allele that differed from B*1502 by a single nucleotide change. This change occurred at position 272, resulting in a C to G substitution at residue 67 in the consensus B15 cDNA sequence. Hence, the Australian Aborigines as an ethnic group show very little primary sequence polymorphism within the class I loci, consistent with the results obtained from previous serological studies.

Hereditary haemochromatosis (HH) is an autosomal recessive disease involving mutations in the recently characterised HFE gene linked to HLA-A in the major histocompatibility complex. The known HFE polymorphisms include the wild-type allele, a G-->A substitution at base 845 (845A) and a C-->G substitution at position 187 (187G). Although most cases of HH are accountable by homozygosity of the 845A allele the exact risk of other HFE genotypes, especially those involving the 187G allele has not been determined. We have compiled estimates of disease risk for all known HFE genotypes by re-analyzing published studies. The data show a hierarchical risk calculated as odds ratio (OR) for each genotype 845A/ 845A (OR=2101); 845A/187G (OR=24); 187G/187G (OR=9); 845A/Wt (OR=5); 187G/Wt (OR=2). Interestingly, the disease risk of 187G-genotypes suggests that subtle functional changes in the HFE product can interact with other genetic factors (e.g. trans allele, gender) and environmental factors (e.g. diet) to manifest either as clinical disease, altered iron stores or a normal phenotype. This paradigm is potentially useful in understanding the contribution of HLA alleles to risk of various disorders especially autoimmunity.

We have used the term supratype to describe combinations of alleles and have examined associations with disease. In RA and insulin-dependent diabetes one or more supratypes appear to be important but their functional significance remains obscure. In MG and SLE the HLA supratype may contain loci involved in immunoregulation, complement synthesis and hormone metabolism. MG induced by D-Pen is associated with Bw35/DR1 rather than A1, B8, DR3. In contrast there is no evidence of a supratype in AS. We have proposed a model for the pathogenesis of sacroiliitis and AS and have postulated two non-linked genes which act stepwise upon HLA-B27. There are cogent reasons for examining the functional effects of known loci within the MHC and particularly those involved in the expression of complement components.

Reactive arthritis associated with Clostridium difficile. J. McCluskey, T. V. Riley, E. T. Owen and D. R. Langlands. Aust. N.Z. J. Med., 1982, 12, pp. 535–537. A case of reactive arthritis in a patient with a previously documented history of Reiter's syndrome is described. The precipitating agent appears to have been Clostridium difficile. High levels of toxin were demonstrable in the faeces and neutralising antitoxin was detected in the patient's serum but not synovial fluid. Resolution of the poly arthropathy was slow despite successful eradication of the C. difficile with a course of vancomycin.