The gammadelta T-cell receptors (TCRs) are limited in their diversity, suggesting that their natural ligands may be few in number. Ligands for gammadeltaTCRs that have thus far been determined are predominantly of host rather than foreign origin. Correlations have been noted between the Vgamma and/or Vdelta genes a gammadelta T cell expresses and its functional role. The reason for these correlations is not yet known, but several different mechanisms are conceivable. One possibility is that interactions between particular TCR-V domains and ligands determine function or functional development. However, a recent study showed that at least for one ligand, receptor specificity is determined by the complementarity-determining region 3 (CDR3) component of the TCR-delta chain, regardless of the Vgamma and/or Vdelta. To determine what is required in the TCR for other specificities and to test whether recognition of certain ligands is connected to cell function, more gammadeltaTCR ligands must be defined. The use of recombinant soluble versions of gammadeltaTCRs appears to be a promising approach to finding new ligands, and recent results using this method are reviewed.

To evaluate the role of the TCR in the αβ/γδ lineage choice during human thymocyte development, molecular analyses of the TCRβ locus in γδ cells and the TCRγ and δ loci in αβ cells were undertaken. TCRβ variable gene segments remained largely in germline configuration in γδ cells, indicating that commitment to the γδ lineage occurred before complete TCRβ rearrangements in most cases. The few TCRβ rearrangements detected were primarily out-of-frame, suggesting that productive TCRβ rearrangements diverted cells away from the γδ lineage. In contrast, in αβ cells, the TCRγ locus was almost completely rearranged with a random productivity profile; the TCRδ locus contained primarily nonproductive rearrangements. Productive γ rearrangements were, however, depleted compared with preselected cells. Productive TCRγ and δ rearrangements rarely occurred in the same cell, suggesting that αβ cells developed from cells unable to produce a functional γδ TCR. Intracellular TCRβ expression correlated with the up-regulation of CD4 and concomitant down-regulation of CD34, and plateaued at the early double positive stage. Surprisingly, however, some early double positive thymocytes retained γδ potential in culture. We present a model for human thymopoiesis which includes γδ development as a default pathway, an instructional role for the TCR in the αβ/γδ lineage choice, and a prolonged developmental window for β selection and γδ lineage commitment. Aspects that differ from the mouse are the status of TCR gene rearrangements at the nonexpressed loci, the timing of β selection, and maintenance of γδ potential through the early double positive stage of development.

Rationale: Lymphocytic alveolitis in HIV-1-infected individuals is associated with multiple pulmonary complications and a poor prognosis. Although lymphocytic alveolitis has been associated with viremia and an increased number of CD8 1 T cells in the lung, its exact cause is unknown.Objectives: To determine if HIV-1-specific T cells are associated with lymphocytic alveolitis in HIV-1-infected individuals.Methods: Using blood and bronchoalveolar lavage (BAL) cells from normal control subjects and untreated HIV-1-infected individuals, we examined the frequency and functional capacity of HIV-1-specific T cells. Measurements and Main Results:We found that HIV-1-specific T cells were significantly elevated in the BAL compared with blood of HIV-1-infected individuals and strongly correlated with T-cell alveolitis. Expression of Ki67, a marker of in vivo proliferation, was significantly reduced on HIV-1-specific T cells in BAL compared with blood, suggesting a diminished proliferative capacity. In addition, HIV-1-specific CD4 1 and CD8 1 T cells in BAL had higher expression of programmed death 1 (PD-1) and lower cytotoxic T-lymphocyte antigen 4 (CTLA-4) expression than those in the blood. A strong correlation between PD-1, but not CTLA-4, and HIV-1-specific T-cell proliferation was seen, and blockade of the PD-1/PD-L1 pathway augmented HIV-1-specific T-cell proliferation, suggesting that the PD-1 pathway was the main cause of reduced proliferation in the lung.Conclusions: These findings suggest that alveolitis associated with HIV-1 infection is caused by the recruitment of HIV-1-specific CD4 1 and CD8 1 T cells to the lung. These antigen-specific T cells display an impaired proliferative capacity that is caused by increased expression of PD-1.

sera of PANDAS subjects, who had 76-89% positive association with elevated individual autoantibody titers and positive CaMKII activity. At 6 months follow-up, symptoms improved for more than 80% of PANDAS subjects, and serum autoantibody titers also significantly decreased. Results reported herein and previously published studies in our laboratory suggest the antibody biomarkers may be a useful adjunct to clinical diagnosis of SC, PANDAS, and related disorders and are the first known group of autoantibodies detecting dopamine receptor-mediated encephalitis in children.

As only a handful of ligands have been identified, the general nature of the ligands recognized by gammadelta T cells remains unresolved. In this study, soluble multimerized gammadelta T cell receptors (smTCRs) representing the TCRs of two gammadelta T cell subsets common in the mouse were used to detect and track their own ligands. Ligands for both subsets were found on resident peritoneal macrophages taken from untreated mice, and the expression of both was further induced by Listeria monocytogenes infection. Nevertheless, the two types of ligand differ from one another in abundance, in the kinetics of their induction following Listeria infection, and in their ability to be induced by in vitro culture with lipopolysaccharide (LPS). Surprisingly, because both are detectable on normal macrophages, these host-derived ligands are likely expressed constitutively, but are induced to higher levels of expression by stress or inflammation. In contrast to T22 and other known cell surface ligands for gammadelta T cells in mice and humans, expression of these smTCR-defined ligands does not depend on beta2-microglobulin, suggesting that they are not MHC class I or class I-like molecules.

γδ T cells express adaptive antigen receptors encoded by rearranging genes. Their diversity is highest in the small region of TCR V-J junctions, especially in the δ chain, which should enable the γδ TCRs to distinguish differences in small epitopes. Indeed, recognition of small molecules, and of an epitope on a larger protein has been reported. Responses to small non-peptides known as phospho-antigens are multi-clonal yet limited to a single γδ T cell subset in humans and non-human primates. Responses to small peptides are multi-clonal or oligo-clonal, include more than one subset of γδ T cells, and occur in rodents and primates. However, less effort has been devoted to investigate the peptide responses. To settle the questions of whether peptides can be ligands for the γδ TCRs, and whether responses to small peptides might occur normally, peptide binding will have to be demonstrated, and natural peptide ligands identified.

Chronic beryllium disease (CBD) is an occupational lung disorder characterized by granulomatous inflammation and the accumulation of beryllium-responsive CD4+ T cells in the lung. These differentiated effector memory T cells secrete IL-2, IFN-γ, and TNF-α upon in vitro activation. Beryllium-responsive CD4+ T cells in the lung are CD28 independent and have increased expression of the coinhibitory receptor, programmed death 1, resulting in antigen-specific T cells that proliferate poorly yet retain the ability to express Th1-type cytokines. To further investigate the role of coinhibitory receptors in the beryllium-induced immune response, we examined the expression of CTLA-4 in blood and bronchoalveolar lavage cells from subjects with CBD. CTLA-4 expression was elevated on CD4+ T cells from the lungs of study subjects compared to blood. Furthermore, CTLA-4 expression was greatest in the beryllium-responsive subset of CD4+ T cells that retained the ability to proliferate and express IL-2. Functional assays show that the induction of CTLA-4 signaling in blood cells inhibited beryllium-induced T cell proliferation while having no effect on the proliferative capacity of beryllium-responsive CD4+ T cells in lung. Collectively, our findings suggest a dysfunctional CTLA-4 pathway in the lung and its potential contribution to the persistent inflammatory response that characterizes CBD.

Analyzing the status of T-cell receptor (TCR) gene rearrangements has been an essential part of deciphering the stages of thymocyte development, understanding the αβ vs. γδ lineage decision, and characterizing T-cell leukemias. Methods such as PCR and quantitative Southern blotting provide useful information, but also have significant shortcomings such as lack of quantitation in the case of PCR and technical challenges in the case of Southern blotting. Here we describe a real-time PCR method that overcomes many of these shortcomings. This new method shows comparable results for the fraction of unrearranged TCRγ and TCRβ genes in human thymocytes and peripheral blood T cells as Southern blotting, and has the advantages of being simple to perform, highly quantitative, and requiring nanogram quantities of DNA. We also describe a real-time PCR method to quantitate T-cell receptor excision circles formed during TCRβ rearrangements.