Adam P. Uldrich

Activation of thymic regeneration in mice and humans following androgen blockade

The thymus undergoes age-related atrophy, coincident with increased circulating sex steroids from puberty. The impact of thymic atrophy is most profound in clinical conditions that cause a severe loss in peripheral T cells with the ability to regenerate adequate numbers of naive CD4+ T cells indirectly correlating with patient age. The present study demonstrates that androgen ablation results in the complete regeneration of the aged male mouse thymus, restoration of peripheral T cell phenotype and function and enhanced thymus regeneration following bone marrow transplantation. Importantly, this technique is also applicable to humans, with analysis of elderly males undergoing sex steroid ablation therapy for prostatic carcinoma, demonstrating an increase in circulating T cell numbers, particularly naive (TREC+) T cells. Collectively these studies represent a fundamentally new approach to treating immunodeficiency states in humans.

A Natural Killer T (NKT) Cell Developmental Pathway Involving a Thymus-dependent NK1.1−CD4+ CD1d-dependent Precursor Stage

The development of CD1d-dependent natural killer T (NKT) cells is poorly understood. We have used both CD1d/α-galactosylceramide (CD1d/αGC) tetramers and anti-NK1.1 to investigate NKT cell development in vitro and in vivo. Confirming the thymus-dependence of these cells, we show that CD1d/αGC tetramer-binding NKT cells, including NK1.1+ and NK1.1− subsets, develop in fetal thymus organ culture (FTOC) and are completely absent in nude mice. Ontogenically, CD1d/αGC tetramer-binding NKT cells first appear in the thymus, at day 5 after birth, as CD4+CD8−NK1.1−cells. NK1.1+ NKT cells, including CD4+ and CD4−CD8− subsets, appeared at days 7–8 but remained a minor subset until at least 3 wk of age. Using intrathymic transfer experiments, CD4+NK1.1− NKT cells gave rise to NK1.1+ NKT cells (including CD4+ and CD4− subsets), but not vice-versa. This maturation step was not required for NKT cells to migrate to other tissues, as NK1.1− NKT cells were detected in liver and spleen as early as day 8 after birth, and the majority of NKT cells among recent thymic emigrants (RTE) were NK1.1−. Further elucidation of this NKT cell developmental pathway should prove to be invaluable for studying the mechanisms that regulate the development of these cells.

Glycolipid Antigen Drives Rapid Expansion and Sustained Cytokine Production by NK T Cells

NKT cells are enigmatic lymphocytes that respond to glycolipid Ags presented by CD1d. Although they are key immunoregulatory cells, with a critical role in immunity to cancer, infection, and autoimmune diseases, little is known about how they respond to antigenic challenge. Current theories suggest that NKT cells die within hours of stimulation, implying that their direct impact on the immune system derives from the initial cytokine burst released before their death. Here we show that NKT cell disappearance results from TCR down-regulation rather than apoptosis, and that they expand to many times their normal number in peripheral tissues within 2–3 days of stimulation, before contracting to normal numbers over subsequent days. This expansion is associated with ongoing cytokine production, biased toward a Th1 (IFN-γ+ IL-4−) phenotype, in contrast to their initial Th0 (IFN-γ+IL-4+) phenotype. This study provides critical new insight into how NKT cells can have such a major impact on immune responses, lasting many days beyond the initial stimulation of these cells.

Antigen-loaded MR1 tetramers define T cell receptor heterogeneity in mucosal-associated invariant T cells

Generation of antigen-loaded MR1 tetramers that specifically stain MAIT cells identifies heterogeneity in phenotypes and TCR repertoires in humans and mice.

NKT Cell Stimulation with Glycolipid Antigen In Vivo: Costimulation-Dependent Expansion, Bim-Dependent Contraction, and Hyporesponsiveness to Further Antigenic Challenge

Activation of NKT cells using the glycolipid α-galactosylceramide (α-GalCer) has availed many investigations into their immunoregulatory and therapeutic potential. However, it remains unclear how they respond to stimulation in vivo, which costimulatory pathways are important, and what factors (e.g., Ag availability and activation-induced cell death) limit their response. We have explored these questions in the context of an in vivo model of NKT cell dynamics spanning activation, population expansion, and subsequent contraction. Neither the B7/CD28 nor the CD40/CD40L costimulatory pathway was necessary for cytokine production by activated NKT cells, either early (2 h) or late (3 days) after initial stimulation, but both pathways were necessary for normal proliferative expansion of NKT cells in vivo. The proapoptotic Bcl-2 family member Bim was necessary for normal contraction of the NKT cell population between days 3–9 after stimulation, suggesting that the pool size is regulated by apoptotic death, similar to that of conventional T cells. Ag availability was not the limiting factor for NKT cell expansion in vivo, and a second α-GalCer injection induced a very blunted response, whereby cytokine production was reduced and further expansion did not occur. This appeared to be a form of anergy that was intrinsic to NKT cells and was not associated with inhibitory NK receptor signaling. Furthermore, NKT cells from mice prechallenged with α-GalCer in vivo showed little cytokine production and reduced proliferation in vitro. In summary, this study significantly enhances our understanding of how NKT cells respond to primary and secondary antigenic challenge in vivo.

CD1d-lipid antigen recognition by the γδ TCR

The T cell repertoire comprises αβ and γδ T cell lineages. Although it is established how αβ T cell antigen receptors (TCRs) interact with antigen presented by antigen-presenting molecules, this is unknown for γδ TCRs. We describe a population of human Vδ1+ γδ T cells that exhibit autoreactivity to CD1d and provide a molecular basis for how a γδ TCR binds CD1d–α-galactosylceramide (α-GalCer). The γδ TCR docked orthogonally, over the A′ pocket of CD1d, in which the Vδ1-chain, and in particular the germ line–encoded CDR1δ loop, dominated interactions with CD1d. The TCR γ-chain sat peripherally to the interface, with the CDR3γ loop representing the principal determinant for α-GalCer specificity. Accordingly, we provide insight into how a γδ TCR binds specifically to a lipid-loaded antigen-presenting molecule.

Recognition of [beta]-linked self glycolipids mediated by natural killer T cell antigen receptors

The most potent foreign antigens for natural killer T cells (NKT cells) are α-linked glycolipids, whereas NKT cell self-reactivity involves weaker recognition of structurally distinct β-linked glycolipid antigens. Here we provide the mechanism for the autoreactivity of T cell antigen receptors (TCRs) on NKT cells to the mono- and tri-glycosylated β-linked agonists β-galactosylceramide (β-GalCer) and isoglobotrihexosylceramide (iGb3), respectively. In binding these disparate antigens, the NKT cell TCRs docked onto CD1d similarly, achieving this by flattening the conformation of the β-linked ligands regardless of the size of the glycosyl head group. Unexpectedly, the antigenicity of iGb3 was attributable to its terminal sugar group making compensatory interactions with CD1d. Thus, the NKT cell TCR molds the β-linked self ligands to resemble the conformation of foreign α-linked ligands, which shows that induced-fit molecular mimicry can underpin the self-reactivity of NKT cell TCRs to β-linked antigens.

Thymic regeneration: teaching an old immune system new tricks

Recent studies in mice and humans show that the importance of the thymus extends well beyond the initial seeding of the peripheral T-cell pool. Although peripheral homeostasis can maintain T-cell numbers, the thymus is the major, if not the exclusive, source of new T-cell specificities. With age, thymus atrophy dramatically reduces the export of new T cells and predisposes an individual to impaired T-cell function, reduced T-cell immunity, and increased autoimmunity. Thymus atrophy is also the primary obstacle to restoration of the T-cell pool in the aftermath of HIV treatment or lymphoablative therapies. Here, we review thymus T-cell production, with particular attention to the factors that influence thymocyte export, and examine the impact that recent thymic emigrants have on the peripheral pool. In the future, thymic regeneration might become a feasible and potentially powerful approach to rejuvenating a depleted peripheral T-cell pool.

Recognition of CD1d-sulfatide mediated by a type II natural killer T cell antigen receptor

Natural killer T cells (NKT cells) are divided into type I and type II subsets on the basis of differences in their T cell antigen receptor (TCR) repertoire and CD1d-antigen specificity. Although the mode by which type I NKT cell TCRs recognize CD1d-antigen has been established, how type II NKT cell TCRs engage CD1d-antigen is unknown. Here we provide a basis for how a type II NKT cell TCR, XV19, recognized CD1d-sulfatide. The XV19 TCR bound orthogonally above the A′ pocket of CD1d, in contrast to the parallel docking of type I NKT cell TCRs over the F′ pocket of CD1d. At the XV19 TCR–CD1d-sulfatide interface, the TCRα and TCRβ chains sat centrally on CD1d, where the malleable CDR3 loops dominated interactions with CD1d-sulfatide. Accordingly, we highlight the diverse mechanisms by which NKT cell TCRs can bind CD1d and account for the distinct antigen specificity of type II NKT cells.

Identification of phenotypically and functionally heterogeneous mouse mucosal-associated invariant T cells using MR1 tetramers

Rahimpour et al. use MR1 tetramers to characterize the heterogeneous population of mouse MAIT cells and find a close resemblance to their human counterparts. These findings will provide the foundation for further investigation of MAIT cells in health and disease.