Barbara A. Sullivan

Mechanisms for Glycolipid Antigen-Driven Cytokine Polarization by Vα14i NKT Cells

Certain glycolipid Ags for Vα14i NKT cells can direct the overall cytokine balance of the immune response. Th2-biasing OCH has a lower TCR avidity than the most potent agonist known, α-galactosylceramide. Although the CD1d-exposed portions of OCH and α-galactosylceramide are identical, structural analysis indicates that there are subtle CD1d conformational differences due to differences in the buried lipid portion of these two Ags, likely accounting for the difference in antigenic potency. Th1-biasing C-glycoside/CD1d has even weaker TCR interactions than OCH/CD1d. Despite this, C-glycoside caused a greater downstream activation of NK cells to produce IFN-γ, accounting for its promotion of Th1 responses. We found that this difference correlated with the finding that C-glycoside/CD1d complexes survive much longer in vivo. Therefore, we suggest that the pharmacokinetic properties of glycolipids are a major determinant of cytokine skewing, suggesting a pathway for designing therapeutic glycolipids for modulating invariant NKT cell responses.

Activation or anergy: NKT cells are stunned by α-galactosylceramide

Invariant natural killer T (iNKT) cells are T lymphocytes that behave similarly to cells of the innate immune system. The glycolipid α-galactosylceramide (α-GalCer) is a potent and specific activator of mouse and human iNKT cells and has been used in cancer clinical trials to drive NKT cell–mediated immune responses. However, little is known about the dynamics of the iNKT cell response to α-GalCer in vivo. In this issue of the JCI, Parekh and colleagues demonstrate that administration of α-GalCer causes iNKT cells to become unresponsive, for at least 1 month, in mice. This leads us to ask, should sequential administration of α-GalCer still be used to activate iNKT cells given the anergic state it has been shown here to induce? This intriguing article raises the issue of the avoidance of anergy induction in the design of treatment regimens that use α-GalCer as a specific activator of iNKT cells.

Co-receptor choice by Vα14i NKT cells is driven by Th-POK expression rather than avoidance of CD8-mediated negative selection

Mouse natural killer T (NKT) cells with an invariant Vα14-Jα18 rearrangement (Vα14 invariant [Vα14i] NKT cells) are either CD4+CD8− or CD4−CD8−. Because transgenic mice with forced CD8 expression in all T cells exhibited a profound NKT cell deficit, the absence of CD8 has been attributed to negative selection. We now present evidence that CD8 does not serve as a coreceptor for CD1d recognition and that the defect in development in CD8 transgene homozygous mice is the result of a reduction in secondary T cell receptor α rearrangements. Thymocytes from mice hemizygous for the CD8 transgene have a less severe rearrangement defect and have functional CD8+ Vα14i NKT cells. Furthermore, we demonstrate that the transcription factor Th, Poxviruses and Zinc finger, and Krüppel family (Th-POK) is expressed by Vα14i NKT cells throughout their differentiation and is necessary both to silence CD8 expression and for the functional maturity of Vα14i NKT cells. We therefore suggest that Th-POK expression is required for the normal development of Vα14i NKT cells and that the absence of CD8 expression by these cells is a by-product of such expression, as opposed to the result of negative selection of CD8-expressing Vα14i NKT cells.

A CD1d-Dependent Antagonist Inhibits the Activation of Invariant NKT Cells and Prevents Development of Allergen-Induced Airway Hyperreactivity

The prevalence of asthma continues to increase in westernized countries, and optimal treatment remains a significant therapeutic challenge. Recently, CD1d-restricted invariant NKT (iNKT) cells were found to play a critical role in the induction of airway hyperreactivity (AHR) in animal models and are associated with asthma in humans. To test whether iNKT cell-targeted therapy could be used to treat allergen-induced airway disease, mice were sensitized with OVA and treated with di-palmitoyl-phosphatidyl-ethanolamine polyethylene glycol (DPPE-PEG), a CD1d-binding lipid antagonist. A single dose of DPPE-PEG prevented the development of AHR and pulmonary infiltration of lymphocytes upon OVA challenge, but had no effect on the development of OVA-specific Th2 responses. In addition, DPPE-PEG completely prevented the development of AHR after administration of α-galactosylceramide (α-GalCer) intranasally. Furthermore, we demonstrate that DPPE-PEG acts as antagonist to α-GalCer and competes with α-GalCer for binding to CD1d. Finally, we show that DPPE-PEG completely inhibits the α-GalCer–induced phosphorylation of ERK tyrosine kinase in iNKT cells, suggesting that DPPE-PEG specifically blocks TCR signaling and thus activation of iNKT cells. Because iNKT cells play a critical role in the development of AHR, the inhibition of iNKT activation by DPPE-PEG suggests a novel approach to treat iNKT cell-mediated diseases such as asthma.

Immediate antigen-specific effector functions by TCR-transgenic CD8+ NKT cells.

Only recently have natural antigens for CD1d‐dependent, invariant Vα14+ natural killer T (iNKT) cells been identified. Similar data for CD1d‐independent and CD8+ NKT cell populations are still missing. Here, we show that the MHC class I‐restricted CD8+ TCR‐transgenic mouse lines OT‐I, P14 and H‐Y contain a significant proportion of transgenic CD8+ NK1.1+ T cells. In liver, most of NK1.1+ T cells express CD8αα homodimers. Transgenic NKT cells did not bind invariant Vα14‐to‐Jα18 TCR rearrangement (Vα14i)‐specific CD1d/α‐galactosylceramide tetramers and the frequency of iNKT cells was severely reduced. The activated cell surface phenotype and the distribution of transgenic NKT cells in vivo were similar to that reported for iNKT cells. The OT‐I and P14 CD8+ NKT cells recognized their cognate antigen in the context of H2‐Kb and produced cytokines shortly after TCR stimulation. Importantly, transgenic NKT cells exerted immediate antigen‐specific cytotoxicity in vitro and in vivo. Our results demonstrate the presence of transgenic CD8+ NKT cells in MHC class I‐restricted TCR‐transgenic animals, which are endowed with rapid antigen‐specific effector functions. These data imply that experiments studying naive T cell function in TCR‐transgenic animals should be interpreted with caution, and that such animals could be utilized for studying CD8+ NKT cell function in an antigen‐specific manner.

Differential Requirement for Tapasin in the Presentation of Leader- and Insulin-Derived Peptide Antigens to Qa-1b-Restricted CTLs

The loading of MHC class I molecules with peptides involves a variety of accessory proteins, including TAP-associated glycoprotein (tapasin), which tethers empty MHC class I molecules to the TAP peptide transporter. We have evaluated the role of tapasin for the assembly of peptides with the class Ib molecule Qa-1b. In normal cells, Qa-1b is predominantly bound by a peptide, the Qa-1 determinant modifier (Qdm), derived from the signal sequence of class Ia molecules. Our results show that tapasin links Qa-1b to the TAP peptide transporter, and that tapasin facilitates the delivery of Qa-1b molecules to the cell surface. Tapasin was also required for the presentation of endogenous Qdm peptides to Qdm-specific, Qa-1b-restricted CTLs. In sharp contrast, tapasin expression was dispensable for the presentation of an insulin peptide to insulin-specific, Qa-1b-restricted CTL isolated from TCR transgenic mice. However, tapasin deficiency significantly impaired the positive selection of these insulin-specific, Qa-1b-restricted transgenic CD8+ T cells. These findings reveal that tapasin plays a differential role in the loading of Qdm and insulin peptides onto Qa-1b molecules, and that tapasin is dispensable for retention of empty Qa-1b molecules in the endoplasmic reticulum, and are consistent with the proposed peptide-editing function of tapasin.

Acid Test: Lipid Antigens Get into the Groove

How do CD1 molecules load lipid antigens? In this issue of Immunity, Relloso et al. (2008) uncover how lysosomal pH targets amino acids in CD1b, causing it to open and attain a conformation more receptive to lipid antigens.