Sungyoo Cho

Selective Loss of Natural Killer T Cells by Apoptosis following Infection with Lymphocytic Choriomeningitis Virus

ABSTRACT Natural killer T (NKT) cells, a unique subpopulation of T cells, coexpress markers also present on NK cells and recognize the major histocompatibility complex class I-like CD1d1 molecule. We studied the effect of an acute virus infection on NKT cells. Mice were infected with the nonhepatotropic Armstrong strain of lymphocytic choriomeningitis virus (LCMV), and at various times postinfection, mononuclear cells from the liver, peritoneum, and spleen were isolated. It was found that within 2 to 3 days, there was a selective loss of NKT cells from the liver with an apparent rapid recovery within 8 to 14 days. There was no increase in peritoneal or splenic NKT cells, indicating that NKT cells did not traffic to these tissues. This loss of NKT cells was independent of gamma interferon (IFN-γ) and interleukin 12 (IL-12) production, but did occur in mice treated with poly(I-C), a classical inducer of IFN-α/β. The reduction in NKT cells was CD28 and fas/fasL independent and occurred via apoptosis. It was not observed in LCMV-infected DNA fragmentation factor 45-deficient mice, and an increase in active caspase 3-specific staining was found in liver NKT cells from LCMV-infected and poly(I-C)-treated mice compared to uninfected wild-type mice. Interestingly, it was also found that liver NKT cells from LCMV-infected mice were themselves infected. These results suggest that the loss of NKT cells following an acute LCMV infection could be due to the induction of IFN-α/β resulting in NKT-cell apoptosis and is important for the hosts immune response to LCMV.

Inhibition of glycolipid shedding rescues recognition of a CD1+ T cell lymphoma by natural killer T (NKT) cells.

Neoplastic transformation of cells is accompanied by an aberration of cell surface glycolipid composition. These tumor-associated, altered glycosphingolipids are often shed into the tumor cell microenvironment and mediate immunosuppressive activity. The nature and form of glycolipids shed by a variety of tumor cell lines and the mechanism(s) of shedding have been well characterized. The murine T cell lymphoma line, L5178Y-R, is known to shed a tumor-associated glycolipid, gangliotriaosylceramide, into the culture medium. We analyzed the effect of glycolipids from L5178Y-R on antigen presentation by murine CD1d1 molecules. CD1d1 molecules present glycolipid antigens to a specialized class of T cells called natural killer T (NKT) cells that mainly express a T cell receptor α chain (Vα14Jα281) associated with Vβ chains of limited diversity. In the current report, we found that L5178Y-R cells express CD1 on their cell surface yet are unable to stimulate CD1d1-specific NKT cells. We hypothesized that the glycolipid(s) shed by L5178Y-R inhibited antigen presentation by CD1d1. Pretreatment of CD1d1+ cells with conditioned medium from L5178Y-R inhibited CD1-specific stimulation of canonical (Vα14+) but not noncanonical (Vα5+) NKT cells. Exogenous addition of lipids extracted from L5178Y-R cells as well as purified gangliotriaosylceramide mimicked this effect. Inhibition of glycolipid shedding in L5178Y-R cells with d-1-phenyl-2-hexadecanoylamino-3-morpholino-1-propanol resulted in the rescue of CD1d1 recognition by canonical (but not noncanonical) NKT cells. These results suggest that one means by which certain tumor cells can evade the hosts innate antitumor immune response is by shedding glycolipids that inhibit CD1-mediated antigen presentation to NKT cells.

Myeloid marker expression on antiviral CD8+ T cells following an acute virus infection

CD11b, CD11c, and F4/80 are normally used to define dendritic cell and/or macrophage populations. In this study, the expression of all three markers was observed on CD8+ T cells following infection of mice with several distinct viruses. Using lymphocytic choriomeningitis virus as a model virus, it was found that relatively more CD11b+CD8+ and CD11c+CD8+ T cells were present in the periphery than in primary lymphoid organs; in contrast, the F4/80+CD8+ T cell population was more prevalent in the spleen. All three myeloid markers were detected on virus‐specific CTL. The expression of CD11b and CD11c on CD8+ T cells correlated with their level of CTL activity, whereas the F4/80+CD8+ T cell population increased after the peak of the CTL response but did not have higher CTL activity. These data suggest that there is a differential induction of CD11b, CD11c, and F4/80 on virus‐specific CD8+ T cells following an acute virus infection.

Long‐term loss of canonical NKT cells following an acute virus infection

NKT cell activation plays an important role in regulating innate and adaptive immunity during infection. We have previously found that there is a dramatic reduction in the NKT cell population on day 3 after an acute lymphocytic choriomeningitis virus (LCMV) infection. In this study, we report that this loss continued for at least 3 months and was not simply due to internalization of the TCR. Concomitant with the decrease in NKT cells was an increase in the percentage of Annexin V+ NKT cells that remained in vivo, suggesting that the reduction in NKT cells at these late stages post‐infection occurred by activation‐induced cell death. Interestingly, APC from LCMV‐infected mice could activate NKT cells in vitro at higher levels than those from uninfected mice and was concomitant with an increase in apoptosis in NKT cells. However, this could not be blocked by mAb to murine CD1d, and APC from LCMV‐infected (but not uninfected) CD1d1‐deficient mice could also stimulate NKT cells. Collectively, our data suggest that the activation and subsequent long‐term loss of NKT cells is a normal component of the hosts antiviral immune response, and this occurs in a CD1d‐independent manner.

A threonine-based targeting signal in the human CD1d cytoplasmic tail controls its functional expression.

CD1d molecules are MHC class I-like molecules that present lipids to a unique subpopulation of T cells called NKT cells. The cytoplasmic tail of human CD1d possesses a tyrosine-based endosomal targeting motif (YXXZ). As such, these molecules traffic through the endocytic pathway, where it is believed that they are loaded with the antigenic lipid that stimulates NKT cells. In the current study, it was found that the T322 residue in the human CD1d tail is a major signal controlling transport to the cell surface and thus its functional expression. Mimicking the phosphorylation of this residue or removal of the entire cytoplasmic tail negates its ability to regulate CD1d trafficking, resulting in lysosomal targeting and degradation. These results demonstrate an important role of a heretofore unknown signal in the cytoplasmic tail of CD1d that may have relevance to other type I integral membrane proteins that traverse through the endocytic pathway.