The Function of Thymic Innate TCRαß+ CD8+ T Cells Is Regulated by Constitutive Expression of B7-H1

Abstract

AbstractAbstract 955Innate T cells display features of Ag-experienced T cells (e.g., high levels of CD44) before any encounter with non-self antigens. They include NKT cells, CD8α/α T cells, as well as an undefined (and presumably very small) proportion of naïve CD4 and CD8 α/β T cells. We used the RAG2-GFP transgenic mouse model to differentiate naïve innate single-positive thymocytes (GFP+CD44high) from re-circulating effector/memory T cells (GFP-CD44high) T cells and naïve conventional (GFP+CD44low) thymocytes. We found that innate T cells represent about 10% of TCRαβ+ single-positive GFP+ thymocytes. To evaluate their proliferative ability under lymphopenic conditions, conventional and innate T cells were sorted and injected in TCRβKO mice. Four weeks later, expansion of conventional T cells was 2.3-fold greater than that of innate T cells. Comprehensive phenotypic analyses of GFP+CD8+CD44low vs GFP+CD8+CD44hi thymocytes revealed that B7-H1 was absent on conventional thymocytes but constitutively expressed on innate CD8+ thymocytes. Moreover, in vitro stimulation of innate T cells with anti-CD3 and anti-CD28 antibodies induced a strong proliferation and secretion of inflammatory mediators such as interferon (IFN)-gamma. The addition of anti-B7-H1 neutralizing antibody, however, inhibited proliferation and IFN-gamma production in a dose-dependent manner. To see whether B7-H1 ligation influenced the viability of CD8+ thymocytes, GFP+CD8+CD44low and GFP+CD8+CD44hi thymocytes were treated for 48 hrs with anti-B7-H1 antibody in vitro then stained for annexin-V and propidium iodine. Under these conditions, 70% of innate GFP+CD8+CD44hi T cells underwent apoptosis as opposed to 30% with conventional thymocytes. To test their bystander properties, we added conventional or innate T cells as third-party cells to an in vitro antigen stimulation assay. Briefly, B6 peritoneal macrophages pulsed with recombinant chicken ovalbumin (rOVA) presented OVA-derived peptides in the context of MHCI, and served as antigen-presenting cells (APCs). For responder cells, we added naive T cells from T cell receptor-transgenic OTI mice that recognize OVA. Interestingly, innate thymocytes were capable of promoting the activation of responding T cells; an effect that was lost upon treatment of innate CD8 T-cells with anti-B7-H1 neutralizing antibody. The sum of these data suggests a role for B7-H1 in the development, survival and proliferation of innate α/β T cells. AcknowledgmentsMoutih Rafei holds a Fellowship from the CIHR. Claude Perreault holds a Canada Research Chair in Immunobiology. This work was supported by grant # 42384 from the CIHR. Disclosures:No relevant conflicts of interest to declare.