Characterization of Bovine Intraepithelial T Lymphocytes in the Gut

Intraepithelial T lymphocytes (T-IELs), which constitute over 50% of the total T lymphocytes in the animal, patrol the mucosal epithelial lining to defend against pathogen invasion while maintaining gut homeostasis. In addition to expressing T cell markers such as CD4 and CD8, T-IELs display T cell receptors (TCR), including either TCRαβ or TCRγδ. Both humans and mice share similar T-IEL subsets: TCRγδ(+), TCRαβ(+)CD8αα(+), TCRαβ(+)CD4(+), and TCRαβ(+)CD8αβ(+). Among these subsets, human T-IELs are predominantly TCRαβ(+) (over 80%), whereas those in mice are mostly TCRγδ(+) (~60%). Of note, the majority of the TCRγδ(+) subset expresses CD8αα in both species. Although T-IELs have been extensively studied in humans and mice, their profiles in cattle have not been well examined. Our study is the first to characterize bovine T-IELs using flow cytometry, where we identified several distinct features. The percentage of TCRγδ(+) was comparable to that of TCRαβ(+) T-IELs (both ~50% of CD3(+)), and the majority of bovine TCRγδ(+) T-IELs did not express CD8 (CD8(−)) (above 60%). Furthermore, about 20% of TCRαβ(+) T-IELs were CD4(+)CD8αβ(+), and the remaining TCRαβ(+) T-IELs were evenly distributed between CD4(+) and CD8αβ(+) (~40% of TCRαβ(+) T-IELs each) with no TCRαβ(+)CD8αα(+) identified. Despite these unique properties, bovine T-IELs, similar to those in humans and mice, expressed a high level of CD69, an activation and tissue-retention marker, and a low level of CD62L, a lymphoid adhesion marker. Moreover, bovine T-IELs produced low levels of inflammatory cytokines such as IFNγ and IL17A, and secreted small amounts of the immune regulatory cytokine TGFβ1. Hence, bovine T-IELs’ composition largely differs from that of human and mouse, with the dominance of the CD8(−) population among TCRγδ(+) T-IELs, the substantial presence of TCRαβ(+)CD4(+)CD8αβ(+) cells, and the absence of TCRαβ(+)CD8αα(+) T-IELs. These results provide the groundwork for conducting future studies to examine how bovine T-IELs respond to intestinal pathogens and maintain the integrity of the gut epithelial barrier in animals.