Thymic precursors of TCRαβ+CD8αα+ intraepithelial lymphocytes are negative for CD103

CD5+TCRαβ+ cells in CD4−CD8− double negative (DN) thymocytes are generally regarded as the thymic precursors of TCRαβ+CD8αα+ intestinal intraepithelial lymphocytes (IELs). However, this population is not homogenous and can be subdivided based on the expression of cell surface markers such as CD103. In this study, we aimed to define a cell population that is enriched in thymic IEL precursors. Here we report that only CD103− but not CD103+cells in the CD5+TCRαβ+ DN thymocyte population can give rise to TCRαβ+CD8αα+ IELs or IEL-like cells in in vivo injections and in vitro cultures, respectively. In addition, we demonstrate that IL-15 stimulation alone is sufficient for upregulation of CD8αα in CD103−CD5+TCRαβ+ DN thymocytes. We also found that the CD103−CD5+TCRαβ+ DN population can be further separated into two fractions: CD69−/lo and CD69+. Of these two fractions, only CD69−/lo cells can give rise to CD8αα IEL-like cells in the presence of IL-15 in in vitro cultures. Based on these results, we conclude that a CD69−/loCD103−CD5+TCRαβ+ DN population is highly enriched in thymic IEL precursors.