Developmental status of CD4-CD8+ and CD4+CD8- thymocytes with medium expression of CD3

In normal mice, more than 10% of thymocytes in the CD4+CD8- and CD4-CD8+ single-positive (SP) subsets express a medium level of CD3 on the cell surface. However, the fate of CD3medium cells is unclear. The CD3medium SP subpopulations might contain (i) cells in an immature stage of the pathways leading to CD3high cells, (ii) cells in developmental pathways that do not lead to CD3high cells, or (iii) cells that have been negatively selected. We found that sorted CD3medium CD4+CD8- thymocytes from adult mice up-regulated CD3 to high levels in reaggregation thymus organ culture. Unlike their CD3high counterparts, CD3medium CD4+CD8- thymocytes were unable to undergo chemotaxis towards the chemokines CCL19 and CCL21. CD3medium thymocytes of both CD4+CD8- and CD4-CD8+ subsets were also considerably more responsive than CD3high SP cells to apoptotic signals induced in vitro by ligation of CD95 (Fas/APO-1) or by dexamethasone. In both SP subsets, a higher frequency of thymocytes expressing forbidden Vbeta+ T cell receptors reactive with endogenous mammary tumor virus superantigens was found in CD3medium subpopulations than in CD3high subpopulations. These findings argue that the CD3medium SP thymocyte subpopulations contain apoptosis-susceptible precursor cells of CD3high SP cells and are subject to negatively selecting pressures.