T cell receptor expression on immature thymocytes with in vivo and in vitro precursor potential

Immature CD8^-CD4^- double-negative (DN) thymocytes differentiate intrathymically into CD8⁺CD4⁺ and CD8^-CD4⁺ thymocytes and migrate to the periphery. This differentiation proceeds through several intermediate phenotypic changes in the expression of CD8 and CD4. We have recently established the existence of a CD8^loCD4^lo cell population in murine thymus that can repopulate the irradiated thymus in vivo and differentiate rapidly in vitro to CD8⁺CD4⁺ double-positive (DP) cells. The CD8^loCD4^lo cells score as DN upon direct cytofluorometric analysis, yet are distinct from true DN cells by various criteria. Experimental evidence strongly suggests that they are descendants of true DN in the maturation pathway. In the experiments presented here, we further characterize this CD8^loCD4^lo thymocyte population. Northern blot and RNA protection analysis reveal that these cells transcribe full length mRNA for the T cell receptor (TcR)α chain, unlike the less mature interleukin 2 receptor-positive DN thymocytes. Surface expression of the TcR-associated CD3 molecule occurs on ∼ 15% of these cells at low levels characteristic of immature cells. In the course of in vitro differentiation a vast majority (∼ 80%) of these cells convert to CD8⁺CD4⁺ and significant numbers of the brightly staining DP convertants (11%–34% on day 1 and 48%–68% on day 2) express immature levels of CD3. Our results indicate that CD8^lo, CD4^lo cells might be the first thymic subset to rearrange TcR α chain genes and express TcR α/β heterodimer on the surface at levels characteristic of immature cells. Furthermore, the surface expression of TcR persists on the in vitro progeny of these thymocytes.