T cell development in mouse thymus : studies on lymphostromal interactions

T lymphocytes, the effectors of cell-mediated immunity, are concerned with the control of intracellular infections: cytotoxic T lymphocytes recognize and destroy vi rally infected cells, whereas helper T lymphocytes, through lymphokines, may activate macrophages in killing intracellular parasites. The parasitaljviral antigens are recognized by the T lymphocytes in association with molecules of the major histocompatibility complex (MHC). This ability, to recognize antigens in the context of self-MHC molecules, also termed MHC-restriction, is learned by the T lymphocytes during their development in the thymus. On their pathway to development (maturation), T cells are differentially influenced by various thymic stromal cells. The stromal cells constitute microenvironments, where developing T cells, through cell-cell interactions and locally secreted cytokines, receive signals to proliferate and mature. This thesis focuses on the specific role of thymic microenvironments in promoting sequential stages in T cell development in the murine thymus. Our understanding of these lymphostromal interactions will be introduced in this chapter, containing three major sections. The first section on "Development and architecture of thymic microenvironments" discusses the variety of thymic microenvironments that have presently been identified. The phenotypic characterization of thymic microenvironments, presented in this section, is more extensively reviewed in chapter 2. The second section on ''T cell differentiation" introduces a major pathway of T cell differentiation, including the a/3 and the yo T cell lineages, showing the complexity of this process. The third section on "Lymphostromal interactions" links the different types of stromal cells to distinct stages ofT cell development. The chapter ends with an "Introduction to the experimental work"

Inhibition of proliferation and differentiation during early T cell development by anti-transferrin receptor antibody

Proliferating cells require iron and, therefore, express the transferrin receptor (CD71) that mediates cellular iron uptake. Cycling thymocytes, which have the CD4−8−3−, CD4−8+3−, or CD4+8+3− phenotypes, also express CD71. The importance of CD71-mediated iron uptake for proliferation and maturation of thymocytes was studied using fetal thymus organ cultures at day 14 of gestation and treating them for 7 days with a CD71 monoclonal antibody (mAb). The intracellular iron deficiency caused by this treatment, inhibits both proliferation and maturation of the thymocytes. Cell recovery was reduced by 60%, but cells still expanded tenfold during the culture. Remarkably, the final maturation of αβ T cells was completely blocked as no thymocytes with low or high CD3/αβTcR expression developed. Moreover, only few cells reached the CD4+8+3− stage of T cell development. CD4−8−3− thymocytes, however, as well as its CD44−25+ subset developed in normal numbers, suggesting that CD44−25+ CD4−8−3− cells, or their immediate progeny, were most vulnerable to CD71 mAb treatment. The development of γδ T cells, which also express CD71, was not affected in these cultures. This suggests that γδ T cells are either less iron-dependent or possess alternative iron-uptake mechanisms. Thus, our observation

Transferrin receptor expression as a marker of immature cycling thymocytes in the mouse

Dividing cells require iron and, therefore, express the transferrin receptor (CD71) on the cell surface to enable internalization of transferrin-bound iron. Since early T cell development is marked by intense proliferation, we questioned whether CD71 might serve as a marker of immature T cells. Therefore, we analyzed the expression of CD71 on fetal, neonatal, and adult thymocytes in correlation with cell size, cell cycle status, and expression of CD3, CD4, CD8, αβTcR, and γδTcR. Phenotypic analysis showed that only the large, immature CD4-8-3-, CD4-8+3-, and CD4+8+3- cells in fetal, neonatal, and adult thymus expressed CD71. In addition, DNA analysis showed that all CD71+ large adult thymocytes were cycling. Downregulation of CD71 occurs when proliferation ceases, i.e., within the CD4+8+3- thymocyte subpopulation. The gradual changes in size and CD71 expression suggest a sequential development within this CD4+8+3- subpopulation from large CD71+ via small CD71± to small CD71- cells. As a consequence, CD71 expression is downregulated, in adult T cell development as well as in ontogeny, before the αβTcR appears on the cell surface of the thymocyte. Together, our findings show that CD71 is a marker of immature, proliferating T cells