Fibroblast dependency during early thymocyte development maps to the CD25⁺ CD44⁺ stage and involves interactions with fibroblast matrix molecules

Abstract

We have investigated the role of specific components of the thymic stroma during development of CD4-8-T cell precursors by separating and reaggregating precursor subsets with individual or combinations of stromal cells. We show that while the development of CD25+ 44+ precursors is dependent upon a combination of major histocompatibility complex (MHC) class II+ thymic epithelial cells and fibroblasts, their direct descendants, CD25+ 44- precursors, develop to the CD4+ 8+ stage in the presence of MHC class II+ thymic epithelial cells alone. Thus, CD25+ 44+ precursors are the last developmental stage to be dependent upon fibroblast support. In addition, while metabolically inactive, 1-ethyl-3-(3'-dimethylaminopropyl) carbodiimide (ECDI)-treated fibroblasts retain the ability to promote T cell development, prior treatment with hyaluronidase abrogates this effect, suggesting that fibroblast-associated extracellular matrix components are the key elements involved. In support of this, we show that fibroblasts are located in cortical regions of the thymus where T cell precursors are known to reside, and that these fibroblasts are associated with an extensive extracellular matrix not found on thymic epithelial cells. Finally, antibodies to alpha 4 integrin and CD44 interfere with the efficiency with which CD4+ 8+ cells are generated from CD25+ 44+ precursors in reaggregate cultures and also reduce the binding of the latter to 3T3 fibroblasts, suggesting these molecules play a role in bringing T cell precursors into contact with fibroblast-associated extracellular matrix.

A role for CD44 in T cell development and function during direct competition between CD44+ and CD44– cells

“…However, no obvious defect in progression from the the DN1 to the DN4 stage was detected. Nevertheless, CD44 is reported to play a role in mediating the interaction between early thymocytes and thymic stroma [27,28], while contact with notch ligands expressed on stromal cells at both DN1 and DN2 stages has been shown to be required for induction and maintenance of T cell lineage specification [29]. Hence, CD44-mediated cell movement or adhesion could contribute to the delivery of signals involved in early thymocyte development.…”

Regulation of thymic epithelium by keratinocyte growth factor

“…Although not demonstrated here, fibroblasts also are a source of KGF in the thymus as they are in other tissues (Revest et al 14 and vida infra). The well-established requirement for fibroblasts in thymic organogenesis and thymic function in vitro 5,54 likely reflects their elaboration of mediators of epithelial/mesenchymal interaction, including KGF. Thymocyte production of KGF may provide a feedback mechanism to more precisely regulate levels of this factor within the intrathymic milieu.…”

Stromal Cells Provide the Matrix for Migration of Early Lymphoid Progenitors Through the Thymic Cortex

“…3), it is conceivable that DN and DP cells may use quite different stromal cell types, despite being present in the same cortical regions. Likewise, the ECM plays a required role in the differentiation, proliferation, and/or survival of lymphoid progenitors (39), either by direct signaling to the progenitors themselves, or through the organization of other stromal cells in their corresponding microenvironments.…”