Adhesion of immature thymocytes to thymic stromal cells through fibronectin molecules and its significance for the induction of thymocyte differentiation.

Abstract

Only 10-15% of unseparated thymocytes adhered to culture plates precoated with fibronectin (FN), but 60-70% of the CD4-8-(double-negative) thymocyte population did. This population bound to FN but not to collagen, laminin, or vitronectin. Its binding to FN was inhibited by anti-FN antibody or a mixture of synthetic peptides corresponding to two different sites of EN, termed the V10 sequence and the RGDS (Arg-Gly-Asp-Ser) sequence, which interact, respectively, with the VLA-4 and VLA-5 FN receptors expressed on T-lineage cells. CD4-8-thymocytes also adhered to a monolayer of a thymic stromal cell clone, MRL104.8a, that induces growth-maintenance and differentiation of such thymocytes. . Most important, blocking the adhesion of CD4-8-thymocytes to the thymic stromal cell monolayer resulted in potent inhibition of the differentiation of these thymocytes, which was otherwise induced toward the expression of CD4 and/or CD8 molecules. These results indicate that immature CD4-8-thymocytes adhere to thymic stromal cells preferentially through FN-FN receptor interaction and that such adhesion has a critical role in inducing and/or supporting the differentiation of these thymocytes.Lymphocytes express a number of surface molecules that mediate the adhesion of cells to one another or to extracellular matrix (ECM) components (1-4). For example, cell-cell interaction molecules such as CD2 and LFA-3 are used to augment specific adhesion of T cells to antigen-presenting cells (5), and other cell-cell interaction molecules function as homing receptors (6, 7). Thus, these cell surface molecules have been recognized as regulating the migration of lymphocytes as well as the interactions of activated lymphocytes during immune responses.Another type of cell adhesion mechanism involves the interaction of cells with ECM (2, 4). While the importance of such cell-cell interactions in developmental biology has been documented (8-12), few studies have investigated the role of cell-ECM adhesion mechanisms in lymphocyte development in lymphopoietic microenvironments such as the bone marrow and thymus. This may be ascribed to the lack of in vitro systems for such analyses and to the complexity arising from the expression of various forms of ECM molecules regulated in a tissue-and cell-specific fashion, as exemplified by fibronectin (FN) molecules (13-15). In this context, several lines of marrow and thymic stromal cells have been isolated to provide tools for analyzing the interaction between developing lymphocytes and stromal cells (16). We have also established a thymic stromal cell clone that is capable of providing an in vitro model for intrathymic T-cell development (17-19).The present study investigates the role of FN molecules expressed on thymic stromal cells in thymocyte-stromal cell adhesion and thymocyte differentiation. The results demonstrate that CD4-8-("double-negative") thymocytes adhere to thymic stromal cells through FN molecules on the stromal cells. Molecular analyses revealed that two adhesion sites, the classic...

Direct association of pp125FAK with paxillin, the focal adhesion-targeting mechanism of pp125FAK.

“…Several laboratories including ours have shown that [31 integrins synergize with the TCR-CD3 pathway to promote T cell proliferation (4-6). [31 integrins are also reported to be involved in T or B cell differentiation through interaction with fibronectin expressed by stromal cells in thymus or bone marrow, respectively (7,8). Furthermore, it was also reported that integrin-ECM binding induced increases of cytoplasmic Ca 2+ and cytoplasmic pH (9, 10).…”

Role of beta 1 and beta 2 integrins in the adhesion of human CD34hi stem cells to bone marrow stroma.

“…The influence of fibronectin on proliferation and differentiation of erythroid cells has been documented (75,76). Adhesion of CD4-CD8 -thymocytes to thymic stromal cells through FN-FN receptor interaction is known to be critical for their differentiation (77 Disruption ofnormal adhesion pathways between stem and stromal cells has been implicated as a critical factor in pathogenesis of chronic myeloid leukemia (81,82). Furthermore, after chemotherapy and/or total body irradiation, the observed extravascation of stem cells into peripheral blood (83) …”

Integrin Functions Play a Key Role in the Differentiation of Thymocytes In Vivo

“…Previously, in vitro systems have shown a requirement for integrins and integrin-related proteins for the maturation of CD4 Ϫ CD8 Ϫ to CD4 ϩ CD8 ϩ cells (12)(13)(14). Most of the in vitro data in mice point to the CD4 Ϫ CD8 Ϫ cells as being adhesive cells with activated integrins.…”

“…A number of in vitro studies have demonstrated that the engagement of integrins is required for the differentiation of CD4 Ϫ CD8 Ϫ cells to become CD4 ϩ CD8 ϩ cells (12)(13)(14). Furthermore, the Abmediated engagement of integrin receptors in addition to the engagement of the CD3 complex on CD4 ϩ CD8 ϩ thymocytes is required for cell proliferation in vitro (15,16).…”