Rosa Sacedón

Bone Morphogenetic Protein 2/4 Signaling Regulates Early Thymocyte Differentiation

Bone morphogenetic protein (BMP)2 and BMP4 are involved in the development of many tissues. In this study, we show that BMP2/4 signaling is involved in thymocyte development. Our data suggest that termination of BMP2/4 signaling is necessary for differentiation of CD44+CD25−CD4−CD8− double negative (DN) cells along the T cell lineage. BMP2 and BMP4 are produced by the thymic stroma and the requisite BMP receptor molecules (BMPR-1A, BMPR-1B, BMPR-II), and signal transduction molecules (Smad-1, -5, -8, and -4) are expressed by DN thymocytes. BMP4 inhibits thymocyte proliferation, enhances thymocyte survival, and arrests thymocyte differentiation at the CD44+CD25− DN stage, before T cell lineage commitment. Neutralization of endogenous BMP2 and BMP4 by treatment with the antagonist Noggin promotes and accelerates thymocyte differentiation, increasing the expression of CD2 and the proportion of CD44−CD25− DN cells and CD4+CD8+ double-positive cells. Our study suggests that the BMP2/4 pathway may function in thymic homeostasis by regulating T cell lineage commitment and differentiation.

Sonic Hedgehog Is Produced by Follicular Dendritic Cells and Protects Germinal Center B Cells from Apoptosis

The Hedgehog (Hh) signaling pathway is involved in the development of many tissues during embryogenesis, but has also been described to function in adult self-renewing tissues. In the immune system, Sonic Hedgehog (Shh) regulates intrathymic T cell development and modulates the effector functions of peripheral CD4+ T cells. In this study we investigate whether Shh signaling is involved in peripheral B cell differentiation in mice. Shh is produced by follicular dendritic cells, mainly in germinal centers (GCs), and GC B cells express both components of the Hh receptor, Patched and Smoothened. Blockade of the Hh signaling pathway reduces the survival, and consequently the proliferation and Ab secretion, of GC B cells. Furthermore, Shh rescues GC B cells from apoptosis induced by Fas ligation. Taken together, our data suggest that Shh is one of the survival signals provided by follicular dendritic cells to prevent apoptosis in GC B cells.

The role of morphogens in T-cell development

The Hedgehog (Hh) and Wnt family proteins, and the bone morphogenetic proteins (BMPs) 2 and 4, act as morphogens during vertebrate embryogenesis and organogenesis by regulating patterning and cell fate. They have recently been found to have a role in regulating cell fate and determination in self-renewing tissues in adults, such as the immune system and haematopoietic system. This Review presents studies on the role of Sonic Hh (Shh), Wnts and BMP2/4 in the regulation of thymocyte development. Shh and BMP2/4 act as negative regulators of thymocyte development. By contrast, Wnt signalling, through beta-catenin, has a positive role in the control of T-cell development, such that an absence or reduction in the Wnt signal leads to a reduction in cell number and cell proliferation rate and differentiation to the CD4+CD8+ double-positive stage.

Expression of hedgehog proteins in the human thymus.

The Hedgehog (Hh) family of secreted proteins includes intercellular signaling molecules that specify cell fate and patterning during the development of many tissues. In this study we show that the different components of the Hh signaling pathway are expressed in human thymus. The three mammalian Hh proteins, Sonic (Shh), Indian (Ihh), and Desert (Dhh) hedgehog, are produced by thymic epithelial cells. Shh-expressing epithelial cells are restricted to the thymic subcapsula and medulla, whereas Ihh- and Dhh-producing epithelial cells are distributed throughout the thymus. The requisite Hh receptors, Patched 1(Ptc1) and Smoothened (Smo), and the Gli transcription factors are expressed by thymocytes and also by epithelial cells. Ptc1 is expressed in most thymocyte subsets, whereas Smo expression is mainly associated with immature thymocytes. The isoform of the Ptc receptor, Ptc2, is expressed only by intrathymic progenitor cells and epithelial cells. Other Hh-binding proteins with modulating functions, such as Hedgehog-interacting protein (Hip) and growth arrest-specific gene-1 (Gas-1), are also expressed in human thymus. Our study shows that the intrathymic expression pattern of the Hh signaling pathway components is complex and suggests that Hh proteins may regulate human thymocyte differentiation from the earliest developmental stages, as well as thymic epithelial cell function.

Expression and Function of the Eph A Receptors and Their Ligands Ephrins A in the Rat Thymus

Thymus development and function are dependent on the definition of different and graded microenvironments that provide the maturing T cell with the different signals that drive its maturation to a functional T lymphocyte. In these processes, cell-cell interactions, cell migration, and positioning are clues for the correct functioning of the organ. The Eph family of receptor tyrosine kinases and their ligands, the ephrins, has been implicated in all these processes by regulating cytoskeleton and adhesion functioning, but a systemic analysis of their presence and possible functional role in thymus has not yet been conducted. In this regard, the current study combines different experimental approaches for analyzing the expression of four members of the Eph A family and their ligands, ephrins A, in the embryonic and adult rat thymus. The patterns of Eph and ephrin expression in the distinct thymic regions were different but overlapping. In general, the studied Eph A were expressed on thymic epithelial cells, whereas ephrins A seem to be more restricted to thymocytes, although Eph A1 and ephrin A1 are expressed on both cell types. Furthermore, the supply of either Eph A-Fc or ephrin A-Fc fusion proteins to fetal thymus organ cultures interferes with T cell development, suggesting an important role for this family of proteins in the cell mechanisms that drive intrathymic T cell development.

Distinct Mechanisms Contribute to Generate and Change the CD4:CD8 Cell Ratio During Thymus Development: A Role for the Notch Ligand, Jagged1

In adult life, the high CD4:CD8 cell ratio observed in peripheral lymphoid organs originates in the thymus. Our results show that the low peripheral CD4:CD8 cell ratio seen during fetal life also has an intrathymic origin. This distinct production of CD4+CD8− and CD4−CD8+ thymocytes is regulated by the developmental age of the thymic stroma. The differential expression of Notch receptors and their ligands, especially Jagged1, throughout thymus development plays a key role in the generation of the different CD4:CD8 cell ratios. We also show that the intrathymic CD4:CD8 cell ratio sharply changes from fetal to adult values around birth. Differences in the proliferation and emigration rates of the mature thymocyte subsets contribute to this change.

Sonic Hedgehog Regulates Early Human Thymocyte Differentiation by Counteracting the IL-7-Induced Development of CD34+ Precursor Cells

The Hedgehog (Hh) family of signaling molecules normally functions in the development of numerous tissues by regulating cellular differentiation and proliferation. Recent results have demonstrated that the different components of the Hh signaling pathway are expressed in the human thymus. In this study, we investigate the potential role of Sonic hedgehog (Shh) in human intrathymic T cell maturation. Results show that the expression of the two components of the Hh receptor, Patched and Smoothened, is mostly restricted to CD34+ precursor cells that are committing to the T cell lineage. Shh significantly increased the viability of CD34+ T cell precursors modulating bcl-2 and bax protein expression, and also inhibited their proliferation. The treatment of chimeric human-mouse fetal thymus organ cultures with Shh resulted in an arrested thymocyte differentiation and an accumulation of CD34+ progenitor cells. This effect was mainly attributed to the ability of Shh to counteract the IL-7-induced proliferation and differentiation of CD34+ cells. Shh down-regulated in the precursor cell population the expression of IL-7R as well as stromal-derived factor-1 chemokine receptor, CXCR4, and inhibited IL-7-dependent STAT5 phosphorylation. Therefore, Shh may function as a maintenance factor for intrathymic CD34+ precursor cells.

Rat Peripheral CD4+CD8+ T Lymphocytes Are Partially Immunocompetent Thymus-Derived Cells That Undergo Post-Thymic Maturation to Become Functionally Mature CD4+ T Lymphocytes

CD4+CD8+ double-positive (DP) T cells represent a minor subpopulation of T lymphocytes found in the periphery of adult rats. In this study, we show that peripheral DP T cells appear among the first T cells that colonize the peripheral lymphoid organs during fetal life, and represent ∼40% of peripheral T cells during the perinatal period. Later their proportion decreases to reach the low values seen in adulthood. Most DP T cells are small size lymphocytes that do not exhibit an activated phenotype, and their proliferative rate is similar to that of the other peripheral T cell subpopulations. Only 30–40% of DP T cells expresses CD8β chain, the remaining cells expressing CD8αα homodimers. However, both DP T cell subsets have an intrathymic origin since they appear in the recent thymic emigrant population after injection of FITC intrathymically. Functionally, although DP T cells are resistant to undergo apoptosis in response to glucocorticoids, they show poor proliferative responses upon CD3/TCR stimulation due to their inability to produce IL-2. A fraction of DP T cells are not actively synthesizing the CD8 coreceptor, and they gradually differentiate to the CD4 cell lineage in reaggregation cultures. Transfer of DP T lymphocytes into thymectomized SCID mice demonstrates that these cells undergo post-thymic maturation in the peripheral lymphoid organs and that their CD4 cell progeny is fully immunocompetent, as judged by its ability to survive and expand in peripheral lymphoid organs, to proliferate in response to CD3 ligation, and to produce IL-2 upon stimulation.

Prolactin affects both survival and differentiation of T-cell progenitors.

We have analysed the in vitro effects of prolactin on thymocyte development concluding that PRL favours the survival and differentiation of T-cell progenitors. Fetal, adult thymocytes and CD45(+) fetal liver lymphoid progenitors express PRL-R. PRL induces survival, proliferation and differentiation of lymphoid progenitors whereas both an anti-PRL antiserum and an anti-PRL-R mAb block T-cell development accumulating CD25(+)DN (CD4(-)CD8(-)) cells. Furthermore, IL2 rescues the blockade of T-cell development in FTOC treated with anti-PRL antiserum but PRL does not recover cultures treated with an anti-IL2R alpha chain mAb, which drastically blocks the T-cell development. These results support IL2/IL2R mediation of PRL effects on developing thymocytes.

Partial blockade of T-cell differentiation during ontogeny and marked alterations of the thymic microenvironment in transgenic mice with impaired glucocorticoid receptor function.

Glucocorticoids (GCs) are widely known to be potent modulators of the immune system. The role of GCs in thymopoiesis as well as the integration of the thymus with the neuroendocrine system is, however, poorly understood. In the present work, we have studied, in transgenic mice with an impaired GC function, the alterations which occur in both T-cell differentiation and thymic stroma maturation, throughout ontogeny as well as in adult condition, analyzing their possible rebounding on the status of adult splenic T lymphocyte populations. These transgenic mice have been described to present a significant decrease (60-70%) of thymic and splenic GC receptor binding capacity but maintain normal their basal plasma ACTH and corticosterone levels. The animals showed a partial blockade of T-cell differentiation and decreased percentages of apoptotic cells during fetal development but not in adult life, when thymic cellularity was significantly increased although thymocyte apoptosis response was not affected. In contrast, thymic stroma was profoundly altered from early fetal stages and large epithelium-free areas appeared in adult thymus. On the other hand, our study revealed a reduction of the splenic TcRalphabeta population accompanied by an increase in the CD4/CD8 ratio. The analysis of different adhesion molecules as well as activation markers demonstrated that most of them (CD5, CD11a, CD11b, CD69 and MHC Class II) were normally expressed in transgenic lymphocytes, whereas CD44 and CD62L expression was altered indicating the existence of an increased proportion of primed T-cells in these animals. In view of the mutual interdependence of thymic stroma and thymocyte maturation, the partial blockade of T-cell differentiation during ontogeny and the profound alterations of the stromal cell compartment in transgenic mice with impaired GR function suggest a key role for GCs in coordinating the physiological dialogue between the developing thymocytes and their microenvironment.