Epithelial-mesenchymal interactions and the role of NCCs in thymus development

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Epithelial-mesenchymal interactions are a common scenario during organogenesis. Mutually inductive interactions between the endoderm and neural crest (NC)-derived mesenchyme are essential during thymus development (reviewed in (7, 31). Using tissue recombination experiments, Auerbach originally demonstrated the importance of NC-derived mesenchyme in development of epithelial thymus rudiment explants (45). This concept was supported by neural crest ablation experiments in chicks that resulted in variable defects in thymus development (46). However, heterotopic transplant experiments using chick:quail chimeras indicated that NC-derived signals do not induce initial organ formation, since endodermal explants taken prior to neural crest cell (NCC) migration were capable of forming a functional thymus in an ectopic location (47). Similar studies in mice using lineage tracing and transplantation experiments also support an entirely endodermal origin for TECs (1). Nevertheless, NC-derived signals are essential for thymus development. The specific role played by NCCs varies throughout ontogeny and in the postnatal thymus. At the outset of thymus organogenesis NCCs are involved in patterning third pharyngeal pouch endoderm by setting the border between thymus and parathyroid fated domains resulting in appropriate allocation of endodermal progenitors to each domain (48). Signals from NC-derived mesenchyme promote separation of the thymus rudiment from pharyngeal endoderm as well as detachment of the developing thymus from the parathyroid (48). Subsequently, NC-derived cells play a role in migration of fetal thymus lobes into the thoracic cavity. Specifically, epithelial-mesenchymal interactions involving BMP signaling are required for thymic capsule formation, thymus-parathyroid separation and organ migration (49). A recent report demonstrated that EphB-ephrinB2 interactions regulate NCC mobility, and that deletion of ephrinB2 from NCCs results in failure of thymus organ migration and ectopic positioning of thymic lobes (50).

NCCs are essential for TEC proliferation and outgrowth of the thymus rudiment, primarily via fibroblast growth factor (FGF) signaling. Reciprocal FGF signaling between third pouch epithelium-derived FGF8 and NCCs expressing FGF10 has been implicated in 3^rd pouch formation and initial outgrowth of the organ primordium. In the developing limb bud epithelial cells produce FGF8 that regulates expression of FGF10 in the underlying mesenchyme (51, 52). Similarly, Fgf10 expression in the perithymic mesenchyme is dependent on Fgf8 expression in the pouch endoderm and/or ectoderm (53). After pouch formation, FGF7 and FGF10 produced by perithymic NC-derived mesenchyme activate the corresponding receptor (FgfR2IIIb) on fetal TECs to promote their proliferation (54-56). FGFR2-IIIb mutants develop severe thymus hypoplasia after E12.5, and Fgf10^-/- mutants display reduced TEC proliferation, indicating that NCC-derived FGF7 and -10 signals are required for thymic epithelial cell (TEC) proliferation (56). Removing the mesenchymal capsule from E12 fetal thymi prior to transplantation inhibits thymus growth in vitro and results in hypoplastic thymi after transplantation under the kidney capsule (55, 57). However, in the absence of NCCs, the transplanted thymuses developed TEC subsets that support thymocyte maturation indicating that NCC-derived signals are required for TEC proliferation but not differentiation. These results also suggest that expansion of the TEC compartment is necessary to provide sufficient intrathymic niches to support thymocyte progenitors.

NC cells regulate thymus size and morphogenesis in part via secretion of BMP4 and WNT family proteins (49, 58, 59). A recent study reported reduced expression of Bmp4 and Wnt3 in mesenchymal cells from MafB deficient embryos (60). The MafB transcription factor is predominantly expressed by NC-derived mesenchyme, and its absence indirectly affects epithelial function. Epithelial cells in the MafB-deficient thymus rudiment express low levels of CCL21 and CCL25, chemokines known to attract thymocyte progenitors to the fetal thymus (57, 61). As result the number of hematopoietic cells in the MafB-deficient fetal thymus is reduced (60).

Thymic mesenchyme has also been proposed to directly participate in thymocyte development, although this role is less well supported and molecular mechanisms have not been identified (62). Although the question of whether NC-derived mesenchymal cells directly or indirectly regulate thymocyte development is unresolved, the overall question of NCC function in the thymus is an important issue that will affect strategies designed to restore thymus function and T cell output after age or disease associated involution. The proportion of mesenchymal cells increases as TEC numbers decrease during aging-related thymic involution (63). As a result, their contribution to the microenvironment increases, and could contribute to changes in the function of the microenvironment with age.