The thymus is the principal site of T cell development and therefore is of central importance wit... more The thymus is the principal site of T cell development and therefore is of central importance within the immune system: congenital athymia results in profound immunodeficiency, while perturbed thymic function can lead to autoimmunity. Although highly active in early life, the thymus undergoes premature involution, such that de novo T cell development diminishes significantly with age. This has implications for immune function in the aging population, and in clinical procedures such as bone marrow and solid organ transplantation, where thymic function is required for T cell reconstitution and/or tolerance induction. Interest therefore exists in enhancing immune reconstitution through regenerative or cell therapies for boosting thymus activity in vivo, or providing customized in vitro generated T cell repertoires for adoptive transfer. The success of such strategies is likely to depend on a detailed knowledge of the mechanisms regulating thymus development and homeostasis. Here, we review current understanding of cellular and molecular regulation of thymus organogenesis, focusing on the epithelial component of the thymic stroma which provides many of the specialist functions required to mediate T cell differentiation and T cell repertoire selection.
The thymus is required for generation of a self-tolerant, self-restricted T-cell repertoire. The ... more The thymus is required for generation of a self-tolerant, self-restricted T-cell repertoire. The capacity to manipulate or replace thymus function therapeutically would be beneficial in a variety of clinical settings, including for improving recovery following bone marrow transplantation, restoring immune system function in the elderly and promoting tolerance to transplanted organs or cells. An attractive strategy would be transplantation of thymus organoids generated from cells produced in vitro, for instance from pluripotent stem cells. Here, we review recent progress toward this goal, focusing on advances in directing differentiation of pluripotent stem cells to thymic epithelial cells, a key cell type of the thymic stroma, and related direct reprogramming strategies.
Abstract 835 The establishment of the thymic microenvironment early in life is crucial for the pr... more Abstract 835 The establishment of the thymic microenvironment early in life is crucial for the production functional T cells. Conversely, thymic involution results in a decreased T cell output. Thymic involution has important health implications especially following bone marrow transplant. Our objective is to determine molecular and cellular mechanisms that will allow for regeneration of involuted thymic tissue, restore production of naïve T cells, and improve immune function while improving our understanding of immunobiology. In this pursuit, we have focused on the Retinoblastoma family of tumor suppressor proteins. The main function of the RB pathway is to restrict passage through the G1/S transition of the cell cycle. RB and its two family members, p107 and p130, mediate the action of a broad range of cellular signals to control the proliferation, survival, and differentiation status of a large number of mammalian cell types. We found that inactivation of the RB pathway in the thymus by early deletion of RB family genes prevents thymic involution, promotes expansion of functional thymic epithelial cells (TECs), and increases thymic T cell output. Moreover, we have identified a direct regulatory relationship between RB and the Foxn1 transcription factor Via E2F transcription factors, where RB/E2F complexes directly repress the Foxn1 promoter, thereby promoting involution. Thus, the RB family is a critical mediator of extra- and intra-cellular signals to regulate thymic epithelial cells and thymus function, and decreasing RB pathway function may promote regeneration of the involuted thymus and restoration of naïve T cell production in patients. Disclosures: No relevant conflicts of interest to declare.
The thymus and parathyroids originate from bilateral 3rd pharyngeal pouch (pp) endoderm. At E11.5... more The thymus and parathyroids originate from bilateral 3rd pharyngeal pouch (pp) endoderm. At E11.5, thymus- and parathyroid-fated domains express Foxn1 and Gcm2 respectively, transcription factors that regulate differentiation but do not specify organ fate. Tbx1, which encodes a T-box transcription factor required for segmentation of the pharyngeal endoderm is initially widely expressed in the 3rd pp, but by E10.5 is excluded from the ventral, thymus-fated domain. We recently reported that Tbx1 is a negative regulator of thymus development (Reeh et al. Development 141:2950, 2014). Ectopic expression of Tbx1 in the thymus-fated domain of the 3rd pp suppresses Foxn1 and inhibits thymic epithelial cell proliferation and differentiation. MicroRNAs (miRs) are important regulators of gene expression. We find that members of the miR-17-92 cluster are expressed in the ventral domain of the 3rd pp in wildtype embryos. Moreover, miR-17-92 downregulates Tbx1 in cardiac progenitor cells to promote their differentiation (Wang et al. Developmental Cell 19:903, 2010). Therefore, we proposed that miR-17-92 regulates Tbx1 expression in 3rd pp endoderm. In support of this hypothesis, we find that deletion of miR-17-92 enhances TBX1 and reduces FOXN1 in 3rd pp endoderm. The data support a model in which miR-17-92 plays an essential role in thymus development by regulating Tbx1 expression in the 3rd pp.
SCI-50 The postnatal thymus is the primary source of T cells in vertebrates, and many if not all ... more SCI-50 The postnatal thymus is the primary source of T cells in vertebrates, and many if not all stages of thymocyte development require interactions with thymic epithelial cells (TECs). There is abundant and growing evidence that the cellular and compartmental organization of the thymus is crucial for optimal organ function. Changes in the organization and stromal composition of the thymus as a result of genetic changes or during aging-associated involution can negatively impact both T cell production and function. The TEC-specific transcription factor Foxn1 is a critical regulator of these processes. I will present data from our Foxn1 allelic series showing that Foxn1 regulates TEC proliferation and differentiation at both fetal and adult stages, and that Foxn1-dependent TEC differentiation is required to assemble the multiple cell types that make up the complex structure of the functional thymus. Thus, Foxn1 is a central regulator throughout the life cycle of the thymus, and orch...
Thymic epithelial cells (TEC) are essential for thymocyte differentiation and repertoire selectio... more Thymic epithelial cells (TEC) are essential for thymocyte differentiation and repertoire selection. Despite their indispensable role in generating functional T cells, the molecular mechanisms that orchestrate TEC development from endodermal progenitors in the third pharyngeal pouch (3rd PP) are not fully understood. We recently reported that the T-box transcription factor TBX1 negatively regulates TEC development. Although initially expressed throughout the 3rd PP, Tbx1 becomes downregulated in thymus-fated progenitors and when ectopically expressed impairs TEC progenitor proliferation and differentiation. Here we show that ectopic Tbx1 expression in thymus fated endoderm increases expression of Polycomb repressive complex 2 (PRC2) target genes in TEC. PRC2 is an epigenetic modifier that represses gene expression by catalyzing trimethylation of lysine 27 on histone H3. The increased expression of PRC2 target genes suggests that ectopic Tbx1 interferes with PRC2 activity and implicat...
The thymus is the principal site of T cell development and therefore is of central importance wit... more The thymus is the principal site of T cell development and therefore is of central importance within the immune system: congenital athymia results in profound immunodeficiency, while perturbed thymic function can lead to autoimmunity. Although highly active in early life, the thymus undergoes premature involution, such that de novo T cell development diminishes significantly with age. This has implications for immune function in the aging population, and in clinical procedures such as bone marrow and solid organ transplantation, where thymic function is required for T cell reconstitution and/or tolerance induction. Interest therefore exists in enhancing immune reconstitution through regenerative or cell therapies for boosting thymus activity in vivo, or providing customized in vitro generated T cell repertoires for adoptive transfer. The success of such strategies is likely to depend on a detailed knowledge of the mechanisms regulating thymus development and homeostasis. Here, we review current understanding of cellular and molecular regulation of thymus organogenesis, focusing on the epithelial component of the thymic stroma which provides many of the specialist functions required to mediate T cell differentiation and T cell repertoire selection.
The thymus is required for generation of a self-tolerant, self-restricted T-cell repertoire. The ... more The thymus is required for generation of a self-tolerant, self-restricted T-cell repertoire. The capacity to manipulate or replace thymus function therapeutically would be beneficial in a variety of clinical settings, including for improving recovery following bone marrow transplantation, restoring immune system function in the elderly and promoting tolerance to transplanted organs or cells. An attractive strategy would be transplantation of thymus organoids generated from cells produced in vitro, for instance from pluripotent stem cells. Here, we review recent progress toward this goal, focusing on advances in directing differentiation of pluripotent stem cells to thymic epithelial cells, a key cell type of the thymic stroma, and related direct reprogramming strategies.
Abstract 835 The establishment of the thymic microenvironment early in life is crucial for the pr... more Abstract 835 The establishment of the thymic microenvironment early in life is crucial for the production functional T cells. Conversely, thymic involution results in a decreased T cell output. Thymic involution has important health implications especially following bone marrow transplant. Our objective is to determine molecular and cellular mechanisms that will allow for regeneration of involuted thymic tissue, restore production of naïve T cells, and improve immune function while improving our understanding of immunobiology. In this pursuit, we have focused on the Retinoblastoma family of tumor suppressor proteins. The main function of the RB pathway is to restrict passage through the G1/S transition of the cell cycle. RB and its two family members, p107 and p130, mediate the action of a broad range of cellular signals to control the proliferation, survival, and differentiation status of a large number of mammalian cell types. We found that inactivation of the RB pathway in the thymus by early deletion of RB family genes prevents thymic involution, promotes expansion of functional thymic epithelial cells (TECs), and increases thymic T cell output. Moreover, we have identified a direct regulatory relationship between RB and the Foxn1 transcription factor Via E2F transcription factors, where RB/E2F complexes directly repress the Foxn1 promoter, thereby promoting involution. Thus, the RB family is a critical mediator of extra- and intra-cellular signals to regulate thymic epithelial cells and thymus function, and decreasing RB pathway function may promote regeneration of the involuted thymus and restoration of naïve T cell production in patients. Disclosures: No relevant conflicts of interest to declare.
The thymus and parathyroids originate from bilateral 3rd pharyngeal pouch (pp) endoderm. At E11.5... more The thymus and parathyroids originate from bilateral 3rd pharyngeal pouch (pp) endoderm. At E11.5, thymus- and parathyroid-fated domains express Foxn1 and Gcm2 respectively, transcription factors that regulate differentiation but do not specify organ fate. Tbx1, which encodes a T-box transcription factor required for segmentation of the pharyngeal endoderm is initially widely expressed in the 3rd pp, but by E10.5 is excluded from the ventral, thymus-fated domain. We recently reported that Tbx1 is a negative regulator of thymus development (Reeh et al. Development 141:2950, 2014). Ectopic expression of Tbx1 in the thymus-fated domain of the 3rd pp suppresses Foxn1 and inhibits thymic epithelial cell proliferation and differentiation. MicroRNAs (miRs) are important regulators of gene expression. We find that members of the miR-17-92 cluster are expressed in the ventral domain of the 3rd pp in wildtype embryos. Moreover, miR-17-92 downregulates Tbx1 in cardiac progenitor cells to promote their differentiation (Wang et al. Developmental Cell 19:903, 2010). Therefore, we proposed that miR-17-92 regulates Tbx1 expression in 3rd pp endoderm. In support of this hypothesis, we find that deletion of miR-17-92 enhances TBX1 and reduces FOXN1 in 3rd pp endoderm. The data support a model in which miR-17-92 plays an essential role in thymus development by regulating Tbx1 expression in the 3rd pp.
SCI-50 The postnatal thymus is the primary source of T cells in vertebrates, and many if not all ... more SCI-50 The postnatal thymus is the primary source of T cells in vertebrates, and many if not all stages of thymocyte development require interactions with thymic epithelial cells (TECs). There is abundant and growing evidence that the cellular and compartmental organization of the thymus is crucial for optimal organ function. Changes in the organization and stromal composition of the thymus as a result of genetic changes or during aging-associated involution can negatively impact both T cell production and function. The TEC-specific transcription factor Foxn1 is a critical regulator of these processes. I will present data from our Foxn1 allelic series showing that Foxn1 regulates TEC proliferation and differentiation at both fetal and adult stages, and that Foxn1-dependent TEC differentiation is required to assemble the multiple cell types that make up the complex structure of the functional thymus. Thus, Foxn1 is a central regulator throughout the life cycle of the thymus, and orch...
Thymic epithelial cells (TEC) are essential for thymocyte differentiation and repertoire selectio... more Thymic epithelial cells (TEC) are essential for thymocyte differentiation and repertoire selection. Despite their indispensable role in generating functional T cells, the molecular mechanisms that orchestrate TEC development from endodermal progenitors in the third pharyngeal pouch (3rd PP) are not fully understood. We recently reported that the T-box transcription factor TBX1 negatively regulates TEC development. Although initially expressed throughout the 3rd PP, Tbx1 becomes downregulated in thymus-fated progenitors and when ectopically expressed impairs TEC progenitor proliferation and differentiation. Here we show that ectopic Tbx1 expression in thymus fated endoderm increases expression of Polycomb repressive complex 2 (PRC2) target genes in TEC. PRC2 is an epigenetic modifier that represses gene expression by catalyzing trimethylation of lysine 27 on histone H3. The increased expression of PRC2 target genes suggests that ectopic Tbx1 interferes with PRC2 activity and implicat...
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