FOXN1
Forkhead box protein N1 (Winged-helix transcription factor nude) [RONU] [WHN]
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Age-associated systemic, chronic inflammation is partially attributed to increased self-autoreactivity, resulting from disruption of central tolerance in the aged, involuted thymus. This involution causally results from gradually decreased expression of the transcription factor FOXN1 in thymic epithelial cells (TECs), whereas exogenous FOXN1 in TECs can partially rescue age-related thymic involution. TECs induced from FOXN1-overexpressing embryonic fibroblasts can generate an ectopic de novo thymus under the kidney capsule, and intrathymic injection of naturally young TECs can lead to middle-aged thymus regrowth. Therefore, as a thymic rejuvenation strategy, we extended these 2 findings by combining them with 2 types of promoter-driven (Rosa26CreERT and FoxN1Cre) Cre-mediated FOXN1-reprogrammed embryonic fibroblasts (FREFs). We engrafted these FREFs directly into the aged murine thymus. We found substantial regrowth of the native aged thymus with rejuvenated architecture and function in both males and females, exhibiting increased thymopoiesis and reinforced thymocyte negative selection, along with reduced senescent T cells and autoreactive T cell-mediated inflammation in old mice. Therefore, this approach has preclinical significance and presents a strategy to potentially rescue decreased thymopoiesis and perturbed negative selection to substantially, albeit partially, restore defective central tolerance and reduce subclinical autoimmune symptoms in elderly people.
Keywords
- Aging
- Immunology
- Immunotherapy
- T cell development
Thymic involution is associated with age-related changes of the immune system. Utilizing our innovative technique of transplantation of a thymus as an isolated vascularized graft in MHC-inbred miniature swine, we have previously demonstrated that aged thymi are rejuvenated after transplantation into juvenile swine. Here we have studied the role of insulin-like growth factor (IGF) and forkhead-box protein-N1 (FOXN1) as well as bone marrow (BM) in thymic rejuvenation and involution. We examined thymic rejuvenation and involution by means of histology and flow cytometry. Thymic function was assessed by the ability to induce tolerance of allogeneic kidneys. Aged thymi were rejuvenated in a juvenile environment, and successfully induced organ tolerance, while juvenile thymi in aged recipients involuted and had a limited ability to induce tolerance. However, juvenile BM inhibited the involution process of juvenile thymi in aged recipients. An elevated expression of both FOXN1 and IGF1 receptors (IGF-1R) was observed in juvenile thymi and rejuvenated thymi. Juvenile BM plays a role in promoting the local thymic milieu as indicated by its ability to inhibit thymic involution in aged animals. The expression of FOXN1 and IGF-1R was noted to increase under conditions that stimulated rejuvenation, suggesting that these factors are involved in thymic recovery.
MeSH Terms
- Aging
- Animals
- Bone Marrow
- Cell Differentiation
- Forkhead Transcription Factors
- Graft Survival
- Immune Tolerance
- Receptor, IGF Type 1
- Rejuvenation
- Swine
- Swine, Miniature
- Thymus Gland
Keywords
- animal models: porcine
- basic (laboratory) research/science
- bone marrow/hematopoietic stem cell transplantation
- immunobiology
- kidney transplantation/nephrology
- thymus/thymic biology
- tolerance: experimental
- translational research/science
Thymus function requires extensive cross-talk between developing T-cells and the thymic epithelium, which consists of cortical and medullary TEC. The transcription factor FOXN1 is the master regulator of TEC differentiation and function, and declining Foxn1 expression with age results in stereotypical thymic involution. Understanding of the dynamics of Foxn1 expression is, however, limited by a lack of single cell resolution data. We have generated a novel reporter of Foxn1 expression, Foxn1G, to monitor changes in Foxn1 expression during embryogenesis and involution. Our data reveal that early differentiation and maturation of cortical and medullary TEC coincides with precise sub-lineage-specific regulation of Foxn1 expression levels. We further show that initiation of thymic involution is associated with reduced cTEC functionality, and proportional expansion of FOXN1-negative TEC in both cortical and medullary sub-lineages. Cortex-specific down-regulation of Foxn1 between 1 and 3 months of age may therefore be a key driver of the early stages of age-related thymic involution.
MeSH Terms
- Aging
- Animals
- Cell Differentiation
- Cell Lineage
- Down-Regulation
- Embryonic Development
- Epithelial Cells
- Forkhead Transcription Factors
- Mice
- Thymus Gland
An autosomal recessive syndrome characterized by congenital hypotrichosis and short life expectancy has been described in the Birman cat breed (Felis silvestris catus). We hypothesized that a FOXN1 (forkhead box N1) loss-of-function allele, associated with the nude phenotype in humans, mice and rats, may account for the syndrome observed in Birman cats. To the best of our knowledge, spontaneous mutations in FOXN1 have never been described in non-human, non-rodent mammalian species. We identified a recessive c.1030_1033delCTGT deletion in FOXN1 in Birman cats. This 4-bp deletion was associated with the syndrome when present in two copies. Percentage of healthy carriers in our French panel of genotyped Birman cats was estimated to be 3.2%. The deletion led to a frameshift and a premature stop codon at position 547 in the protein. In silico, the truncated FOXN1 protein was predicted to lack the activation domain and critical parts of the forkhead DNA binding domain, both involved in the interaction between FOXN1 and its targets, a mandatory step to promote normal hair and thymic epithelial development. Our results enlarge the panel of recessive FOXN1 loss-of-function alleles described in mammals. A DNA test is available; it will help owners avoid matings at risk and should prevent the dissemination of this morbid mutation in domestic felines.
MeSH Terms
- Amino Acid Sequence
- Animals
- Binding Sites
- Cats
- Forkhead Transcription Factors
- Gene Deletion
- Gene Frequency
- Hypotrichosis
- Life Expectancy
- Molecular Sequence Data
Thymic involution is central to the decline in immune system function that occurs with age. By regenerating the thymus, it may therefore be possible to improve the ability of the aged immune system to respond to novel antigens. Recently, diminished expression of the thymic epithelial cell (TEC)-specific transcription factor Forkhead box N1 (FOXN1) has been implicated as a component of the mechanism regulating age-related involution. The effects of upregulating FOXN1 function in the aged thymus are, however, unknown. Here, we show that forced, TEC-specific upregulation of FOXN1 in the fully involuted thymus of aged mice results in robust thymus regeneration characterized by increased thymopoiesis and increased naive T cell output. We demonstrate that the regenerated organ closely resembles the juvenile thymus in terms of architecture and gene expression profile, and further show that this FOXN1-mediated regeneration stems from an enlarged TEC compartment, rebuilt from progenitor TECs. Collectively, our data establish that upregulation of a single transcription factor can substantially reverse age-related thymic involution, identifying FOXN1 as a specific target for improving thymus function and, thus, immune competence in patients. More widely, they demonstrate that organ regeneration in an aged mammal can be directed by manipulation of a single transcription factor, providing a provocative paradigm that may be of broad impact for regenerative biology.
MeSH Terms
- Aging
- Animals
- Cell Proliferation
- Cellular Microenvironment
- Epithelial Cells
- Forkhead Transcription Factors
- Lymphocyte Count
- Mice
- Mice, Transgenic
- Models, Animal
- Phenotype
- Regeneration
- Stem Cells
- T-Lymphocytes
- Thymus Gland
- Up-Regulation
Keywords
- FOXN1
- Mouse
- Organ regeneration
- Thymic involution