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Reprogrammed Cells Mediate the Development of Functional Organ

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Review of “An organized and functional thymus generated from FOXN1-reprogrammed fibroblasts” from Nature Cell Biology by Stuart P. Atkinson.

While the field of regenerative medicine has matured to a stage where we can derive multiple defined cell types by various means, the generation of an entire organ, such as the thymus, has so far remained tantalizingly out of reach. However, researchers from the laboratory of Clare Blackburn (Medical Research Council Centre for Regenerative Medicine, University of Edinburgh, UK) have now taken this task to hand, and through direct reprogramming of fibroblasts with a lineage specific transcription factor have generated thymic epithelial cells (TECs), a key cell type of the thymic stroma [1], which they how to have the ability to generate a complete thymus on transplantation [2].

The researchers generated TECs through the overexpression of the transcription factor forkhead box N1 (Foxn1), critically required for development of TECs [3, 4], and for the sub-specialisation into cortical (c) and medullary (m) TECs which mediate separate aspects of T cell development [5], in mouse embryonic fibroblasts (MEFs). After ten days of Foxn1 expression, MEF morphology altered from an elongated bipolar shape to a broader epithelial-like polygonal shape, and became positive for epithelial-specific markers (K8 and EpCam) and markers expressed by TECs during early thymus development. Isolated EpCam+ cells expressed TEC marker genes (Dll4, Ccl25 and Kitl), as well as other non-Foxn1 target TEC-associated genes (Pax9 and Trp63) and endogenous Foxn1 itself. This altogether points to the efficient Foxn1-mediated conversion of fibroblasts to TEC-like cells (iTECs).

The researchers then assessed iTECs capacity to support T cell development by seeding early T lineage progenitors (ETPs) [6] onto an iTEC monolayer and assessing CD4 and CD8 cell development. At 12 days thymopoiesis was evident; CD4+ cells, CD8+ cells and CD4+CD8+ double positive cells were present in the co-culture with cells also expressing CD3 (role in antigen recognition) and T cell antigen receptor beta (TCR). Importantly, thymopoiesis did not occur upon co-culture with control MEFs. The group then assessed the potential of iTECs to form an organized and functional thymus through the aggregation of day 5 iTECs with immature thymocytes and fetal thymic mesenchyme (to ensure of growth factor supply) and engraftment of given aggregate under the kidney capsule of syngeneic adult mice. Excitingly, this led to the formation of macroscopic organs exhibiting characteristic thymus morphology, with multiple clearly defined cortical and medullary regions evident. Additionally, haematopoietic and epithelial cells were present in similar proportions to that observed in the adult thymus, and the researchers also observed the correct compartmentalization of functional cTECs and mTECs. Thymocyte subset distribution in the iTEC-derived grafts also matched adult thymus, while peripheral blood analysis demonstrated the population of the peripheral immune system with CD4+ and CD8+ T cells, and Foxp3+ regulatory T cells, whose levels rose with time.

This exciting study demonstrates the ability of a single factor to reprogram a cell across a germ-layer boundary, and the subsequent ability of this cell type to establish a complete, fully organized and functional organ – the first example of its kind. The therapeutic potential of this research is clear; this technique could allow for the rapid and correct generation of patient-matched T cells in the laboratory, while human translation of this work could aid those that suffer from a lack of the thymus or a dysfunctional thymus,

References

  1. Ritter MA and Boyd RL Development in the thymus: it takes two to tango. Immunol Today 1993;14:462-469.
  2. Bredenkamp N, Ulyanchenko S, O'Neill KE, et al. An organized and functional thymus generated from FOXN1-reprogrammed fibroblasts. Nat Cell Biol 2014;16:902-908.
  3. Nehls M, Kyewski B, Messerle M, et al. Two genetically separable steps in the differentiation of thymic epithelium. Science 1996;272:886-889.
  4. Blackburn CC, Augustine CL, Li R, et al. The nu gene acts cell-autonomously and is required for differentiation of thymic epithelial progenitors. Proceedings of the National Academy of Sciences of the United States of America 1996;93:5742-5746.
  5. Manley NR, Richie ER, Blackburn CC, et al. Structure and function of the thymic microenvironment. Front Biosci (Landmark Ed) 2011;16:2461-2477.
  6. Allman D, Sambandam A, Kim S, et al. Thymopoiesis independent of common lymphoid progenitors. Nature immunology 2003;4:168-174.