Abstract
The numbers of thymic epithelial cells (TECs) and thymocytes steadily increase during embryogenesis. To examine this dynamic, we generated several TEC-specific transgenic mouse lines, which express fluorescent proteins in the nucleus, the cytosol and in the membranes under the control of the Foxn1 promoter. These tools enabled us to determine TEC numbers in tissue sections by confocal fluorescent microscopy, and in the intact organ by light-sheet microscopy. Compared to histological procedures, flow cytometric analysis of thymic cellularity is shown to underestimate the numbers of TECs by one order of magnitude; using enzymatic digestion of thymic tissue, the loss of cortical TECs (cTECs) is several fold greater than that of medullary TECs (mTECs), although different cTEC subsets appear to be still present in the final preparation. Novel reporter lines driven by Psmb11 and Prss16 promoters revealed the trajectory of differentiation of cTEC-like cells, and, owing to the additional facility of conditional cell ablation, allowed us to follow the recovery of such cells after their depletion during embryogenesis. Multiparametric histological analyses indicate that the new transgenic reporter lines not only reveal the unique morphologies of different TEC subsets, but are also conducive to the analysis of the complex cellular interactions in the thymus.
Highlights
The thymus can be found in all vertebrates and provides a dedicated micro-environment that is essential for T cell development[1]
When litters of treated mice were analysed at P7, we established that transgenic siblings still exhibited significantly reduced numbers of thymic epithelial cells (TECs) and thymocytes (Fig. 5c), as we had anticipated, and compatible with the notion that the desired reduction of Psmb11- and Prss16-expressing TECs was achieved
For the 4-week time point, our studies indicate that the thymus contains a total of ~1.5 × 106 TECs per thymus
Summary
The thymus can be found in all vertebrates and provides a dedicated micro-environment that is essential for T cell development[1]. The thymopoietic activity of the thymus (expressed as the ratio of haematopoietic cells to thymic epithelial cells) changes over time, rapidly increasing during the embryonic and perinatal periods, and reaching a peak in adolescence before slowly declining[17] While this general pattern seems to be true for rodents as well as for humans, it has been observed that the peak size of the thymus varies between different strains of rats[18] and mice[19]; a recent study ascribed the striking differences in thymopoietic activity of thymic epithelial cells in two mouse strains to the presence of a co-dominantly inherited genetic determinant(s)[20]
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