The thymus is the primary lymphoid organ in which T lymphocytes develop and mature. Its microenvironment relies largely on thymic epithelial cells, which fall into two populations according to their location: cortical epithelial cells, involved in T-lineage commitment and positive selection, and medullary epithelial cells, or mTECs, which complete T-lymphocyte maturation through negative selection and contribute to the generation of regulatory T cells. To fulfil their role in immune tolerance, mTECs express a broad range of tissue-specific antigens under the control of transcription factors such as AIRE, FEZF2 and PRDM1, as well as various cytokines and chemokines essential for T-cell signalling and migration. The function of these cells has been studied mainly in mice, even though significant differences exist in the kinetics of marker expression between humans and rodents, which limits the transferability of murine models.
To address this difficulty, the authors describe a method for culturing primary human mTECs that dispenses with enzymatic procedures and cell sorting by flow cytometry. The cells are obtained from explants of fresh thymic biopsies, an approach that reduces the amount of tissue required, preserves cell viability and increases the number of cells recovered. The enzymatic techniques conventionally used, often coupled with cell sorting, are indeed liable to alter the expression of surface molecules and to compromise cell purity and survival, while rarely allowing prolonged culture.
After seven days of primary culture, the thymus-derived cells display the markers specific to mTECs (K5, K14, claudin 4 and UEA-1), without expression of the cortical markers (K8 and K18). They retain their capacity to express the key players in immune tolerance, including AIRE, tissue-specific antigens, cytokines and chemokines. AIRE expression fluctuates over the course of the culture, reaching its optimum around day six or seven before declining; the addition of RANKL restores this expression, suggesting that the decline reflects the progressive depletion of the lymphoid microenvironment. The cells also adjust their gene expression network in response to their environment, notably responding to oestrogenic and inflammatory signals.
The authors thus propose a stable and reproducible model of human mTECs, derived from normal or pathological thymuses, that can be used to test the effect of various molecules on the homeostasis and physiology of the thymic epithelium. This model, presented as a first step towards the establishment of immortalised human cell lines, should make it possible to explore the specific features of human mTECs under normal or pathological conditions, for example in autoimmune myasthenia gravis, and to move beyond the extrapolations made from murine models.