The recent thymic origin, differentiation and suppressive mechanism of regulatory T cells.

Abstract

Regulatory T cells are a purported lineage of CD4⁺ cells that inhibit the proliferation and effector functions of other T cells to prevent the development of autoimmune disease. However, little is known about how they arise, their lifespan and their patterns of recirculation. Furthermore, the mechanisms through which they inhibit other T cells remain unclear. In order to address these issues, we investigated the relationship between regulatory T cells and recent thymic emigrants (RTE) which are newly formed T cells released into the periphery from the thymus. The CD25⁺ Foxp3⁺ regulatory T cell subset was found to be closely associated with RTE, and generated the CD25⁻ Foxp3⁺ T regulatory T cell subset by unidirectional differentiation. This process was exploited to mature flow sorted CD4⁺ CD25bright[bright in superscript]Foxp3⁺ T cells into CD25⁻ Foxp3⁺ T cells and determine that they retain their functional suppressive activity. The phenotype and physiology of the CD25⁺ Foxp3⁺ and CD25⁻ Foxp3⁺ regulatory T cell subsets were characterised and compared to conventional T cell subsets, revealing the differential expression of numerous key molecules. The high expression of CD62L and LFA-1 by CD25⁺ Foxp3⁺ regulatory T cells was consistent with both their relative enrichment within secondary lymphoid tissues and their sessile nature. The profile of adhesion molecules on the surface of CD25⁻ Foxp3⁺ cells suggested they may tend to localise to sites of inflammation other than the lamina propria, as they have a low expression of CD103 and CD62L, but high expression of LFA-1. However, CD25⁻ Foxp3⁺ regulatory T cells were found to selectively migrate into the intestinal mucosa, where they were enriched, and they also returned back to the thymus, suggesting they may constitute a tissue homing subset of regulatory T cells. We explored the mechanism of regulatory T cell suppression, and found that regulatory T cells condition APC to reduce their ability to activate other T cells. Following the application of this system to differential gene expression microarray analysis, we identified several putative molecular targets of regulation including 10 novel predicted serine/threonine kinases, a novel four point-1, ezrin, radixin and moesin domain containing signalling molecule, the E2F transcription factor 5, and a CD163-like molecule. Aging was found to negatively affect the productivity of the thymus, although it was found to be still generating new T cells into old age. While the number of thymocytes decreased with aging, the number of Foxp3⁺ cells in the thymus was unaffected, possibly their preferential recirculation back to the thymus. The size of the peripheral T cell pool decreased with aging, and the proportion of CD25⁺ Foxp3⁺ regulatory T cells among CD4+ T cells declined. However, the proportion of CD25⁻ Foxp3⁺ regulatory T cells increased with aging in the periphery. The conversion of CD25⁺ Foxp3⁺ regulatory T cells into CD25⁻ Foxp3⁺ T cells may compensate for the declining thymic output of CD25⁺ Foxp3⁺ regulatory T cells that occurs with aging, in order to maintain the regulatory T cell pool.