We used MTS24 expression to sort epithelial cells and deletion of Rac1 blocked the regeneration of a new thymic microenvironment when transplanted under the kidney capsule. Importantly www.selleckchem.com/products/arq-197.html the MTS24 negative cell population used in our controls includes non-epithelial cells and hence differs from that previously used by Rossi and colleagues where a second epithelial marker was used (Epcam1) to ensure MTS24- epithelial cells were used [12]. These experiments combined with the K14 litter mates controls supplied important controls for Cre expression and tamoxifen treatment. A recent paper has questioned whether the MTS24 marker uniquely identifies a thymic progenitor subpopulation with the ability to repopulate functional thymic epithelium [12].
These differences from previous reports [9], [10] are likely due to different sorting strategies and differing numbers of cells implanted. Importantly, in our studies we implanted a low number of cells that were confirmed simply as epithelial based on MTS24 expression. We have demonstrated that Rac1 expression is required for this subset of epithelial cells to regenerate the thymic microenvironment. It has been proposed that Rac and Myc represent a global stem cell regulatory axis [30]. In the epidermis Rac1 deletion leads to loss of stem cells while acute Myc over-expression promotes epidermal and hematopoietic differentiation, possibly through disruption of cellular adhesion [30], [38], [39], [40], [41]. Conversely, in the gut Myc determines self renewal while loss of Rac1 triggers differentiation [42].
In other models sustained induction of Myc leads to tumor development [43]. Hence it is known from other epithelial systems that Myc regulation is tightly controlled to avoid differentiation of stem cells or tumorigenesis. To determine whether a Rac1-Myc axis may be involved in thymic epithelial homeostasis, we showed in Fetal Thymic Organ Cultures derived from K14CreERxRac1flox/flox mice and six week old K5CrexRac1flox/flox thymus that Rac1 deletion leads to an increase in c-Myc expression. While this is not conclusive evidence that the same regulatory pathways are operational in the thymus as the skin we believe this will be Dacomitinib an interesting line of future investigation. In conclusion deletion of Rac1 results in the failure of thymic ontogeny in embryos and thymic atrophy in adults. Understanding mechanisms of thymic stem cell maintenance may help the development of therapies for patients with thymic developmental defects, or reverse damage from aging, chemo or radiotherapies. Further, maintenance of thymic epithelium ex vivo may allow in vitro generation of differentiated T cells. Supporting Information Figure S1 Distribution of keratins in vivo after Rac1 deletion.