Identification And Functional Characterization Of Thymic Mesenchymal Stromal/Stemal/Stem Cells

As one of the central immune organs of the human,the thymus is the place where T cells differentiate and mature,and plays a vital role in the immune system.The thymus,as a multicellular complex structure,is composed of thymic stroma(including dendritic cells,macrophages,epithelial cells,mesenchymal cells and vascular components)and thymic cells,however,the biological significance of the mesenchymal cells(especially the non-hematopoietic mesenchymal stromal/stem cells(MSCs))is still unclear.Therefore,it is improtant to identify thymic MSCs(T-MSCs)and study their biological functions.By examining the basic characteristics of T-MSCs,we found that T-MSCs and bone marrow-derived MSCs(BM-MSCs)had the similar "stemness".They both had the ability of undergoing adipogenic and osteogenic differentiation,and expressed similar cell surface markers.Further,we constructed NestintdTomato mice to explore the distribution of Nestin+cells in the thymus based on the expression of Nestin on T-MSCs.We found that Nestin+cells were mainly distributed in the medulla region and the cortical-medulla junction.Next,we explored the immunomodulatory function of T-MSCs in vitro.Like BM-MSCs,T-MSCs could also exert immunosuppressive effects on T cells under the stimulation of inflammatory factors(IFN-γ/TNF-α)and could ameliorate the inflammatory bowel disease induced by sodium dextran sulfate in mice,but the effects were partially abrogated in Nos2-/-T-MSCs,suggesting that the immunomodulatory effect of T-MSCs was partly dependent on nitric oxide(NO).MSCs can phagocyte apoptotic cells and thus play an immunosuppressive role,and the thymus produces a large number of apoptotic cells every day.Do T-MSCs have the ability of phagocytosing apoptotic cells?We found that T-MSCs could engulf apoptotic cells in vitro co-culture of T-MSCs and apoptotic cells.However,when we induced apoptosis of thymocytes by applying Dexamethasone(DEX)in vivo,the phagocytic effect of T-MSCs was not detected.The scarcity of T-MSCs in vivo made it hard to detect its phagocytic function.Moreover,we speculated that dexamethasone might have an effect on T-MSCs in addition to inducing apoptosis of thymocytes.Indeed,we found that the number of T-MSCs in the thymus was reduced significantly one day after dexamethasone treatment,while the number of other CD45-stromal cells did not change much,suggesting that T-MSCs were sensitive to dexamethasone in the thymus.Interestingly,no reduction in cell viability was observed when T-MSCs was treated with high concentration of dexamethasone in vitro.This in vivo-specific effect might be related to the microenvironment of T-MSCs,but it could not be ruled out that the sensitivity of T-MSCs to Dex was changed during the continuous subculture in vitro.This study confirms the "stemness" of T-MSCs and immunosuppressive ability to T cells.Using Nestin as the identification marker of T-MSCs we determined the distribution of T-MSCs in the thymus.Although T-MSCs can engulf apoptotic cells in vitro and is insensitive to dexamethasone,these properties differ in vivo,which may be related to the complex microenvironment in the thymus.The construction of NestintdTomato mice provides a useful tool for studying the maturation of thymocytes or the interaction of T-MSCs with other stromal cells.