The aim of this work was to characterize multipotent mesenchymal stromal cells (MSCs) in the postnatal human thymus and to localize these MSCs in the organ. Adherent cells isolated from thymus samples were characterized by cell-surface antigen expression. This showed that adherent cells have a MSC profile as assessed by the expression of CD73 and CD105 markers and the lack of CD45 expression. These cells are able to differentiate in vitro into adipocytes, osteoblasts, and chondrocytes and to inhibit mixed lymphocyte reaction. This indicates that isolated cells have all of the characteristics of MSC. The fibroblast colony-forming unit (CFU-F) assay was used to determine their frequency in the postnatal thymus. This frequency was 60.9 +/- 14.8 CFU-F per 1 x 10(5) freshly isolated mononuclear cells. Moreover, taking advantage of CD34 and CD105 expression, immunohistological staining allowed us to localize MSC within interlobular trabeculae in close contact with the outer cortex. Polymerase chain reaction experiments indicated that thymic MSC expressed interleukin-7 and stromal cell-derived factor-1 messenger RNA. Overall, these results confirm previous findings of the presence in the adult human thymus of multipotent MSCs with a phenotype similar to adipose-derived adult stem cells. These results also show for the first time a histological localization of MSC in an organ. This suggests a possible role of thymic MSC in intrathymic differentiation.