Mesenchymal stem cells (MSCs) are multipotent stem cells that can generate various microenvironment components in bone marrow, ensuring a precise control over self-renewal and multilineage differentiation of hematopoietic stem cells. Nevertheless, their spatiotemporal correlation with embryonic hematopoiesis remains rudimentary, particularly in relation to the human being. Here, we reported that human aorta-gonad-mesonephros (AGM) resided with bona fide MSCs. They were highly proliferative as fibroblastoid population bearing uniform surface markers (CD45(-), CD34(-), CD105(+), CD73(+), CD29(+), and CD44(+)), expressed pluripotential molecules Oct-4 and Nanog, and clonally demonstrated trilineage differentiation capacity (osteocytes, chondrocytes, and adipocytes). The frequency and absolute number of MSCs in aorta plus surrounding mesenchyme (E26-E27) were 0.3% and 164, respectively. Moreover, they were functionally equivalent to MSCs from adult bone marrow, that is, supporting long-term hematopoiesis and suppressing T-lymphocyte proliferation in vitro. In comparison, the matching yolk sac contained bipotent mesenchymal precursors that propagated more slowly and failed to generate chondrocytes in vitro. Together with previous knowledge, we propose that a proportion of MSCs initially develop in human AGM prior to their emergence in embryonic circulation and fetal liver.