The human high-mobility group protein A2 (HMGA2) protein is an architectural transcription factor that transforms chromatin structure by binding to DNA. Recently, it has been reported that HMGA2 is highly expressed in fetal neural stem cells and has the capacity to promote stemness. However, there is currently no information available on the functional significance and molecular mechanisms of the cellular in vitro aging and proliferation of human umbilical cord blood-derived stromal cells (hUCBSCs). In the present study, we evaluated the direct effects of HMGA2 on the cellular aging and proliferation of hUCBSCs and investigated potential regulatory mechanisms responsible for the corresponding functions. We found that the overexpression of HMGA2 enhanced proliferation and reduced or even reversed the in vitro aging process of hUCBSCs. This effect was accompanied by the increased expression of cyclin E and CDC25A and the significantly decreased expression of cyclin-dependent kinase inhibitors. Furthermore, HMGA2 inhibition compromised cell proliferation and adipogenic differentiation in early-stage hUCBSCs. From the molecular/cellular functional analysis of microarray data, we found that HMGA2 overexpression induced a PI3K/Akt/mTOR/p70S6K cascade, which in turn suppressed the expression of p16(INK4A) and p21(CIP1/WAF1) in hUCBSCs. These results provide novel insights into the mechanism by which HMGA2 regulates the in vitro aging and proliferation of hUCBSCs.