Mesenchymal stem cells (MSC) have emerged as a new therapeutic tool for a number of clinical applications, because they have multipotency and paracrine effects via various factors. In the present study, we investigated the effects of adipose-derived MSC (Ad-MSC) transplantation via intrathecal injection through the cisterna magna on cell proliferation and differentiation of endogenous stem cells in the hippocampal dentate gyrus (DG) using Ki-67 (a marker for proliferating cells), and doublecortin (DCX, a marker for neuroblasts). The transplanted Ad-MSC were detected in the meninges, not in the hippocampal parenchyma. However, the number of Ki-67-immunoreactive cells was significantly increased by 83% in the DG 2 days after single Ad-MSC injection, and by 67% at 23 days after repeated Ad-MSC treatment compared with that in the vehicle-treated group after Ad-MSC transplantation. On the other hand, the number of DCX-immunoreactive cells in the DG was not changed at 2 days after single Ad-MSC injection; however, it was significantly increased by 62% 9 days after single Ad-MSC injection. At 23 days after repeated Ad-MSC application, the number of DCX-immunoreactive cells was much more increased (223% of the vehicle-treated group). At this time point, DCX protein levels were also significantly increased compared with those in the vehicle-treated group. These results suggest that the intrathecal injection of Ad-MSC could enhance endogenous cell proliferation, and the repeated Ad-MSC injection could be more efficient for an enhancement of endogenous cell proliferation and differentiation in the brain.