Several recent studies demonstrated the potential of bioengineering using somatic stem cells in regenerative medicine. Adult human mesenchymal stem cells (hMSCs) derived from bone marrow have the pluripotency to differentiate into cells of mesodermal origin, e.g., bone, cartilage, adipose, and muscle cells; they, therefore, have many potential clinical applications. On the other hand, stem cells possess a self-renewal capability similar to cancer cells. For safety evaluation of tissue engineered medical devices using normal hMSCs, in this study, we investigated the expression levels of several genes that affect cell proliferation in hMSCs during in vitro culture. We focused on the relationship between the hMSC proliferation and their transforming growth factor β (TGFβ) signaling during in vitro culture. The proliferation rate of hMSCs gradually decreased and cellular senescence was observed for about 3 months. The mRNA expressions of TGFβ1, TGFβ2, and TGFβ receptor type I (TGFβRI) in hMSCs increased with the length of cell culture. The mRNA expressions of Smad3 increased, but those of c-myc and nucleostemin decreased with the length hMSCs were in in vitro culture. In addition, the expression profiles of the genes which regulate cellular proliferation in hMSCs were significantly different from those of cancer cells. In conclusion, hMSCs derived from bone marrow seldom underwent spontaneous transformation during 1—2 months in vitro culture for use in clinical applications. In hMSCs as well as in epithelial cells, growth might be controlled by the TGFβ family signaling.