This study was purposed to investigate the relation of the Notch signaling pathway to senescence of murine bone marrow stromal cells in vitro. Intracellular domain of Notch 1 (ICN) was transfected into cultured murine bone marrow stromal cells by lipofectamine transfection. After transfection for three days the proliferation of transfected cells was measured by MTT, cell cycle distribution was analyzed by flow cytometry. The percentage of senescence associated beta-galactosidase (SA-beta-Gal) positive cells were measured by cytochemical method, and the expression rates of P53 and p21Cip1/Waf1 at gene and protein levels were analyzed by RT-PCR and Western blot respectively. The results showed that after transfection for 3 days the proliferation of murine bone marrow stromal cells was inhibited with induction of G1 arrest, the percentage of SA-beta-gal positive cells increased and the p53 and p21Cip1/Waf1 mRNA and protein expression levels were upregulated. It is concluded that the activated Notch signaling can induce premature senescence of bone marrow stromal cells through the p53-p21Cip1/Waf1 pathway.