Stem cell therapy for repair of myocardial injury has inherent limitations due to the poor attachment of the stem cells after cell transplantation. Ischemia after myocardial infarction (MI) and interaction of Mesenchymal stem cell (MSC) with niche is associated with increased production of reactive oxygen species (ROS). The intracellular ROS plays a key role in the regulation of cell adhesion, migration, and proliferation. The aim of this study is tried to explore the role of ROS on MSC-niche interaction. We examined the adhesion of MSC in 50 關M H2O2 -or non-treated condition coated with fibronectin or three-dimensional matrix. The adhesion rate of MSC was the highest in non-treated H2O2 condition of 3-D matrix. After treatment of H2O2, the detachment of MSC was increased but the increase in cell detachment was inhibited dose-dependently by pretreatment with the free radical scavenger, N-acetyl-L-cysteine(NAC). In addition, extracellular ROS activation enhanced by H2O2 was suppressed after the pretreatment of NAC. FAK phosphorylation and other downstream events, including activation of MAPK and Src, and expression of paxillin were all significantly attenuated by oxidation signaling. To assess the potential involvement of integrin-induced ROS production associated with cell adhesion, we determined whether small GTPase Rac-1 is regulated in MSC adhesion after H2O2 treatment. Under ROS condition, expression of Rac-1 was increased considerably in MSCs. For the cross talk of integrin with receptor tyrosine kinase (RTK), ROS may suppress paxillin-associated protein tyrosine phosphatase (PTP) relieving its negative regulatory effects. To verify the involvement of PTP in adhesion to MSC, PTP decreased in MSC adhesion after H2O2 treatment. Above results indicated that ROS suppress the cell adhesion and downstream signaling molecules in the regulation of cell adhesion and niche interaction. These findings contribute to a better understanding of the transplantation of MSC on infarcted heart.