Previous studies have shown that the injured spinal cord is not a favorable environment for the survival, migration, and differentiation of the donor cells [12,13]. light of recent findings that Rho family GTPases regulate stem cell functions, genetic manipulation of Rho GTPases can potentially control phenotypes of transplanted cells. Therefore CP-466722 we expressed mutant forms of Rho GTPases, Rac, Rho, and Cdc42, in the neural stem/progenitor cells and examined their survival and migration after transplantation. Results Manipulation of the individual Rho GTPases showed differential effects on survival, with little variance in their migratory route and predominant differentiation into the oligodendroglial lineage. Combined suppression of both Rac and Rho activity experienced a prominent effect on promoting survival, consistent with its highly protective effect on drug-induced apoptosis in culture. Conclusion Manipulation of Rac and Rho activities fully rescued suppression of cell survival induced by the spinal cord injury. Our results indicate that precise regulation of cell autonomous factors within the donor cells can ameliorate the detrimental environment created by the injury. Background Neural stem/progenitor cells (NSPCs) are widely present in the developing mammalian central nervous system (CNS) and known for their capability of self-renewal and potential of differentiation into multicellular lineages VCA-2 [1,2]. The protocols of in vitro differentiation and maintenance of NSPCs have been established [3,4], and transplantation of NSPCs is usually thought to be an CP-466722 important approach toward functional restoration of the damaged CNS tissue, including injured spinal cords [5-7]. Previous studies have shown partial functional improvement after spinal cord injury by transplantation of NSPCs derived from the embryonic CNS or embryonic stem cells [8-11]. Although these results suggested the potential of NSPC transplantation in the improvement of spinal cord function, there have been a small number of reports on basic mechanisms of NSPC survival in the host spinal cord environment. Previous studies have shown that the hurt spinal cord is not a favorable environment for the survival, migration, and differentiation of the donor cells [12,13]. Therefore, identification of regulatory mechanisms of NSPC survival and differentiation both in the intact and injured spinal cord environment should be important. The Rho family small GTPases, users of the Ras superfamily, are known to regulate cell shape, movement, and adhesion in multiple mammalian cell types [14]. Rho GTPases also have been demonstrated to activate a number of transmission transduction pathways involved in cell cycle progression, gene expression, and cell survival [15]. In the context of development and maintenance of neurons and glial cells in the CNS, negative functions of Rho family GTPases in cell survival have been implicated. For example, Rac/Cdc42 GTPases promote the apoptotic death of NGF-deprived sympathetic neurons [16,17] and activation of Rac by p75 neurotrophin receptor (p75NTR) induces apoptosis via activation of c-jun N-terminal kinase (JNK) in oligodendrocytes [18]. Rho activation is usually prominently enhanced in the hurt spinal cord and involved in p75NTR-dependent apoptosis [19]. These experimental evidences suggest negative functions of Rho GTPase in survival of transplanted NSPCs in an adverse microenvironment of the injured spinal cord. In this study we recognized Rho and Rac GTPase activity as a potent regulator of cell survival after NSPC transplantation. Recombinant adenovirus-mediated expression of dominant-negative forms of RhoA and Rac1 increased the survival of transplanted cells more then two-fold in the intact spinal cord. Furthermore, expression of RhoDN and RacDN in NSPCs fully rescued down-regulation of cell survival after the transection of spinal cords. Taken CP-466722 together with strong protective effects of the same genetic manipulation against chemically induced apoptosis in vitro, these results show that Rho GTPase is one of the crucial cell- autonomous CP-466722 factors promoting successful integration and survival of NSPCs in the hurt spinal cord. Results Suppression of NSPC survival in the hurt spinal cord environment Our previous study of NSPC transplantation into the intact spinal cord indicated advantage of using hippocampus-derived NSPCs as a donor source in comparison with spinal cord-derived NSPCs [20]. Additionally, hippocampus-derived NSPCs showed more preferential migration toward the white matter of the host spinal cord. This preferential association with the white matter and directed differentiation of NSPCs toward the oligodendroglial lineage are advantageous in facilitating remyelination in the hurt spinal cord. In this study, we first examined whether hippocampus-derived NSPCs show similar survival and migratory behavior in the hurt spinal cord. We utilized total transection of the neonatal spinal cord as a model of the spinal cord injury. In this injury model, the location and extent of the lesion can be precisely controlled. We generated total transection of the spinal cord at the level of T9-10 on postnatal day 7. Transplantation.