Bone Mesenchymal Stem Cell-conditioned Medium Promotes The Effective Differentiation Of Neural Stem Cells By Inhibiting Notch1 Signaling Pathway

BACKGROUND: Spinal cord injury(SCI)can lead to severe motor sensory disturbance,which has a great impact on patients' lives in recent years.According to different stages of trauma,cell therapy has great potential for tissue regeneration after spinal cord injury.(NSCs)is a kind of pluripotent stem cells with inherent differentiation ability,especially related to the promotion and reconstruction of injured neuronal spinal tract.By providing neurotrophic support and restoring synaptic connections,neural stem cells regenerate the microenvironment of the injured spinal cord after spinal cord injury and are not conducive to the growth and differentiation of transplanted cells.limited by a series of inhibitory factors produced by pathological injury,exogenous neural stem cell transplantation can not achieve the best effect,partly because of neuronal differentiation and neural network reconstruction.Previous studies have found that when NSCs is co-cultured with BMSCs,neuroglobular cells produce more neurons and oligodendrocyte astrocyte proliferation is a key factor in glial scar formation.Notch signaling pathway coordinates a large number of cell operations,including cell proliferation,metabolism and differentiation,regulates cell survival in development and adulthood,and plays an important role in nervous system development.Including neuronal differentiation and glial production.OBJECTIVE: To discuss the effect of BMSC-CM on differentiation of NSCs and the role of Notch1 signal pathway in this process.METHODS: NSCs and BMSCs were cultured in vitro and NSCs were cultured in different differentiation media.The experimental cells were divided into four groups: control group(NSCs),Jagged group,BMSC-CM group and BMSC-CM+Jagged group.The differentiation of NSCs in each group was identified by immunofluorescence double staining technique.The expression levels of MAP-2,GFAP and the key proteins of Notch1 pathway,Notch1,NICD and target genes Hes1 and Hes5,were detected by Western blot.RESULTS: The results of immunofluorescence double staining and Western blot showed that compared with the control group,the level of MAP-2was up-regulated and the level of GFAP was down-regulated in BMSC-CM group.At the same time,the production of key proteins Notch1,NICD and target genes Hes1 and Hes5 in Notch1 pathway were reduced.In the Jagged group,the results were completely opposite,down-regulating the level of MAP-2 and up-regulating the level of GFAP,while increasing the production of protein Notch1,NICD and Hes1 and Hes5,while the BMSC-CM+Jagged group was close to the control group and had no significant change.Compared with Jagged group,MAP-2 increased and GFAP decreased in BMSC-CM+Jagged group.And,the production of Notch1,NICD,Hes1 and Hes5 were decreased.CONCLUSION: These results suggest that when the Notch signal is activated,it promotes the fate of differentiation into astrocytes at the cost of reducing neurons.In vitro,BMSC-CM can promote NSCs to differentiate into neurons and reduce the production of astrocytes by inhibiting Notch1 signal pathway.