Investigation Of Stem Cell-like Cells Derived From Spherical Culture Of Astrocytes

It is challenging to obtain a large number of neural precursor cells(NPCs)for the repair of particularly central nerous injuries.This study was to investigate whether the three-dimensional(3D)culture strategy was able to help induce murine astrocytes(ASTs)into NPC-like cells and whether the derived cells are able to promote nerve regeneration and improve neurological function in injuried mice.Mouse astrocytes cultured on agarose spontaneously formed 3D spheroids.The expression levels of stem cell marker genes were characterized by RT-PCR,immunofluorescence staining,flow cytometry,Western blotting and alkaline phosphatase staining.It was found that various stem cell marker genes of spherical astrocytes were enhancing at both gene and protein levels over a period of cultivation time,resulting in NPC-like cells.Sequencing analysis was performed on murine astrocytes,murine astrocyte-derived NPC-like cells,P56-NPCs,and P0-NPCs,and the results confirmed that murine astrocytes were induced into NPC-like cells(termed AST-derived stem cells,A-iSCs)on agarose.KEGG signal pathway analysis suggested that the PI3K-Akt signaling pathway,Rap1 signaling pathway,and Notch signaling pathway were involved in the cell induction process.The in vitro differentiation potential of A-iSCs into functional neurons was chacraterized by immunofluorescence staining,flow cytometry,calcium imaging,and patch-clamp assay.Transplantation of A-iSCs promoted the rapir of nerves and improved neurological functions in mice of MCAO,SCI,and PNI models,suggesting the potantial of A-iSCs in promoting nerve regeneration and improving neurological functions regardless of the sites or types of nerve injuries.Derivitaion of NPC-like cells from astrocytes provides a novel,sufficient and safe source of NPCs for nerve regeneration,and offers a broad spectrum solution for the treatment of neurological injuries.