The Effect of Exosomes Secreted by Astrocytes on the Vitality of Neural Stem Cells

ZHOU Shao-ting, ZHAO Jing, XU Dong-sheng

Abstract

To investigate the effects of exosomes of mouse astrocytes on the viability of neural stem cells.  Methods  Cultured and isolated the mouse astrocytes, and collected the cell supernatant for obtain the exosomes by ultracentrifugation. Neural stem cells that primary cultured for 2nd to 6th generation were obtained and treated with medium contained 0, 20, 40, 60 μg/mL of exosomes respectively. Screening the optimal exosome concentration for culturing neural stem cells by CCK-8 method. The optimal exosome concentration for neural stem cells was 40 μg/mL according to CCK-8 results. Then cells were intervened with 40 μg/mL of exosome in experimental group for 72 h, and the control group was added with the same volume of PBS. After intervention, the positive stem cells were labeled with EdU kit. Using the Transwell model, the number of nucleus stained by DAPI in the lower chamber in 40 μg/mL exosome treatment group and the control group were counted under a fluorescence microscope.  Results  ① Identification of astrocyte exosomes: The successful obtain of exosomes of cell supernatant were confirmed by techniques such as electron microscopy, Western blot, exosome concentration and particle size measurement. ② CCK8 experiment: As the increasement of the concentration of exosomes, cell proliferation of primary neural stem cells gradually increased. Compared with the control group, proliferation of the cells in 40 μg/mL and 60 μg/mL exosome treatment groups was significantly enhanced, but there was no significant difference between the two groups. So, 40 μg/mL was selected as the best intervention concentration. ③ EdU detection: Number of EdU positive labeled cells in the 40 μg/mL exosome group was higher than that in the control group (P<0.05). ④ Transwell experiment: In the Transwell model, more neural stem cells in the 40 μg/mL exosome group migrated from the upper layer to the lower layer of the Transwell membrane, and the number was higher than that of the control group (P<0.05).  Conclusion  Mouse astrocyte exosomes can improve the viability of neural stem cells.

 

Keywords: Exosome, Neural stem cells, Stroke

 

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