The role of repetitive trans-spinal magnetic stimulation in regulating the activation of spinal cord derived neural stem cells through Notch1

English Keywords:Neural stem cells  Repetitive trans-spinal magnetic stimulation  Notch1  Activation  Mice

English Abstract:

  【Abstract】 Objectives: To observe the effect of repetitive trans-spinal magnetic stimulation(rTSMS) on the activation of spinal cord derived neural stem cells(NSCs) through Notch1. Methods: The complete spinal cord of fetal C57BL/6 mice at 16 weeks of gestation was promptly extracted following euthanasia. NSCs obtained from the spinal cord were cultured in vitro. Subsets and rate changes of NSCs were analyzed using single cell sequencing and RNA profiling. The cultured NSCs were divided into control group, intervention(rTSMS) group and experiment(rTSMS+Notch1 shRNA) group. The control group was treated with sham stimulation. The rTSMS group was treated with 20Hz high frequency stimulation with 1500 pulses. The rTSMS+Notch1 shRNA group received the same rTSMS intervention after transfection with Notch1 shRNA lentivirus. The expression of Notch1 was detected by Western blot, and the activation of NSCs was detected by immunofluorescence staining and sphere formation test. Results: Spinal cord derived NSCs cultured in vitro could differentiate into neurons, astrocytes and oligodendrocytes. Single cell sequencing showed that NSCs could be divided into 6 subgroups with special heterogeneity, and Notch1 mainly expressed in the subgroups of dNSCs and pNSCs. The expression of Notch1 in the cells decreased significantly on the 5th day after transfection with Notch1 shRNA lentivirus (Control group: 1.080±0.086 vs Experiment group: 0.520±0.041, P<0.05). rTSMS intervention could significantly up-regulate the expression of Notch1(Control group: 1.030±0.068 vs Intervention group: 1.480±0.087, P<0.05). The diameter of the neurosphere(Control group: 88.5±7.5 vs Intervention group: 106.2±8.8 vs Experiment group: 89.2±8.1, P<0.05) and the rate of BrdU positive cells were significantly increased(Control group: 38.6±3.7 vs Intervention group: 50.3±4.6, P<0.05) after rTSMS intervention. After Notch1 shRNA, the neurosphere diameter and BrdU positive cell rate(Intervention group: 50.3±4.6 vs Experiment group: 35.1±3.5, P＜0.05) significantly decreased in the experiment group. Conclusions: rTSMS can promote activation of NSCs through Notch1 in vitro.