Effects Of Magnetic Stimulation On Astrocytes Migration And Underlying Mechanism

Part I:Effects of magnetic stimulation with different intensities on astrocytes migration and underlying mechanismExp I:Effects of magnetic stimulation with different intensities on astrocytes migration[Abstract] Objective To investigate the effects of magnetic stimulation with different intensities on astrocytes migration.Methods Twenty-four adult healthy Sprague-Dawley rats were selected and injected with 0.5μl of 1% ethidium bromide (EB) in PBS into the dorsal spinal cord funiculus of the left side at T10-11 level to make located spinal cord injury models and were randomly divided into four groups.The rats of four groups were exposed to pulsed magnetic stimulation(1Hz,30 pulses) at the following intensities respectively:0 Tesla(control group),0.76 Tesla (Group 0.76T),1.52 Tesla(Group 1.52T),1.9 Tesla(Group 1.9T). On the day 14 after stimulation, the rats were sacrificed and the expressions of glial fibrillary acidic protein(GFAP),5-Bromo-2'-deoxyuridine (Brdu), microtubule associated protein-2 (MAP-2) and the volume of the holes of injuried area of spinal cord were detected with immunohistochemistry and Western-Blot technique. Quantitative analysis of the expression of GFAP, Brdu and MAP-2 were performed with the image analysis system.Results With the increase of magnetic stimulation intensity, the volume of the holes of injuried area of spinal cord discrease at day 14. In the reduced area of the holes,the expressions of GFAP could be seen,while Brdu and MAP-2 could not be seen. Significant differences were revealed in the expressions of GFAP among the four groups, which were significantly higher in magnetic stimulation groups than that in control group (P< 0.05). Conclusion Under the condition of 1Hz, no larger than 1.9Tesla magnetic stimulation, magnetic stimulation promote astrocytes to migrate. After magnetic stimulation, astrocytes migrate into the injuried spinal cord holes. the ability of astrocytes migration increase with the increase of magnetic stimulation intensities.ExpⅡ:The mechanism of different intensities magnetic stimulation promoting astrocytes migration.[Abstract] Objective To investigate effects of ERK inhibitor U0126 of different dose on the ability of 1Hz magnetic stimulation promoting astrocytes migration, then to select suitable dose of U0126, and study the mechanism of different intensity magnetic stimulation promoting astrocytes migration. Methods 24 adult healthy Sprague-Dawley rats were selected to inject 0.5ml of 1% ethidium bromide (EB) in PBS into the dorsal spinal cord funiculus on the left side at the T10-11 level to make located spinal cord injury models and be randomly devided into four groups.The four groups were exposed to magnetic stimulation(1Hz,1.52Tesla, 30pulses) at the following dose respectively:Omg/kg U0126 (control group), 0.1mg/kg U0126(low-dose group),0.2mg/kg U0126(middle-dose group),0.4mg/kg U0126(high-dose group). On the day 14 after stimulation, the rats were killed and the expression of glial fibrillary acidic protein(GFAP),microtubule associated protein-2 (MAP-2), extracellular signal-regulated kinase1/2 (ERK1/2) and the volume of holes were detected with immunohistochemistry and Western-Blot technique. Quantitative analysis of the expression of GFAP, MAP-2 and ERK1/2 were performed with the image analysis system.Results With the increase of U0126 dose,the volume of holes increased on day 14. In the lesion area, the positive expression of GFAP and ERK1/2 could be seen and had identical change tendency,while MAP-2 could not be seen. Significant difference was revealed in the expression of GFAP and ERK1/2 among the four groups, it was significantly lower in the U0126 groups than that in the control group (P< 0.05). while the middle-dose group had similar effect with the high-dose group (P> 0.05).Conclusion U0126 of different dose all could inhibit astrocytes migration that 1Hz,1.52Tesla magnetic stimulation caused,and 0.2mg/kg was the suitable dose.What's more, the effect of 1 Hz,1.52 Tesla magnetic stimulation promoting astrocytes to migrate is relevant to ERK pathway.Part II:Effects of magnetic stimulation with different frequencies on astrocytes migration and underlying mechanismExp I:Effects of magnetic stimulation with different frequencies on astrocytes migration[Abstract] Objective To investigate the action tendency of magnetic stimulation with different frequencies on astrocytes migration. Methods Thirty adult healthy Sprague-Dawley rats were selected and injected with 0.5μl of 1% ethidium bromide (EB) in PBS into the dorsal spinal cord funiculus of the left side at T10-11 level to make located spinal cord injury models and were randomly divided into five groups.The rats of five groups were exposed to pulsed magnetic stimulation (1.52Tesla,30 pulses) at the following frequencies respectively:0Hz(control group, 1Hz(Group 1HzT),5Hz(Group 5Hz),10Hz(Group 10Hz) and 20Hz(20Hz Group). On the day 14 after stimulation, the rats were sacrificed and the expressions of glial fibrillary acidic protein(GFAP),5-Bromo-2'-deoxyuridine(Brdu), microtubule associated protein-2 (MAP-2) and the volume of the holes of injuried area of spinal cord were detected with immunohistochemistry and Western-Blot technique. Quantitative analysis of the expression of GFAP, Brdu and MAP-2 were performed with the image analysis system. Results With the increase of magnetic stimulation frequency, the volume of the holes of injuried area of spinal cord discrease at day 14. In the reduced area of the holes, the positeve expressions of GFAP could be seen, while Brdu and MAP-2 could not be seen. Significant differences were revealed in the expressions of GFAP among the five groups, which were significantly higher in magnetic stimulation groups than that in control group (P< 0.05). Conclusion Under the condition of 1.52 Tesla, no larger than 20Hz magnetic stimulation, Magnetic stimulation promote astrocytes to migrate. After magnetic stimulation, astrocytes migrate into the injuried spinal cord holes. the ability of astrocytes migration increase with the increase of magnetic stimulation frequencies.Exp II:The mechanism of different frequencies magnetic stimulation promoting astrocytes migration.[Abstract] Objective To investigate the effects of ERK inhibitor U0126 of different doses on the ability of lOHz magnetic stimulation promoting astrocyte migration, then to select suitable dose of U0126, and study the mechanism of different frequencies magnetic stimulation promoting astrocyte migration. Methods Twenty-four adult healthy Sprague-Dawley rats were selected to inject 0.5ml of 1% ethidium bromide (EB) in PBS into spinal cord dorsal funiculus on the left side at T10-11 level to make located spinal cord injury models and randomly divided into four groups.The four groups were exposed to magnetic stimulation (10Hz,1.52Tesla, 30 pulses) at the following doses respectively:Omg/kg U0126(control group), 0.1mg/kg U0126(low-dose group),0.2mg/kg U0126 (middle-dose group),0.4mg/kg U0126(high-dose group). On the 14 after stimulation, the rats were sacrificed and the expression of glial fibrillary acidic protein(GFAP), microtubule associated protein-2 (MAP-2), extracellular signal-regulated kinasel/2 (ERK1/2) and the volume of spinal cord injuried holes were detected with immunohistochemistry and Western-Blot technique. Quantitative analysis of the expression of GFAP, MAP-2 and ERK1/2 were performed with the image analysis system. Results With the increase of U0126 dose, the volume of spinal cord injuried hole increase on 14(P< 0.05). In the lesion area, the positive expression of GFAP and ERK1/2 could be seen and had identical change tendency,while MAP2 could not be found. Significant difference was revealed in the expression of GFAP and ERK1/2 among the four groups, and it was significantly lower in U0126 groups than that in control group (P< 0.05). while the middle-dose group had similar effect with the high-dose group (P> 0.05) Conclusion U0126 of different dose all could inhibit astrocytes migration that 1.52Tesla,10Hz magnetic stimulation caused, and 0.2mg/kg was the suitable dose. What's more, the effect of 1.52Tesla,10Hz magnetic stimulation promoting astrocytes migrate is relevant to ERK pathway.