The Expression Of GFAP Following Radiation Induced Brain Injury And Analysis Of Gfap Expression After CD34~+ Cells Transplantation

Objective In this study,we analyze the dynamic changes of GFAP expression in cerebral cortex in each period,and then explore the feasible that GFAP can be used as a protein marker to evaluate radiation-induced brain injury.Methods1.Animals grouping and radiation-induced brain injury(RIBI)model making Sixty healthy adult male Sprague-Dawley(SD)rats(body weight,210-230g)were randomly divided into sham-operated group(n=12),RIBI control group(n=36)and RIBI treating group(n=12).Rats in the RIBI control group were randomly subdivided into three groups(each n=12)for analysis on days 7,14 and 28 after irradiation.The RIBI treating group which was injected with CD34+ cells on 7 days after the establishment of RIBI models was compared with control group on day 14 and 28 after RIBI(each n=6).RIBI model was established by computed tomography(CT)scanning.2.CD34+ cells preparation and transplantation CD34+ cells were abstracted and purified by flow cytometry(FCM)and were labeled by 5-bromo-2-deoxyuridine(Brd U).Every rat in RIBI treating group received an infusion of CD34+ cells labeled with Brd U via the tail vein at 7 days after radiation.3.The detection of various indicators We observed the capillary permeability between the sham-operated group and the RIBI control group using Microscope vascular camera device and microcirculation video recording system.EB dyeing of intravascular and extravascular were analysed by measuring the IOD using Image-Proplus 5.0 medical image software and the ratio of IOD about EB dyeing(IOD of extravascular/the sum IOD of extravascular and intravascular)was calculated as observation indicator.The expression of GFAP and Brd U(brown staining)in each section were captured in five fields by two independent observers blinded to the experimental conditions.The overall expression level of GFAP was determined by measuring the IOD of each image using Image-Proplus 5.0 medical image analysis software.Results1.After radiation-induced brain injury,the EB extravasation was obvious when viewed through microscope equipped with vascular camera device and Microcirculation video recording system.The EB extravasation ratio in the RIBI control group was obviously higher than that in the sham-operated group at three time points.2.In this study,the expression of GFAP in rat brain sections was evaluated by immunohistochemistry analysis.The level of GFAP was increased at day 7 in the RIBI control group compared with the sham-operated group,and subsequently decreased from day 14 to day 28,but still higher than the sham-operated group.Compared to the RIBI control group,the positive expression of GFAP was significantly reduced in the animals receiving CD34+ cells.3.The presence of CD34+ cells labeled with exogenous Brd U in RIBI treating group was detected through the method of immunohistochemistry staining at 14 and28 days after irradiation.Conclusions1.We use the CT X-ray irradiation to make radiation brain injury model of rats in a particular area,through analysis of this study,which confirm that this model have a good clinical simulation and it can be reused.2.This study using EB as tracer to detect the capillary permeability after radiation-induced brain injury,which can demonstrate visualized radiation brain injury animal models to make success.3.In this experiment,we adopt GFAP as observation indicator,the expression was evaluated by immunohistochemistry,and then it was found that the level of GFAP was obviously increased in the RIBI control group compared with the sham-operated group,which was consistent with the change of EB extravasation viewed by Microscope vascular camera device and microcirculation video recording system after radiation brain injury;and then the expression level was significantly decreased after intravenous delivery of CD34+ cells compared with RIBI control group.This strongly indicates that GFAP expression can be used to evaluate the status of brain injury and repair.4.In the present study,we opted to transplant CD34+ cells via the tail vein on day 7after irradiation and demonstrated that CD34+ cells labeled with Brd U could be detected by immunohistochemistry in the RIBI treating group,this indicates that the implantation technique is feasible.Moreover,this result can serve as the basis for future studies.