The Study Of Neurogenesis Contributed By The Increased Expression Of Survivin In Hippocampus After Traumatic Brain Injury

Objective:To investigate the expression changes of Survivin after traumatic brain injury (TBI); and further to study the relationship between the up-regulation of Survivin and endogenous neurogenesis. Throught this study, we preliminarily demonstrate the role of increased Survivin to promote the endogenous neurogenesis; and detect the possible mechanism in this process. To find the approach that can promote restoration of nervous function and to provide a new idea for the clinical treatment of TBI.Methods:C57BL/6mice,25-30g body weight, total n=146, were randomly divided into sham operation group and TBI group, the mice brains were removed for Quantitative Real-Time Polymerase Chain Reaction, Western blot analysis and immunofluorescence staining at designated times. All mice were fed for1week after purchase. TBI group was subjected to (202±2) KPa lateral fluid percussion to establish models of brain injury, and sham-operation group was subjected to surgery with no lateral fluid percussion.1. Mice used for PCR were sacrificed at12hours,1,2,5,7and14days post-TBI (each time point n=6), and hippocampus tissues from the ipsilateral hemispheres were carried out for analyze the expression changes of Survivin mRNA. Mice in sham-operation group (n=6) were sacrificed at day3after operation, and ipsilateral hippocampus were removed for PCR.2. At each time point as mentioned above (n=6), mice in TBI and sham-operation group used for Western Blot were sacrificed, and hippocampus tissues from the ipsilateral hemispheres were carried out for analyze the expression changes of Survivin protein.3. We adopted immunofluorescence staining at day3post injury, when Survivin protein is steadily expressed after TBI. Mice used for BrdU staining received intraperitoneal injections of BrdU (twice daily) for3days before that been sacrificed, and mice used for DCX, NeuN and GFAP staining were without injection. Each kind of staining group was divided into TBI group (n=4) and sham-operation group (n=4). Survivin/BrdU, Survivin/DCX, Survivin/GFAP and Survivin/NeuN double immunofluorescence staining were carried out respectively. All slices were observed by fluorescence microscopy and confocal microscope and record the number of positive cells.Result:1. The Quantitative Real-Time PCR analysis showed that the expression of Survivin mRNA is increased after TBI and expressed in a time-dependant fashion. Survivin activation was present at12hours following injury and was sustained until day5, with a peak at day1(P<0.01), and then declined to level of sham operation group at day14.2. Western blot analysis revealed that the up-regulation of Survivin protein starting at day1(P<0.05) and lasting to day7, peaking at day2(P<0.01).3. Compared to sham group, the number of Survivin (+) cells in SGZ of ipsilateral hippocampus was significantly increased after TBI by immunofluorescence staining.4. The results of immunofluorescence staining revealed BrdU/Survivin co-labeled cells (93.82±3%) and DCX/Survivin co-labeled cells (61.28±4%) in SGZ of DG Besides, there were minority of GFAP (+) cells expressing Survivin, but no NeuN (+) cells expressing Survivin in adult DG (dentate gyrus) of hippocampus after TBI.5. Immunofluorescence staining results also revealed that the number of BrdU (+) and DCX (+) cells were significantly increased (P<0.001) in SGZ of DG after TBI compared with sham-operation group.Conclusion:1. The expression of Survivin, an antiapoptotic protein, increased in a time-dependence manner in the SGZ of hippocampus after TBI.2. Increased Survivin is expressed in neural stem cells, immature neurons and minority of astrocyte, but not that of mature neuron in adult DG of hippocampus after TBI.3. Increased expression of Survivin correlate to the proliferation of neural stem cell in DG of hippocampus. Survivin gene was likely to promote the proliferation of neural stem cell and restoration of nervous function early after TBI.4. Significant increased expression of Survivin in DCX-positive immature neurons in DG of hippocampus early after TBI. This suggested that Survivin may play a role in the survival of immature neurons, and promote these immature neurons to differentiate into granular neurons.