Experimental Research Of The Expression And Neuroprotection Of Nix After Subarachnoid Hemorrhage In Rats

Objective:1.To investigate the expression of Nix in brain tissue after subarachnoid hemorrhage(SAH)in rats.2.To study the role of Nix in early brain injury(EBI)after SAH in rats.Methods:In experiment 1,healthy adult male Sprague-Dawley(SD)rats were randomly assigned to 8 groups of 12 rats each,a sham group and 7 SAH groups arranged by time:3h,6h,12h,24h,48h,72h and 7d after SAH.A rat SAH model was induced by injecting 0.3 ml of autologous non-heparinized arterial blood into the prechiasmatic cistern.All rats in the experiment were killed at the matching time point after SAH.The brain tissue of six rats in each group was taken for Western blot and immunofluorescence analysis respectively,to investigate the expression of Nix.The experimental results were statistically analyzed.In experiment 2,healthy adult male SD rats were randomly divided into 6 groups of 18 rats each:sham group,SAH group,SAH+vector group,SAH+over-Nix group,SAH+Ctr siRNA group and SAH+Nix siRNA group.The brain tissue of six rats in each group was taken for western blot analysis to test the expression level of Nix and mitochondrial marker protein TOMM20.Another six rats were killed for terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling(TUNEL)staining and Fluoro-Jade B(FJB)staining,to evaluate the neuro apoptosis and degeneration.The last six rats were used for neurological score and spatial learning and cognitive capacity,which was tested using the Morris water maze.Results:In experiment 1,the level of Nix expression was higher than sham group after SAH and peaked at 24h,and Nix was mainly expressed in neurons.Based on these findings,subsequent experiments focused on the 24h after SAH.In experiment 2,Nix overexpression plasmid and siRNA can separately increase and decrease the expression level of Nix after SAH;separately decrease and increase the protein level of TOMM20 after SAH;separately increase and decrease the number of neuronal apoptosis and cell degeneration;separately ameliorate and exacerbate the neurological deficits and cognitive capacity.Conclusions:The expression of Nix is increased in neurons after experimental SAH in rats,and may play a neuroprotective role in EBI following subarachnoid hemorrhage.