| Objective:To explore the changes of exploration ability,neurological functional defects,pathophysiologic,and infarct volume in the ischemia-reperfusion(IR)rats.And To observe the dynamic expression and cellular localization of Kalirin and Rac1 in the hippocampus nerve cells,to investigate the correlation of rats'exploration ability and neurological functional defects between the levels of Kalirin,Rac1 in rats'hippocampus tissue after ischemia-reperfusion.Methods:SD rats were randomly divided into normal group,sham operation group,ischemia reperfusion(IR)group(6h?12h?24h?72h?1W?2W),and the IR injury model was established by using middle cerebral artery occlusion(MCAO)method.The modified Zea-Longa score was used to evaluate the neurological defects in rats.Smart3.0 behavioral test system automatically collects the motion track and moving distance of rats in the open-field experiment to observe the change of exploration ability of rats.The triphenyltetrazolium chloride(TTC)staining was used to determine the content of ischemic brain tissue,and the infarct volume was calculated by image-pro plus(IPP)software.Histopathologic changes were detected by hematoxylin-eosin(HE)staining.Immunohistochemistry(IHC)were used to localize the expression of Kalirin and Rac1.Then the Western blotting(WB)assay was used to make a quantitative analysis of the expression of Kalirin and Rac1.Correlation analysis of Kalirin and Rac1 with the behavioral changes(neurological functional defects and exploration ability)was also performed by WB in rats.Results:1.The results of modified Z-Longa score showed that the neurological deficit scores of normal group,sham operation group,6h group,24h group,72h group,1W group and2W group were(0.00),(0.20±0.45),(2.40±0.54),(2.20±0.45),(1.60±0.54),(0.80±0.45),(0.6±0.54),respectively.The rats in IR model groups had neurological deficit symptoms at different time points,and the modified Z-Longa scores in 6h,24h groups were significantly different with those in normal group and sham operation group(P<0.05).There was no significant difference in Z-Longa scores between the and 72h,1W and 2W groups and the sham operation group(P>0.05).2.The results of open-field test showed that the moving distance of normal group,sham operation group,6h group,24h group,72h group,1W group and 2W group were(188.65±8.60 cm),(182.35±8.60cm),(81.26±11.42cm),,(115.26±16.27cm),(153.31±11.94cm),(183.68±10.96cm)and(188.65±18.91cm),respectively.The moving distance decreased in the 6 h,24 h and 72 h groups compared with sham operation group.The difference was statistically significant(P<0.05).There was no significant difference in moving distance between the 1W,2W groups and the sham operation group(P>0.05).The results suggested that the spatial exploration ability of rats decreased after 6 h of IR injury,and then following an ascend trend.Finally,the moving distance of IR 2W group approximated to the sham operation groups.3.The results of TTC staining showed that there was no infarction in normal group and sham operation group,and infarction was found in all time points after IR injury.The proportion of infarct area at 6h group,24h group,72h group,1W group and 2W group was(50.84±6.95%),(45.80±3.15%),(45.41±14.65%),(49.72±4.31%)and(51.69±5.55%),respectively.There was no significant difference in infarct volume ratio(P>0.05).4.The results of HE staining showed that there were edema of pericellular space,nuclear pyknosis and cell necrosis in the ischemic tissue of IR rats.The structure of hippocampal neurons in normal group and sham operation group was clear,the size of nucleus was regular,and no obvious pathological changes were observed.5.The results of IHC showed that Kalirin was expressed in the nucleus of hippocampal neurons and Rac1 was expressed around the nuclei of hippocampal neurons in rats.6.The results of WB showed that Kalirin and Rac1 expressed in the brain tissue of rats in each group.The basic expression of Kalirin and Rac1 were found in the normal group and sham group.,There was no significant difference between the two groups(P>0.05).Compared with the sham group,the expression level of Kalirin increased in IR 6h and 24h group(P<0.05).and decreased back to the basic level.in IR 72h,1W,2W.there were no significant difference in the expression of Kalirin between the IR 72 h,1 W,2 W groups and sham operation group(P>0.05).However,the expression of Rac1 decreased at IR 6 h and 24 h groups,which were significantly different from that in the sham operation group(P<0.05).With the passage of time,the expression of Rac1 increased,There was no significant difference in the expression of Rac1 between IR 72h,1W,2W groups and sham operation group(P>0.05).7.The results of correlation analysis showed that the expression level of Kalirin in MACO model rats was positively correlated with the neurological function score of rats(r_s=0.74,P<0.05).It was negatively correlated with the total distance of moving in rats(r=?0.76,P<0.05).The expression level of Rac1 was negatively correlated with the neurological function score of rats(r_s=?0.81,P<0.05).It was positively correlated with the total distance of moving in rats(r=0.78,P<0.05).Conclusion:The Kalirin and Rac1 is dynamically expressed in the MACO rats,and it is correlated with praxiology.The changes of exploration ability in rats may be related to the increase of Kalirin expression and the decrease of Rac1 protein level.Kalirin and Rac1proteins may be involved in the process of IR injury. |
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