| Objective: To observe the changes of ultrastructure of Neurovascular Unit(NVU)at different time points in the peri-ischemic brain tissue of rats after cerebral ischemia reperfusion(CIR)and the expression changes of neuronal nuclear antigen(NeuN),glial fibrillary acidic protein(GFAP)and laminin(LN)at different time points,to investigate the effects of Panax Notoginseng Saponins(PNS)on the ultrastructure of NVU in brain tissue of rats and related protein expression after cerebral ischemia,and further clarify that PNS has a protective effect on cerebral ischemia.Methods:(1)Male Sprague-Dawley rats were randomly divided into 3 groups: sham operation group,model group and treatment group,and each group was divided respectively into four subgroups at 24 h,72 h,7 d,and 3 w after cerebral ischemia-reperfusion,12 rats in each subgroup.The middle cerebral artery occlusion(MCAO)model was made by the suture-occluded method.The neurological function scores in every rat at 4 h after operation were recorded according to Longa's standard,after intervention in each group,and the neurological scores were recorded respectively on 24 h,72 h,7 d and 3 w after MCAO.(2)The expression changes of NeuN,GFAP and LN were detected by western blot(WB).(3)The ultrastructural changes of NVU in the peri-ischemic brain tissue of rats in each group were observed by transmission electron microscope at different time points.Results:(1)Neurological function score: there was no significant difference in the neurological scores between treatment group and model group postoperative 4 h;the neurological function score in treatment group improved gradually after PNS intervention,on24 h after MCAO,compared with the model group,the difference was not statistically significant(P>0.05);however,on 72 h,7 d and 3 w after MCAO,there was statistical difference significantly compared with the model group at the same time(P<0.05).(2)Visual observation of the rat brain with MCAO model: IR 3w after MCAO showed obvious cerebral infarction in the model group,the surface of cerebral cortex was pale and swollen,infarcted central area with varying degrees of brain tissue atrophy,the branches of blood vessels were small and unclear;the cerebral cortex area of the treatment group was not obvious pale on the surface,and the tissue swelling was lighter than that of the model group,the central area of infarct was smaller than the model group,and the blood vessel distribution was more abundant than that of model group,its branches were numerous and the lines were clearly visible.(3)Western blotting results of NeuN,GFAP and LN: there was no significant change in the expression of NeuN,GFAP,and LN proteins in the sham operation group at each time point.The expression of NeuN protein in the model group was lower than that in the sham operation group at all time points,and the expression of IR on 24 h,7 d,and 3 w was significantly different from that of the sham operation group(P<0.05);the expression of NeuN in the treatment group gradually increased,its expression levels at each time point were higher than those in the model group,after IR 72 h,the NeuN expression level was almost normal,and on 7 d and 3 W compared with the model group at the same time point,the difference was statistically significant(P<0.05).The relative expression level of GFAP protein in the model group increased gradually and was still lower than that in the sham group,the GFAP expression at each time point was significantly different from that in the sham operation group(P<0.05);the relative expression level of GFAP at each time point in the treatment group was higher than that in the model group,its expression on 72 h,7 d,and 3 w was statistically significant compared with that of the model group(P<0.05),after IR 7 d,the expression of GFAP protein in the treatment group was almost normal.The expression of LN in the model group increased gradually and still lower than the sham operation group at eachtime point,its expression on 24 h,72 h,7 d was statistically significant compared with the sham operation group(P<0.05),However,there was no significant difference between 3 w and the sham operation group(P>0.05);the expression level of LN protein in the treatment group was higher than that in the model group at each time point,and its expression on 7 d and 3 w was significantly different from that in the model group(P<0.05).(4)Respectively on 24 h,72 h,7 d and 3 w after MCAO,electron microscopy showed that the pathological damage changes of the ultrastructure of NVU in the peri-ischemic brain tissue of rats in the each treatment group were all significantly lightened than that in the model group at the same time.Conclusion:(1)The protective effect of PNS on NVU after cerebral ischemia may be related to the up-regulation of NeuN,GFAP and LN protein expression after cerebral ischemia.(2)PNS can reduce the pathological changes of NVU injury after cerebral ischemia.(3)It indicates that PNS can protect cerebral ischemia by integrationing to promote the repairment of NVU after cerebral ischemia,and improving neurological deficit symptoms. |
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