| Objective: To observe the pathological changes of brain tissue, the infarct volume and the changes of EAAT2’s(excitatory amino acidstransporter-2)expression after middle cerebral artery occlusion followed ischemiare perfusion in rats and clarify the effect of NBTI(S-4-Nitrobenzyl-6-thioinosine), an ENT1(equilibrativenucleoside transporter-1) inhibitor, on the expression of EAAT2.Methods: SD rats were randomly divided into normal group, sham operation group, various time points(3h, 6h, 24 h, 72 h, 1week) ischemia reperfusion groups and ENT1 inhibitor group. Cerebral ischemia reperfusion model of rats were built by middle cerebral artery occlusion(MCAO). In this study, TTC(triphenyltetrazolium chloride) staining was used to stain brain tissue, analyzed the infarct volume by IPP(image-pro plus) software and calculated the volume ratio of cerebral infarction. HE(hematoxylin--eosin) staining was used to observe the pathological changes of brain tissue in rats. Immunohistochemistry and Western blotting assay were used to make a location and quantitative analysis of the EAAT2’s expression of brain tissue.Results:The results of the TTC staining showed that the normal group and sham operation group were not found infarct sections, and infarct sections were found in all various time points’ ischemia reperfusion groups. There was no statistical significance in the difference of infarct volume ratio of each various time points’ ischemia reperfusion group(p>0.05).The results of the HE staining showed that the structure of hippocampus in non-infarct side was clear, the size of nucleus was regular, and no abnormal pathological changes were found. However, most nerve cells of brain tissue in the ischemic area of the infarct side were dead and showed nuclear condensation. The nuclear morphology was irregular. Western Blotting showed that EAAT2 had basic expression in normal group and sham operation group. Compared with the normal group and the sham operation group, the expression of EAAT2 was significantly increased after 3 hours or 6 hours of ischemia reperfusion. And the difference has statistical significance(p<0.05).With the prolongation of the time of ischemia reperfusion(24hours, 72 hours or one week), the expression of EAAT2 decreased gradually. Immunohistochemistry results showed that EAAT2 is mainly expressed on the membrane of nerve cells. in the hippocampus CA1,CA3 and DG area. Finally, the volume of cerebral infarction was significantly reduced after the use of NBTI(an ENT1 inhibitors). And there is a significant difference compared with the control group.and the result of Western Blotting showed that the expression of EAAT2 of the inhibitor group was significantly increased compared with reperfusion group(24h hours). The difference was statistically significant(p<0.05).Conclusion: ENT1 inhibitor can reduce the volume of cerebral infarct, and the mechanism of ENT1 inhibitor may increases the expression of EAAT2. |
PDF Full Text Request