The Cerebral Protection Research Of ENT1 Involved In The Ischemia-reperfusion Rats

Objective: To observe the dynamic expression and cellular localization of equilibrativenucleoside transporter-1(ENT1)of the ischemia-reperfusion rats.And to investigate the effect of equilibrativenucleoside transporter-1(ENT1)inhibitor,S-4-Nitrobenzyl-6-thioino sine(NBTI),on the degree of the neurological functional defects and the infarct volume of the ischemia-reperfusion(IR)rats.Methods: Male Sprague-Dawley(SD)rats were randomly divided into normal group,sham-operated group,model groups,dimethyl sulfoxide(DMSO)-treated group and NBTI-treated group.The model groups were further studied randomly at five time points,3 h,6 h,24 h,72 h,and 1 w,following the reperfusion.Middle cerebral artery occlusion was used to establish the reperfusion models of rats.Immunohistochemistry,immunofluorescence and Western Blot were used to detect the expression of ENT1 in each group.Zea-Longa score and TTC staining were used to evaluate the neurological deficits and detect the infarct area of the ischemic brain respectively.Results:1.The Western Blot results showed that in the normal group and sham-operated group,ENT1 was expressed in the basic level.Compared with the normal group and sham-operated group,ENT1 expression in the model group 3 h after reperfusion were significantly decreased(P<0.05).The expression of ENT1 was increased at 6 h and 24 h after reperfusion,compared with normal group and sham operation group,there was no significant difference(P>0.05).With time went,ENT1 expression decreased gradually.And with reperfusion time extended,ENT1 expression in the model group 72 h after reperfusion and the model group 1 w after reperfusion were significantly decreased compared with the normal group and sham-operated group(P < 0.05).2.The results of immunohistochemistry showed that ENT1 was widely expressed in the hippocampus CA3 area.And Immunofluorescence results showed that ENT1 immunoreactivity was located in neuronal and astrocytic membrane and cytoplasm.3.The Zea-Longa score results showed that scores of normal group,sham-operated group,IR groups(3 h,6 h,24 h,72 h,1 w),NBTI group and DMSO group were respectively(0)score,(0)score,(2.42+0.51)score,(2.50+0.52)score,(2.67+0.49)score,(2.58+0.51)score,(2.42+0.51)score,(2.20+0.45)score,(2.60+0.55)score.Different degree of neurological deficit was found in IR groups,NBTI group and DMSO group.There was no statistical significance in the difference of Zea-Longa score of IR groups and DMSO group(P>0.05).Take the model group 24 h after reperfusion's sample using NBTI intervention,NBTI decreased Zea-Longa score significantly(P< 0.05).4.The results of the TTC staining showed that infarct volume of normal group,sham-operated group,IR groups,NBTI group and DMSO group respectively were(0)%,(0)%,(52.84±6.95)%,(45.80±3.15)%,(45.41±14.65)%,(49.72±4.31)%,(51.69±5.55)%,(18.00±3.09)%,(50.66±7.47)%.Infarct area was not found in the normal group and sham-operated group,but it was found in IR groups,NBTI group and DMSO group.With no statistical significant among the seven groups(P>0.05).Take the model group 24 h after reperfusion's sample using NBTI intervention,the infarction volume of NBTI group was significantly decreased(P<0.05).Conclusions: The decrease in ENT1's expression in the IR models of rats.ENT1 inhibitor,NBTI,reduces the neurological deficit score and cerebel infarction volume,indicating that ENT1 may be involved in the protection of cerebral ischemia-reperfusion.