| Objective:Neonatal Hypoxic-Ischemic Brain Damage (HIBD) caused in most cases by perinatal asphyxia. Brain edema is an important element of pathophysiology changes after HIBD, so to understand the mechanism of brain edema and to give timely intervention is important. This experiment set up neonatal rats hypoxic-ischemic brain damage (HIBD) model, to observe the effect of edaravone on expression of AQP-4 of brain tissue after neonatal rats HIBD and the effect of the treatment of HIBD. To further explore protective effect of edaravone on neonatal rats HIBD and its possible protection mechanisms. Our experiment may provide theoretical basis for clinical application of neonatal HIE.Methods:A total of 120 seven-day-old Wistar rats were randomly divided into three groups (n=40):sham-operated group, hypoxic-ischemic brain damage (HIBD) group and edaravone-treated group. HIBD group and edaravone-treated group were first prepared by HIBD model. Each group was divided into five sub-groups (n=8 in each subgroup) based on different time points after HIBD (6h,12h,24h,3d,5d). HIBD model rats were prepared by keeping rats into hypoxic environment of 8% O2 concentration for 2 hours after clamping left common carotid artery. Rats in edaravone-treated group were given edaravone treatment (2mg/kg, intraperitoneal injection) instantly after HIBD, and were given four subsequent days (five total injections). Sham-operated group rats were given median neck incision and freed left common carotid artery, non hypoxic-ischemic. Rats were sacrificed at different time points, to observe morphological changes in brain tissue in general. Changes of brain water content were determined. The pathological changes of left brain tissue were observed under light microscope, and immunohistochemical technique was used to determine the expression of AQP-4.Results:(1) Neonatal rats had various abnormal behaviors after HIBD. (2) After HIBD, various morphologic abnormality of brain tissue at different time points were observed; Above-mentioned changes weakened after edaravone intervention. (3) Brain tissue water content:Cerebral edema can be seen at 6h after HIBD, the peak reached on 3d, afterward brain tissue water content decreased gradually. There was significant difference at different time points between HIBD group and sham-operated group (P<0.01). Brain tissue water content in edaravone-treated group was significantly lower than HIBD group at each time point, and there was statistically significant difference between them (P<0.01). (4) HE staining:In sham-operated group, brain tissue had normal size and morphology, brain tissue structure and cell-level clearly, neurons arranged closely. Different extent cellular swelling, degeneration, necrosis, cellular nuclear fragmentation, solution, neuron loss, gliocyte proliferation were observed in brain tissue of HIBD model group at different time points. In edaravone-treated group, the pathological changes of brain tissue lessened significantly compared with HIBD model group, and cell arranged regularly, more nerve cells survival, only saw a small amount of nerve cells degeneration and necrosis. (5)Immunohistochemical staining:AQP-4 positive staining showed buffy fine grain deposition, positive expression was mainly seen in the ependyma, pia mater and blood vessels surrounding the astroglia cell membrane. AQP-4 protein in sham-operated group showed a low level of expression, no significant change at different time. AQP-4 positive cells were observed in brain tissue in HIBD group at each time point, and the expression of AQP-4 positive cells was increased significantly compared with sham-operated at the same time point. There was statistically significant difference between them (P<0.01). The expression of AQP-4 increased significantly on 6h after HIBD, the peak reached on 3d, afterward positive expression decreased gradually. The level of AQP-4 expression in edaravone-treated group was significantly lower than HIBD group at each time point, and there was statistically significant difference between them (P<0.01).Conclusion:(1) The expression of AQP-4 after HIBD is increased, and there is positive correlation with the changes of brain tissue water content. It confirmed that AQP-4 involved in the occurrence and development process of cerebral edema after HIBD. (2) Edaravone treatment after HIBD in neonatal rats can reduce pathological changes of brain tissue. Edaravone can decrease brain tissue water content and reduce the expression of AQP-4, so as to alleviate brain edema after HIBD, which maybe one of the neuroprotective effect mechanism of edaravone.
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