| Objective: Applying focal ischemia-reperfusion model of SD rat through observing neurologic impairment score, cerebral infarction volume, cerebral pathomorphology and the expression of Interleukine-1β(IL-1β) and Caspase-3 to discuss whether pretreated with fluoxetine can relieve the injury of local cerebral ischemia-reperfusion in SD rats and possible mechanism.Method: 72 healthy male SD rats, body weight 180-220g, were randomly divided into 3 groups, sham-operated group (n=24), ischemia- reperfusion group (namely control group, n=24), fluoxetine group (n=24). Each group was divided into 4 subgroups according to reperfusion 0h (n=6), 2h (n=6), 6h (n=6), 24h (n=6) after ischemia for 2 hours. Sodium chloride solution (10ml/kg/d), sodium chloride solution (10ml/kg/d) and fluoxetine (2.0mg/kg/d) was given respectively to 3 groups once a day by gastric gavage 14 days before ischemia-reperfusion.Sham-operated group: sewing up the skin after exposing the common carotid artery (CCA) in the right side. Control group and fluoxetine group: using the improved Longa-Zea method to establish ischemia- reperfusion model in right cerebral middle artery of SD rat, pull out the string in 2nd hour after ischemia then performing reperfusion. Evaluate the neurologic impairment score by referring to Zea-Longa's standards of five grades. All removed brains were stained respectively with 2% Tripheny1 Tetrazalium Chloride (TTC). Judging the ischemic location, measuring the infarction volume with the pathology image analysis system, observing cerebral pathomorphology through Hematoxylin-eosin (HE). Observing the expression of IL-1βand Caspase-3 with immunohistochemical technique. Comparison:(1) Dynamic change of cerebral infarction volume, IL-1βand Caspase-3 of infarcted brain tissue at every point of time in control group;(2) Dynamic change of cerebral infarction volume, IL-1βand Caspase-3 of infarcted brain tissue at every point of time in fluoxetine group;(3) The change of neurologic impairment score,cerebral infarction volume, cerebral pathomorphology and the expression of IL-1βand Caspase-3 at the same point of time in control group and sham-operated group;(4) The change of neurologic impairment score,cerebral infarction volume, cerebral pathomorphology and the expression of IL-1βand Caspase-3 at the same point of time in fluoxetine group and control group.Results:1. The neurologic impairment score in the control group was higher than that in the sham-operated group at every subgroup and the result has significant difference (p<0.01); Zea longa evaluation showed no difference between the fluoxetine group and the control group except in the group of 24h reperfusion following 2h cerebral ischemia (p<0.05).2. Sham-operated group and the left hemisphere of the other two groups had no infarction areas. The infarction area appeared at 2h after ischemia and increased in 24h in both control group and fluoxetine group. Compared every reperfusion time point in control group and fluoxetine group p<0.01. The infarction volume showed no difference between the fluoxetine group and the control group except in the group of 24h reperfusion following 2h cerebral ischemia (p<0.01).3. Morphology of cerebral tissue stained by HESham-operated group and the left hemisphere of the other two groups had no evident pathological changes.In control group: the number of neurons at ischemic area decreased, nerve cells shrinked and degenerated with clear boundary from peripheral tissue. Karyopycnosis, apparent edema between tissues and vacuolization,with the reperfusion time extending, damage on the ischemic neuron became severe showing irreversible neuron damage change and most neurons appeared necrosis.In fluoxetine group: the injured degree of cerebral tissue was in positive proportion to reperfusion time, however if compared with control group the number of neuron which appeared degeneration and necrosis decreased in the ischemia side of hippocampus tissue, vacuolization and intertissue edema became relieved.4. Immunohistology result:(1) Dynamic change of expression of IL-1βand Caspase-3 in the control group: There were a few of IL-1βpositive cells at 2h after ischemia in the ischemia side of hippocampus tissue. The expression increased since 2h after reperfusion, and reached the peak at 6h after reperfusion, then it decreased gradually, there was significant difference among each point of time (p<0.01). There were few of Caspase-3 positive cells at 2h after ischemia in the ischemia side of hippocampus tissue. The expression increased since 2h after reperfusion and increased gradually within 24h, there was significant difference among each point of time (p<0.01).(2) Dynamic change of expression of IL-1βand Caspase-3 in the fluoxetine group: There were a few of IL-1βpositive cells at 2h after ischemia in the ischemia side of hippocampus tissue. The expression increased since 2h after reperfusion, and reached the peak at 6h after reperfusion, then it decreased gradually, there was significant difference among each point of time (p<0.01). There were few of Caspase-3 positive cells at 2h after ischemia in the ischemia side of hippocampus tissue. The expression increased since 2h after reperfusion and increased gradually within 24h, there was significant difference among each point of time (p<0.01).(3) Comparison between control group and sham-operated group at the same point of time: expression of IL-1βand Caspase-3 in the ischemia side of hippocampus tissue increased (p<0.01). There was significant difference between control group and sham-operated group except in the group of 2h after ischemia (p>0.05).(4) Comparison between fluoxetine group and control group at the same point of time: expression of IL-1βin the ischemia side of hippocampus tissue decreased, p<0.05 at reperfusion 2h and 6h, p<0.01 at reperfusion 24h; expression of Caspase-3 in the ischemia side of hippocampus tissue decreased, p<0.05 at reperfusion 2h and 6h, p<0.01 at reperfusion 24h. The expression of IL-1βand Caspase-3 showed no difference between the fluoxetine group and the control group at 2h after ischemia (p>0.05)Conclusions:1. There were a few of IL-1βpositive cells at 2h after ischemia in the ischemia side of hippocampus tissue. The expression increased since 2h after reperfusion, and reached the peak at 6h after reperfusion, then it decreased gradually. There were few of Caspase-3 positive cells at 2h after ischemia in the ischemia side of hippocampus tissue. The expression increased since 2h after reperfusion and increased in 24h. It suggested us that the expression of IL-1βand Caspase-3 took part in the process of the ischemia-reperfusion and increased ischemia-reperfusion injury.2. Prevented administration of fluoxetine could decrease focal ischemia-reperfusion injury, relieve the neurological functional defect, reduce infarction volume, reduce the neuronal degeneration, necrosis and tissue edema, reduce the expression of IL-1βand Caspase-3 in the hippocampus tissue in SD rats.3. The protective effect of fluoxetine on cerebral injury induced by ischemia-reperfusion may be related to inhibiting the expression of IL-1βand Caspase-3. Fluoxetine has a significant neuroprotective effect since the acute phase of cerebral ischemia-reperfusion.
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