| Background:Acute cerebral infarction often leads to neurological dysfunction, which causes disability or death. Comparing with ischemia preconditioning, non-ischemia preconditioning shows a greater practical value. It has been demonstrated that one-time short-term isoflurane preconditioning possesses neuroprotection effect against ischemia-reperfusion injury, but the effects of repetitive isoflurane preconditioning have not been tested. It was also proposed that ginsenoside has neuroprotective effects on focal cerebral ischemia-reperfusion injury. Ginsenoside-Rd is the only saponin monomer, which has been used in the clinical trial, however, the neuroprotective effects of ginsenoside-Rd pretreatment, has not been tested. The aims of the present study were to investigate the effects of repetitive isoflurane perconditoning and Ginsenoside Rd pretreatment on focal cerebral ischemia-reperfusion injury. Part 1The effect of repetitive isoflurane preconditioning on focal cerebral ischemia-reperfusion injury in ratsObjectives1. To studyt the neuroprotective effects of repetitive isoflurane preconditioning on focal cerebral ischemia-reperfusion injury in rats2. To investigate the influence of isoflurane preconditioning on exression of COX-2 during transient focal cerebral ischemia-reperfusion injury in rats.Methods1. Neuroprotective effects of repetitive isoflurane preconditioning on focal cerebral ischemia-reperfusion injury in rats50 male SD rats were randomly divided into control (CON), ISO5 days, ISO10 days, ISO15 days and ISO20 days groups (n=10 each). In all rats, MCAO was induced by an occlusion of right internal carotid artery with a nylon monofilament. The neurological behavior scores (NBS) were assessed at 24h, 48h and 72h after reperfusion respectively, and the animals were then decapitated to determine the brain infarct volume.2. The effect of isoflurane preconditioning on COX-2 exression during cerebral cortex after transient focal cerebral ischemia-reperfusion injury in rats.20 male SD rats were randomly divided into normal group, control group, Iso10 days and Iso20 days groups. At 24h after reperfusion, the rats were perfused with polyoxymethylene, and then the brains were cut into frozen sections and immunohistochemistry stained.Results1. Neuroprotective effects of repetitive isoflurane preconditioning on focal cerebral ischemia-reperfusion injury in ratsThe neurological behavior scores in Iso5 days group, Iso10 days group and Iso15 days group were significantly higher than those in control group and Iso20 days group at 24h, 48h and 72h after reperfusion, and their brain infarct volumes were obviously smaller than CON group and Iso20 days group as well. The brain infarct volumes in Iso10 days group were significantly smaller than Iso5 days and Iso15 days group.2. The effect of isoflurane preconditioning on COX-2 expression in cerebralcortex after transient focal cerebral ischemia-reperfusion injury in rats. The number of COX-2 positive cells in CON and Iso20 days groups was significantly higher than those in normal and Iso10 days groups (P<0.05). There were no significant difference between CON group and Iso20 days group (P>0.05).Part 2Neuroprotective effects of ginsenoside Rd pretreatment on focal cerebral ischemia-reperfusion injury in ratsObjectives1. To study the dose-dependent responses of ginsenoside Rd pretreatment.2. To investigate the effect of ginsenoside Rd pretreatment on exression of COX-2 in transient focal cerebral ischemia-reperfusion injury in rats.Methods1. Dose-dependent responses of ginsenoside Rd pretreatment.70 male SD rats were randomly divided into 7 groups(n=10 each): CON, Rd5mg, Rd10mg, Rd20mg, Rd40mg, Rd80mg and VEC. In all rats, the middle cerebral artery occlusion (MCAO) was induced by an occlusion of right internal carotid artery with a nylon monofilament. According to the protocol, Rd or propylene glycol solvent(the vehicle of Rd)was intraperitoneally administered at 1h before cerebral ischemia. The neurological behavior scores(NBS)were measured by Garcia Scale at 24h,48h and 72h after I/R. The brain infarct volume percentage was then assessed by TTC staining following the last neurological outcome evaluation.2. The effect of isoflurane pretreatment on COX-2 expression in cerebral cortex after transient focal cerebral ischemia-reperfusion injury in rats.20 male SD rats were randomly divided into 4 groups: normal group, Rd40'(Rd40mg/kg administration without I/R)group, CON group, and Rd40 (Rd40mg/kg+I/R)group Rd40mg/kg were intraperitoneally administered just 1h before MCAO in group Rd40'and group Rd40. 24h after reperfusion, the rats were perfused with polyoxymethylene, and then the brains were cut into frozen sections and immunohistochemistry stained.Results1. Dose-dependent relationship of ginsenoside Rd pretreatment.The values of NBS in Group Rd 5mg, Rd10mg, Rd 20mg, Rd 40mg and Rd 80mg at 24h, 48h and 72h after I/R were significantly higher than those in Group I/R and VEC(P<0.05), their infarct volume percentage were smaller than those of in I/R and VEC groups(P<0.05). A dose-dependent neuroprotection of the Rd was found when the dosage was lower than 40mg/kg. The infarct volume percentage in Group Rd 80mg was bigger than that in Group Rd 20mg and Rd 40mg, and the NBS was lower. However, compared with Group Rd 5mg and Rd 10mg, Group Rd 80mg had no difference in infarct volume percentage and NBS(P>0.05). 2. The effect of isoflurane pretreatment on COX-2 expression in cerebral cortex after transient focal cerebral ischemia-reperfusion injury in rats.The number of COX-2 positive cells in CON group was significantly higher than that in normal and Rd40 group (P<0.05). The immune response intensity and the number of COX-2 positive cells in group Rd40'were significantly smaller than that in CON group (P<0.05) .Conclusion1. Repetitive isoflurane preconditioning for 10days has the best neuroprotective effects on focal cerebral ischemia-reperfusion injury in rats. The effect disappeares when it prolongs to 20 days.2. The neuroprotective effect of Ginsenoside Rd pretreatment is dose dependent; Rd40mg/kg is the optimal dosage.3. Isoflurane preconditioning and ginsenoside Rd pretreatment reduces the expression of COX-2 in cerebral cortex after focal cerebral ischemia- reperfusion in rats. |
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