| ObjectiveTo investigate the neuroprotective effect of resveratrol on improving memory and inhibiting the translocation of Aβ1-42 in the hippocampus, and the effect on the proteins of the NF-κB signal pathway in AD rats.MethodsPart 1 Resveratrol improves memory and neuronal structures in AD ratsRats were allowed 1 week to adapt to their environment before being used for experiments, 60 famale Wistar rats(200 ~ 220g) were randomly divided into 6 groups: 1) OVX + D-gal 100 mg/kg group(AD model group); 2-4) OVX, D-gal and Res 20, 40, 80 mg/kg treated groups; 5) OVX, D-gal and estradiol valerate 0.8 mg/kg treated group(ET); and 6) Sham control group. After the initial wound healing, the intervention began. The AD model group: daily intraperitoneal injection of D-gal + distilled water; Res 20 mg/kg, 40 mg/kg and 80 mg/kg in the intervention group: daily intraperitoneal injection of D- gal, respectively, with a dose of Res 20, 40 and 80 mg/kg; ET group: intraperitoneal injection of D- gal + 0.8 mg/kg orally estradiol valerate; the Sham group: intraperitoneal injection of saline + distilled water. The intervention lasted for 12 weeks. All rats were tested by Morris water maze(MWM) for memory. Hippocampus was fixed by glutaraldehyde after cardiac perfusion forultrastructure changes of neuronal mitochondria, synaptic and nucleus under transmission electron microscope(TEM).Part 2 Effect of resveratrol on the translocation and aggregation of Aβ1-42 and the effect on the integrity of the blood-brain barrier in AD ratsRats were divided into 6 groups: AD model group, Res 20 mg/kg, 40 mg/kg and 80 mg/kg group, ET group and the S ham group. Blood and hippocampus were collected. The concentration of Aβ1-42 was measured by ELISA assay. Western blot assay was conducted for the expressions of β-APP, RAGE, MMP-9 and C laudin-5, and the IHC was used for dectetion of the distribution of RAGE and Claudin-5.Part 3 Effect of resveratrol on inflammation mediated by signaling pathway of NF-κB in the hippocampus of AD ratsRats were divided into 6 groups: AD model group, Res 20mg/kg, 40mg/kg, 80mg/kg group, ET group and the Sham group. Blood and hippocampus were collected. Western blot assay was conducted for the e xpression of NF-κBp65. ELISA assay was conducted for concentration of TNF-α and IL-6. The activity of the i NOS and the concentration of the NO were also detected in hippocampus.ResultsPart 1 Resveratrol improves memory and neuronal structures in AD rats1. Compared with the AD model group, Res intervention groups had no significant difference in body weight, weight gain, food consumption and food utilization of rats(P > 0.05).2. In the results of Morris water maze(MWM) test, the escape latency to find the submerged platform significantly declined every day in each group, moreover, the latency in 80 mg/kg Res and ET groups decreased significantly, while the number of times rats crossed over the platform site in 80 mg/kg Res and ET groups was increased markedly compared with AD model group(P < 0.05). The time remained in the target quadrant in all of the intervention groups was longer compared with the AD model group(P < 0.05).3. Compared with the AD model group, the mean rank of mitochondrial vacuolar degeneration area ratio in Res 80 mg/kg group was significantly smaller(P < 0.05) under TEM of 20,000 times; ET group had no significant difference compared with the AD model group(P > 0.05). Under TEM of 30,000 times, number of the synaptic structure reduced and the synaptic gap was not clear. Synapse composition before and after the electron density decreased and synaptic vesicles reduced in the AD model group; Number of synaptic structure increased in Res 80 mg/kg group compared with the AD model group. Under TEM of 12,000 times, compared with the Sham control group, nuclear deformed and shattered in the AD model group; Res 80 mg/kg group improved compared with the AD model group.Part 2 Effect of resveratrol on the translocation and aggregation of Aβ1-42 and the effect on the integrity of the blood-brain barrier in AD rats1. The expression of β-APP in hippocampus in Res 20, 40 and 80 mg/kg treated groups was decreased compared with the model group(P < 0.05). Compared with the ADmodel group, the level of Aβ1-42 in Res 80 mg/kg group was lower in hippocampus(P < 0.05), and Res 20 mg/kg group was significantly higher in serum(P < 0.05). Expression of RAGE in AD model group was increased compared with the Sham group(P < 0.05), and the attribution of RAGE protein around vessels was significantly increased in rat hippocampus(P < 0.05). The expression of RAGE in Res 20, 40, 80 mg/kg and ET groups was significantly decreased compared with the AD model group(P < 0.05), and the attribution of RAGE around vessels in Res 80 mg/kg and ET groups was significantly decreased compared with the AD model group(P < 0.05).2. Expression of MMP-9 in AD model group was increased compared with the Sham group(P < 0.05). The expression of MMP-9 in Res 20, 40, 80 mg/kg and ET groups was significantly decreased compared with the AD model group(P < 0.05). Expression of Claudin-5 in AD model group was decreased compared with the Sham group(P < 0.05), and the attribution of C laudin-5 protein around vessels was significantly decreased in rat hippocampus(P < 0.05). The expression of C laudin-5 in Res20, 40, 80mg/kg and ET groups was significantly increased compared with the AD model group(P < 0.05), and the attribution of C laudin-5 protein around vessels in Res 80 mg/kg and ET groups were significantly increased compared with the AD model group(P < 0.05).Part 3 Effect of resveratrol on the inflammation mediated by signaling pathway of NF-κB in the hippocampus of AD rats1. In Western blot assay, level of NF-κBp65 in the hippocampus of AD model group was increased compared with the Sham control group(P < 0.05), expression of NF-κBp65 protein in Res 40, 80mg/kg and ET groups had significantly decreased compared with the AD model group(P < 0.05).2. Compared with the Sham control group, levels of TNF-α and IL-6 in the serum of AD model group rats was significantly higher(P < 0.05). Contents of TNF-α and IL-6 in Res 20, 40, 80 mg/kg and ET groups were significantly lower than those of AD model(P < 0.05).3. The i NOS activity and the level of NO in Res 20, 40, 80 mg/kg groups, the Sham control group and ET group were significantly lower than those of the AD model group(P < 0.05).Conclusions1. Memory in AD rats can be proved by Res(80 mg/kg). Also, the structure of neuron can be protected significantly.2. Res can protect Aβ1-42 from translocating to the brain by protecting the integrity of BBB.3. Res can inhibit the inflammation in the brain mediated by NF-κB inflammatory pathway. |
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