Effects of Gingko biloba leaf extract on learning, memory, and hippocampal amyloid precursor protein mRNA expressions in diabetic rats

Xiaofan Zhang; Bo Liang; Zhifeng Liang; Jun Lin

摘要：BACKGROUND: The mechanisms of brain injury Following diabetes could be related to amyloid precursor protein (APP) mRNA overexpression. Studies have shown that Gingko biloba leaf extract (Egb) is effective in promoting functional recovery of the brain after traumatic injury. Egb is also effective in improving central nervous system plasticity and learning and memory functions of the elderly.OBJECTIVE: To study the effects of Egb on learning and memory, as well as hippocampal APP mRNA expression in the brains of diabetic rats, using Morris water maze behavioral testing and reverse transcription polymerase chain reaction (RT-PCR), respectively.DESIGN: Complete random design, controlled experimental study.SETTING: Department of Pharmacology, Pharmaceutical School, Guangxi Medical University.MATERIALS: A total of 70 male Wistar rats (180-220 g), 8 weeks old and specific pathogen free, were used for this study. GbE (containing 24.8% flavone glycosides and 6.2% diterpene lactone) was purchased from Guilin Sitejia Natural Plants Pharmaceutical Factory (Guangxi Province, Lot NO. 200405). Streptozotocin was purchased from Sigma (USA). Protamine zinc insulin injection was purchased from WANBANG Biochemical Pharmaceutical Co., Ltd. (Xuzhou Jiangsu, China).METHODS: The experiment was performed in the Experimental Center of Guangxi Medical University from March to October 2005. ① Experimental intervention: 70 rats were divided randomly into normal control group, diabetic model group (DM group), diabetic model +10 μg/kg insulin group (DM + Ins group), diabetic model + 100 mg/kg ginkgo leaf extract group (DM + Egb high-dose group), and diabetic model + 50 mg/kg ginkgo leaf extract group (DM + Egb low-dose group); there were 14 rats in each group. Rats with an intraperitoneal (I.p.) injection of citrate buffer solution (pH 4.4) served as the control group. To establish the diabetes model, rats were treated with I.p. Injection of 55 mg/kg streptozotocin. Insulin (10 U/kg) was injected subcutaneously (s.c.) every day for 6 months in the DM group. Egb (100 mg/kg) and Egb (50 mg/kg) was administered intragastrically every day for 6 months in the DM + Egb high-dose group and DM + Egb low-dose group,respectively. The DM group and control group were administered distilled water intragastrically every day for 6 months. Drugs were administered once every morning. ② Experimental evaluation: Six month after intervention, learning and memory of diabetic rats was tested by Morris water maze. Rats were allowed to train for 4 days, and the escape latency and platform-searching score were measured at days 5 and 8. Changes in hippocampal APP mRNA expression were measured with RT-PCR.MAIN OUTCOME MEASURES: Morris water maze performances and hippocampal APP mRNA expression in rats.RESULTS: A total of 70 Wistar rats were included in the final analysis, without any loss. ① Learning and memory dysfunction of diabetic rats: After 4 days of Morris water maze training, escape latency was longer in the DM group on days 5 and 8, and the platform-searching score was lower in the DM group compared to the control group. In the DM + Egb group, the escape latency score was shorter and platform-searching score was significantly increased compared to the DM group. ② APP mRNA expression: in the hippocampus of diabetic rats, a 340 bp mRNA product was amplified, which is comparable to the APP mRNA amplification length of design. The expression of APP mRNA from the hippocampus of diabetic rats with learning and memory dysfunctions was significantly increased. Egb extract significantly inhibited the APP mRNA expression in these rats.CONCLUSION: Egb not only ameliorated the learning and memory dysfunctions in diabetic rats, but also significantly inhibited APP mRNA expression. Results from this study led to the hypothesis that diabetes could be one of the risk factors for AD.