Improving cognitive impairment by Tongxinluo via inhibiting expression of beta-secretase 1/beta-amyloid peptide in experimental vascular dementia

BACKGROUND: Tongxinluo has been clinically proven to be effective in improving memory and cognitive function in patients with post-stroke vascular dementia. Is the mechanism related to the deposition of beta-amyloid peptide (Aβ) in hippocampus OBJECTIVE: To observe the effect of Tongxinluo on cognitive impairment in a mouse model with vascular dementia and the changes of Aβ deposition andβ-secretase 1 (BACE1) expression. DESIGN: Randomized controlled study.SETTING: State Key Laboratory of Pharmaceutical Biotechnology of Nanjing University and Affiliated Drum Tower Hospital of Nanjing University Medical School.MATERIALS: The experiment was carried out in the State Key Laboratory of Pharmaceutical Biotechnology of Nanjing University and Affiliated Drum Tower Hospital of Nanjing University Medical School from March 2006 to January 2007. A total of 36 healthy Kunming mice, 18 of each gender, were chosen. The study was conducted in accordance with the National Regulations of Experimental Animal Administration, and all animal experiments were approved by the Committee of Experimental Animal Administration of Nanjing University. Tongxinluo was provided by Shijiazhuang Yiling Pharmaceutical Co., Ltd.METHODS: All mice were randomly divided into 6 groups, including naive control (n=6), sham-operated control (n=6) and experimental groups treated with different doses of Tongxinluo (0.2, 0.4, and 0.6 g/kg/d; n=6 for each group) or vehicle (n=6). Five groups were subjected to bilateral common carotid arteries (2-VO) occlusion to produce a vascular dementia model (no occlusion was performed in sham-operated group). The mice in the Tongxinluo treatment groups were intragastricly administered daily with a Tongxinluo suspension (40 g/L in distilled water) at doses of 0.2, 0.4 or 0.6 g/kg/d from day 1 to day 30 post-surgery. The animals in vehicle, sham-operated and naive groups were administered an equal volume of distilled water. MAIN OUTCOME MEASURES: ①Escape latency time determined in all groups of mice before and after 2-VO occlusion by Morris water maze. ②Changes in BACE1 mRNA expression in the hippocampi of mice among the six groups by RT-PCR assay, and BACE1 and Aβ protein expression in the hippocampi of mice by Western blot.RESULTS: All 36 mice were involved in the final analysis. ① No difference was detected in escape latency time to a hidden platform among all groups in water maze test before surgery (P > 0.05) At 30 days after 2-VO occlusion, the vehicle animals exhibited a significantly longer latency in finding the hidden platform compared to that of sham-operated and naive animals (P < 0.01). The prolonged escape latency was significantly reduced by oral administration of 0.4 g or 0.6 kg/day (P < 0.01, P < 0.05). BACE1 mRNA and protein expression in vehicle animals were much higher than in sham-operated and naive animals (P < 0.01). The ischemia-induced increases in BACE1 mRNA and protein level were attenuated by all three doses of Tongxinluo treatment (P < 0.01), and the 0.4 g/kg/d treatment was the most effective. Aβ protein expression in vehicle animals after 2-VO occlusion were much higher than in sham-operated and na?ve animals (P < 0.01). 2-VO occlusion-induced Aβ generation was significantly attenuated by all doses of Tongxinluo treatment, with the most effective dose being 0.4 g/kg/d (P < 0.01).CONCLUSION: BACE1 mRNA levels and protein levels of BACE1 and Aβare reduced in the hippocampi of vascular dementia model mice by all three doses of Tongxinluo treatment, with the most effective dose being 0.4 g/kg/d. The results suggest that inhibition of post-ischemia BACE1 expression and Aβ generation in brain might underlie Tongxinluo's effects in improving cognitive impairment.