Danfeng Tian1, Yangyang Guo1, Dandan Zhang1, Qiang Gao1, Ganlu Liu1, Jingfeng Lin1, Ze Chang1, Yuchun Wang1, Rui Su2, Zhenyun Han3. 1. Beijing University of Chinese Medicine, Beijing, 100029, China. 2. Department of Scientific Research, Beijing Hospital of Traditional Chinese Medicine, Capital Medical University, Beijing, 100010, China. 3. Shenzhen Hospital of Beijing University of Chinese Medicine (Longgang), No. 1 Dayun Road, Longgang District, Shenzhen, 518172, China. tohanzhenyun@126.com.
Abstract
BACKGROUND: Synaptic damage and glutamate excitotoxicity have been implicated in the pathogenesis of vascular dementia (VD). Clathrin, RAB5B and N-methyl-D-aspartic acid receptor 1 (NMDAR1) proteins play a vital role in endocytosis of synaptic vesicles in neurons and glutamate over accumulation. Previous researches have been confirmed that Shenzhi Jiannao (SZJN) formula has an anti-apoptotic and neuroprotective effect in VD, but the underlying mechanisms are still unclear. In this study, we aimed to explore the effect of SZJN formula on cognitive impairment and glutamate excitotoxicity via clathrin-mediated endocytosis (CME) in vivo and in vitro. METHODS: SZJN formula consists of Panax ginseng C.A.Mey., Anemarrhena asphodeloides Bunge, and Paeonia anomala subsp. veitchii (Lynch) D.Y.Hong & K.Y.Pan. All herbs were prepared into granules. Both common carotid arteries were permanent occluded (2-vessel occlusion, 2VO) in male Sprague Dawley (SD) rats to model VD. One day after operation, the rats began daily treatment with SZJN formula for 2 weeks. The neuroprotective effects of SZJN formula was subsequently assessed by the novel object recognition test, Morris water maze, hematoxylin-eosin (HE) staining and Nissl staining. Glutamate cytotoxicity was assessed by detecting cell viability and cell death of PC12 cells. Immunohistochemistry, immunofluorescence, Western blot, and quantitative real-time PCR were used to detect the expression levels of clathrin, RAB5B, and NMDAR1. RESULTS: Administration of SZJN formula effectively improved short-term memory and spatial memory. SZJN formula treatment significantly reduced hippocampal neuronal loss, and recovered the arrangement and morphology of neurons and Nissl bodies. Moreover, SZJN formula promoted the proliferation of PC12 cells and inhibited glutamate-induced cell death. The down-regulation of clathrin and RAB5B, as well as the upregulation of NMDAR1 in the brain induced by 2VO or glutamate was also notably reversed by SZJN formula at both the protein and mRNA levels, which may contribute to SZJN formula induced improved neurological function. CONCLUSIONS: Taken together, our findings provide evidence that the neuroprotective effects of SZJN formula in experimental VD maybe mediated through promoting the expression of clathrin-mediated endocytosis and reducing NMDARs-associated glutamate excitotoxicity. SZJN formula serves as a promising alternative therapy and may be a useful herbal medicine for preventing progression of VD.
BACKGROUND: Synaptic damage and glutamate excitotoxicity have been implicated in the pathogenesis of vascular dementia (VD). Clathrin, RAB5B and N-methyl-D-aspartic acid receptor 1 (NMDAR1) proteins play a vital role in endocytosis of synaptic vesicles in neurons and glutamate over accumulation. Previous researches have been confirmed that Shenzhi Jiannao (SZJN) formula has an anti-apoptotic and neuroprotective effect in VD, but the underlying mechanisms are still unclear. In this study, we aimed to explore the effect of SZJN formula on cognitive impairment and glutamate excitotoxicity via clathrin-mediated endocytosis (CME) in vivo and in vitro. METHODS:SZJN formula consists of Panax ginseng C.A.Mey., Anemarrhena asphodeloides Bunge, and Paeonia anomala subsp. veitchii (Lynch) D.Y.Hong & K.Y.Pan. All herbs were prepared into granules. Both common carotid arteries were permanent occluded (2-vessel occlusion, 2VO) in male Sprague Dawley (SD) rats to model VD. One day after operation, the rats began daily treatment with SZJN formula for 2 weeks. The neuroprotective effects of SZJN formula was subsequently assessed by the novel object recognition test, Morris water maze, hematoxylin-eosin (HE) staining and Nissl staining. Glutamatecytotoxicity was assessed by detecting cell viability and cell death of PC12 cells. Immunohistochemistry, immunofluorescence, Western blot, and quantitative real-time PCR were used to detect the expression levels of clathrin, RAB5B, and NMDAR1. RESULTS: Administration of SZJN formula effectively improved short-term memory and spatial memory. SZJN formula treatment significantly reduced hippocampal neuronal loss, and recovered the arrangement and morphology of neurons and Nissl bodies. Moreover, SZJN formula promoted the proliferation of PC12 cells and inhibited glutamate-induced cell death. The down-regulation of clathrin and RAB5B, as well as the upregulation of NMDAR1 in the brain induced by 2VO or glutamate was also notably reversed by SZJN formula at both the protein and mRNA levels, which may contribute to SZJN formula induced improved neurological function. CONCLUSIONS: Taken together, our findings provide evidence that the neuroprotective effects of SZJN formula in experimental VD maybe mediated through promoting the expression of clathrin-mediated endocytosis and reducing NMDARs-associated glutamate excitotoxicity. SZJN formula serves as a promising alternative therapy and may be a useful herbal medicine for preventing progression of VD.
Authors: Kätlin Silm; Jing Yang; Pamela F Marcott; Cedric S Asensio; Jacob Eriksen; Daryl A Guthrie; Amy H Newman; Christopher P Ford; Robert H Edwards Journal: Neuron Date: 2019-04-16 Impact factor: 17.173