核桃蛋白水解肽改善大鼠酒精性认知下降的作用机制

Mechanism of walnut proteolytic peptide in improving alcoholic cognitive decline in rats

中文关键词:  酒精  认知  核桃肽  海马  突触

英文关键词:alcohol  cognition  walnut peptide  hippocampus  synapse

中文摘要:

      目的 评价核桃蛋白水解肽(walnut protein hydrolysates, WPH)对酒精性认知下降的改善效应并探讨其作用机制。方法 40只雄性SD大鼠随机分为空白对照组(Control)、酒精模型组(Model)、WPH干预低、中、高剂量组(WPH-L、WPH-M、WPH-H)。Morris水迷宫检测空间学习记忆能力, 分析免疫相关血细胞指标, 组织病理学分析海马区神经元细胞密度, 超高效液相色谱-串联质谱法(ultra performance liquid chromatography-tandem mass spectrometry, UPLC-MS/MS)分析大脑皮质内记忆相关神经递质含量, 蛋白质印迹法(Western blotting)和逆转录-聚合酶链反应(reverse transcription-polymerase chain reaction, RT-PCR)检测突触信号传递标记蛋白及其基因表达水平。结果 与Control相比, Model组平台潜伏期、运动总路程显著延长, 其目标象限平台穿越次数与停留时间显著下降(P<0.05); WPH干预后平台潜伏期、运动总路程均剂量依赖性降低, 目标象限平台穿越次数、停留时间均剂量依赖性增加, 且在WPH-H组达到显著性水平(P<0.05), 表明WPH干预后能有效缓解长期酒精摄入导致的认知下降; 长期酒精摄入诱导炎性细胞(白细胞和淋巴细胞)水平显著升高(P<0.05), 海马CA1区神经元细胞明显减少, 细胞核出现固缩; 高剂量WPH干预后炎性细胞水平显著降低(P<0.05), 神经元数目明显上升, 细胞排列较为整齐, 表明WPH可有效减轻长期酒精摄入诱导的海马区神经元丢失; 此外, 高剂量WPH可明显改善长期酒精摄入引起的大脑皮质神经递质紊乱(乙酰胆碱、多巴胺、5-羟色胺和γ-氨基丁酸), 且可显著逆转神经细胞突触信号传递关键蛋白分子BDNF、CREB、PSD95和ERK1/2及其基因表达水平的下降(P<0.05)。结论 本研究初步揭示WPH改善酒精性认知下降的海马结构基础和突触分子机制, 为核桃蛋白的高值化利用奠定了理论基础。

英文摘要:

      Objective To evaluate the ameliorative effects and explore the underlying mechanism of walnut protein hydrolysates on the cognitive impairments induced by chronic alcohol abuse in SD rats. Methods Forty healthy SD rats were randomly divided into control group, alcohol model group, and WPH administration group. The memory capacity, immune-related indicators, neuron density in hippocampus region, neurotransmitter content in cerebral cortex, and expression levels of marker proteins and genes related to synaptic transduction efficiency were systematically determined with Morris water maze, blood cell profiles analysis, pathological observation, ultra performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method, Western blotting and reverse transcription-polymerase chain reaction (RT-PCR) procedures, respectively. Results Compared with the control group, the platform latency and total exercise distance of the model group was significantly prolonged, and the number of target quadrant platform crossings and stay time decreased significantly (P<0.05); after WPH intervention, the platform latency and total exercise distance decreased in a dose-dependent manner, and the number of target quadrant platform crossings and stay time increased in a dose-dependent manner, and reached a significant level in the WPH-H group (P<0.05), it showed that WPH intervention could effectively alleviate the cognitive decline caused by long-term alcohol intake; long term alcohol intake induced a significant increase in the level of inflammatory cells (leukocytes and lymphocytes) (P<0.05), a significant decrease in neurons in hippocampal CA1 region, and pyknosis in the nucleus; after the intervention of high dose WPH, the level of inflammatory cells was significantly reduced (P<0.05), the number of neurons was significantly increased, and the cells were arranged orderly, indicating that WPH could effectively reduce the loss of neurons in hippocampus induced by long-term alcohol intake; in addition, high dose WPH could significantly improve the neurotransmitter disorder of cerebral cortex caused by long-term alcohol intake (acetylcholine, dopamine, 5-hydroxytryptamine and γ-aminobutyric acid), and significantly reversed the decrease of BDNF, CREB, PSD95, ERK1/2 and their gene expression levels (P<0.05). Conclusion These findings preliminarily reveals the synaptic mechanism of WPH reversing the cognition degradation and lays the experiment basis for high value utilization of walnut protein resource.