益生菌和二十二碳六烯酸对Aβ_(25-35)诱导阿尔茨海默症小鼠学习记忆及脑损伤的作用

Effects of probiotics and docosahexaenoic acid on learning memory and brain damage in Aβ_(25-35)-induced Alzheimer's disease mice

探究并比较益生菌及二十二碳六烯酸(docosahexaenoic acid,DHA)与阿尔茨海默症(Alzheimer's disease,AD)治疗药物多奈哌齐对AD小鼠学习认知及脑损伤相关指标的影响,为其治疗AD提供实验依据。所有动物实验经河南中医药大学伦理委员会批准(伦理号为DWLL2018080003)。将50只C57BL/6J雄性小鼠随机分为假手术组、模型组、多奈哌齐组(10 mg·kg^(-1))、益生菌组(2.7×10^(9)CFU·d^(-1))、DHA组(0.104 g·kg^(-1)),除假手术组外,其他组采用侧脑室注射Aβ_(25-35)(200μmol·L^(-1))的方法构建AD动物模型,之后灌胃给药4周。检测各组小鼠学习记忆能力、海马区神经元形态、原代海马细胞凋亡和免疫细胞水平;同时检测小鼠脑组织Aβ_(1-42)、Aβ_(1-40)、p-Tau、Ach E、Ach、氧化应激、胶质细胞活化及炎症因子IL-1β、IL-10、TNF-α的水平;并通过16S r DNA测序进一步探究多奈哌齐和益生菌对AD的干预作用。结果显示,多奈哌齐、益生菌及DHA通过增加Aβ_(25-35)诱导小鼠运动时间、运动距离、自主交替率,缩短到达平台时间,改善了其认知障碍并增强学习记忆能力;通过降低脑组织中Aβ_(1-42)、Aβ_(1-40)、p-Tau、Ach E、IL-1β、TNF-α、MDA及增加Ach、IL-10、GSH-Px、T-SOD水平,同时减少胶质细胞活化,显著减轻Aβ_(25-35)诱导小鼠脑损伤和神经炎症,并且对免疫细胞具有调节作用;16S r DNA测序显示多奈哌齐和益生菌均可恢复菌群稳态,并且差异菌与认知、AD病理、神经炎症等存在密切相关。综合所有指标,多奈哌齐与益生菌效果优于DHA。综上所述,多奈哌齐、益生菌及DHA通过调节免疫细胞、减少脑内细胞凋亡和胶质细胞活化数量,降低氧化应激及炎症因子表达水平来改善Aβ_(25-35)诱导小鼠的认知功能障碍和脑损伤,其中多奈哌齐及益生菌效果优于DHA,且多奈哌齐及益生菌对AD的治疗与调节菌群密切相关。

The study aims to investigate and compare the effects of probiotics and docosahexaenoic acid(DHA)with the Alzheimer's disease(AD)therapeutic drug donepezil on the learning cognition and brain damage related indexes in AD mice,and to provide experimental basis for its treatment of AD.All animal experiments were approved by the Ethics Committee of the Henan University of Chinese Medicine(ethics number DWLL2018080003).Fifty male C57BL/6J mice were randomly assigned to one of five groups:sham-operated,model,donepezil(10 mg·kg^(-1)),probiotic(2.7×10^(9) CFU·d^(-1)),and DHA(0.104 g·kg^(-1)).Except for the sham-operated group,the AD animal model was established by injecting Aβ_(25-35)(200μmol·L^(-1))in the lateral ventricle,followed by gavage administration for4 weeks.In all mouse groups,learning memory ability,neuronal morphology in the hippocampus,apoptosis of primary hippocampal cells,and immune cell levels were detected.The levels of Aβ_(1-42),Aβ_(1-40),p-Tau,Ach E,Ach,oxidative stress,glial cell activation,and the inflammatory factors IL-1β,IL-10,and TNF-αin the brain tissue of mice were also detected.16S r DNA sequencing was used to further investigate the effects of donepezil and probiotics on AD.Donepezil,probiotics and DHA improved cognitive deficits and enhanced learning memory in Aβ_(25-35)-induced mice by increasing locomotion time,locomotion distance,autonomic alternation rate,and shortening the time to reach the plateau;it significantly attenuated Aβ_(25-35)-induced brain injury and neuroinflammation in mice by decreasing Aβ_(1-42),Aβ_(1-40),p-Tau,Ach E,IL-1β,TNF-α,and MDA and increasing the levels of Ach,IL-10,GSH-Px,and T-SOD in brain tissues,as well as decreasing the activation of glial cells,and had a modulatory effect on immune cells.16S r DNA sequencing shows that both donepezil and probiotics restore flora homeostasis and that differential bacteria are strongly associated with cognition,AD pathology,and neuroinflammation.Combining all indicators,donepezil and probiotics were more effective than DHA.All in all,donepezil,probiotics and DHA ameliorate Aβ_(25-35)-induced cognitive dysfunction and brain damage in mice by modulating immune cells,reducing the number of apoptotic cells and glial cell activation in the brain,and decreasing the levels of oxidative stress and inflammatory factor expression,among which the effects of donepezil and probiotics were better than those of DHA,and the therapeutic effects of donepezil and probiotics on AD were closely related to the modulation of gut microbiome.