洗心汤对快速老化模型小鼠肠道菌群多样性及脑肠组织Aβ1-42、LPS、SAA、ACH水平的影响

Effects of Xixin Decoction(洗心汤)on the Diversity of Intestinal Flora and Levels of Aβ1-42,LPS,SAA,and ACH in Brain and Intestinal Tissues of Rapidly Aging Model Mice

目的探讨洗心汤防治阿尔茨海默病(AD)的可能作用机制。方法将50只快速老化模型小鼠(SAMP8)随机分为模型组,益生菌组,洗心汤高、中、低剂量组,每组10只,另取10只同源对抗快速老化小鼠(SAMR1)设为对照组。喂养10周后,洗心汤高、中、低剂量组以洗心汤5.08、2.54、1.27 g/(kg·d)灌胃,益生菌组以复合益生菌冻干粉0.39 g/(kg·d)灌胃,对照组和模型组以蒸馏水10 ml/(kg·d)灌胃,各组每天灌胃1次。灌胃10周后,采用Morris水迷宫试验检测各组小鼠空间学习记忆能力;HE染色观察小鼠海马CA3区、结肠组织病理学改变;ELISA法检测小鼠海马组织和结肠区β淀粉样蛋白_(1-42)(Aβ_(1-42))、脂多糖(LPS)、血清淀粉样蛋白A(SAA)、乙酰胆碱(ACH)水平;16S rRNA测序技术检测小鼠粪便中肠道菌群多样性的变化。结果与对照组比较,模型组海马和结肠组织Aβ_(1-42)、LPS、SAA含量明显升高,ACH含量明显降低(P<0.05或P<0.01)。与模型组比较,益生菌组第2、5天的逃避潜伏期明显缩短,洗心汤高剂量组第2~5天逃避潜伏期明显缩短,目标平台象限滞留时间显著增加,洗心汤中剂量组第5天逃避潜伏期明显缩短(P<0.05或P<0.01),洗心汤高、中剂量组海马神经元细胞排列较为紧密,海马组织及结肠组织SAA、Aβ_(1-42)、LPS含量明显降低,ACH含量明显升高,Simpson、Shannon指数明显升高(P<0.05或P<0.01);益生菌组和洗心汤低剂量组海马神经元细胞排列较为松散,益生菌组和洗心汤高剂量组的拟杆菌门、拟普雷沃氏菌属明显降低,厚壁菌门、乳酸杆菌占比明显升高(P<0.05或P<0.01)。与益生菌组和洗心汤高剂量组比较,洗心汤中剂量组杯状细胞数目减少,洗心汤低剂量组腺体数目减少并伴有萎缩。与洗心汤中、低剂量组比较,洗心汤高剂量组海马区Aβ_(1-42)含量明显降低,海马区及结肠组织ACH含量明显升高,结肠组织SAA含量降低(P<0.05或P<0.01),且洗心汤低剂量组海马区SAA含量明显高于高、中剂量组(P<0.01)。结论洗心汤可以改善SAMP8的空间学习记忆能力,减轻海马、结肠组织中LPS、SAA、Aβ_(1-42)的生成与沉积,增加脑肠区ACH的含量,其防治AD的作用机制可能与调节肠道微生态、影响菌群多样性、改善炎症反应有关。

Objective To observe the possible mechanism of Xixin Decoction(洗心汤,XXD)in the prevention and treatment of Alzheimer's disease(AD).Methods Fifty rapid aging model mice(SAMP8)were randomly divid⁃ed into model group,probiotic group,high-,moderate-and low-dose group of XXD,with 10 mice in each group.An⁃other 10 homologous anti-rapid aging mice(SAMR1)were set as control group.After 10 weeks of feeding,the control group and the model group were given 10 ml·kg^(-1)·d^(-1)of distilled water by gavage,while the probiotic group(0.39 g·kg^(-1)·d^(-1)),the high-dose group of XXD(5.08 g·kg^(-1)·d^(-1)),the moderate-dose group of XXD(2.54 g·kg^(-1)·d^(-1)),and the low-dose group of XXD(1.27 g·kg^(-1)·d^(-1))were given corresponding drugs or decoctions by gavage,once a day in all groups.After 10 weeks of intragastric administration,Morris water maze was used to detect the spatial learning and memory ability of mice in each group.HE staining was used to observe the pathological changes of hippocampal CA3 region and colon.The levels ofβ-amyloid 1-42(Aβ_(1-42)),lipopolysaccharide(LPS),serum amyloid A(SAA)and acetylcholine(ACH)in hippocampus and colon were detected by ELISA.The diversity of intestinal flora in mouse feces was detected by 16S rRNA sequencing.Results Compared to those in the control group,the levels of Aβ_(1-42),LPS,SAA increased,while the level of ACH decreased in the model group(P<0.05 or P<0.01).Compared to those in the model group,the escape latency period of the probiotic group was significantly shortened on the 2nd and 5th days,while the escape latency period was shortened,and the residence time in the target platform quadrant in⁃creased in the high-dose XXD group during the 2nd to 5th days;the escape latency period was shortened significantly in the moderate-dose XXD group on the 5th day(P<0.05 or P<0.01).Compared to those in the model group,the hippocampal neuron cells in the high-and moderate-dose XXD groups were arranged more closely,with decreased levels of SAA,Aβ_(1-42)and LPS,increased ACH level,Simpson and Shannon index(P<0.05 or P<0.01);the arrange⁃ment of hippocampal neuron cells in the probiotic group and the low-dose XXD group was relatively loose;the proportions of Bacteroidetes and Prevotella were significantly reduced in the probiotic group and the high-dose XXD group,while that of Firmicutes and Lactobacillus significantly increased(P<0.05 or P<0.01).Compared to those in the probiotic group and the high-dose XXD group,the number of goblet cells in the moderate-dose XXD group decreased,and the number of glands in the low-dose XXD group decreased with atrophy.The high-dose XXD group had decreased Aβ_(1-42)level in the hippocampus,increased ACH level in thehippocampus and colon tissue,and decreased SAA in the colon tis⁃sue than the moderate-and low-dose XXD groups(P<0.05 or P<0.01);moreover,the SAA level in the hippocampus was significantly higher in the low-dose XXD group than the high-and moderate-dose groups(P<0.01).Conclusion XXD can improve the spatial learning and memory ability of SAMP8,reduce the production and deposition of LPS,SAA and Aβ_(1-42)in brain and intestine,and increase the content of ACH.The mechanism of its prevention and treatment of AD maybe related to regulating intestinal microecology,affecting flora diversity and improving inflammatory response.