目的 探讨n-3 PUFAs缺乏对糖尿病小鼠认知功能的影响及其机制。方法 3周龄自发糖尿病KKay雄性小鼠随机分为n-3 PUFAs正常组（normal n-3 PUFAs content diet,ND）和n-3 PUFAs缺乏组（n-3 PUFAs deficient diet,DD）;实验期间,每周监测体重、进食量;实验结束前一周,检测空腹血糖水平,开展葡萄糖耐量试验,并采用Y迷宫和新物体识别实验评估小鼠认知功能;实验结束后,处死小鼠,留取红细胞、结肠内容物及脑组织。气相色谱检测红细胞膜和脑组织中n-3 PUFAs相对含量;蛋白质免疫印迹检测海马中氧化应激相关蛋白表达水平;宏基因组学和非靶向代谢组学检测结肠内容物中菌群及代谢物。结果 ND组和DD组体重和进食量无显著差异（P>0.05）;DD组空腹血糖高于ND组（P<0.05）,且其葡萄糖耐量曲线下面积大于ND组（P<0.05）;Y迷宫实验结果显示DD组新异臂进入次数显著低于ND组（P<0.05）,新物体识别实验结果显示DD组新物体认知指数、辨别率、差异分数等指数均低于ND组（P<0.05）;DD组海马氧化应激相关蛋白表达水平显著高于ND组;与ND组相比,DD组n-3指数降低（P<0.05）,但脑组织中n-3 PUFAs水平未见明显差异（P>0.05）;非靶向代谢组学和宏基因组分析显示,与ND组相比,DD组代谢物组成发生明显改变,厚壁菌门与拟杆菌门比值降低,一些营养代谢途径发生显著改变。结论 n-3 PUFAs缺乏可能通过影响糖尿病小鼠肠道菌群稳态及代谢物组成加重认知功能障碍。
Abstract
Objective To investigate the impact of dietary n-3 PUFAs deficiency on diabetic cognitive function in mice and its underlying mechanisms. Methods 3-week-old male spontaneous diabetic KKay mice were randomly divided into two groups: the n-3 PUFAs normal group (normal n-3 PUFAs content diet, ND) and the n-3 PUFAs deficient group (n-3 PUFAs deficient diet, DD). Body weight and food intake were monitored weekly. One week before the end of this experiment, the fasting blood glucose detection and glucose tolerance tests were performed. Meanwhile, the cognitive function was assessed using the Y-maze and novel object recognition tests. After an 18-week intervention, all mice were euthanized to collect red blood cells, colonic contents and brain tissues. The relative contents of n-3 PUFAs in red blood cell membranes and brain tissues were determined by gas chromatography. The expression levels of oxidative stress-related proteins in the hippocampus were analyzed by Western blotting. Metagenomics and untargeted metabolomics were used to examine the gut microbiota and metabolites in the colonic contents. Results There were no significant differences in body weight and food intake between the ND and DD groups. Compared with the ND group, the DD group developed increased fasting blood glucose level and exacerbated glucose intolerance. The functional studies showed that n-3 PUFAs deficiency reduced the entries into the new arm, recognition index, discrimination ratio and discrimination score. n-3 PUFAs deficiency increased oxidative stress in the hippocampus. Interestingly, n-3 PUFAs deficiency did not lead to a decrease in the brain n-3 PUFAs level, despite the significant reduction in red blood cell membrane n-3 PUFAs level. Metagenomic and untargeted metabolomicanalyses showed significant alterations in the intestinal metabolic composition in the DD group compared to the ND group, with a decreased ratio of Firmicutes to Bacteroidetes. Significant alterations in the pathways of nutrition metabolism were demonstrated. Conclusion Dietary deficiency of n-3 PUFAs exacerbates cognitive dysfunction in diabetic mice possibly by disrupting gut microbiota homeostasis and altering the composition of metabolites.
关键词
n-3多不饱和脂肪酸 /
糖尿病 /
认知功能障碍 /
肠道菌群 /
小鼠
Key words
n-3 polyunsaturated fatty acids /
diabetes /
cognitive function /
gut microbiota /
mice
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