高尿酸血症脾虚湿盛证病证结合小鼠模型的建立及其肠道菌群特征分析

Establishment of hyperuricemia mouse model with spleen deficiency and dampness abundance syndrome and analysis of its intestinal flora characteristics

目的:建立一种契合临床高尿酸血症(HUA)脾虚湿盛证患者发病特点的病证结合小鼠模型,分析其肠道菌群特征,探究病证结合造模对肠道菌群的影响。方法:40只SPF级2月龄雄性KM小鼠按随机数表法分为正常组、HUA组、脾虚湿盛证组、病证结合组(HUA脾虚湿盛证),每组10只。分别予普通饲料+0.9%氯化钠溶液、普通饲料+次黄嘌呤联合氧嗪酸钾、高脂高糖饲料联合劳倦过度+0.9%氯化钠溶液、高脂高糖饲料联合劳倦过度+次黄嘌呤联合氧嗪酸钾灌胃干预。通过各组小鼠血尿酸水平、脾虚湿盛证候积分评价模型建立的效果,并于干预结束后收集各组小鼠粪便,采用16SrRNA测序技术检测粪便中菌群结构,分析模型小鼠肠道菌群特征。结果:与正常组及HUA组比较,病证结合组小鼠血尿酸水平、脾虚湿盛证候积分显著升高(P<0.01,P<0.05);肠道菌群结构发生明显改变,拟杆菌门、放线菌门相对丰度显著上升,厚壁菌门相对丰度显著下降,肠道有益菌乳杆菌属相对丰度显著下降(P<0.05)。与脾虚湿盛证组比较,病证结合组小鼠血尿酸水平显著升高(P<0.05);拟杆菌门相对丰度显著上升,厚壁菌门相对丰度显著降低(P<0.05);两组脾虚湿盛证候积分差异无统计学意义。结论:高脂高糖饲料联合劳倦过度+次黄嘌呤联合氧嗪酸钾灌胃可较好构建HUA脾虚湿盛证病证结合小鼠模型,并导致小鼠肠道菌群结构发生显著变化,提示HUA脾虚湿盛证的发生可能与肠道菌群的改变高度相关。

Objective:Establish a disease syndrome combined mouse model that fits the clinical characteristics of hyperuricemia(HUA)patients with spleen deficiency and dampness abundance syndrome,further more,analyze the features of its intestinal flora,explore the effect of this modeling method on intestinal flora.Methods:Forty SPF 2-month-old male KM mice were randomly divided into normal group,HUA group,spleen deficiency and dampness abundance syndrome group,and HUA plus spleen deficiency and dampness abundance group,with 10 mice in each group.They were given normal diet+normal saline,normal diet+hypoxanthine combined with potassium oxazinate,high-fat and high sugar diet combined with overwork+normal saline,highfat and high sugar diet combined with overwork+hypoxanthine combined with potassium oxazinate.The blood uric acid level and the score of spleen deficiency and dampness abundance syndrome were used to evaluate the effect of model establishment.The feces of mice in each group were collected,and the bacterial structure composition in feces was detected by 16S rRNA sequencing technology to analyze the characteristics of intestinal flora of model mice.Results:Compared with the normal group and HUA group,the level of blood uric acid and the score of spleen deficiency and dampness abundance syndrome in the HUA plus spleen deficiency and dampness abundance group were significantly higher(P<0.01,P<0.05).The structure and composition of intestinal flora were significantly changed,the relative abundance of Bacteroidea and Actinobacteria increased,the relative abundance of Firmicutes decreased,and the relative abundance of beneficial intestinal Lactobacillus decreased significantly(P<0.05).Compared with the spleen deficiency and dampness abundance syndrome group,the serum uric acid level of mice in the HUA plus spleen deficiency and dampness abundance group was significantly higher(P<0.05).The relative abundance of Bacteroidetes increased and Firmicutes decreased significantly(P<0.05).There was no significant difference in the syndrome score of spleen deficiency and dampness abundance.Conclusion:High fat and high sugar diet combined with overwork+hypoxanthine combined with potassium oxazinate by gavage can successfully construct the mouse model of HUA combined with spleen deficiency and dampness abundance syndrome,and leads to the significant changes of the intestinal flora suggesting that the occurrence of spleen deficiency and dampness abundance syndrome in HUA is highly related to the changes of intestinal flora.