基于肠道菌群-LPS/TLR4通路探讨野菊花提取物对“过食膏粱厚味”致高尿酸血症大鼠的作用

Based on Intestinal flora-LPS/TLR4 to Research the Effect of Chrysanthemi Indici Flos Extract on the Model of Hyperuricemia Induced by Overeating Greasy

[目的]研究野菊花提取物(Chrysanthemi Indici Flos extract,CYM.E)对“过食膏粱厚味”致高尿酸血症(hyperuricemia,HUA)大鼠的治疗作用及机制。[方法]将64只大鼠随机分为正常对照组,模型对照组,别嘌醇10 mg·kg^(-1)组,苯溴马隆5 mg·kg^(-1)组,CYM.E15、30、60、90 mg·kg^(-1)组,每组8只。除正常对照组外,其余各组均灌胃脂肪乳剂,建立拟人“过食膏粱厚味”致HUA大鼠模型。实验期间测定血清尿酸(uric acid,UA)、总胆固醇(total cholesterol,TC)、甘油三酯(triglyceride,TG)、高密度脂蛋白胆固醇(high density lipoprotein cholesterol,HDL-c)和低密度脂蛋白胆固醇(low density lipoprotein cholesterol,LDL-c)水平;酶联免疫吸附测定(enzyme linked immunosorbent assay,ELISA)法测定血清白细胞介素-6(interleukin-6,IL-6)、肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)和血浆脂多糖(lipopolysaccharide,LPS)水平;实时荧光定量聚合酶链式反应(Real-time quantitative polymerase chain reaction,qRT-PCR)法测定小肠中TNF-α、Toll样受体4(Toll like receptor 4,TLR4)的mRNA表达;免疫印迹法检测小肠中TLR4蛋白表达;应用16S rDNA测序技术,对肠道菌群进行操作分类单元(operational taxonomic units,OTU)分类,选择合适的OTU进行结构分析、LEfSe差异贡献分析等。[结果]CYM.E能显著降低模型大鼠血清UA、TC、LDL-c、IL-6和TNF-α水平;显著降低血浆LPS水平;同时显著下调小肠组织TNF-αmRNA、TLR4 mRNA表达和TLR4蛋白表达,改善肠道的微炎症状态;增加乳酸杆菌、双歧杆菌的丰度与种类,降低拟杆菌、大肠杆菌的丰度与种类。[结论]CYM.E具有降低血清UA水平、调节血脂、减低炎症损伤、减少LPS释放等作用,可能通过调节肠道微生态发挥作用。

[Objective]To study the effect and mechanism of Chrysanthemi Indici Flos extract(CYM.E)on hyperuricemia in rats induced by“overeatinggreasy.”[Methods]Sixty-four rats were randomly dividedinto normal control group,model control group,all opurinol 10mg·kg^(-1)group,Benzbromarone 5 mg·kg^(-1)group,and CYM.E 15,30,60 and 90 mg·kg^(-1)group,with 8 rats in each group.Except for the normal control group,the other groups used fat emulsion to establish a rat model of anthropomorphic hyperuricemia based on“overeating greasy”.During the experiment,serum uric acid(UA),total cholesterol(TC),triglyceride(TG),high density lipoprotein cholesterol(HDL-c)and low density lipoprotein cholesterol(LDL-c)levels were measured.Interleukin-6(IL-6),tumor necrosis factor-α(TNF-α)and lipopolysaccharide(LPS)were determined by enzyme linked immunosorbent assay(ELISA).The mRNA expression of TNF-α and Toll like receptor 4(TLR4)in the small intestine was measured by Real-time quantitative polymerase chain reaction(qRT-PCR);and the protein expression of TLR4 in the small intestine was measured by Western blot.Intestinal microecology was classified with operational taxonomic units(OTU),and OTU was analyzed with structure,and LEfSe difference,etc.[Results]CYM.E significantly reduced the serum UA,TC,LDL-c,IL-6 and TNF-α levels of model rats;and reduced plasma LPS level.Moreover,CYM.E could reduce the expression of TNF-αmRNA and TLR4 mRNA in the small intestine,and reduced the protein expression of TLR4 in the small intestine,thus improving the microinflammatory state of the intestinal tract,increased the abundance and species of Lactobacillales and Bifidobacteriales,and reduced the abundance and species of Bacteroidales and Escherichia coli.[Conclusion]CYM.E has the effects of decreasing serum UA levels,regulating blood lipids,decreasing inflammatory damage,and decreasing endotoxin release,which may exert an UA-lowering effect by regulating intestinal microecology.