Study On The Mechanisms Of Astilbin For Accelerating Uric Acid Excretion In Renal Injury Mice Based On Gut Microbiota

Objective To identify gut microbiota is the target of astilbin(the main active ingredient of the anti-gout traditional Chinese medicine Smilax glabra Roxb.)to accelerate uric acid excretion by comparing its effects in SPF and pseudo-germ-free mice with renal injury,and explore its mechanisms.Methods Male C57BL/6 mice were divided into SPF control group(S-CN),SPF model group(S-MD)and SPF astilbin low dose group(S-AL),SPF astilbin high dose group(S-AH),pseudo-germ-free control group(P-CN),pseudo-germ-free model group(P-MD)and pseudo-germ-free astilbin low dose group(P-AL),pseudo-germ-free astilbin high dose group(P-AH),10 mice per group.In addition to S-CN and P-CN groups,all the other groups were fed AIN-93M diet containing 0.2%adenine for 3 weeks,and all mice were fed AIN 93M diet from the 4th week to the end of the experiment.All pseudo-germ-free group was given drinking water with antibiotic mixture(vancomycin 200 mg/kg,metronidazole 200 mg/kg,neomycin 200 mg/kg)for 6 weeks from the 4th week,and the SPF mice were given distilled water.At the same time,the mice in the S-AL group and the P-AL group were given astilbin(25 mg/kg),and the S-AH group and the P-AH group were given astilbin(50 mg/kg)for 6 weeks from the beginning of the 4th week.At the end of the experiment,serum of the mice was collected to detected creatinine(CRE)and blood urea nitrogen(BUN).Uric acid(UA)levels in serum,intestinal contents,feces and urine were determined.The mice feces were detected by 16S rDNA sequencing,the contents of short chain fatty acids(SCFAs)and the metabolites such as hippuric acid(HA),indole-3-acetic acid(IAA),p-Cresyl sulfate(p-CS)and indoxyl sulfate(IS)were measured in serum,feces and urine.Determination of glucose transporter 9(GLUT9),organic anion transporter 1(OAT1),organic cation transporter 2(OCTN2),urate anion exchanger 1(URAT1)protein expression levels in renal,and the expression levels of ABCG2 in intestinal(duodenum,ileum,colon)and kidney.Results 1.Compared to model group,astilbin reduced serum uric acid and increased uric acid in urine and intestinal cavity,and the effect of uric acid excretion in SPF renal injury mice was stronger than that in pseudo-germ-free renal injury mice.2.The expression of ABCG2,OCTN2 and OAT1 in kidney with renal injury mice were promoted by astilbin,and the expression of URAT1 and GLUT9 were reduced.At the same time,it can also up-regulated the expression of the uric acid transporter ABCG2 in the intestine(duodenum,ileum and colon),thus excretion of uric acid was promoted.In addition,the effect of astilbin on uric acid transporter in pseudo-germ-free mice was significantly lower than that in SPF mice.3.Astilbin reduced the gut microbiota which prone to toxic metabolites in renal injury mice,such as Bacteroides,Akkermansia,Barnesiella,Parabacteroides,Eisenbergiella,Peptococcus,Oscillibacter,Anaerotruncus,Pseudoflavonifractor,Eubacterium,etc.And the number of bacteria groups of Clostridiaceae,Enterobacteriaceae and Micrococcaceae which can secrete uricase and urease were reduced.Besides,the Allobaculum,Bifidobacterium,Olsenella and Lactobacillus were increase by astilbin,which promote the formation of short-chain fatty acids.4.Compared to model group,the contents of IS,p-CS,HA and IAA in feces and serum of renal injury mice were significantly reduced,the contents of IS,HA and IAA in urine were reduced,but the contents of p-CS in urine were increased by astilbin.At the same time,astilbin significantly increased the content of SCFAs in feces and serum of renal injury mice.Conclusion Astilbin could regulated the composition of gut microbiota and metabolites of bacteria,the functions of kidney and intestinal uric acid transporters in renal injury mice were improved,and the excretion of uric acid in renal and intestinal were further improved.