| Objective:To observe the effect of Tangnaikang on metabolic syndrome,insulin resistance and renal damage in SHR/CP rats by biochemical detection,16S sequencing,protein chip,histopathological staining and protein imprinting,and to provide evidence for clinical applicationMethods:1.Effects of Tangnaikang on metabolic syndrome in SHR/CP rats:according to FBG and body weight level,SHR/CP rats with spontaneous hypertension and obesity were selected.The rats were randomly divided into two groups(n=8):(1)the rats treated with 3.24g/kg TNK and(2)the untreated model group(CON).TNK was prepared with sterile water and administered by gavage once a day for 8 weeks.The model group was given the same volume of sterile water.Age matched male WKY rats were used as normal control group.During the whole experiment,weight,food intake and water intake were recorded every three days.The tail blood pressure was measured at the end of two weeks.The urine volume was recorded in the 8th week.After 8 weeks of treatment,blood samples were collected to detect blood glucose,blood lipid and renal function indexes,and 24-hour urine protein content was detected2.The effect of Tangnaikang on intestinal flora of SHR/CP rats:before the establishment of the model and administration of Tangnaikang,samples were taken 8 weeks later,the rectal contents of rats were taken,and fresh feces were stored at-80?.The diversity of intestinal microorganisms was studied by 16S rDNA sequencing and sub genome analysis to reflect the changes of functional gene expression.Serum biomarker analysis was used to study the level of blood glucose and lipid in order to determine the changes of sub genome and microbial community abundance.3.The mechanism of Tangnaikang on improving renal damage in SHR/CP rats:the kidney tissue was fixed and frozen at-80? after 8 weeks of administration.The mechanism of action was further studied by renal histopathology staining and 67 kinds of cytokine antibody chipResults:1.Effects of Tangnaikang on metabolic syndrome in SHR/CP rats:after 8 weeks of treatment,the body weight,BM1,food intake and urine volume of WKY group were lower than those of model group,and the water volume was higher than that of model group.TNK could reduce the body weight and urine volume(P<0.01).The body weight,BMI,blood glucose,OGTT,aucogtt,triglyceride,cholesterol,systolic blood pressure and diastolic blood pressure in the model group were significantly higher than those in the normal group(P<0.001).After TNK administration,the indexes of blood glucose,blood pressure,OGTT,aucogtt,triglyceride,cholesterol,systolic blood pressure and diastolic blood pressure in TNK group were all improved compared with those in the model group,and the results were statistically different(P<0.01,P<0.05).2.The effect of Tangnaikang on the intestinal flora of SHR/CP rats:753 OTUs were produced by sequencing the paired ends,representing 865 ecological groups,and 123 genera and 10 phyla were identified.Calculate the ecological diversity index of each sample,such as chaos,PD whole tree,Shannon,etc.There was significant difference between WKY group and model group,but there was no signi ficant difference between other groups ? diversity analysis showed that there was significant difference in bacterial abundance between model group,TNK group and WKY group.The abundance of akkermansiaceae in the model group was lower than that in the normal group.However,after TNK treatment,the abundance of akkermansiaceae did not increase significantly compared with the model group.The abundances of clostridiaceae,erysipelototrichaeae,defluviitaleaceae and family ?? were higher than those of model group It is worth noting that the number of clostridiaceae 1 decreased significantly after TNK treatment.TNK caused the imbalance of intestinal microflora and changed the abundance of some rare genera,including Clostridium-sensu-stricto-1 and eisenbergiella.TNK also regulates metabolic genes in the genome.The regulation of Clostridium-sensu-stricto-1 and eisenbergella and the changes of their subgenomes are related to the changes of blood glucose and lipid levels,especially the changes of triglycerides and cholesterol.3.The mechanism of Tangnaikang improving renal damage in SHR/CP rats:the serum creatinine,urea nitrogen and urine protein in model group were significantly higher than those in normal group(P<0.01,P<0.05).After 8 weeks of administration,TNK significantly reduced serum creatinine,urea nitrogen and urinary protein(P<0.01,P<0.05).Pathological images showed that the renal lesions in TNK group decreased.Compared with the normal group,the expression of galectin-3,TIMP-1,P-cadherin,activin A,ctack,nope,VEGF,prolactin,SCF,EphA5 and adiponectin in the model group were significantly up-regulated(P<0.01)and fractalkine was significantly down regulated(P<0.001).Compared with the model group5 the expression of ICAM-1 and TIMP-1 in TNK group was significantly down regulated(P<0.05),while the expression of CINC-2,Ifng,IL-1B,IL-2 and prolactin r was significantly up regulated(P<0.01,<0.05),which were mainly enriched in TNF,NF kappa B,JAK-STAT and IL-17 signaling pathways.WB results showed that TNK decreased the phosphorylation level of JAK2,STAT1 and STAT3,and the expression of ICAM-1 and IL-1B protein compared with the model group.Conclusion:1.Effect of Tangnaikang on metabolic syndrome in SHR/CP rats:TNK can improve metabolic disorder in SHR/CP rats2.The effect of Tangnaikang on intestinal microflora of SHR/CP rats:TNK can promote the production of SCFAs and improve insulin resistance and obesity of T2DM by regulating intestinal microflora,which may be related to amino acid pathway.3.Study on the mechanism of Tangnaikang improving renal damage in SHR/CP rats:TNK can improve the metabolic disorder and renal damage in shr-cp rats.The most important mechanism of TNK therapy is to regulate JAK stat,TNF,NF kappa B,IL-17 signaling pathway.TNK can significantly reduce the expression of inflammatory factors and the phosphorylation activity of JAK/STAT pathway protein. |
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