| Objective: Type 2 diabetes mellitus is a metabolic syndrome caused by insulin resistance and insulin secretion.It has become the third largest chronic non-communicable disease after malignant tumor and cardiovascular disease.Its pathogenesis is complex,and the clinical curative effect is not remarkable.In recent years,intestinal microbes have become the focus of many diseases in medical field.The purpose of this study was to investigate the effects of intestinal microorganism transplantation on the formation and improvement of insulin resistance,and the relationship between microorganism and plasma amino acid level in the development of insulin resistance.Methods: 1)The fecal microorganism group of Uygur type 2 diabetes mellitus was transplanted into db/m mice to establish insulin resistance model.Six mice were randomly selected for index detection and 3 mice were killed at 0 week.The remaining mice were randomly divided into db/m+PBS and db/m+UT2DM groups according to their body weight.The animals were gavaged with the fecal fluid of the carbocyanine fluorescent dye tracer chlormethylbenzamido1,1-dioctade-cyl-3,3,3'3'-tetramethylindo-carbocyamine used for cell staining.Fecal samples were collected after intragastric administration with sterile cryopreserved tubes on days 3,7 and 14 respectively.The colonization of intestinal bacteria in mice was observed by fluorescence microscopy.Blood glucose levels were detected by glucose oxidase method,blood lipid level was detected by enzyme-linked immunosorbent assay,and the abundance of target bacteria in fecal samples of mice was detected by real-time fluorescence quantitative PCR at 4,6,8 and 10 weeks,respectively.2)Establishment of insulin resistance model in mice by feeded with high fat feed and perfused fecal bacteria from Uygur patients with type 2 diabetes mellitus.At the same time,to observe the preventive effect of faecal bacteria of Kazak normal glucose tolerance human on the formation of insulin resistance.the mice were gavaged with fecal bacteria of Kazak normal glucose tolerance human.Ten mice were randomly selected as 0-week control.The remaining mice were randomly divided into NCD,NCD+UT2DM,HFD,HFD+UT2DM,HFD+KNGT.The detection of colonization ability of intestinal microflora,glycolipid levels and the abundance of target bacteria in mice were the same as method 1).3)According to the criteria of insulin resistance in mice,insulin resistance mice were selected from mice fed with high fat diet and fecal bacteria solution of Uygur type 2diabetes mellitus for 8 weeks.The mice were divided into groups after testing the index of0 week.The faecal bacteria of Kazakh with normal glucose tolerance were gavaged into db/db diabetic mice and insulin resistant mouse using fecal bacteria transplantation technique.Metformin was used as positive drug to observe the effect of fecal bacteria transplantation on metabolism of mice.4)High performance liquid chromatography was used to detect the levels of plasma amino acids in mice at different time points.Pearson correlation analysis was used to analyze the relationship between intestinal microflora and amino acid metabolic profile and its correlation with the changes of glycolipid level related to insulin resistance.The effects of intestinal microorganism and plasma amino acid levels on the development of insulin resistance in mice were analyzed by multivariate stepwise regression method.Result: 1)The results of variable region sequencing of Illumina16 Sr RNAgene showed that the abundance of intestinal Clostridiaceae in db/db mice was significantly higher than that of db/m mice at the level of science.The abundance of Clostridium in intestinal tract of mice treated with fecal bacteria of Uygur type 2 diabetes mellitus for 6 weeks was significantly increased at the generic level.Compared with the db/m mice,the abundances of the intestinal Desulfovibrio in the db/db diabetic mice were significantly higher at the species level.2)Fecal microbial community transplantation from Uygur patients with type 2 diabetes could lead to the disorder of glycolipid metabolism in normal glucose tolerance mice,and the changes of the abundance of Desulfovibrio,Clostridium coccoides and Akkermansia muciniphila in the intestinal tract,and leading to the occurrence of insulin resistance in mice.3)The fecal microbial community transplantation of normal glucose tolerance in Kazakh nationality could significantly improve the glycolipid metabolism and increase the abundance of Akkermansia muciniphila in the intestinal tract of db/db2 diabetic mice.It also inhibited the increase of the abundance of Desulfovibrio and Clostridium coccoides in the intestinal tract of mice.4)Transplantation of faecal microbial community in normal glucose tolerance population of Kazakh can significantly inhibit the increase of brain-body ratio in insulin-resistant mice,which is superior to metformin.But the effect on lipid body ratio in insulin resistant mice was significantly lower than that of metformin,and the effect of intestinal target microbial was later than metformin.5)High fat diet and microbial transplantation in feces of Kazakh with normal glucose tolerance can cause the changes of plasma amino acids in mice.And the changes were significantly correlated with intestinal flora and insulin resistance phenotype.6)The results of multiple stepwise regression analysis showed that the effect of Desulfovibrio on plasma glycine in mice was significantly positive correlation.The effect of Akkermansia muciniphila on plasma isoleucine was negatively correlated.The effects of valine on the abundance of desulfovibrio and fasting blood glucose were negatively correlated,while aspartic acid and isoleucine had a negative correlation on the abundance of Akkermansia muciniphila.7)Microbial transplants in faeces of normal glucose tolerance in Kazakh nationality had a significant negative correlation with glycolipid metabolism during insulin resistance in mice.Clostridium coccoides has a significant positive correlation with plasma total cholesterol and triglyceride.The microbial transplants in faeces of normal glucose tolerance in Kazak nationality have a significant negative correlation with glucose and lipid metabolism during insulin resistance formation in mice.Clostridium coccoides has a significant positive correlation with plasma total cholesterol and triglyceride.Conclusion:1)Fecal microbial community transplantation in Uygur patients with type 2 diabetes can cause disorder of glycolipid metabolism and change of intestinal microorganism abundance in normal glucose tolerance mice,and promote and lead to insulin resistance in mice.2)The faecal microbial community transplantation from Kazakh normal glucose tolerance people effectively inhibited the disorder of glycolipid metabolism and the change of intestinal microbial structure in the process of insulin resistance induced by high-fat diet,and had a preventive effect on the production of insulin resistance.It also significantly improved the glucose and lipid metabolism and regulated the intestinal flora abundance in db/db2 diabetic mice.3)Microbial transplants from normal glucose tolerant of Kazakh significantly inhibited the increase of glycolipid levels during insulin resistance in mice.The decrease of the abundance of Clostridium coccoides can inhibit the increase of plasma total cholesterol and triglyceride.In addition,valine may inhibit the increase of fasting blood glucose by reducing the abundance of Desulfovibrio. |
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