| Objective:Polygonati Rhizoma is a medicinal and edible Chinese medicinal materials.Polygonatum kingianum is the genuine medicinal materials in Yunnan Province,and showed the largest production in three varieties contained in the Chinese pharmacopoeia.Early studies showed that polysaccharide of Polygonatum kingianum has a strong inhibitory effect on alpha glucosidase,which can reduce TC and TG contents of serum in the high fat diet and streptozotocin induced type 2 diabetic rats.These results indicated that the polysaccharide from Polygonatum sibiricum could regulate the metabolism of glucose and lipid.In order to explore the polysaccharide from Polygonatum kingianum on the regulation of lipid metabolism disorders and its mechanism.this study separated and purified the polysaccharide,and conducted its chemical composition analysis and quality control.Experiments were explored the effects of polysaccharide from Polygonatum kingianum lipid metabolism disorder of rats,its regulation on intestinal microflora and molecular messenger SCFAs,and the effects of polysaccharide on LPS-TLR4/NF?B signaling pathway.This research may provide more scientific evidences on the development and utilization of “Yunyao”.Methods:The gel permeation chromatography?GPC?,liquid chromatography?HPLC?and chemical analysis were used to determine the molecular weight,monosaccharide composition,protein,polysaccharide and uronic acid content of the polysaccharide from Polygonatum kingianum,respectively.The lipid metabolism disorder of Spraque-Dawley rats were induced by high fat diet.At the same time,SD rats were intervened by the high molecular weight polysaccharide?>100KDa??PSF?and total polysaccharide?PS?of different concentration of aqueous solution.TC,TG,HDL-C,LDL-C contents in blood samples were measured every two weeks,evaluating of blood lipids regulation activity of polysaccharide.The stool samples were used by 16 S rDNA V4 highly variable region measurement method to regulate the role of polysaccharide in the intestinal microflora.The content of short chain fatty acid?SCFAs?in feces was determined by gas chromatography?GC?to investigate the effect of polysaccharides on the content of SCFAs in the intestinal tract of rats.The expression of tight junction protein ZO-1 and occludin of intestinal tissue,and detection of the key protein expression in the LPS-TLR4/NF?B signaling pathway were detected by Western blot technique.Results:The gel permeation chromatography?GPC?analysis showed that the average molecular weights of PSF and PS were 1.786×105Da and 1.347×105Da,respectively,which were composed of mannose,galacturonic acid,galactose and fucose,in which the content of total sugar were 30.80%?39.17% respectively,the content of uronic acid were 25.06%?27.81%,the total protein content of were 10.91%,7.65%.High fat diet SD rats were given polysaccharide solution by intragastric gavage,PSF and PS could reduce body weight,reduce epididymal fat and liver index,inhibited epididymal fat cells growth,alleviate hepatic steatosis,reduce TC,TG,LDL-C level and increased HDL-C levels in serum.PSF and PS could reduce fasting blood glucose and insulin levels in serum,inhibited insulin resistance,which provided oral glucose tolerance and promote the ADP secretion in fat cells,inhibited the secretion of LEP,and activate PPAR? expression.It is indicated that the polysaccharide could enhance insulin sensitivity,which regulate the energy metabolism in obese rats.The result of V4 highly variable region of 16 S rDNA showed that The polysaccharide from Polygonatum kingianum could downregulate Firmicutes to Bacteroidetes ratio,decrease the relative abundance of Proteobacteria,while reducing the relative abundance of Lactobacillus and Psychrobacter,improve the composition of intestinal microflora.115 OTUs and host metabolic markers were measured by Pearson correlation analysis in PSF groups,103 OTUs and host metabolic markers were measured by Pearson correlation analysis in PS groups.The results suggested that OTUs belong to Firmicutes and Bacteroidetes showed positive correlation with FINS and FBG,respectively;OTUs belong to Proteobacteria showed a positive correlation with LPS,a negative correlation with serum HDL-C in PSF groups.However,OTUs belong to Firmicutes showed a negative correlation with FINS and TG of liver,respectively;and showed a positive correlation with HDL-C of serum in PS groups.On the contrary,OTUs belong to Bacteroidetes showed a positive correlation with FINS and TG of liver,respectively;and showed a negative correlation with HDL-C of serum in PS groups.Meanwhile,those belonging to putative short-chain fatty acids?SCFA?-producing bacteria were increased by both PSF and PS,promote the production of SCFAs.The relative abundance of Clostridiumsensustricto1,Roseburia,Bifidobacterium,Streptococcus and Allobaculum was positively correlated with the content of SCFAs.Obevious degeneration and even necrosis in intestinal mucosa epithelial tissues were observed in high fat diet rats.The expression of intestinal tight junction protein ZO-1 and occludin were significantly decreased compared with normal rats,so that LPS enters the blood circulation and activates TLR4/NF?B signaling pathways that induce chronic low-grade inflammatory states.The expression of ZO-1 and occludin protein in the intestine was significantly higher than that in the model group,and the LPS content in the portal vein was significantly decreased,which decreased the expression of TLR4 protein,reduce the degradation of I?B-?,inhibit the release of TLR4/NF?B signaling pathway mediated proinflammatory cytokines IL-1? andTNF-?,promote the production of anti-inflammatory factor IL-10,alleviate chronic low-grade inflammation,and improve lipid metabolism disorder status.Conclusion:The polysaccharide from Polygonatum kingianum could activate the PPAR? to promote the differentiation of adipocytes,reduce the volume of cells,alleviate liver fat steatosis,increase fat factor,enhance insulin sensitivity,reduce insulin resistance.Both PSF and PS could effective improve the intestinal microflora,enhance the content of SCFAs,promote the repair of intestinal epithelial cells,reduce intestinal permeability,and alleviate the chronic low-grade inflammation by reduce the LPS into the blood circulation,inhibit TLR4/NF?B signaling pathway,improve lipid metabolism disorders.PS regulates lipid accumulation better than PSF.PSF regulates the composition and function of intestinal flora,reduces intestinal permeability and inhibits TLR4/NF?B signaling pathway better than PS. |
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