Molecular Mechanism Of Mulberry Branch Bark Improving Metabolic Imbalance In Diabetic Mice

Unlike the traditional Chinese medieine mulberry branches,the research object of this subject is the ainual bark of mulberry trees.The skin and phloem tissue of mulberry branches can be obtained by the traditional Chinese medicine peeling machine.In recent years,mulberry bark has been proved to have antioxidant,hypoglycemic,hypolipidemic and other effects.In this study,we first explored the chemical composition of mulberry bark,then established the diabetic model of C57/6J mice,and then analyzed the effect of mulberry bark on diabetes mellitus in mice by non-targeted metabonomics using GC-MS platform.At the same time,we also examined the effects of mulberry bark on immune cells and inflammatory factors in mice,and selected the potential target Foxol for protein and gene detection.The main contents of this paper are as follows:(1)a large number of compounds in mulberry branch bark were detected by LC-MS method,including amino acids,polysaccharides,vitamins,organic acids,flavonoids,alkaloids,steroids,phenylpropanoids and coumarins:a variety of characteristic amino acids and derivatives,dipeptides,such as threonine,O-phosphate-L-threonine;monosaccharides,disaccharides,oligosaccharides;a variety of nutrients such as inositol,vitamin A,B,D;A variety of isopentenyl,phenolic hydroxyl,methoxy substituted flavones,flavonols,chalcones,isoflavones,flavanols,anthocyanins and other compounds;N-heterocyclic alkaloids such as 1-deoxynojirimycin and its derivatives,scopolamine,bowlflower anthocyanin,etc;steroids such as rapeseed sterol,birch acid,isoergosterol,etc;common small molecular organic acids and fatty acids such as tartaric acid,palmitic acid and acidic compounds rich in phenolic hydroxyl groups,such as resveratrol and resveratrol oxide;phenylpropanoids such as aescin and curcumin,ferulic coumarin and other coumarins;in addition,there are other compounds containing benzofuran(ketone),benzopyran(ketone),diphenyl ethylene and other structural types.(2)The polysaccharides in mulberry bark are mainly composed of rhamnose,xylose,arabinose,mannose,glucose,galactose and galactUronic acid.The main chains are mainly composed of(1?3)linked glucose or(1?3)linked rhalnose and(1?2,4)linked xylose.(3)A diabetic mice model with insufficient insulin secretion and insulin resistance was established by injecting streptozotocin(STZ)and inducing high-fat diet to simulate the complex diabetes which has been found in clinic case.(4)Diabetic mice were fed with three different doses of mulberry bark powder.Metabolites in serum samples of mice were analyzed by metabolomics method based on GC-MS,and were verified by biochemical evaluation,histological examination and enzyme protein detection.Through the analysis of the difference of endogenous metabolites and the enrichment of biological functions,it was found that the metabolic disorders of model mice included the activation of sorbitol pathway and the enhancement of galactose metabolism,the enhancement of gluconeogenesis pathway,fatty acid oxidation pathway and protein decomposition pathway,the weakening of pentose phosphoric acid pathway,glycolysis pathway and aerobic oxidation pathway,the synthesis of cholesterol and the production of inositol.Compared with model mice,these abnormal metabolic changes were alleviated in mulberry bark feeding group,and showed in a dose dependent way.More specifically,it is found that mulberry bark promotes glucose metabolism by increasing inositol;the pentose phosphate pathway providing NADPH is strengthened,which can alleviate oxidative stress at the material level;inhibit the activity of carnitine palmitoyl transferase I,and promote the reduction of fatty acid oxidation level and ketone body formation.In conclusion,mulberry bark feeding increased the level of sugar metabolism in model mice,which reduced the energy demand for lipid metabolism and protein metabolism,alleviated oxidative stress,changed the unreasonable regulation of tricarboxylic acid cycle by too much NADH and too little ATP,and normalized energy metabolism.(5)The composition and proportion of lymphocyte in the spleen of immune organs of diabetic eice changed to some extent compared with the normal group.The CD3+T cells increased slightly,the ratio of CD4+/CD8+decreased,the ratio of B cells decreased,and the NK cells also decreased slightly.These changes were inhibited to some extent by mulberry branch bark therapy.The expression levels of pro-inflammatory factors such as NF-?B p65?IL-6?IL-1??TNF-? increased in the model group,suggesting that there was systemic inflammation in the model group.The expression of pro-inflammatory factors in mulberry branch bark treatment group was decreased,which indicated that mulberry branch bark could improve the inflammation of diabetic mice.(6)Foxol,the upstream target of glycometabolism,was selected to test the effect of mulberry bark on the pancreas of mice with streptozotocin injection injury.The results showed that prophylactic feeding of mulberry bark could alleviate STZ-induced pancreatic beta cell dysfunction and injury,and improve sugar metabolism.In PI3K/Akt/Fox01 signaling pathway,activation of Akt and inhibition of Foxol were involved in the protection of mulberry bark.The translocation of FoxOl from nucleus to cytoplasm was enhanced,and the target of Foxol gene in nucleus was inhibited.At the same time,mulberry branch bark could increase the gene expression of PI3K,PDK1,AKT and FoxO1.In this paper,the chemical composition of mulberry bark of Husang was comprehensively analyzed.The effects of mulberry bark on endogenous metabolism in diabetic mice were systematically detected and analyzed on the platform of GC-MS.An upstream target of controlling sugar metabolism,FoxO1,was selected and validated.The results of this study pave the way for the further development of the molecular mechanism of hypoglycemia and diabetes mellitus.Follow-up work can continuously follow the clues of metabolite changes to explore the target protein and gene levels.The changes of CD4+T cells,CD8+T cells and related inflammatory factors in immune cells also indicate that more detailed changes of inflammatory cell subtypes can be explored.This topic also provides a basis for expanding the medicinal resources of mulberry and macing full use of mulberry bark,a waste of agricultural cultivation.