| Diabetic nephropathy(DN)is one of the most serious complications of diabetes and has become the main cause of end-stage renal disease.At present,angiotensin-converting enzyme inhibitors have some adverse reactions.Therefore,it is necessary to study new anti-DN drugs.Paecilomyces cicada is a fungus that can reduce urine protein,serum creatinine and protect the kidneys.Radix astragli is a Chinese medicine,which can protect the glomerular podocytes and delay DN.However,However,radix astragli could not be completely absorbed by human body due to its large dosage.The use of microorganisms to ferment traditional Chinese medicine can enhance the efficacy of microorganisms through biotransformation.Using Paecilomyces cicadae as a fermentation strain to ferment radix astragli.The growth and metabolism of Paecilomyces cicadae can be used to transform the components contained in radix astragli,so as to improve the efficacy of radix astragli.In this paper,Paecilomyces cicadae was used for solid-state fermentation of radix astragli(RPF).Frist,DN mice were used as the model to evaluate the efficacy of fermentation products and compare with radix astragli.Then,explore the effect of RPF on intestinal flora and the mechanism of action on podocytes.These studies provide reference for the research and development of anti DN drugsMain research:(1)The components in RPF water extract were determined by HPLC and compared with those in radix astragli.The results showed that compared with radix astragli,the chemical composition of RPF increased significantly.In addition,compared with radix astragli,the content of Calycosin-7-glucoside and Ononin in RPF decreased significantly,but the content of Calycosin had no significant change Cordycepic acid and N-6(2-hydroxyethyl)adenosine in RPF come from the growth and metabolism of Paecilomyces cicadae.These results indicate that Paecilomyces cicadae transformed the substances in radix astragli during the fermentation process(2)DN model mice were established and metformin hydrochloride was used as a positive drug.Measure the body weight,diet,drinking water,urinary protein,fasting blood glucose,serum creatinine,blood urea nitrogen and other physiological indicators of the mice,and evaluate the improvement of DN by RPF.To further evaluate the effect of RPF on mouse kidney tissue function.We used HE and PAS staining to check renal pathology,Western blot to detect renal autophagy activity,and immunohistochemistry to detect the expression of podocyte marker proteins Synaptopodin and Podocin.The results show that RPF can significantly increase the body weight of DN mice,and reduce the amount of diet intake and water intake.At the same time,RPF can significantly reduce urinary protein,fasting blood glucose,serum creatinine,blood urea nitrogen in DN mice,improve glomerular tissue lesions,increase renal tissue autophagy activity.In addition,the effect of RPF against DN is better than radix astragli.Furthermore,compared with the metformin hydrochloride,there were no significant differences in renal function indexes such as urine protein,fasting blood glucose,serum creatinine,and blood urea nitrogen in DN mice in the RPF group,and RPF delayed DN by targeting podocytes.(3)In order to explore the protective mechanism of RPF on podocytes,a high glucose-induced podocyte injury model was established.we detected the cell viability,apoptosis rate,reactive oxygen level,the expression of podocyte marker protein,autophagy marker protein and PI3K/AKT/mTOR signaling pathway related protein.The results show that RPF can increase the cell viability of podocytes,reduce the apoptosis rate and reactive oxygen level,and increase the expression of podocyte marker protein.In addition,RPF can increase the expression of PTEN,inhibit the activation of PI3K/AKT/mTOR signaling pathway,improve podocyte autophagy,and alleviate podocyte damage.This indicates that RPF protects podocytes by reducing reactive oxygen species,enhancing autophagy,and inhibiting the activation of PI3K/AKT/mTOR signaling pathway.(4)Illumina MiSeq high-throughput sequencing technology was used to analyze the effect of RPF on the structure and composition of intestinal micro flora in DN mice.The results showed that the intestinal microbial diversity of DN mice decreased compared with normal mouse,and the abundance of the bacteria changed.For example,the abundance of Lachnospiraceae increased significantly,and the abundance of Prevotellaceae,Ruminococcaceae,Erysipelotrichaceae decreased significantly.After the treatment with RPF,the diversity of intestinal flora and the abundance of specific bacteria were restored to a certain extent.In addition,the renal function index of DN mice are closely related to intestinal flora.For example,Helicobacteraceae,unclassified f Lachnospiraceae,Alisipes are positively correlated with renal function indexes.Prevotellaceae_Ga6Al_group are negatively correlated with renal function indexes.RPF can delay the DN by regulating the abundance of these bacteria in the intestine of DN mice,improve the renal function indexIn conclusion,we investigated the changes of chemical composition of radix astragli before and after fermentation,explored the effect of RPF on DN mice and the mechanism of podocyte action through pharmacological experiments in vitro and in vivo,analyzed the effect of RPF on intestinal bacteria in DN mice by high throughput sequencing,which laid a foundation for the development of anti DN drugs. |
PDF Full Text Request