| Objective: To study the protective effect and mechanism of Astragalosides IV(AS-IV)on early renal damage in type 2 diabetes mellitus by regulating phosphatidylinositol 3-kinase(PI3K)/protein kinase B(Akt)/forkhead box O1(Fox 21)signaling pathway in rats.Method: After 6 weeks of high-sugar and high-fat diet for SPF SD rats,a diabetic rat model of type 2 diabetes was established by intraperitoneal injection of streptozotocin(STZ)(35 mg·kg-1).The rats were randomly divided into normal group,model group,astragaloside(20,40,80 mg·kg-1)group and metformin hydrochloride(positive)group.The changes of body mass,renal index,blood sugar,24-hour urinary protein,urinary microalbumin,urinary creatinine,serum creatinine,blood urea nitrogen,malondialdehyde(MDA)content and activity of GSH-Px were detected after 8 weeks of continuous administration.Hematoxylin and eosin(HE)staining and Masson trichrome staining were performed on renal tissue sections.The renal histopathological changes were observed by periodic acid-sliver methenamine(PASM)staining;the expression of phospho-Fox O1 protein in rat kidney tissue was detected by immunohistochemistry;the expression levels of PI3K/Akt/Fox O1 signal pathway protein and autophagic marker protein were analyzed by Western blot(WB).Result: 1.Compared with the normal group,the model group rats showed typical symptoms of polydipsia,polydipsia,polydipsia and polydipsia.After treatment,the symptoms of polydipsia and polydipsia in AS-IV(20,40,80 mg.kg-1)groups were improved,and the urine volume was reduced to some extent.2.Compared with the normal group,the weight of rats in the model group decreased significantly,the kidney index and blood sugar increased significantly(p <0.01);compared with the model group,the weight of rats in the astragaloside group(40,80 mg·kg-1)increased(p < 0.05,p< 0.01),the blood sugar in the astragaloside group(40,80 mg·kg-1)decreased significantly(p< 0.01),and the kidney index in the astragaloside group(20,40,80 mg·kg-1)increased significantly.The difference was statistically significant(p< 0.01).3.Compared with the normal group,the urinary creatinine in the model group decreased significantly,and the ratio of 24 h UAER,urinary protein creatinine and urinary microalbumin increased significantly(p< 0.01);compared with the model group,the content of urinary creatinine in the astragaloside group(40,80 mg·kg-1)increased(p<0.05,p<0.01),24 h UAER and urinary microalbumin decreased significantly(p< 0.05,p< 0.01).The urinary protein creatinine ratio also decreased(p< 0.01).4.Compared with the normal group,the serum creatinine and urea nitrogen levels in the model group were increased,with significant difference(p< 0.01);the serum creatinine and urea nitrogen levels in the astragaloside group(40,80 mg·kg-1)were decreased in varying degrees compared with the model group,and the difference was significant(p< 0.01).5.The level of MDA in model group was significantly higher than that in normal control group(p< 0.01);the level of MDA in astragaloside A group(40,80 mg·kg-1)was significantly lower than that in model group(p< 0.01).Compared with the normal group,the level of GSH-Px in the model group was significantly lower(p< 0.01);the level of GSH-Px in the astragaloside A group(40,80 mg·kg-1)was significantly higher than that in the model group(p< 0.05,p< 0.01).6.HE,Masson and PASM staining results showed that the normal group had clear and complete renal structure,regular shape,normal glomerulus,tubule and interstitial structure,no proliferation of mesangial cells and matrix,no thickening of basement membrane and no proliferation of fibrous tissue.In the model group,glomerular hypertrophy,basement membrane thickening,mesangial dilatation and mesangial fineness were observed.Cell and endothelial cell proliferation and atrophy,extracellular matrix proliferation,part of renal tubule atrophy,etc.Compared with the model group,the treatment group reversed these changes to a certain extent,and astragaloside(80 mg·kg-1)group improved the pathological changes most significantly,with the best effect.7.Immunohistochemical results showed that the protein level of phospho-Fox O1 in the model group was significantly higher than that in the normal group(p< 0.01);compared with the model group,the protein level of phospho-Fox O1 in the kidney tissue of astragaloside(40,80 mg·kg-1)group was significantly lower(p< 0.01).8.Western blot results showed that the levels of phospho-PI3K/total-PI3 K,phospho-Akt/total-Akt,phospho-Fox O1/total-Fox O1 in the model group were significantly higher than those in the normal group(p<0.01);compared with the model group,the levels of phospho-PI3K/total-PI3 K in the astragaloside group(40,80 mg·kg-1)were significantly lower(p<0.01),and the levels of phospho-Akt/total-Akt and phospho-Fox O1/total-Fox O1 were also lower(p<0.05,p<0.01).Compared with the normal group,Bnip3,Lc3II/Lc3 I and Belcin1 in the model group were significantly lower(p< 0.01);compared with the model group,Bnip3,Lc3II/Lc3 I and Belcin1 in the astragaloside A group(40,80 mg·kg-1)were significantly higher(p< 0.05,p< 0.01).Conclusion:In the model of type 2 diabetes mellitus,astragaloside may promote the expression of downstream genes and enhance the autophagic activity of renal tissue cells by regulating PI3K/Akt/Fox O1 signaling pathway,thus alleviating the early renal function damage of type 2 diabetes mellitus and delaying its development process. |
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