Dihydromyricetin Prevents Against Diabetic Nephropathy Through The Activation Of AMPK Pathway

Diabetic nephropathy(DN)is a common microvascular complications of diabetes,and also one of the main causes of end-stage kidney disease.and the development of end-stage renal disease by DN is more tricky than other causes?With the continuous improvement of people's living standard,the accelerated pace of life,and changes in diet structure and lifestyle,More and more people are suffering from diabetes,and the incidence of DN accounts for about 30% of all diabetics,so this led directly to the increase in the number of DN patients,therefore,it is important to study theControl measures of DN.DN is due to Diabetes patients' abnormal glucose,protein and lipid metabolism caused by renal hemodynamic changes.The pathogenesis of DN is extremely complex,and has not been completely identified now.It is thought that it is mainly associated with metabolic disorders,hemodynamic disorders,oxidative stress injury,inflammation and genetic factors.Glomerular hypertrophy and sclerosis is the early pathological changes of DN,the main reason is the thickening of glomerular basement membrane and increased mesangial matrix precipitation.Those can gradually develop into collagen fibrosis.A large number of studies have shown evidence that the pathogenesis of DN is closely related to the inhibition of renal cell autophagy and extracellular matrix deposition.In the state of diabetes the renal cells are in the high glucose environment,this resulting in damage to the renal cell organelles and cytotoxic substances can not be timely and effective removal,then accumulated in the calls,affecting the normal function of cells to accelerate cell damage and senility,and then cause kidney damage.Adenosine 5'-monophosphate activatedprotein kinase(AMPK)as an important receptors in the cells,not only regulate the body's glucose,lipid metabolism,but also regulate the cell autophagic activity.Studys has found that the activity of AMPK in DN kidney cells is reduced.Therefor the activation of the AMPK pathway enhances autophagic activity of kidney cells,thereby reducing the kidney damage from high glucose and preventing the occurrence and development of diabetic nephropathy.Dihydromyricetin(DHM)is an active ingredient of the tTengcha flavonoids,extracted from the leaves of the vine,with anti-inflammatory,anti-oxidation,antibacterial,hpyerglycemic,regulating blood lipids,anti-tumor and other pharmacological efficacy.Recent researches show that,DHM is a natural activator of AMPK,can regulate autophagic signal pathway mediated by AMPK,suggesting that DN has a potential prevention and treatment application value.Matedals and methods: Male Sprague-Dawley rats were employed in present study,and rats model of DN were established by intraperitoneal injection of streptozotocin(STZ)60 mg/kg.Then,all rats were divided into normal group(CON),diabetic nephropathy group(DN group)and the DHM group(DHM 50 mg/kg?100 mg/kg).The experimental groups were treated with DHM(50 mg/kg or 100 mg/kg)by lavage administration for 8 weeks,while the DN group was given the equal distilled water.The pathological morphology of the kidney was observed by using H&E and Masson staining.The contents of type I collagen(COLI),type IV collagen(COL IV)and fibronectin(FN)in rat kidney were detected by ELISA.The protein levels of phosphorylated AMPK(p-AMPK),transforming growth factor-?(TGF-?1),?-smooth actin(?-SMA)and rapamycin mammalian target protein(m TOR),autophagy-associated microtubule-associated protein 1 light chain 3(LC3),and Beclin-1 in rat kidney were determined by Western Blotting analysis.To observe the effect of DHM on DN kidney and to explore whether DHM can prevent the rat kidney,and explore the role of autophagy signal pathway activated by AMPK.The purpose o f this study is providing basic theoretical support for the development and application of DHM for prevention and treatment of DN.Main results and conclusions:(1)H&E staining showed less mesangial matrix proliferation,more narrow-mesangial,as well as wider renal capsule space and reduced capsule wall adhesion in the DHM group when compared with the DN group.Masson staining displayed that the DHM group had higher degree of renal interstitial blue shallow,and less renal fibrosis.This suggested that DHM has a certain prevent effect on renal pathological damage of DN rats.(2)The result of ELISA showed less expression level of COL I,COL IV and FN in the DHM group than DN group.(P<0.05).In addition,DHM also up-regulated the protein level of AMPK,P-AMPK/AMPK,LC3?/LC3?and Beclin-1,and the 100 mg/kg group increased more than 50 mg/kg(P<0.01).It suggested that DHM can reduce the content of extracellular matrix in kidney of DN rats and reduce the deposition of extracellular matrix in a dose-dependent manner.(3)The result of Western Blotting analysis showed that DHM also up-regulated the protein level of AMPK,p-AMPK/AMPK,LC3?/LC3?and Beclin-1,and the 100 mg/kg group increased more than 50 mg/kg(P<0.01).However,DHM reduced the expression level of TGF-?1,?-SMA and mTOR in rat kidney(P<0.01),and more significant decline of TGF-?1 and m TORwas detected with the 100 mg/kg group(P<0.05).It was suggested that DHM can activate AMPK pathway in renal tissue of diabetes rats,inhibited the expression of m TOR and promote cell autophagic activity.Meanwhile,it inhibited the expression of TGF-?1 and down-regulated ?-SMA level,and has the advantages of glomerular sclerosis and renal fibrosis in diabetes rats.Conclusion:DHM can inhibite glomerular extracellular matrix deposition in diabetes rats,improving glomerular sclerosis and reducing renal fibrosis.Its mechanism may be through activating AMPK/mTOR pathway,enhancing cell autophagic activity and inhibiting expression of TGF?-1 and ?-SMA.