The Role Of Endoplasmic Reticulum Stress In Diabetic Nephropathy And The Intervention Mechanism Of Berberine

Objective:To investigate whether endoplasmic reticulum stress（ERS）is involved in the pathogenesis of Diabetic Nephropathy（DN）by mediating activation of ATF6、IRE1、PERK/CHOP/Caspase-12 pathway and explore how Berberine（BBR）regulates endoplasimic reticulum stress and participate in the protective mechanism of diabetic nephropathy.Methods:1.Modeling,grouping and intervention in rats:The DN RAT model was induced by high glucose and high fat diet combined with intraperitoneal injection of Streptozocin.The experiment was divided into 3 groups:normal group（NC group）,model group（DN group）and berberine intervention group（DN+BBR group）（n=6）.DN+BBR group was treated with 200 mg/(kg·d^-1)on the basis of the model of DN.NC group and DN group were treated with the same dose of sodium carboxymethyl cellulose.2.Structure and function damage of renal tissue and ERS related protein expression in rats:After 6 weeks treatment measured various indicators（include body weight,renal index（KI=kidney weight/body weight）,fasting blood glucose（FBG）,blood creatinine（SCR）,urea nitrogen（BUN）and 24h urine protein（24h Upro））.The pathological changes of kidney were observed by HE,PAS and Masson staining.The changes of glomerulus and renal interstitium were observed by transmission electron microscope.The expression of PERK,IRE1,ATF6,CHOP and Caspase-3 were detected by immunohis to chemical staining.Western Blotting was used to detected the expression of ATF6 and Caspase-12 in rat kidney tissue.Results:1.SCR,Bun,FBG and 24h Upro in DN group were significantly higher than those in NC group and the renal function of DN group severely impaired.Compare with DN group the renal function of DN+BBR group was significant improved.There were significant differences in SCR,Bun,FBG and 24h Upro in each group（P<0.01）.2.The results of HE,PAS and Masson staining showed that the glomerulus in DN group was more irregular and larger than that in NC group;the lumen of glomerulus became narrowed,diffuse mesangial matrix was increased and renal tubule was edematous.The glycogen deposits and collagen fibers in the renal interstitium were increased and inflammatory cells were infiltrated.After the berberine rescued,the glomerular condition of DN+BBR group was obviously improved;the edema of renal tubules was alleviated;the deposition of glycogen was decreased and the collagenous fibers accumulation is relatively reduced.3.The results of transmission electron microscope showed that the podocytes of DN Group were irregular and a large number of podocytes fused and broken.The basement membrane was inhomogeneous and thickened.Whereas,the morphological function of podocyte of DN+BBR Group was improved.And the basement membrane was slightly thickened.4.Immunohistochemistry shows that the expression of CHOP,PERK,IRE1,ATF6 and Caspase-3 in DN group was significantly increased and the DN+BBR group was contrarily.The difference of protein expression was statistically significant（P<0.05）.5.Western blotting showed that the expression of endoplasmic reticulum stress-related protein ATF6 and the apoptosis-related protein Caspase-12 were increased in the DN group.After berberine intervention,protein expression decreased in the DN+BBR group.The difference of protein expression was statistically significant（P<0.05）.Conclusion:Endoplasmic reticulum stress is involved in the pathogenesis of Diabetic Nephropathy（DN）by mediating activation of ATF6、IRE1、PERK/CHOP/Caspase-12 pathway.And BBR can significantly improve the renal function and alleviate renal structural damage in DN rats.The hypothesis is that BBR can suppress endoplasmic reticulum stress and reduce the apoptosis of kidney cells,thus prevent the progress of DN and protect the kidney tissue.