Effects Of Shenshao Oral Liquid On Apoptosis Induced By PPERK Pathway In Diabetic Nephropathy

Objectives Through observe the changes of biochemical indexes and morphological changes in each group rats as well as the expression changes of endoplasmic reticulum stress-related factors GRP78,PERK,e IF2?,and CHOP in renal tissues and renal cell apoptosis,to investigate the effect of Shenshao oral liquid(SS)on ERS-mediated apoptosis in rats with diabetic nephropathy and the possible mechanisms of protective effects on diabetic nephropathy.Methods 50 healthy male SPF Wistar rats were fed adaptively in the experimental animal house for 7 days.The rats were randomly divided into normal control group(NC),model group(DN),and low dose Shenshao Oral liquid group(LSS),medium dose Shenshao oral liquid group(MSS)and high dose Shenshao oral liquid group(HSS)were used in 10 rats in each group.The Sham group was fed with common diet,and the DN group and SS different dose groups were fed with high-fat diet and fed for one month.After DN group and SS different dosage groups were fasted for 12 hours,a single dose of 25 mg/kg Streptozotocin(STZ)was intraperitoneally injected to prepare DN models,which were monitored 3 times on days 3,7,and 14 respectively.Blood glucose(GLU)?16.7 mmol/L,urine collection of urine glucose was 3 + to 4 +(strong positive),24 h urinary protein(24h UA)> 30 mg/24 h,successfully constructed for DN rat model.After successful modeling,drug intervention was performed.The LSS group,MSS group,and HSS group were given intragastric administration of Shenqi Oral Liquid 4ml/kg.d,8ml/kg.d,and ml/kg.d.The NC group and DN group were given1 ml distilled water gavage once a day.After 12 weeks of drug gavage intervention,24 hours urine was collected from the rats and blood was collected from the abdominal aorta.Blood glucose,blood urea nitrogen(BUN),creatinine(Scr)and 24-hour urinary protein(24h UA)were measured in each group.HE staining and Masson staining were performed:The histopathological changes and renal fibrosis were observed under light microscope.The expression of ERS-related factors GRP78 and CHOP were detected by immunohistochemical staining.The expression ERSrelated factors GRP78,PERK,e IF2?,CHOP were detected by Western blot.Td T-mediated d UTP Nick-End Labeling(TUNEL)was used to detect apoptosis in rat renal tissue.Results 1 General condition: NC group rats had normal blood glucose levels,demeanor,normal intake of water,and sensitive response;compared with NC group,blood glucose level of DN group was significantly higher(P <0.05),and the coat was dull and dark.Partially detached accompanied by shedding,eating and drinking increased significantly,increased urination,mental fatigue;SS different dose groups and DN group compared rats eating water and mental and activity status improved significantly,blood glucose levels decreased(P <0.05).2 Detection of renal function: BUN,Scr and 24 h UA in DN group were all higher than those in NC group(P <0.05).BUN,Scr and 24 h UA in different SS groups were lower than those in DN group(P <0.05).3 HE observation of renal pathological conditions: compared with the NC group,the glomerular volume was significantly enlarged in the DN group,the narrowing of the tubular capillary lumen was narrowed,the glomerular mesangial area was widened,and the mesangial matrix was significantly increased.The tubular basement membrane became thicker,part of the glomerulus was reduced in size and hardened,and some renal tubular epithelial cells became hypertrophic and vacuolar degeneration was observed in the cytoplasm.The above-mentioned lesions were significantly reduced in the SS different dose groups compared with the DN group.4 Masson observed the renal tissue fibrosis: compared with NC group,there was a large amount of collagen in the glomeruli and tubulointerstitial tissue in DN group.The blue collagen fibers were arranged in disorder and connected to each other to form a reticular structure.Compared with the DN group,the SS of different dose groups showed significant improvement in renal fibrosis.5 Immunohistochemistry results: Compared with NC group,the expression of GRP78 and CHOP protein in kidney tissue of DN group was significantly increased(P <0.05).Compared with DN group,the expression of GRP78 and CHOP protein in kidney tissue of different dose SS groups was significantly reduced(P <0.05),of which the HSS group showed the most significant reduction in expression.6 Westernblot results:Compared with NC group,the expression of GRP78,PERK,e IF2? and CHOP protein in kidney tissue of DN group was significantly higher(P <0.05).Compared with DN group,The expressions of GRP78,PERK,e IF2?,CHOP in kidney tissue of different dose SS groups were significantly decreased(P <0.05),of which the HSS group showed the most significant reduction in expression.7 TUNEL results: A small amount of apoptosis was observed in NC group.Compared with NC group,a large amount of apoptosis was observed in kidney tissue of DN group.Compared with DN group,apoptosis was significantly decreased in SS group,and the situation of apoptosis was found in HSS group is closer to the NC group.Conclusions 1 The high expression of endoplasmic reticulum stress-related factors GRP78,PERK,e IF2?,and CHOP in kidney tissue of rats with diabetic nephropathy and a large number of renal cell apoptosis,indicating that the presence ofendoplasmic reticulum stress activation and the occurrence of apoptosis induced by the PERK-e IF2?-CHOP signaling pathwayin the DN kidney group.2 Shenshao oral liquid can reduce the pathological damage of renal tissue in DN rats,delay the decline of renal function,and have a protective effect on the kidneys,the mechanism may be to decrease the expression of ERS-related factors GRP78,PERK,e IF2? and to decrease renal cells apoptosis.