Protective Effect Of Grape Seed Proanthocyanidin Extract On The Endoplasmic Reticulum Stress In Kidney Of Mice And Rats

Backgroud and Objective:Cisplatin (CP) is used as an antineoplastic drug in the clinic, but its nephrotoxicity limits its use. Grape seed proanthocyanidin extract (GSPE) is a powerful antioxidant. It is shown to possess a variety of potent properties. However, whether GSPE can protect cisplatin induced acute kidney injury is not clear. In this study, GSPE was used to treat the mouse model of CP-induced nephropathy, we observed the general change, renal tubules pathological changes, cell apoptosis of mice, GRP78^P-ERK、Caspase-12protein expression, in order to examine its potential protective effect(s) and examine whether these effects are mediated by the inhibition of ER stress-induced apoptosis via Caspase-12pathway occurring in tubular cells.Methods:Sixty five male C57/BL6mice, after one week adaptation feeding, were randomly divided into four groups:control group (N, n=10), CP group (C, n=20),which received an intraperitoneal (ip) injection of20mg/kg CP, GSPE group (G, n=15), which received an intragastric (ig) administra-tion of500mg/kg GSPE, and CP+GSPE group (C+G, n=20), ig administration of500mg/kg GSPE was performed30min prior to ip injection of CP, followed by an additional ig administration of GSPE72h later. Blood and kidney samples were collected120h after treatment. The pathological changes in the kidney were examined by peri-odic acid-Schiff (PAS) staining, while the protein levels of glucose-regulated protein78(GRP78), phosphorylated-extra-cellular signal-regulated kinase (p-ERK) and Caspase-12were examined by western blotting and immunohistochemical staining. Apoptosis was examined by a terminal deoxy-nucleotidyl transferase dUTP nick-end labeling (TUNEL) assay.Results:(1) General states changes of mice in each group:The levels of BUN, Scr and RI were significantly increased in the CP group compared to the control group (P<0.05). Compared to the CP group, the levels of BUN, Scr and RI were significantly decreased in the CP+GSPE group (P<0.05). The levels of BUN, Scr and RI in the GSPE group did not show significant differences compared to the control group.(2) Pathological changes of mice in each group:In the control and GSPE groups, the kidneys maintained normal structure.While we observed brush border damage, renal tubular epithelial cell swelling, degeneration, necrosis, tubular casts and cell vacuole degeneration in the proximal tubules of kidney in the CP group, Compared to the CP group, tubular damage was greatly decreased in the CP+GSPE group.(3) TUNEL results:There was limited apoptosis was detected in the control and the GSPE group.While the CP group showed a high number of TUNEL-positive cells (P<0.05). The TUNEL-positive cells were significantly reduced in the CP+GSPE group compared to the CP group (P<0.05).(4) Western blot results:Compared to the N group,the protein expression of GRP78, p-ERK and caspase-12are highly expressed in the CP group, while their levels are significantly reduced in the CP+GSPE group (P<0.05). There are no difference between the control group and GSPE group.(5) Immunohistochemistry results:The protein expression of GRP78, p-ERK and caspase-12are highly expressed in the CP group compared to the N group, while their levels are significantly reduced in the CP+GSPE group (P<0.05). There are no difference between the control group and GSPE group. Conclusion:GSPE can protect the renal function from CP-induced nephrotoxicity and can attenuate the endoplasmic reticulum (ER) stress induced apoptosis via regulation of the caspase-12pathway. Background:In developed countries, Diabetic nephropathy(DN) is the most commoncause of endstage renal disease (ESRD)..In the United States,DN making up about45%of all ESRD cases. In China, DN is the second leading cause of ESRD, the high morbidity and mortality received more and more people’s attention. Grape seed proanthocyanidin extracts (GSPE) are powerful antioxidants,its ability of antioxidant is50times that of vitamin E,20times that of vitamin C.Studies have shown that GSPE has protective effect on the DN, and GSPE has functions of prevention and treatment of various diseases, including the protection of cisplatin-induced nephrotoxicity, which may be associated with inhibiting endoplasmic reticulum stress-induced apoptosis.We take this as a starting point, research that whether GSPE can protects from Streptozotocin-induced diabetic nephropathy by inhibiting endoplasmic reticulum stress-induced apoptosis.Methods:We randomly selected ten rats rom the fifty five male SD rats as the normal control group, the rest forty five rats were given the high fat and sugar diet for one month as the model group.Then the model group intraperitoneal injected with40mg/kg Streptozotocin, successful modeling rats were randomly divided into diabetes mellitus group(D, n=20), and GSPE treatment group(G,n=20)(ig250mg/kg/d GSPE for12weeks after rats were induced into,. the N and D group received intragastric of saline.The D and G group were continue given high fat and sugar diet. The rats were sacrificed at the16th week..Blood and kidney samples were collected after treatment.The renal pathological changes were examined with periodic acid-Schiff(PAS) staining and electron microscope, the protein expression of GRP78, p-ERK and Caspase-12were examined by Western blotting and immunohistochemical staining. Apoptosis was examined by TUNEL kit.Results:(1)General states changes of rats in each group:Compared to the N group,the D group showed significantly increased in the renal index (RI),24h urine protein.Compared with the D group, the G group had a significant decline in the level of the renal index (RI),24h urine protein.(P<0.05).While the level of blood urea nitrogen (BUN),serum creatinine (Scr) has no significant changes(table1).Compared with the D group,the G group had no change in the level of the blood glucose.(2) Pathological changes of rats in each group:The N group showed no obvious abnormality,the D group showed significantly increased glomerular volume, glomerular basement membrane thickening, mesangial widening, mesangial cell proliferation.The above histopathology change was slighter in G group than group DN (P<0.05).(3)TUNEL results:There was limited apoptosis detected in the N and the G group.While the D group showed a high number of TUNEL-positive cells (P<0.05).(4) Western blot results:Compared to the N group,the protein expression of GRP78, p-ERK and Caspase-12are highly expressed in the D group, while their levels are significantly reduced in the G group (P<0.05).(5) Immunohistochemistry results:The protein expression of GRP78, p-ERK and Caspase-12are highly expressed in the D group compared to the N group, their levels are significantly reduced in the G group compared to the D group (P<0.05).Conclusion:In this study we conclude GSPE can protect the renal function and attenuate endoplasmic reticulum stress-induced apoptosis by the Caspase-12pathway in Streptozotocin-induced nephrotoxicity.