mRNA expression of insulin receptor and leptin receptor in liver of nonalcoholic fatty liver disease from diabetic rats

AIM: To observe the pathologic changes of liver in diabetic rats and to investigate the role of mRNA expression of insulin receptor and leptin receptor in the pathogenesis of nonalcoholic fatty liver disease (NAFLD). METHODS: Twenty male Sprague-Dawley rats were divided randomly into two groups: normal control group and diabetic group. After fed with high-fat diet for 4 weeks, diabetic rats were injected with streptozotocin at a dosage of 30 mg/kg intraperitoneally to induce NAFLD model of type 2 diabetes mellitus. Then the diabetic animals were fed with high-fat diet continuously for 12 weeks. At the end of the experiment, the rats were sacrificed, the concentrations of blood glucose, serum lipid, ALT and AST were measured biochemically. The levels of serum leptin and serum insulin were detected by enzyme-linked immunosorbent assay (ELISA) and radio immunoassay (RIA), respectively. The pathologic changes of liver were observed under light microscopy (LM) stained with HE, Sudan Ⅲ and Masson trichrome staining, respectively. The ultra-structural changes of liver were observed under transmission electron microscopy (TEM). Additionally, the mRNA expressions of PEPCK, G6Pase, insulin R and leptin R from rat livers were assayed by semi-quantitative RT-PCR. RESULTS: The levels of blood glucose, serum insulin, serum TG, ALT and AST increased significantly (P<0.01), serum TC elevated (P<0.05), and the levels of serum leptin decreased (P<0.01) in diabetic group compared to those in normal control group. Obvious liver fatty degeneration, piecemeal necrosis with accompanying inflammatory infiltration and fibrosis were found under LM. Hepatocytes pyknosis, lots of lipid deposits in cytoplasm of hepatocytes, proliferation of collagen in space of Disse were observed under TEM in diabetic group. The expression of insulin R and leptin R mRNA in liver from diabetic rats increased significantly (P<0.01) while the expression of PEPCK and G6Pase mRNA remained unchanged. CONCLUSION: Insulin resistance plays an important role in the pathogenesis of NAFLD. Low level of serum leptin, up-regulation of mRNA expression of insulin R and leptin R in liver caused by insulin resistance may be involved in this process.     

Effects of puerarin combined with saxagliptin on renal fibrosis in type 2 diabetic rats

AIM：To observe the effects of puerarin combined with saxagliptin on renal fibrosis in type 2 diabetic rats. METHODS：Fifty male Wistar rats were used, of which 8 rats were randomly chosen as normal control group, and the remaining rats were used to establish the type 2 diabetic model. The rats that met the criterion for the diabetic mo-del were randomly divided into model group, puerarin treatment group, saxagliptin treatment group, puerarin combined with saxagliptin treatment group and metformin combined with saxagliptin treatment group. The above-mentioned drugs were administered for 8 weeks. After that period, all rats were sacrificed. The kidney index (kidney weight/body weight),and blood glucose and HbA1c were examined in all the rats. The morphological changes were observed by HE and Masson staining. The levels of TNF-α and macrophage migration inhibitory factor (MIF) in the serum were measured by ELISA. The mRNA expression of TNF-α, MIF and CD68 was examined by RT-PCR. RESULTS：Compared with normal group, the kidney index, blood glucose and HbA1c, the levels of TNF-α and MIF in the serum and the mRNA expression of TNF-α, MIF and CD68 were increased (P<0.05) in the kidney tissues of model group. Compared with model group, the kidney index, blood glucose and HbA1c, the levels of MIF and TNF-α in the serum and the mRNA expression of TNF-α, MIF and CD68 were decreased (P<0.05) in puerarin combined with saxagliptin treatment group. CONCLUSION：Puerarin combined with saxagliptin reduces blood glucose, decreases MIF and TNF-α, and down-regulates the mRNA expression of TNF-α, MIF and CD68 in the kidney tissues of type 2 diabetic rats, which may contribute to the inhibition of renal fibrosis.     

PPAR-α involves in cardiomyocyte hypertrophy induced by high glucose and insulin

AIM: To study the role of peroxisome proliferator-activated receptor-α (PPAR-α) signal transduction pathway in cardiac hypertrophy induced by high glucose and insulin (HGI). METHODS: The cultured neonatal rat cardiomyocytes were used to observe the effect of fenofibrate (FF), a selective PPAR-α agonist, on cardiomyocyte hypertrophy induced by HGI (glucose at concentration of 25.5 mmol/L and insulin at 0.1 μmol/L). The cardiomyocyte hypertrophic responses were assayed by measuring the cell surface area, protein content, and mRNA expression of atrial natriuretic factor (ANF). The expressions of mRNA and protein were assayed by real -time PCR and Western blotting. RESULTS: In cultured cardiomyocytes, HGI induced profound change of hypertrophic morphology, the significant increase in cell surface area, protein content and ANF mRNA expression compared to those in vehicle control (P<0.01), but the expressions of PPAR-α mRNA and protein decreased significantly (P<0.05). At the same time, the expression of cyclooxygenase 2 (COX-2), one of the PPAR-α downstream effectors was obviously elevated (P<0.05). However, FF (0.1, 0.3 and 1 μmol/L) inhibited the cardiomyocyte hypertrophy induced by HGI in a concentration-dependent manner (P<0.01). FF at concentration of 0.3 μmol/L increased the expressions of PPAR-α in both mRNA and protein levels (P<0.05) and inhibited the expressions of COX-2 (P<0.05), which were abolished by MK 886 (0.3 μmol/L), a selective PPAR-α antagonist (P<0.05). CONCLUSION: PPAR-α signal transduction pathway and its downstream effector COX-2 might involve in the cardiomyocyte hypertrophy induced by HGI.     

Expression of OPN and macrophage colony stimulating factor in diabetic rats and intervention of mycophenolate mofetil

AIM: To explore the effect of mycophenolate mofetil (MMF) on expression of osteopotin (OPN) and macrophage colony stimulating factor (M-CSF) in diabetic rats with renal tubulo-interstitial injury. METHODS: Diabetes was induced in uninephrectomized male Wistar rats by peritoneal injection of streptozotocin (65 mg/kg). The rats were randomly divided into three groups: control group (NC), diabetic group (DM) and MMF treated group ［DM+MMF, treatment of MMF (15 mg·kg^-1·d^-1) by gavage from the next day of the induction for 8 weeks］. Serum biochemistry, 24 h urinary protein and the ratio of left kidney weight/body weight were determined after 8 weeks. The renal tubulo-interstitial morphological change was observed, immunohistochemical method was used to analyze the expression of OPN, M-CSF and CD68. The mRNA of OPN in renal tissue was amplified by quantitative real-time PCR. RESULTS: Compared with control group, serum glucose level, 24 h urinary protein and the ratio of left kidney weight/body weight were significantly increased (P<0.01), and the relative area of interstitial fibrosis was also significantly enlarged in DM group (P<0.01).Compared with NC group, the expressions of OPN, M-CSF, CD68 protein and OPN mRNA were significantly upregulated in DM group (P<0.01). After intervention with MMF, the upregulations of the above-mentioned parameters, except blood glucose and serum creatinine, were all significantly inhibited (P<0.05 or P<0.01). CONCLUSION: The expressions of OPN, M-CSF and CD68 in renal tubulointerstitial decrease in diabetic rats treated with MMF. MMF also inhibits the level of OPN mRNA, reduces proteinuria and prevents renal injury. MMF plays an apparently protective role in renal tubulointerstitial injury, probably associated with inhibiting chemokine and proliferation on macrophages.     

Effect of EGB on pituitary-testicular axis and expressions of LHR and StAR in type II diabetic rats

AIM: To observe the effect of the extract of Ginkgo biloba(EGB) on pituitary-testicular axis and the mRNA expressions of luteinizing hormone receptor (LHR) and steroidogenic acute regulatory protein (StAR). METHODS: Thirty male Sprague-Dauley rats were divided into three groups randomly: normal control group, type II diabetic group and EGB treatment group. After fed with high-fat diet for 4 weeks, the later two groups were injected with strepozotocin intraperitoneally to induce type II diabetes mellitus. The EGB treatment group was given EGB at the dose of 50 mg/kg once a day for 12 weeks by intragastric administration. The normal control and diabetic group were given normal saline of equal volume per day for 12 weeks. The indices of blood glucose, insulin and low-density lipoprotein-cholesterol (LDL-c) were measured. The morphologic change of testicular tissue was observed under light microscopy (LM) and transmission electron microscopy (TEM) respectively. The concentrations of blood luteinizing hormone (LH) and testosterone (T) were assayed by the technique of enzyme linked immunosorbent assay (ELISA). The mRNA expressions of LHR and StAR from Leydig cells were detected by RT-PCR. RESULTS: The concentrations of blood glucose, insulin and LDL-c increased obviously, and the testis weights lessened obviously in type II diabetic groups compared to those in normal control groups. Rare spermatogenic cells of seminiferous tubule and germinal arrest were observed in diabetic group under LM. Ultrastructural analysis of testicular tissue by TEM showed dilation of the endoplasmic reticulum and mitochondrial swelling in Leydig cell and sertoli cell in diabetic group. The level of blood LH and T decreased in type II diabetic groups in comparison with that in the normal control group. Compared to normal groups, the mRNA expression of StAR in type II diabetic groups decreased, while the mRNA expression of LHR increased. After the treatment of EGB, the pathological change of testis was relieved, the concentrations of blood glucose, insulin and LDL-c were decreased, the level of blood LH and T, and the mRNA expression of StAR were increased, and the mRNA expression of LHR descended compared to type II diabetic groups. CONCLUSION: EGB may increase the LH-induced testosterone production by correcting metabolic disorder of glucose and lipid, improving the function of pituitary-testicular axis and regulating the expression of LHR and StAR mRNA.     

Effect of advanced glycation end products on inflammation in cultured cardiomyocytes

AIM: To investigate the effect of advanced glycation end products on inflammation in cultured cardiomyocytes.METHODS: Primary cardiomyocytes were isolated from Sprague-Dawley neonatal (1 to 2 days old) rats ventricles.The insulin resistant cardiomyocyte model was established.Neonatal rat ventricular myocytes were exposed to AGEs for 24 hours.TNF-α mRNA and PPAR-γ mRNA expressions were determined by RT-PCR.Activation of NF-κB in the cells was examined by using immunocytochemistry.The ultrastructure of the cells was detected by transmission electron microscope.RESULTS: The exprssion of TNF-α mRNA and the activation of NF-κB increased,the expression of PPAR-γ mRNA decreased compared with control group (P<0.05).The differences among different AGE-BSA groups were significant (P<0.05).The numbers of chondriosome and smooth endoplasmic reticulum increased.CONCLUSION: AGEs significantly increase TNF-α mRNA expression and NF-κB activation,and restrain the expression of PPAR-γ mRNA.These data suggest that AGEs play an important role in the onset of diabetic cardiomyopathy.     

Effects of Astragalus injection combined with puerarin injection on expression of BMP-7 and TGF-β1 in type 2 diabetic KKAy mouse kidney

AIM: To observe the effects of Astragalus injection combined with puerarin injection on the expression of transforming growth factor beta 1 (TGF-β₁) and bone morphogenetic protein 7 (BMP-7) in the kidney of type 2 diabetic KKA^y mouse. METHODS: The male KKA^y mice of 14 weeks old were randomly divided into model group and Astragalus injection combined with puerarin injection treatment (Astragalus+puerarin) group. The age-matched male C57BL/6J mice were selected as normal group. The general conditions and body weight of the mice were observed. Blood glucose (BG), triglyceride (TG), cholesterol (TC) and serum creatinine (SCr) were examined at the 20th, 24th and 28th week. The protein expression of renal TGF-β₁ was determined by immunohistochemical method. The mRNA expression of BMP-7 and TGF-β₁ was detected by RT-PCR. RESULTS: Compared with normal group, the body weight, BG, TG, TC and SCr increased significantly in model group. TGF-β₁ expression at protein and mRNA levels was increased, while mRNA expression of BMP-7 was decreased in KKA^y mice. Compared with model group, the body weight, BG, TG, TC and SCr reduced in Astragalus+puerarin group. The mRNA expression of BMP-7 in the renal tissues was higher, and TGF-β₁ expression at mRNA and protein levels was significantly lower in Astragalus+puerarin group than those in model group. CONCLUSION: Astragalus injection combined with puerarin injection has renal protective effects on type 2 diabetic KKA^y mice. The mechanism may be related to restoring BMP-7 expression and reducing the overexpression of TGF-β₁ in renal tissues.     

Expression of cathepsin B and cystatin C and its potential role in diabetic rats

AIM: To observe the expressions of cathepsin B (CB) and cystatin C (CC) in different stage of diabetic rats and to investigates their potential roles.METHODS: Sixty rats were divided into diabetes mellitus group induced by intravenous injection of streptozotocin (55 mg/kg) and normal group injected with citrate buffer. Ten rats were sacrificed respectively at the end of fourth week, eighth week and sixteenth week in both groups. 24 h urine excretion was collected in rats before sacrifice. The blood and the kidney were also collected. The mRNA and protein expressions of CB and CC in kidney were detected by real time PCR and immunohistochemical staining, respectively.RESULTS: At the end of eighth week, the expression of Ccr, 24 h urinary protein excretion, CB, CC in diabetic rats increased significantly, compared to the results at the fourth week (P<0.01 or P<0.05). With the aggravation of diabetic nephropathy, the expressions of CC, colⅣ, FN and 24 h urinary protein excretion were up-regulated significantly (P<0.01 or P<0.05). The expression of CB in diabetic rats was up-regulated at eighth week significantly (P<0.01), whereas, at the end of sixteenth week it was down-regulated significantly (P<0.01). The 24 h urinary protein excretion, the expressions of colⅣ at protein and mRNA levels and FN were negatively correlated with CB (P<0.01).CONCLUSION: The unbalance of CB and CC exists in diabetic nephropathy renal tissue, which is likely to lead to the accumulation of extracellular matrix.     

Down-regulation of PPAR-γ expression by p300-mediated histone acetylation induces glucose-lipid metabolism disorder in cardiomyocytes of GDM offspring mice

AIM: To investigate the effect of gestational diabetes mellitus (GDM) on glucose-lipid metabolism in the offspring mice and the underlying mechanisms. METHODS: Wild-type female mice were intraperitoneally injected with streptozotocin at 30 mg/kg in the second trimester of pregnancy to establish GDM model. Normal saline was used as control. F1 offspring mice were fed for 8 weeks after birth. The blood glucose and lipid levels were detected randomly. The mRNA levels of p300 and p300/CBP-associated factor (PCAF) were detected by qPCR. The expression of peroxisome proliferator-activated receptor-γ (PPAR-γ), glucose transporter typer 4 (GLUT-4) and medium-chain acyl-CoA dehydroge-nase (MCAD) at mRNA and protein levels was determined by qPCR and Western blot. ChIP-qPCR was employed to analyze the binding status of p300 with the promoter of PPAR-γ and the acetylation level of histone H3 in the promoter region of PPAR-γ. RESULTS: Blood glucose and total cholesterol levels were significant increased in the offspring mice (P<0.05). The expression levels of p300, PPAR-γ, GLUT-4 and MCAD were decreased compared with the control group (P<0.05). Binding affinity of p300 with the promoter of PPAR-γ was reduced (P<0.05). The level of acetylated histone H3 in the promoter region of PPAR-γ was decreased significantly (P<0.05). CONCLUSION: Regulation of PPAR-γ expression by p300 may induce glucose-lipid metabolism disorder in the cardiomyocytes of GDM offspring mice.     

Effect of puerarin on myocardial injury in KKAy diabetic mice

AIM:To observe the effect of puerarin on myocardial injury according to the time of occurrence of myocardial injury in the development of type 2 diabetic mice. METHODS: The serum levels of glucose (GLU), triglyceride (TG), total cholesterin (TC), low density lipoprotein-cholesterol (LDL-C) and high density lipoprotein-cholesterol (HDL-C) in 17-week, 20-week, 24-week, 28-week KKAy mice were detected by automatic biochemical methods. The apoptotic percentage of cardiomyocytes was examined by flow cytometry. The expressions of bax and bcl-2 mRNA in cardiomyocytes were detected by RT-PCR. Caspase-3 expression in cardiomyocytes was determined by immunohistochemical staining. RESULTS: Compared to normal control mice, not only GLU level increased, but also the levels of TG, TC, LDL-C, HDL-C in 20-week, 24-week and 28-week KKAy mice increased apparently (P<0.01). Apoptosis of cardiomyocytes in KKAy mice began to show up at 28-week. The expression of bax mRNA increased and expression of bcl-2 mRNA reduced. At the same time, the expression of caspase-3 increased. Puerarin obviously decreased the apoptotic percentage of cardiomyocytes, reduced the expression level of bax mRNA, improved the expression level of bcl-2 mRNA (P<0.01), inhibited the expression of caspase-3 (P<0.01). CONCLUSION:Myocardial injury exits in type 2 diabetic mice. Mitochondria apoptotic pathway might participate in the hurting course.     

Protective effect of curcumin on myocardium in diabetic rats

AIM: To study the effects of curcumin (Cur) on diabetic cardiomyopathy (DCM) in rats. METHODS: Male Wistar rats (n=75) were divided into control group and diabetes model group, in which the rats were fed with high-fat diet and then intraperitoneally injected with streptozotocin (STZ, 40 mg/kg). Fasting blood glucose was measured 72 h and 1 week after STZ injection. The diabetic rats were diagnosed when sustained fasting blood glucose levels ≥ 11.6 mmol/L. The diabetic rats were randomly divided into DCM group, DCM+Cur 100 mg/kg group and DCM+Cur 200 mg/kg group. After treatment for 16 weeks, glutathione peroxidase (GSH-Px) activity and malondialdehyde (MDA) level were measured, and the level of cardiac troponin I (cTnI) in the serum was determined by enzyme-linked immunosorbent assay. The protein expression of protein kinase C (PKC) was detected by Western blotting. RESULTS: Curcumin significantly decreased the blood glucose level, increased the body weight, inhibited MDA content and up-regulated the GSH-Px activity in the diabetic rats. Furthermore, curcumin treatment inhibited the diabetes-induced protein expression of PKC. CONCLUSION: Curcumin may have a protective effect on diabetic cardiomyopathy by attenuating oxidative stress.     

Dietary soy isoflavones decrease methylglyoxal formation and prevent the diabetic cataracts in streptozotocin-induced severe diabetic rats

AIM: Methylglyoxal (MG) is considered as a very active free radical, which is an important contributor to diabetic complications such as cataracts. The purpose of this study was to investigate the serum MG levels and protective effects of soybean isoflavones on streptozotocin induced diabetes. METHODS: Diabetes was induced in male Sprague-Dawley rats by intraperitoneal injection of 100 mg/kg streptozotocin (STZ). Diabetic rats were then randomly divided into 3 groups and received a special diet supplemented with casein (diabetes), low-isoflavone soy protein (diabetes+LIS), or high-isoflavone soy protein (diabetes+HIS) for 8 weeks, respectively. RESULTS: Compared to the diabetes or diabetes+LIS groups, diabetes+HIS diet significantly increased serum insulin levels, and reduced serum glucose, HbA1c and methylglyoxal levels (P<0.05 or P<0.01). Serum GSH levels were also increased in diabetes+HIS-fed rats as compared to the diabetes or diabetes+LIS rats (P<0.01). Significant insulin production by the β-cells of the islets was observed in diabetic rats treated with diabetes+HIS protein as compared to that in diabetes group or diabetes+LIS group. More importantly, the incidence of cataracts in the diabetic rats was markedly decreased in diabetes+HIS group. No difference of above parameters between the rats in diabetes+LIS group and diabetes group was observed. CONCLUSION: Injection of high isoflavones soy protein not only lowers the glucose levels but also reduce the incidence of cataracts in diabetic rats. The underlying mechanism of soy isoflavones are attributed to the increase in insulin secretion, the decrease in MG formation, and the antioxidant effect.     

Expression and significance of thrombospondin-1 in myocardium of type 2 diabetic cardiomyopathy rats

AIM: To investigate the expression and significance of thrombospondin-1 (TSP-1) in left ventricular myocardium of type 2 diabetic cardiomyopathy (DCM).METHODS: The rat model of DCM was established by eating a high-fat diet together with injection of low dose streptozocin (30 mg/kg) intrapertoneally.After 12 weeks,the content of collagen was quantified by Masson staining.The mRNA level of TSP-1 was determined by quantification real-time RT-PCR,while the protein level of TSP-1 was analyzed by Western blotting and immunohistochemistry.RESULTS: Compared with the control group,the content of collagen in the DCM group was increased greatly (11.01±3.05 vs 16.92±3.18,P<0.01).The mRNA and protein expressions of TSP-1 were significantly higher than those in control group (0.0089±0.0034 vs 0.0141±0.0037,P<0.05；96.38±16.80 vs 129.98±16.96,P<0.05).In DCM group,the mRNA and protein expressions of TSP-1 showed significantly positive correlations with the levels of fasting blood glucose and collagen (r=0.762,P<0.01; r=0.717,P<0.05; r=0.735,P<0.01; r=0.750,P<0.01).There was a significantly positive correlation of TSP-1 mRNA level with LVEDP (r=0.658,P<0.05).In contrast,there was a significantly negative correlation of TSP-1 protein with LVSP and -dp/dtmax (r=-0.605,P<0.05; r=-0.694,P<0.05).There was a significantly positive correlation of TSP-1 protein with LVEDP (r=0.716,P<0.05).There was a significantly negative correlation of TSP-1 protein with LVSP and -dp/dtmax (r=-0.633,P<0.05; r=-0.669,P<0.05).CONCLUSION: The increased expression of TSP-1 may play an important role in the development of myocardial interstitial fibrosis in DCM.     

Effect of interlukin-22 antibody on diabetic nephropathy via inhibition on Snail1 expression

AIM:To investigate the effect of interlukin-22 (IL-22) on diabetic nephropathy (DN) and its possible mechanism. METHODS:C57BL/6 mice were randomized to normal control (NC) group,DN group, DN+recombinant IL-22 (rIL-22) group and DN+IL-22 antibody (anti-IL-22) group. After successful establishment of diabetes model for 8 weeks, the mice in DN+rIL-22 group and DN+anti-IL-22 group were intraperitoneally injected with rIL-22 (200 μg/kg) and anti-IL-22 (200 μg/kg), respectively, and the mice in NC group and DN group were intraperitoneally injected with 0.1% bovine serum albumin, twice a week for 4 weeks. After the intervention, blood glucose, kidney function, 24 h urine microalbumin (m-Alb) and 24 h urine creatinine (UCr) were measured. The pathological changes of renal tissues were observed under light microscope. The mRNA expression of Snail1 was detected by qPCR. The protein levels of fibronetin (FN) and E-cadherin were determined by Western blot. RESULTS:After the intervention, the ratio of 24 h m-Alb/UCr increased significantly in other model groups compared with NC group (P<0.05). The levels of 24 h m-Alb and 24 h UCr increased significantly in DN+rIL-22 group compared with DN group (P<0.05). However, in DN+anti-IL-22 group, the levels of 24 h m-Alb, 24 h UCr and 24 h m-Alb/UCr ratio were significantly lower than those in DN group and DN+rIL-22 group (P<0.05). The tubular epithelial cell vacuolar degeneration, protein cast formation and glomerular mesangial expansion in the renal tissues from diabetic mice were observed under light microscope. The lesions were more severe in DN+rIL-22 group, but attenuated in DN+anti-IL-22 group. The mRNA expression of Snail1 increased significantly in diabetic mice (P<0.05), but decreased significantly after a 4-week intervention by anti-IL-22 (P<0.05). The expression of FN, an extracellular matrix protein, increased significantly in DN+rIL-22 group (P<0.05). The expression of E-cadherin, an epithelial-mesenchymal transition marker, decreased significantly in DN+rIL-22 group as well (P<0.05). CONCLUSION:IL-22 neutralizing antibody may attenuate microalbuminuria and delay the progression of DN via inhibition of Snail1 expression in the renal tubular epithelial cells.     

Effects of pyrrolidine dithiocarbamate on synthesis of hepatic glycogen in diabetic rats

AIM: To determine the effect of pyrrolidine dithiocarbamate on hepatic glycogen synthesis and its mechanism in diabetic rats. METHODS: Male Wistar rats were randomly divided into normal diet group and high-fat diet group. After 8 weeks of feeding, the rats in high-fat diet group were injected intraperitoneally with a single dose of streptozotocin (27 mg/kg) to induce type 2 diabetes. The diabetes rats were randomly divided into 3 groups: diabetes mellitus group (DM), PDTC-treated group (DM+PDTC) and insulin-treated group (DM+INS). The rats in PDTC-treated group were injected with PDTC (50 mg/kg) intraperitoneally daily. At the same time, the rats in normal diet group, DM group and insulin-treated group were injected with equivalent volume of saline in the same way. The rats in insulin-treated group were injected with insulin (1 U/kg) 1 h before killed. After the treatment was taken for 1 week, the levels of blood glucose were measured, then the animals in all groups were killed. The liver glycogen content was detected, and the levels of GSK-3β and Akt phosphorylation in the liver tissues were analyzed by Western blotting. RESULTS: The blood glucose level and liver glycogen content were significantly higher, and the levels of GSK-3β and Akt phosphorylation were lower in DM group than those in normal-diet group (P<0.01). Compared with DM group, the glycogen content, the phosphorylation of Akt and GSK-3β in the liver tissues in DM+PDTC group and DM+INS group increased significantly (P<0.01), and the blood glucose levels decreased (P<0.01). CONCLUSION: PDTC increases the synthesis of liver glycogen and decreases the level of blood glucose by regulating the activity of Akt and GSK-3β in the liver.     

Effects of astragalus injection combined with puerarin injection on process of endoplasmic reticulum stress through PERK pathway in dabe-tic nephropathy mice

AIM: To investigate the effects of astragalus injection combined with puerarin injection on endoplasmic reticulum stress through PERK pathway in diabetic nephropathy mice. METHODS: Male KKA^y mice were randomly divided into model group (injected with normal saline) and treatment group (injected with astragalus and puerarin). The male C57BL/6J mice served as normal group. The mice were sacrificed 4 weeks after treatments for observing morphological changes under electron microscope. The renal tissues were collected to determine the expression of protein kinase R-like endoplasmic reticulum kinase (PERK), eukaryotic initiation factor 2α (eIF2α) and glucose-regulated protein 78 (GRP78) at mRNA and protein levels by real-time PCR and Western blot. RESULTS: Under electron microscope, the renal tubular epithelial cells in model group and treatment group showed the swelling of the nucleus, endoplasmic reticulum and mitochondria. The results of real-time PCR and Western blot showed that the expression of PERK, eIF2α and GRP78 at mRNA and protein levels in model group was higher than that in normal group (P<0.05), while that in treatment group was lower than that in model group. CONCLUSION: Astragalus injection combined with puerarin injection reduces the mRNA and protein expression of PERK, eIF2α and GRP78, thus inhibiting the endoplasmic reticulum stress in type 2 diabetic mice to protect the kidney function.