Astragalus polysaccharides affects multidrug resistance gene 1 and P-glycoprotein 170 in adriamycin nephropathy rats via regulating microRNA-16/NF-κB axis / 中南大学学报(医学版)

OBJECTIVES@#Nephrotic syndrome is a common disease of the urinary system. The aim of this study is to explore the effect of astragalus polysaccharides (APS) on multidrug resistance gene 1 (MDR1) and P-glycoprotein 170 (P-gp170) in adriamycin nephropathy rats and the underlying mechanisms.@*METHODS@#A total of 72 male Wistar rats were divided into a control group, a model group, an APS low-dose group, an APS high-dose group, an APS+micro RNA (miR)-16 antagomir group and an APS+miR-16 antagomir control group, with 12 rats in each group. Urine protein (UP) was detected by urine analyzer, and serum cholesterol (CHOL), albumin (ALB), blood urea nitrogen (BUN), and creatinine (SCr) were detected by automatic biochemical analyzer; serum interleukin-6 (IL-6), IL-1β, tumor necrosis factor α (TNF-α) levels were detected by ELISA kit; the morphological changes of kidney tissues were observed by HE staining; the levels of miR-16 and MDR1 mRNA in kidney tissues were detected by real-time RT-PCR; the expression levels of NF-κB p65, p-NF-κB p65, and P-gp170 protein in kidney tissues were detected by Western blotting; and dual luciferase was used to verify the relationship between miR-16 and NF-κB.@*RESULTS@#The renal tissue structure of rats in the control group was normal without inflammatory cell infiltration. The renal glomeruli of rats in the model group were mildly congested, capillary stenosis or occlusion, and inflammatory cell infiltration was obvious. The rats in the low-dose and high-dose APS groups had no obvious glomerular congestion, the proliferation of mesangial cells was significantly reduced, and the inflammatory cells were reduced. Compared with the high-dose APS group and the APS+miR-16 antagomir control group, there were more severe renal tissue structure damages in the APS + miR-16 antagomir group. Compared with the control group, the levels of UP, CHOL, BUN, SCr, IL-6, IL-1β, TNF-α, and MDR1 mRNA, and the protein levels of p-NF-κB p65 and P-gp170 in the model group were significantly increased (all P<0.05); the levels of ALB and miR-16 were significantly decreased (both P<0.05). Compared with the model group, the levels of UP, CHOL, BUN, SCr, IL-6, IL-1β, TNF-α, and MDR1 mRNA, and the protein levels of pNF-κB p65 and P-gp170 in the low-dose and high-dose APS groups were significant decreased (all P<0.05); and the levels of ALB and miR-16 were significantly increased (both P<0.05). Compared with APS+miR-16 antagomir control group, the UP, CHOL, BUN, SCr, IL-6, IL-1β, and TNF-α levels, MDR1 mRNA, and the protein levels of p-NF-κB p65 and P-gp170 were significantly increased (all P<0.05). The levels of ALB and miR-16 were significantly decreased in the APS+miR-16 antagomir group compared with the APS+miR-16 antagomir control group (both P<0.05).@*CONCLUSIONS@#APS can regulate the miR-16/NF-κB signaling pathway, thereby affecting the levels of MDR1 and P-gp170, and reducing the inflammation in the kidney tissues in the adriamycin nephropathy rats.

Protective effect of leonurine on adriamycin-induced kidney injury in mice / 医学研究生学报

Objective Leonurine(LEO) has shown biological effects such as reducing fat, lowering blood viscosity, and improving microcirculation. The article aimed to investigate the protective effect of LEO on the kidney of mice with adriamycin nephropathy(AN).Methods A total of 30 male mice were randomly assigned into normal control group(n=10), model group(n=10), and LEO group(n=10). Mice in model group and LEO group were injected with 5.0mg/kg adriamycin via caudal vein to induce nephropathy. The same volume of isotonic saline was injected in control group. At 1d after modeling, mice in LEO group were given 50mg/(kg·d) LEO by gavage, and the same volume of distilled water was given by gavage in control group and model group for 6 consecutive weeks. Measurement was made on 24 hour urinary albumin, plasma triglyceride, creatinine, blood urea nitrogen (BUN), SOD, GSH, MDA. Elisa was applied in testing serum TGF-β, IL-18 and IFN-γ and western blot was used to examine the phosphorylation of SMAD2/3.Results Compared with control group, the significant exaltation in the expression of TGF-β、IL-18 and IFN-γ, urinary albumin, plasma triglyceride, creatinine, blood urea nitrogen (BUN), MDA and the phosphorylation of SMAD3 and the target protein of SMAD3 (α-SMA, fibronectin) were elevated in model group(P＜0.05) ; while the indexes were obviously decreased in LEO group than those in model group (P＜0.05).Conclusion LEO can improve the renal pathological damage induced by adriamycin, whose mechanism may be related to the inhibition of oxidative stress and inflammation.

Research progress on adriamycin nephropathy model and its pathological mechanism / 中草药

Adriamycin-induced nephritic model is one of the well-recognized models in simulating human nephropathy, whose clinical manifestations of animal models are similar to human nephrotic syndrome, and its cell models are widely used in the pathological mechanism study of nephropathy because of the well-simulated basic function of glomerular podocyte. Based on the animal models and cell models in vitro of adriamycin nephropathy, this study aims at reviewing the modelling method and pathological mechanism of adriamycin-induced nephritic model in order to provide a scientific and rational pharmacological experimental model reference for drug research and development based on kidney disease, and provide evidence for the research of the mechanism of Chinese herbal compound in the treatment of nephropathy.

Protective effects of astragaloside in rats with adriamycin nephropathy and underlying mechanism / 中国天然药物

The present study was designed to determine the mechanism underlying the treatment of nephrotic syndrome using astragaloside by observing the effects of astragaloside on the expression of nephrin and podocin proteins and genes in kidneys of rats with adriamycin nephropathy. The rats were injected with adriamycin and, after successful model establishment, randomly divided into a model group, a Methylprednisolone (MP) group, and an astragaloside group. The 24-h complete urine samples were collected. Biochemical indicators were monitored, and kidney tissues were collected for pathological analysis using light microscopy and electron microscopy. The mRNA expression of nephrin and podocin was measured in the kidney tissues using the real-time qPCR, and the protein expression levels of nephrin and podocin were detected using Western blot analysis. At the end of 12 weeks of drug intervention, the urinary protein level was lower in the MP and astragaloside groups than that in the model group (P = 0.008 and P = 0.01, respectively). Serum albumin was higher in the MP and astragaloside groups than in the model group (P < 0.001 and P = 0.012, respectively). Podocytes in the MP group were nearly normal, and fusion of podocytes in the astragaloside group was significantly less than that in the control group. The nephrin and podocin mRNA and protein expression levels in the intervention groups were higher (P < 0.05) than that in the model group. Due to the increased expression of podocyte-related nephrin and podocin proteins, astragaloside maintained slit diaphragm integrity and decreased the level of proteinuria in rats with adriamycin nephropathy.

Pathological changes of the kidneys in mouse models of adriamycin-induced-nephrosis / 中国实验动物学报

Objective Our purpose was to observe the renal pathological changes in the mouse modells of adriamy -cin-induced nephropathy in different periods .Method 48 healthy male BALB/c mice were randomly divided into control group and model group .The model group received a disposable tail vein injection of adriamycin 10.5 mg/kg body weight , and the control group received the same amount of saline .24-hour urinary protein , serum biochemical indexes and kidney pathological changes were dynamically observed for 12 weeks.Results Proteinuria of model mice appeared in the 2th week after ADR injection, which lasted to the end of the 12-week experiment, At the 8th week, the amount of urine protein reached a peak (P<0.05);The serum albumin was decreased at the 4th week, cholesterol was increased at 8th week.At the end of experiment, serum creatinine was also increased (P<0.05).Minimal change nephrotic syndrome (MCNS) was observed in model mice at the 4th week;the lesions in renal tissues at 8th weeks were more serious than that at 4th weeks, but glomerular sclerosis was unconspicuous .Focal segmental glomerulonephritis ( FSGS) was seen at the 12th week.The GSI of the model mice was(2.81 ±0.84)%, significantly higher than that of the control mice ((0.33 ±0.21)%) at 12th week(P<0.01).Conclusions A mouse model with adriamycin-induced-nephrosis can be successfully established by a disposable tail vein injection of adriamycin in a dose of 10.5 mg/kg body weight .The early manifest ation of this model is MCNS, and at a late stage , it may be changed into FSGS .

Effects of Qufengtongluo Recipe on anionic site in glomerular basement membrane in adriamycin-induced nephropathy rats / 西安交通大学学报(医学版)

Objective To investigate the expression of anionic site in adriamycin-induced nephropathy (AN) rats, and to further explore the effects of Qufengtongluo Recipe on charge barrier in glomerulus in AN rats. Methods Adriamycin nephropathy was induced by a single tail intravenous injection of adriamycin. Totally 80 rats were randomly divided into normal control group and nephropathy model group. Three weeks later, the nephropathy model was established, and 50 AN rats were randomly divided into five groups: nephropathy model group (B, n=10), Qufeng group (C, n=10), prednisone and Qufeng group (D, n=10), prednisone group (E, n=10) and benazepri group (F, n=10), and they were treated respectively. With treatment being given respectively, renal tissue samples in each group were collected at week 3 and 7, respectively. The ultrastructure and expression of anionic site were examined by electron microscope observation. Results ① After adriamycin injection, a significant increase of the 24-hour urinary protein was observed at week 3 (P<0.01). In AN rats, serum albumin was decreased (P<0.01) while serum TCH and TG were increased (P<0.01). ② In AN rats the diffuse fusion and effacement of foot processes and decrease of anionic sites in GBM were observed at week 3. ③ At week 7, the average intensity of AS dramatically increased in C and E groups (P<0.01) compared with that in nephropathy model group. Conclusion The abnormal expression of AS is the important mechanism that leads to the occurrence and development of proteinuria in AN rats. It is possible that Qufengtongluo Recipe has effects on nephrotic syndrome through altering the charge barrier in GBM in glomerulus.

Analysis of differential urinary proteome in experimental minimal change nephropathy / 基础医学与临床

Objective To screen early urine protein markers for minimal change nephropathy.Methods Adriamycin nephropathy was employed as minimal change nephropathy model.Urinary protein and ConA captured glycoproteins were respectively profiled.Results By profiling urine proteome,25 differential proteins were identified.These differential proteins were from leaked plasma proteins,secreted proteins from immuno-and inflammatory cells,specifically asecreted proteins from urinary tract,and so on.They took part in different pathogenic process,eg.hemodynamic changes,podocytes injury,immunological disorder and so on.By profiling ConA-enriched urinary glycoproteome,21 differential proteins were identified,among which 12(57%) were different from the above 25 differential proteins.This indicates that the knowledge of urine glycoproteome is complementary to urine proteome in understanding kidney condition.Conclusion These differential proteins can be potential indicators of minimal change nephropathy,and can help better understand the pathogenesis by further studying their functions.

Expression of nephrin, TGF-β1 and WT1 in adriamycin-induced-nephropathy rat model and its significance / 西安交通大学学报(医学版)

Objective To investigate podocyte number, the expression of nephrin and transforming growth factor-β1(TGF-β1) in adriamycin-induced-nephropathy rat model and its significance. Methods The rat adriamycin nephrosis model was constructed to detect blood and urine biochemical indicators and observe the pathological changes of renal tissues by light microscope and electron microscope. The expression levels of nephrin and TGF-β1 as well as the podocyte number were examined at different time points by immunohistochemistry. Results The pathological changes of the renal tissues were obvious. Nephrin presented a weak signal at the end of the first week (P<0.05). TGF-β1 started to increase (P<0.05) while the podocyte number started to decrease at the end of the eighth week (P<0.05). Expression of nephrin was negatively correlated with the P<0.05) and serum creatinine (r=-0.71, P<0.05). Expression of TGF-β1 was blood urea nitrogen (r=0.62, P<0.05) and serum creatinine (r=0.59, urinary protein (r=-0.63, P<0.05), blood urea nitrogen (r=-0.72, P<0.05) and serum creatinine (r=-0.76, P<0.05); it was positively correlated with nephrin (r=0.78, P<0.01) but negatively correlated with TGF-β1 (r=-0.64, P<0.05). Conclusion The acute and chronic adriamycin nephrosis models were twice every two weeks. The genesis and development of proteinuria are closely related to the abnormal expression of nephrin. Focal segmental glomerulosclerosis occurs when the podocyte number decreases and TGF-β1 accelerates it.

Effect of Qufengtongluo recipe on expression of nephrin mRNA in adriamycin-induced nephropathy in rats / 中南大学学报(医学版)

Objective To investigate the expression of nephrin mRNA in adriamycin-induced nephropathy (AN) in rats, and to explore the effect of Qufengtongluo recipe on proteinuria in AN rats and on the expression of nephrin mRNA. Methods Adriamycin nephropathy was induced by a single tail intravenous injection of adriamycin. We randomly divided 140 rats into a normal control group (n=32) and a nephropathy model group (n=108). Three weeks later, 90 AN rats were randomly divided into 5 groups: a model group, a qufeng group, a qufeng and prednisone group, a prednisone group, and a benazepri group (18 rats in each group). They were treated respectively, and renal tissue samples were collected at week 0, 3, 5, and 7, respectively. The distribution and expression of nephrin mRNA were examined by indirect immunofluorescence and semi-quantity RT-PCR. Results In the AN rats, the diffuse fusion and effacement of foot processes were observed at week 3. The fluorescence intensity of nephrin and the expression of nephrin mRNA significantly increased in the qufeng group and the prednisone group compared with the model group at the week 7 (P＜0.01).Conclusion Abnormal expression of nephrin is the important molecular mechanism that leads to the occurrence and development of proteinuria in AN rats. Qufengtongluo recipe has effect on nephrotic syndrome through altering the expression and distribution of nephrin in glomerulus.

Protective Effect of Sulodexide on Podocyte Injury in Adriamycin Nephropathy Rats / 华中科技大学学报（医学）（英德文版）

This study examined the effect of sulodexide on podocyte injury in rats with adriamycin nephropathy (AN). A total of 36 healthy male SD rats were randomly assigned to three groups:control group,AN group and sulodexide treatment group. Rat models of AN were established by a single tail intravenous injection of adriamycin (6.5 mg/kg) in both AN group and sulodexide treatment group. Sulodexide (10 mg/kg) was administered the rats in the treatment group once daily by garage from the fast day of model establishment until the 14th day or the 28th day. Samples of 24-h urine and renal cortex tissues were harvested at day 14,28 after the model establishment. Excretion of 24-h urinary protein was measured by Coomassie brilliant blue method. The pathological changes in renal tissues were observed by light microscopy and electron microscopy respectively. Heparanase mRNA was detected by RT-PCR. Expressions of desmin,CD2AP and heparanase were determined by immunohistological staining. The results showed that the expressions of heparanase mRNA and proteinwere increased in the glomeruli of AN rats at day 14 and 28 after the model establishment,which was accompanied by the increased expression of desmin and CD2AP. The mRNA and protein expression of heparanase was decreased in the sulodexide-treated rats as compared with AN rats at day 14 and 28. And,the protein expression of desmin and CD2AP was reduced as with heparanase in the sulodexide-treated rats. Proteinuria and podocyte foot process effacement were alleviated in the AN rats after sulodexide treatment. There was a positive correlation between the expression of heparanase and the expression of desmin and CD2AP (as well as 24-h urinary protein excretion). It was concluded that increased heparanase is involved in podocyte injury. Sulodexide can maintain and restore podocyte morphology by inhibiting the expression of heparanase in AN.

Effects of Mycophenolate Mofetil on Minimal Change Nephrotic Syndrom / 医学研究杂志

Objective To detecte the changes of nephrin in adriamycin nephropathy rats, study the effects of mycophenolate mofetil on minimal change nephrotic syndrome, investigate its possible mechanism. Methods 18 SD rats were randomly divided into three groups: control group(CG, n=6), adriamycin model group(AG, n=6), MMF treated group(TG, n=6). Rats in MG and TG were given adriamycin 7.5mg/kg through vena caudalis. At the same time, an equal volume of normal saline was given to the rats in CG by the same method. The rats in TG received MMF 20mg/(kg?d) by daily gastric gavage from the second day. Urinary protein excretion of each rat were measured at the day before adriamycin injection,and the 14th day and the 28th day after adriamycin injection. Six rats of each group were killed at the 28th day. Serum urea nitrogen, creatinine, cholesterol, triglyceride and albumin were measured at the end of the study. Immunohistochemistry and western blot were used to examine the expression of nephrin. Results The urinary protein excretion of AG at 28th day was the highest. Serum albumin decreased markedly at the 14th day(p

THE EXPRESSION OF CYCLOPHILIN mRNA IN ADRIAMYCIN NEPHROPATHY / 解放军医学杂志

We have established adriamycin(ADR) nephropathy model, the expression of cyclophilin(CyP)mRNA was measured with RT PCR, and its relationship with the levels of total protein, albumin, cholesterol, 24 hour urine protein, BUN, and creatinine were determined.After four weeks,the levels of urine protein in ADR nephropathy animals were higher than 100mg/24h, whereas the levels in the control animals were lower than 6mg/24h. It was showed that the expression of CyP gene of kidney tissue in ADR nephropathy rats was significantly higher than that of the control animals. Meanwhile, it was found that there was negative correlation was found between the expression of Cyp gene and the levels of total protein and albumin,but positive correlation between the expression of CyP gene and the levels of 24 hour urine protein and cholesterol,and the expression of CyP gene had no correlation with the levels of BUN and creatinine. Our results suggested that the high levels of Cyp mRNA expression might contribute to renel tissue damage in ADR nephropathy,and it might reflect the degree of severity and prognosis in ADR nephropathy.

Astragali radix extract ameliorates renal resistance to atrial natriuretic peptide in rats with experimental nephrotic syndrome / 中国病理生理杂志

AIM:To study the effects of astragali radix extract(ARE)on renal resistance to atrial natriuretic peptide(ANP)in rats with experimental nephrotic syndrome.METHODS:Male Sprague-Dawley rats were randomly divided into normal control,adriamycin nephropathy(ADR),ADR treated with ARE(2.5 g? kg-1? d-1)and ADR treated with benazepril(10 mg? kg-1? d-1).After 6 weeks,rats received intravenous infusion of 2% body weight isotonic saline.Urinary cGMP excretion(UcGMPV),plasma ANP level,renal PDE5 activity and protein expression were also detected.RESULTS:ARE increased UNaV while ACEI was not natriuretic.Nephrotic rats had a blunted natriuretic response and reduced rate of UcGMPV after volume expansion despite higher plasma ANP concentration.ARE increased UcGMPV and restored partly natriuretic response to volume expansion.The activity and protein abundance of renal PDE5 were high in nephrotic rats.ARE significantly reduced the PDE5 activity and protein expression.CONCLUSION:ARE may ameliorate the renal resistance to ANP in rats with adriamycin nephropathy by inhibiting the PDE5.

Effect of benazepril on mesangial matrix of adriamycin nephrotic rats / 中华肾脏病杂志

Objective To investigate the effect of angiotensin convening enzyme inhibitor (ACEI), benazepril, on the clinicohistological changes of normal and adriamycin nephrotic rats. Methods When proteinria was present for 2 weeks after injection of adriamycin, these rats were treated with saline、predsone、 amlodipine and benazepril. Urinary protein excretion was measured in 0, 3rd, 6th week after treatment. Mesangial cell index and mesangial matrix index were assessed in the 6th week. Moreover immunohistochenical stain for type Ⅳ collagen, fibronectin and laminin were performed before and after treatment, and RNA transcription of laminin was evaluated with slit hybridization. Results Proteinuria of benazepril-treated rats was significantly reduced after 6-week treatment compared with those before treatment[ (44 .7 ? 6.0)mg／２４h vs(56.8 ?23 .4)mg/24h. The proliferation of mesangial cells and the synthesis of mesangial matrix were inhibited in the benazepril-treated rats. Similar results were achieved in the predisone-treated rats, but not in amlodipine-treated rats. Conclusion Benazepril can sigificantly reduce the synthesis of mesangial matrix in adriamycin nephrosis rats by inhibiting RNA transcription.

Effect of different levels of the dietary protein on long term prognosis of experimental renal damage

We examined the effect of various levels of dietary protein on long term prognosis of Adriamycinnephropathy of S-D rat, fed with high protein (30%), intermediately low (10%), and strictly low (5%) protein diet for 15 weeks. 1) In rats fed with strictly low protein diets (5%), proteinuria and serum creatinine decreased and creatinine clearance and histological changes were relatively well preserved. But hypoproteinemia and weight loss were more marked and 2 rats died due to severe ascites and pleural effusion in cachexic state. 2) In rats fed with high protein diets (30%), general health condition and weight gain were relatively well preserved. But there were massive proteinuria, progressive increase in serum creatinine and progressive decrease in creatinine clearance. Focal glomerular sclerosis and severe tubulointerstitial change on histologic examination were marked. 3) With intermediately low protein diet (10%), renal function and pasma protein levels were relatively well preserved compared with high protein diet group. But weight gain did not increase normally. 4) We tentatively conclude that appropriately restricted dietary protein can prevent functional and histological renal damage. But too strict protein restriction aggravate nutitional state and general condition.

The expression of glomerulosclerosis by benazepril and its relationship with apoptosis of kidney cells / 实用儿科临床杂志

Objective To study the effects of angiotensin converting enzyme inhibitor benazepri1 on apoptosis and the expression of Fas and FasL in the kidney of rats with adriamycin-indued nephritic glomeruosclerosis.Methods After uninephrectomy and the injection of adriamycin induced rats model with glomerulosclerosis, benazapril(6 mg/kg) was delivered daily by gavage to the rats in therapeutic groups for 12 weeks.Apoptosis was examined by means of terminal-deoxynucleotidyl trans ferase mediated d-UTP nick end label ling(TUNEL) and immunohistochemistry was utlized to detect the expression of Fas and FasL.Software of pathological analysis quantitated the level of Fas and FasL.Results Compared with those of the control group, the kidney of model group had moresevere glomerulosclerosis, much more apoptotic cells and higher level of exprssion of Fas and FasL. The degree of glomeruloscleroais, the nuxner of apoptotic cells and the level of expression of Fas and FasL were ameliofated by benazepril treatment.Conclusion Benazepril may suppress the excessive apoptosis of kidney cell by lowering the expression of the protin correlatng apoptosis Fas and FasL,so as to postpone the process of glomeruosclerosis.

Establishment of a modified model of adriamycin nephrosis in rats / 西安交通大学学报(医学版)

Objective To study the adriamycin nephrosis model in different development pathological stages. Methods The rat adriamycin nephrosis model was established by injecting adriamycin into tail-vein twice every two weeks to detect blood and urina biochemical indicators and to observe the pathological changes of the renal tissues. Results The model showed serious edema,proteinuria,hypoproteinemia,and hyperlipidemia. Podocytes were swollen and mesangial cells developed mild hyperplasia at the end of the fourth week. The nephric tubule atrophied at the end of the eighth week accompanied with adhesion between glomeruli and Bowman's capsule. Glomeruli sclerosis of mild or medium degree was observed at the end of the twelfth week with obvious lymphocyte infiltration in the renal interstitium as well as the formation of collagen fibers. Conclusion The adriamycin nephrosis model was successfully developed by injecting adriamycin 4 mg/kg into tail-vein twice every two weeks. The acute model is similar to human minimal change disease,and the chronic model is similar to human focal segmental glomerulosclerosis.

Expression of nephrin,TGF-?1 and WT1 in adriamycin-induced-nephropathy rat model and its significance / 西安交通大学学报(医学版)

Objective To investigate podocyte number,the expression of nephrin and transforming growth factor-?1(TGF-?1) in adriamycin-induced-nephropathy rat model and its significance.Methods The rat adriamycin nephrosis model was constructed to detect blood and urine biochemical indicators and observe the pathological changes of renal tissues by light microscope and electron microscope.The expression levels of nephrin and TGF-?1 as well as the podocyte number were examined at different time points by immunohistochemistry.Results The pathological changes of the renal tissues were obvious.Nephrin presented a weak signal at the end of the first week(P