OBJECTIVETo investigate RANK-RANKL expression in the kidneys of a rat model of puromycin aminonucleoside nephropathy (PAN).

METHODSThirty-six SD rats were randomly divided into PAN model group and normal control group. PAN was induced by a single intravenous injection of 100 mg/kg puromycin aminonucleoside. Serum creatinine and 24-hour urinary protein were measured on days 3, 7, and 14 after the injection, and renal pathologies were assessed with optical and immune transmission electron microscopy. The expression of RANK and RANKL in the kidneys was examined using reverse transcription-polymerase chain reaction (RT-PCR) and Western blotting.

RESULTSThe PAN model rats showed massive proteinuria and elevated serum creatinine on day 3, which peaked on day 7. RANK-RANKL protein and mRNA expressions in PAN model group was higher than those in the control group. In the PAN rats, RANK was expressed mainly on the top cell membrane and in the cytoplasm of renal podocytes with a significantly increased expression level compared with that in the control group.

CONCLUSIONThe PAN rat model shows aberrant RANK and RANKL expressions in the podocytes, indicating their contribution to podocyte injury in PAN.

Animals ; Creatinine ; blood ; Female ; Kidney ; drug effects ; metabolism ; Kidney Diseases ; chemically induced ; metabolism ; pathology ; Male ; Podocytes ; drug effects ; metabolism ; Proteinuria ; pathology ; Puromycin Aminonucleoside ; adverse effects ; RANK Ligand ; metabolism ; Rats ; Rats, Sprague-Dawley ; Receptor Activator of Nuclear Factor-kappa B ; metabolism

AIMTo investigate the effect and mechanism of ascorbic acid on podocyte, last barrier of glomerular filtration, in diabetic rats.

METHODSDiabetic rats induced by streptozotocin injection intraperitoneally were treated by ascorbic acid for 5 weeks. The levels of blood glucose (BG), HbA1c, urinary albumin excretion rate (UAER) and superoxide diamutase (SOD), catalase (CAT) and malondialdehyde (MDA) in renal cortex were measured. The podocyte ultrastructure was observed while the expression of desmin protein, a marker of podocyte injury, was examined.

RESULTSCompared with control group, BG and HbA1c were increased markedly in diabetic group. The activities of SOD and CAT were decreased and the concentrations of MDA were increased significantly in diabetic renal cortex. There were the increased proteinic expression of desmin, foot process effacement in podocytes and UAER markedly in diabetic rats. Compared with diabetic rats, foot process effacement and the changes of UAER were ameliorated markedly while the activities of SOD were increased, the levels of MDA and proteinic expression of desmin were decreased markedly although BG, HbA1c and the activities of CAT were no significant difference in the diabetic rats by ascorbic acid treatment.

CONCLUSIONThe findings suggest that there are marked injury in podocyte, last barrier of glomerular filtration, in diabetic rats and administration of ascorbic acid can protect podocyte by increasing antioxidative capacity and ameliorating the renal oxidative stress.

Animals ; Ascorbic Acid ; pharmacology ; Catalase ; metabolism ; Desmin ; metabolism ; Diabetes Mellitus, Experimental ; metabolism ; pathology ; physiopathology ; Diabetic Nephropathies ; metabolism ; pathology ; physiopathology ; Male ; Oxidative Stress ; drug effects ; Podocytes ; drug effects ; metabolism ; ultrastructure ; Random Allocation ; Rats ; Rats, Wistar ; Superoxide Dismutase ; metabolism

OBJECTIVETo study the effect of arctiin on mouse podocyte epithelial-mesenchymal transition (EMT) induced by advanced oxidation protein products (AOPP).

METHODSMouse podocytes were stimulated by 200 µg/ml AOPP for 24 h in the presence of 50, 100, 200, and 400 µmol/L arctiin. The expressions of α-smooth muscle actin, Grp78 and CHOP were detected using Western blotting.

RESULTThe expressions of α-SMA, Grp78 and CHOP were inhibited by arctiin, showing a dose-dependent effect within a given range of arctiin concentration.

CONCLUSIONAOPP causes endoplasmic reticulum stress to induce EMT of mouse podocytes, and arctiin can decrease EMT by alleviating the stress. This finding sheds light on a new scope of research of renal fibrosis.

Actins ; metabolism ; Advanced Oxidation Protein Products ; adverse effects ; Animals ; Cell Line ; Endoplasmic Reticulum Stress ; drug effects ; Epithelial-Mesenchymal Transition ; drug effects ; Furans ; pharmacology ; Glucosides ; pharmacology ; Heat-Shock Proteins ; metabolism ; Mice ; Podocytes ; metabolism ; pathology ; Transcription Factor CHOP ; metabolism

OBJECTIVETo investigate the morphological changes and expressions of desmin and podocin in podocytes of rats with diabetic nephropathy (DN) rats and renal protection mechanism of Shenkangwan.

METHODSDN model was established in rats by a single injection of streptozotocin. The rats were then randomly divided into model group, Shenkangwan treatment group, irbesartan treatment group, and Shenkangwan plus irbesartan treatment group, with normal rats as the control group. All the rats received daily gavage for 8 weeks. The urinary protein quantity in 24 h were detected, and the morphological changes of the kidneys were observed with optic and transmission electron microscopes. The expressions of desmin and podocin in the podocytes were detected by immunohistochemistry.

RESULTSShenkangwan and irbesartan reduced the urinary protein quantity in 24 h and alleviated the renal damage in DN rats, and the expression of desmin was significantly attenuated while podocin expression increased in the podocytes.

CONCLUSIONSShenkangwan can provide renal protection against DN in rats and alleviate the structural and functional damages of podocytes possibly by reducing desmin expression and increasing podocin expression in the podocytes.

Animals ; Desmin ; biosynthesis ; Diabetic Nephropathies ; drug therapy ; pathology ; Drugs, Chinese Herbal ; pharmacology ; therapeutic use ; Immunohistochemistry ; Kidney ; drug effects ; pathology ; ultrastructure ; Male ; Microscopy, Electron, Transmission ; Phytotherapy ; Podocytes ; drug effects ; metabolism ; pathology ; Random Allocation ; Rats ; Rats, Sprague-Dawley

OBJECTIVETo evaluate the effect of bone morphogenic protein 7 (BMP-7) on nephrin expression and distribution in diabetic rat kidneys.

METHODSTwenty rats with diabetes mellitus (DM) induced by streptozotocin (STZ) injection were randomly divided into DM model group and BMP-7 treatment group, with another 10 normal rats serving as the normal control group. The rats in BMP-7 group received intraperitoneal human recombinant BMP-7 injections at 30 microg/kg twice a week for 24 consecutive weeks, while normal saline was administered in rats of the other two groups. Blood glucose and 24 hour urinary protein and creatinine (Ccr) were measured at 8, 16 and 24 weeks, and the rats were sacrificed at 24 weeks to obtain the renal tissues for detecting the expression and distribution of nephrin using immunofluorescence assay and RT-PCR and for examining the expressions of transforming growth factor-beta1 (TGF-beta1) and WT1 using immunohistochemistry.

RESULTSCompared with the normal control group, the DM model group showed significantly increased 24 hour urinary protein, kidney to body weight ratio and TGF-beta1 expression, but had lowered Ccr, glomerular podocyte number and nephrin expression. The linear distribution of nephrin along the capillary loops as found in the normal control group became granular in the kidney of diabetic rats. The rats in BMP-7 group showed less urinary protein excretion, lower TGF-beta1 expression and greater glomerular podocyte number than those in the DM group, and the expression and distribution of nephrin remained normal in the kidney.

CONCLUSIONAdministration of BMP-7 can significantly suppress the down-regulation of nephrin expression and maintain its normal distribution in the podocytes in diabetic rats possibly in association with a direct suppression of TGF-betasignaling.

Animals ; Blood Glucose ; metabolism ; Body Weight ; drug effects ; Bone Morphogenetic Protein 7 ; pharmacology ; Cell Count ; Diabetes Mellitus ; genetics ; metabolism ; pathology ; Gene Expression Regulation ; drug effects ; Humans ; Kidney ; drug effects ; metabolism ; pathology ; ultrastructure ; Male ; Membrane Proteins ; genetics ; metabolism ; Microscopy, Electron, Transmission ; Organ Size ; drug effects ; Podocytes ; drug effects ; pathology ; RNA, Messenger ; genetics ; metabolism ; Rats ; Rats, Sprague-Dawley ; Transforming Growth Factor beta1 ; metabolism

OBJECTIVETo observe the effect of Wenyang Huoxue Lishui Recipe (WHLR) containing serum on the expression of cathepsin L (CatL) in puromycin aminonucleoside-induced injury of mouse glomerular podocytes.

METHODSMouse podocyte cells (MPCs) in vitro cultured were divided into the normal control group, the model group, the dexamethasone (DEX) group, 10% WHLR containing serum group, 20% WHLR containing serum group, the vehicle serum control group. MPCs in the normal control group were cultured at 37 degrees C culture solution for 24 h. 45 mg/L puromycin was acted on MPCs in the model group for 24 h. On the basis of puromycin intervention, 1 limol/L DEX was co-incubated in MPCs of the DEX group for 24 h; 10% or 20% WHLR containing serum was co-incubated in MPCs of the 10% WHLR containing serum group and 20% WHLR containing serum group for 24 h. The vehicle serum control group was also set up by incubating with WHLR containing serum alone for 24 h. The expression of CatL and its substrate Synaptopodin in podocytes were detected by cell immunofluorescence staining. FITC-conjugated phalloidin was used to stain F-actin. A cortical F-actin score index (CFS index) was designed to quantify the degree of cytoskeletal reorganization in cultured podocytes.

RESULTSCompared with the normal control group, the expression of synaptopodin significantly decreased and the expression of CatL significantly-increased in the model group. F-actin arranged in disorder, gradually forming pericellular F-actin ring. CFS index was obviously elevated (P < 0.01). Compared with the model group, the epression of synaptopodin increased, the expression of CatL decreased, and CFS index also decreased in the DEX group, 10% WHLR containing serum group, and 20% WHLR containing serum group (P < 0.05, P < 0.01). Compared with the DEX group, the expression of synaptopodin decreased in 10% WHLR containing serum group, CFS index also decreased in 20% WHLR containing serum group (P < 0.05).

CONCLUSIONSWHLR could up-regulate the expression of synaptopodin, down-regulate the expression of CatL, and alleviate cytoskeletal reorganization of F-actin. It was helpful to stabilize the cytoskeleton of F-actin and improve the merging of podocytes.

Actins ; metabolism ; Animals ; Cathepsin L ; metabolism ; Cells, Cultured ; Down-Regulation ; Drugs, Chinese Herbal ; pharmacology ; Kidney Glomerulus ; cytology ; Mice ; Microfilament Proteins ; metabolism ; Podocytes ; drug effects ; pathology ; Puromycin Aminonucleoside ; adverse effects ; Up-Regulation

PURPOSE: The aim of this study was to investigate whether pathologic changes in zonula occludens-1 (ZO-1) are induced by interleukin-13 (IL-13) in the experimental minimal-change nephrotic syndrome (MCNS) model and to determine whether montelukast, a leukotriene receptor antagonist, has an effect on ZO-1 restoration in cultured human podocytes. MATERIALS AND METHODS: Human podocytes cultured on bovine serum albumin-coated plates were treated with different doses of IL-13 and montelukast and then examined for distribution using confocal microscopy and for ZO-1 protein levels using Western blotting. RESULTS: ZO-1 was internalized and shown to accumulate in the cytoplasm of human podocytes in an IL-13 dose-dependent manner. High doses (50 and 100 ng/mL) of IL-13 decreased the levels of ZO-1 protein at 12 and 24 h (both p<0.01; n=3), which were significantly reversed by a high dose (0.5 microM) montelukast treatment (p<0.01; n=3). CONCLUSION: Our results suggest that IL-13 alters the expression of ZO-1, and such alterations in the content and distribution of ZO-1 may be relevant in the pathogenesis of proteinuria in the MCNS model.
Acetates/*pharmacology ; Blotting, Western ; Dose-Response Relationship, Drug ; Humans ; Interleukin-13/*pharmacology ; Leukotriene Antagonists/*pharmacology ; Microscopy, Confocal ; Podocytes/*drug effects/metabolism ; Proteinuria/pathology ; Quinolines/*pharmacology ; Tight Junctions ; Zonula Occludens-1 Protein/*metabolism

PURPOSE: The aim of this study was to investigate whether pathologic changes in zonula occludens-1 (ZO-1) are induced by interleukin-13 (IL-13) in the experimental minimal-change nephrotic syndrome (MCNS) model and to determine whether montelukast, a leukotriene receptor antagonist, has an effect on ZO-1 restoration in cultured human podocytes. MATERIALS AND METHODS: Human podocytes cultured on bovine serum albumin-coated plates were treated with different doses of IL-13 and montelukast and then examined for distribution using confocal microscopy and for ZO-1 protein levels using Western blotting. RESULTS: ZO-1 was internalized and shown to accumulate in the cytoplasm of human podocytes in an IL-13 dose-dependent manner. High doses (50 and 100 ng/mL) of IL-13 decreased the levels of ZO-1 protein at 12 and 24 h (both p<0.01; n=3), which were significantly reversed by a high dose (0.5 microM) montelukast treatment (p<0.01; n=3). CONCLUSION: Our results suggest that IL-13 alters the expression of ZO-1, and such alterations in the content and distribution of ZO-1 may be relevant in the pathogenesis of proteinuria in the MCNS model.
Acetates/*pharmacology ; Blotting, Western ; Dose-Response Relationship, Drug ; Humans ; Interleukin-13/*pharmacology ; Leukotriene Antagonists/*pharmacology ; Microscopy, Confocal ; Podocytes/*drug effects/metabolism ; Proteinuria/pathology ; Quinolines/*pharmacology ; Tight Junctions ; Zonula Occludens-1 Protein/*metabolism

Diabetic nephropathy is the most serious complication in diabetes mellitus. It is known that oxidative stress and inflammation play a central role in the development of diabetic nephropathy. In this study, we investigated that ferulic acid (FA) known as anti-oxidative agent could effect on diabetic nephropathy by anti-oxidative and anti-inflammatory mechanism. We examined the effects of FA in obese diabetic Otsuka Long-Evans Tokushima Fatty (OLETF) rats and non-diabetic control Long-Evans Tokushima Otsuka (LETO) rats. We treated FA to experimental rats from 26 to 45 weeks of age. We evaluated ACR, MDA and MCP-1 in 24 h urine and examined renal histopathology and morphologic change in extracted kidneys from rats. Also, we evaluated the ROS production and MCP-1 levels in cultured podocyte after FA treatment. In the FA-treated OLETF rats, blood glucose was significantly decreased and serum adiponectin levels were increased. Urinary ACR was significantly reduced in FA-treated OLETF rats compared with diabetic OLETF rats. In renal histopathology, FA-treated OLETF rats showed decreased glomerular basement membrane thickness, glomerular volume, and mesangial matrix expansion. FA treatment decreased oxidative stress markers and MCP-1 levels in 24 h urine of rats and supernatants of cultured podocyte. In conclusion, it was suggested that FA have protective and therapeutic effects on diabetic nephropathy by reducing oxidative stress and inflammation.
Animals ; Anti-Inflammatory Agents, Non-Steroidal/*therapeutic use ; Antioxidants/*therapeutic use ; Cells, Cultured ; Chemokine CCL2/genetics/urine ; Collagen/analysis ; Coumaric Acids/*therapeutic use ; Diabetes Mellitus, Type 2/*complications ; Diabetic Nephropathies/*complications/*drug therapy/pathology ; Gene Expression/drug effects ; Kidney/drug effects/pathology ; Malondialdehyde/urine ; Podocytes/drug effects/metabolism ; Rats ; Rats, Inbred OLETF ; Reactive Oxygen Species/metabolism ; Transforming Growth Factor beta1/analysis

Diabetic nephropathy (DN) is a progressive kidney disease that is caused by injury to kidney glomeruli. Podocytes are glomerular epithelial cells and play critical roles in the glomerular filtration barrier. Recent studies have shown the importance of regulating the podocyte actin cytoskeleton in early DN. The phosphoinositide 3-kinase (PI3K) inhibitor, wortmannin, simultaneously regulates Rac1 and Cdc42, which destabilize the podocyte actin cytoskeleton during early DN. In this study, in order to evaluate the reno-protective effects of wortmannin in early DN by regulating Rac1 and Cdc42, streptozotocin (STZ)-induced proteinuric renal disease (SPRD) rats were treated with wortmannin. The albuminuria value of the SPRD group was 3.55 +/- 0.56 mg/day, whereas wortmannin group was 1.77 +/- 0.48 mg/day. Also, the albumin to creatinine ratio (ACR) value of the SPRD group was 53.08 +/- 10.82 mg/g, whereas wortmannin group was 20.27 +/- 6.41 mg/g. Changes in the expression level of nephrin, podocin and Rac1/Cdc42, which is related to actin cytoskeleton in podocytes, by wortmannin administration were confirmed by Western blotting. The expression levels of nephrin (79.66 +/- 0.02), podocin (87.81 +/- 0.03) and Rac1/Cdc42 (86.12 +/- 0.02) in the wortmannin group were higher than the expression levels of nephrin (55.32 +/- 0.03), podocin (53.40 +/- 0.06) and Rac1/Cdc42 (54.05 +/- 0.04) in the SPRD group. In addition, expression and localization of nephrin, podocin and desmin were confirmed by immunofluorescence. In summary, we found for the first time that wortmannin has a reno-protective effect on SPRD rats during the early DN. The beneficial effects of wortmannin in SPRD rats indicate that this compound could be used to delay the progression of the disease during the early DN stage.
Albumins/metabolism ; Androstadienes/*administration & dosage/pharmacology ; Animals ; Creatinine/blood ; Desmin/genetics/metabolism ; Diabetes Mellitus, Experimental/*drug therapy/metabolism/pathology ; Diabetic Nephropathies/*drug therapy/metabolism/pathology ; Disease Models, Animal ; Humans ; Intracellular Signaling Peptides and Proteins/genetics/metabolism ; Kidney/*pathology ; Membrane Proteins/genetics/metabolism ; Phosphatidylinositol 3-Kinases/*antagonists & inhibitors ; Podocytes/*drug effects/metabolism/pathology ; Rats ; Rats, Inbred Strains ; cdc42 GTP-Binding Protein/genetics/metabolism ; rac1 GTP-Binding Protein/genetics/metabolism