Effects of Houttuynia Cordata Thumb on Expression of BMP-7 and TGF-β1 in the Renal Tissues of Diabetic Rats

Objective： To explore the effects of Houttuynia Cordata Thumb （HCT 鱼腥草 Yu Xing Cao） on expression of transforming growth factor-β1 （TGF-β1） and bone morphogenetic protein-7 （BMP-7） in the renal tissues of diabetic rats. Methods： The diabetic rats induced by intravenous injection of streptozotocin（STZ） were randomly divided into a model group, a HCT group and a lotensin group, with normal rats designated as the controls. 8 weeks later, the ratio of kidney weight to body weight, the glomerular area, the excretion of β 2-microglobin （β2-MG） in 24-hr urine, the albumin excretion in 24-hr urine, and creatinine clearance rate （CCR） were investigated. The expression of TGF- β 1, BMP-7 and collagen I in the renal tissues was observed with the immunohistochemical method and by the semi-quantitative assay. Results： The overgrowth of glomerulus, the excretion of β 2-MG in 24-hr urine, the albumin excretion rate in 24-hr urine and CCR in the HCT group significantly reduced （P〈0.05）, and the expression of TGF-β1 and collagen I significantly decreased （P〈0.05）, but BMP-7 significantly increased （P〈0.05） in the HCT group as compared with those in the model group, with no significant difference as compared with the lotensin group （P〉0.05）. Conclusion： HCT has a protective effect on the renal tissues in diabetic rats, which is probably correlated with the decrease of the expression of TGF-β1 and collagen I and with the increase of the expression of BMP-7 in the renal tissues.     

Expression of Serum and Glucocorticoid-inducible Kinase1 in Diabetic Rats and Its Modulation by Fluvastatin

The expression of serum and glucocorticoid-induced protein kinase in the renal cortex of diabetic rats was examined, and the function of signal transduction mediated by SGK1 in diabetic nephropathy and its modulation by fluvastatin were also investigated. 24 male Wistar rats were randomly divided into normal control group （n = 8）, diabetic nephropathy group （n = 8） and fluvastatin-treated diabetic nephropathy group （15 mg/kg/d, n=8）. The metabolic parameters were measured at the 8th week. The expression of transforming growth factor β1 （TGF-β1） and fibronectin （FN） was immunohistochemically examined. The expression of SGK1 was detected by RT-PCR and Western blot, and CTGF mRNA was assessed by RT-PCR. As compared to DN, blood glucose, 24-h urinary protein, Cer and kidney weight index were all decreased and the weight was increased obviously in group F. At the same time, mesangial cells and extracellular matrix proliferation were relieved significantly. The levels of cortex SGK1 mRNA and protein were up-regulated, and both TGF-β1 and FN were down-regulated by fluvastatin. The mRNA of SGK1 was positively correlated with the CTGF, TGF-β1 and FN. SGK1 expression is markedly up-regulated in the renal cortex of DN group and plays an important role in the development and progress of diabetic nephropathy by means of signal transduction. Fluvastatin suppressed the increased SGKlmRNA expression in renal cortex and postponed the development of diabetic nephropathy.     

Effect of ethanol extract of Rhodiola rosea on the early nephropathy in type 2 diabetic rats

This study aimed to investigate the therapeutical effects of Rhodiola rosea extract on rats with type 2 diabetic nephropathy (DN). The rat type 2 DN model was established by high fat and high calorie feeding and intravenous injection of streptozocin (STZ). Wistar rats were randomly divided into normal group, control group, low dose Rhodiola rosea group, high dose Rhodiola rosea group and Captopril group. Oral glucose tolerance test (OGTT) was performed to determine the impairment of glucose tolerance in the established animal model. A series of parameters including fasting blood glucose (FBG), total cholesterol (TC), triglyceride (TG), creatinine clearance rate (Ccr), 24-h urinary albumin (UA), the ratio of kidney mass/body weight (renal index) and glomerular area were examined after 8 weeks. Moreover, the expression of transforming growth factor (TGF)-β1 in renal tissues was detected by using immunohistochemisty. At the end of the eighth week, FBG, TC, TG, Ccr, 24-h urinary albumin, the ratio of kidney mass/body weight and glomerular area were significantly reduced in Rhodiola rosea extract treatment groups as compared with those in control group. TGF-β1 expression in renal tissues of Rhodiola rosea extract treatment groups was also significantly decreased as compared with that of control group. These results indicate that Rhodiola rosea extract may have a protective effect on early nephropathy in diabetic rats, which might be related to the decrease of the renal expression of TGF-β1.     

山茱萸提取物-总三萜烯酸改善链脲佐菌素诱导糖尿病大鼠肾损害进展

Objective:To Investigate whether total triterpene acids(TTAs),isolated from Corpus Fructus,attenuates renal function by reducing oxidative stress and down-regulating the expression of transforming growth factor β_1(TGF-β_1).Methods:Diabetes was induced by an injection of streptozotocin(40 mg/kg intravenously).Thirty rats were randomly divided into three groups:control group,diabetic model group and TTAs treatment group(50 mg/kg,intragastrically)administrated for 8 weeks from 5th to 12th week.All rats were anaesthetized and then were killed to remove kidneys.The renal function and redox enzyme system parameters were tested.Glomerular morphology was observed by a light microscopy.Immunohistochemistry and Western blot assays were employed to determine the protein levels of TGF-β_1.Results:TTAs attenuated the levels of urinary protein,serum creatinine and blood urea nitrogen,although it did not significantly reduce the level of glucose.In addition,TTAs decreased the malondialdehyde while increased superoxide dismutase,catalase and glutathione peroxide activities in diabetic rats.The renal pathological changes in TTAs treatment group were ameliorated.Furthermore,TTAs also ameliorated the expression of TGF-β_1.Conclusion:TTAs improved renal function via reducing oxidative stress and down-regulation the expression of TGF-β_1 in diabetic rats.     

Effect of Calcium Dobesilate on Nephropathy in Type 2 Diabetic Rats

In order to investigate the effects and mechanisms of calcium dobesilate on renal lesions in experimental type 2 diabetic rats, dibetic rats were randomly divided into control group (group C) and experimental group (group D) treated with calcium dobesitate. The serum creatinine (Scr),protein kinase C (PKC), creatinine clearance (Ccr), transforming growth factor-beta, (TGF-β1),type Ⅳ collagen were compared among the groups after 24 weeks. The renal tissues were observed under light microscopy and electron microscopy. The results showed that after 24 weeks, Scr,PKC, TGF-β1 in group D were significantly lower than in group C, meanwhile, renal pathologic changes in group D were improved. Ccr had no difference between group C and group D. It was concluded that calcium dobesilate could ameliorate renal lesions in diabetic rats through inhibiting PKC and TGF-β1.     

Effects of mycophenolate mofetil, valsartan and their combined therapy on preventing podocyte loss in early stage of diabetic nephropathy in rats

Background Podocyte has inflammatory role in the development of diabetic nephropathy (DN). Mycophenolate mofetil (MMF), an anti-inflammatory agent, can suppress macrophage infiltration and reduce renal injury in streptozotocin-induced diabetic rats. Angiotensin II receptor blocker (ARB), another renal protecting agent, can decrease podocyte loss in DN. In this study, we detected the expression levels of monocyte chemoattractant protein-1 (MCP-1) and nephrin to evaluate podocyte’s role in inflammatory reaction in DN, observe and compare the effect of MMF alone and in combination with valsartan, on preventing podocyte loss in streptozotocin (STZ) induced diabetic rats. Methods Diabetic model was constructed in uninephrectomized male Wistar rats by single peritoneal injection of STZ (65 mg/kg). The successfully induced diabetic rats were randomly divided into four groups: diabetes without treatment group (DM), valsartan treated group (DMV), MMF treated group (DMM), and combined therapy group (DMVM). Normal rats of the same sibling were chosen as control (NC). At the end of the 8th week, serum biochemistry, 24-hour urinary protein (UP) and the ratio of kidney weight/body weight (RWK/B) were measured. The rats were sacrificed for the observation of renal histomorphology through light and electron microscope. Nephrin, desmin and MCP-1 levels were detected by semi-quantitative immunohistochemical assays. Real-time quantitative PCR was used to detect the mRNA levels of nephrin and MCP-1.Results Compared with group NC, serum glucose level, 24-hour UP and RWK/B in group DM were significantly higher (P<0.01), and the nephrin mRNA level in DM group was significantly lower (P<0.05). The nephrin mRNA expression levels in group DMV, DMM and DMVM were all higher than that of DM group (P<0.05) and no significant differences were found among the three treatment groups (P>0.05). Treatment with MMF, valsartan or their combination could significantly decrease the 24-hour UP and RWK/B, and suppress glomerulosclerosis and interstitial fibrotic lesions in diabetic rats. In diabetic rats, the high expressions of desmin and MCP-1 in kidney were suppressed by valsartan, MMF or their combination.Conclusions Podocytes are involved in the inflammatory reaction of diabetic rats. MMF could suppress MCP-1 and desmin expression, enhance nephrin expression, and attenuate proteinuria in diabetic rats. The combined therapy of valsartan and MMF did not show any superiority over monotherapies on renal protection. MMF may have renoprotective effect in early stages of diabetic nephropathy through preventing podocytes loss and anti-inflammatory activity.     

Inhibitory Effect of TanshinoneⅡA on TGF-β1-induced Cardiac Fibrosis

This study examined the effect of tanshinoneⅡA (TSNⅡA) on the cardiac fibrosis induced by transforming growth factor β1 (TGF-β1) and the possible mechanisms. Cardiac fibroblasts were isolated from cardiac tissues of neonatal Sprague-Dawley (SD) rats by the trypsin digestion and differential adhesion method. The cells were treated with 5 ng/mL TGF-β1 alone or pretreated with TSNⅡA at different concentrations (10–5 mol/L, 10–4 mol/L). Immunocytochemistry was used for cell identification, RT-PCR for detection of the mRNA expression of connective tissue growth factor (CTGF) and collagen type Ⅰ (COLⅠ), Western blotting for detection of the protein expression of Smad7 and Smad3, and immunohistochemistry and immunofluorescence staining for detection of the protein expression of phosphorylated Smad3 (p-Smad3), CTGF and COLⅠ. The results showed that TGF-β1 induced the expression of CTGF, COLⅠ, p-Smad3 and Smad7 in a time-dependent manner. The mRNA expression of CTGF and COLⅠ was significantly increased 24 h after TGF-β1 stimulation (P<0.01 for all). The protein expression of p-Smad3 and Smad7 reached a peak 1 h after TGF-β1 stimulation, much higher than the baseline level (P<0.01 for all). Pretreatment with high concentration of TSNⅡA resulted in a decrease in the expression of p-Smad3, CTGF and COLⅠ (P<0.01). The protein expression of Smad7 was substantially upregulated after pretreatment with two concentrations of TSNⅡA as compared with that at 2h post TGF-β1 stimulation (P<0.05 for low concentration of TSNⅡA; P<0.01 for high concentration of TSNⅡA). It was concluded that TSNⅡA may exert an inhibitory effect on cardiac fibrosis by upregulating the expression of Smad7, suppressing the TGF-β1-induced phosphorylation of Smad3 and partially blocking the TGF-β1-Smads signaling pathway.     

Effect of expression of TGF-beta and TNF-alpha with Qidan granule treatment in rats by bleomycin-induced pulmonary fibrosis

Objective: To evaluate the therapeutic effects and mechanisms of Qidan granule in blemycinA5-induced pulmonary interstitial fibrosis (PIF)in rats. Methods: PIF models were established by blemycinA5-induced in rats. They were treated by Qidan granule and Hydrocortisone respectively. The pathological changes and collagen protein disposition were observed, and the expression of TGF-β, TNF-α proteins were measured by immunohistochemical technique . Results: The pulmonary alveolitis and fibrosis were alleviated remarkably in Qidan granule group compared with those in the model control group and hydrocortisone group (P <0. 01). The expression of TGF-β and TNF-α protein were higher in Qidan granule group than those in normal group , and were significantly less than those in the model control group and in hydrocortisone group (P < 0. 01). Conclusion: Qidan granule would ameliorate the pulmonary alveolitis and fibrosis. TGF-β and TNF-α might play an important role in the development of alveolitis and fibro     

Different Expressions of Protein Kinase C-α,βⅠ and βⅡ in Glomeruli of Diabetic Nephropathy Patients

In current study, the expressions of protein kinase C （PKC）-α, βⅠ and βⅡ as well as their correlation to the expression of transforming growth factor-βⅠ （TGF-βⅠ） and vascular endothelial growth factor （VEGF） were investigated in glomeruli of normal renal tissues taken from human kidney tumors and kidney tissues from patients with diabetic nephropathy （DN）. The accumulation of glomerular extracelluar matrix （ECM） was determined by PAS staining, the expressions of PKC-α, PKC-βⅠ, PKC-βⅡ, TGF-βⅠ and VEGF were measured by semi-quantitative immunohistochemistry. Our results showed that in glomeruli of normal renal tissues, PKC-α and βⅡ had a strong expression whereas the expression of PKC-βⅠ was weak; in glomeruli of DN patients, the expressions of PKC-α, PKC-βⅠ, VEGF and TGF-βⅠ and the accumulation of ECM increased significantly, but the expression of PKC-βⅡ decreased markedly. Meanwhile, the expressions of PKC-α and βⅠ had a positive correlation to the expressions of VEGF and TGF-βⅠ respectively, whereas PKC-βⅡ showed no correlation to VEGF and TGF-βⅠ. It is concluded that the expressions of PKC-α, βⅠ and βⅡ in glomeruli of normal subjects and DN patients are different. PKC-α seems to play a critical role in human DN by up-regulating VEGF expression, whereas PKC-βⅠ is relatively important for the up-regulation of TGF-βⅠ and the accumulation of ECM under diabetic conditions.     

Protective effects of eplerenone on podocyte injury in adriamycin nephropathy rats

To investigate the protective effects of eplerenone on adriamycin-induced renal injury and the possible mechanisms involved,36 male Sprague-Dawley rats were randomly divided into control group,adriamycin nephropathy(AN) group and eplerenone-treated group(100 mg.kg-1.d-1 eplerenone).Blood pressure,24-h urinary protein,serum potassium,sodium and creatinine were measured 28 days after adriamycin injection(a single tail intravenous injection of 6.5 mg/kg adriamycin).The morphological changes of renal tissues were observed by light and electron microscopy.Immunohistochemistry and Western blotting were performed to examine the expression of TGF-β1 and desmin in renal cortex.The results showed that 28 days after adriamycin injection,there were no significant changes in the level of serum potassium,sodium,creatinine concentrations and blood pressure values in the rats of the three groups.Meanwhile,the 24-h proteinuria excretion in the AN group was significantly higher than that in the control group(P<0.01),but that in the eplerenone-treated group was substantially reduced when compared with that in the AN group(P<0.05).Mild mesangial cell proliferation and matrix expansion,diffuse deformation and confluence of foot processes in podocytes were found in the AN group.By contrast,rats in the eplerenone-treated group exhibited obvious attenuation of these morphological lesions.The protein expression of TGF-β1 and desmin in the AN group was markedly up-regulated in contrast to that in the control group(P<0.01),whereas that in the eplerenone-treated group was much lower than in the AN group(P<0.05).It was concluded that eplerenone may ameliorate the proteinuria and the development of pathological alteration in adriamycin-induced nephropathy presumably via the inhibition of cytokine release,and restore the morphology of podocytes independent of its blood pressure-lowing effects.     

Effect of Yishen Huayu Fang on kidney tissue E-cadherin expression in unilateral ureter ligation in rats

OBJECTIVE:To observe E-calcium sticky protein(E-cadherin) expression in kidney tissues in a rat model of unilateral ureter ligation and the effect of Yishen Huayu Fang(formula of tonifying the kidney and dissolving accumulated blood stasis) on the expression.METHODS:A total of 150 clean grade male rats were randomly divided into a control group,model group,low-dose Yishen Huayu Fang group(low-dose group),high-dose Yishen Huayu Fang group(high-dose group),and Lotensin group.A renal fibrosis model was established with unilateral ureteral obstruction(UUO).Pathological changes of rat renal tissue were observed with light microscopy on days 3,7,14,21,and 28 after UUO.Changes in kidney tissue E-cadherin expression were observed with immunohistochemistry.RESULTS:Three days after modeling,kidney edema appeared followed by gradual inflammatory cell infiltration,and part of the small tubules disappeared while the renal cortex thinned.Meanwhile,the E-cadherin expression level dropped,which was negatively correlated with the obstruction time.After intervention,E-cadherin expression was increased in all treatment groups(P<0.01 or P<0.05),while there were no significant differences between the high-dose and Lotensin groups.CONCLUSION:Yishen Huayu Fang delays the renal fibrosis process by promoting E-cadherin expression in renal tissues and reducing extracellular matrix deposition.     

Effect of Coiquhoumia Root on the Expression of Transforming Growth Factor-β in Mesangiai Proliferation Giomerulonephritis Model

Summary： To study the efficacy and the mechanism of Colquhoumia root （ Tripterygium hypoglaucure （Le,vL） Hutch） in the treatment of mesangial proliferation glomerulonephritis （MsPGN）, SD rats were injected with anti-thymoeyte serum （ATS） to make MsPGN model （anti-Thyl model）. The rats were then divided into 3 groups： normal control group, anti-Thyl model group and treatment group. Histopathologieal （HE, PAS）, immunohistoehemieal, RT-PCR technique and computer imaging analysis system were used to evaluate mesangial matrix production, the expression of TGF-β protein and mRNA in the tissues of kidney. Our result showed that proteinuria and the ratio of extraeellular matrix/glomerular capillaries area （ECM/CA） were increased significantly in model group. The expression of both TGF-β protein and mRNA in glomeruli was much higher in model group than in control group （P〈0.01）. After the treatment with Colquhoumia root, proteinuria, ECM/CA and the expression of both TGF-β1 protein and mRNA in glomeruli were significantly decreased in treatment group as compared with those in model group. It is concluded that Colquhoumia root is effective in reducing proteinuria and mesangial matrix proliferation in MsPGN and it may achieve these effects by inhibiting the expressions of TGF-β1 protein and mRNA of mesangial cells.     

Role of BMP-7 and TGF-β1 expression in renal tubulo-interstitial lesion of Wistar rats induced by unilateral ureteral obstruction

Objective： To explore the role of bone morphorgenetic protein-7 （BMP-7） in the renal tubulo-interstitial lesions induced by unilateral ureteral obstruction （UUO）. Methods： Sixty Wistar rats were equally and randomly divided into normal control, sham operation and UUO groups, and respectively sacrificed on the 1st, 3rd, 7th, 14th day after the time of UUO operation. The mRNA levels of BMP-7 and TGF-β1 in the renal tissues were examined by RT-PCR. The expression sites and levels of BMP-7 and TGF-β1 proteins were detected by immunohistochemistry staining. Results：Compared to control groups, the level of BMP-7 mRNA was significantly decreased, but that of TGF-β1 mRNA was significantly increased in UUO rats. Immunohistochemistry staining indicated that BMP-7 mainly expressed in the renal tubules and interstitum, rarely in the glomeruli. In UUO rats, the expression of BMP-7 protein was decreased, but that of TGF-β1 was increased in an obstruction dependent manner. Conclusion：The downregulation of BMP-7 is observed in the early phase of fibrotic process of the renal interstitium, suggesting it may be involved in the formation and development of the tubulo-interstitial lesions.