Podocyte number predicts progression of proteinuria in IgA nephropathy.

Podocyte injury is a feature of glomerulopathies associated with proteinuria, which in turn has been used as a clinical prognostic factor for glomerular diseases. The goal of this study is to investigate the relationship between podocyte injury found in biopsied renal tissue and change of proteinuria in IgA nephropathy (IgAN). In all, 35 patients with biopsy-proven IgAN and proteinuria (>1.0 g per 24 h) were enrolled in the IgAN group, while 8 patients with excision of renal harmatoma or carcinoma served as kidney controls (Control). Immunohistochemistry was applied to detect the expression of nestin, cell-cycle regulatory protein p27, as well as complement C5b-9 and complement receptor 1 (CR1). Podocyte foot process width (FPW) and podocyte population in renal biopsied samples were measured by morphometric analysis. On the basis of the podocyte density (Nv), the IgAN patients were divided into podocytopenic group (n=17, Nv<57.10 /microm(3) x 10(6)) and normopodocytic group (n=18, Nv> or =57.10 /microm(3) x 10(6)). Changes of proteinuria were followed for 18 months after biopsy. Compared with the Control, IgAN glomeruli had reduced podocyte expression of p27 and nestin along with decreased podocyte number. IgAN glomeruli also showed activation of C5b-9 in mesangial and subepithelial areas with decreased CR1 expression in podocytes. The C5b-9 positivity was inversely correlated with the number of WT-1-positive podocytes. Although the magnitude of proteinuria at biopsy correlated with podocyte FPW (P<0.05), the change in the amount of proteinuria expressed as proteinuria progression rate significantly correlated with the podocyte density. Thus, the normopodocytic group showed significantly lower proteinuria progression rate than the podocytopenic group regardless the comparable clinical features at biopsy and treatment regimen between the two groups. The results of this study indicate that, in IgAN, podocyte injury is involved in development of proteinuria and loss of podocytes predicts progression of the proteinuria. Complement activation may contribute to podocyte damage in IgAN.

Prevention of early liver injury by breviscapine in streptozotocin-induced diabetic rats.

The aim of this study was to investigate the protective effect of breviscapine extracted from the Chinese herb Erigeron breviscapus on liver injury in diabetic rats induced by streptozotocin. Treatment with breviscapine significantly reduced liver weight, liver lipid level, fatty liver and liver fibrosis score in diabetic rats. Treatment with breviscapine also significantly decreased lipid peroxidation malondiadehyde levels and increased the activities of antioxidative enzymes such as superoxide dismutase, catalase and glutathione peroxidase in diabetic liver. Immunohistochemical observations revealed that macrophage (ED-1-positive cells) infiltration in diabetic liver was inhibited by treatment with breviscapine. Western blot analysis showed that the expression of transforming growth factor-beta1 in diabetic liver was lowered by breviscapine treatment. In conclusion, our results indicate that breviscapine has potential as a treatment for diabetic liver injury through attenuating liver lipid accumulation and oxidative stress.

Protein kinase C beta inhibitor LY333531 attenuates intercellular adhesion molecule-1 and monocyte chemotactic protein-1 expression in the kidney in diabetic rats.

In vitro studies have shown that activation of protein kinase C (PKC) is a key mediator of intercellular adhesion molecule-1 (ICAM-1) and monocyte chemoattractant protein-1 (MCP-1) in a range of cell types and in response to high glucose, however, its role in the in vivo setting has not been clearly delineated. Streptozotocin-induced diabetic rats were treated with the PKC-beta isoform inhibitor LY333531 for 8 weeks. LY333531 treatment significantly attenuated increased urinary albumin excretion rate and glomerular volume and tubulointerstitial injury index as well as elevated PKC activity and PKC-beta protein expression in the kidney. Level of malondialdehyde was markedly higher and antioxidant enzyme activity such as superoxide diamutase and catalase as well as glutathione peroxidase were significantly lower in the kidney from diabetic rats than that of the control group. LY333531 administration could remit these changes. Increased macrophages recruitment as well as ICAM-1 and MCP-1 protein expression in the kidney were significantly inhibited by LY333531 in diabetic rats. It is concluded that mechanism of renoprotection of LY333531 may be correlated, at least partly, with suppression of increased macrophages recruitment and overexpression of ICAM-1 and MCP-1 in diabetic rats.

Prevention of early renal injury by mycophenolate mofetil and its mechanism in experimental diabetes.

Previously it was shown that treatment with mycophenolate mofetil (MMF) attenuated renal inflammation and glomerular injury in a model of diabetes. However, the mechanism involved in the renoprotective effects of MMF in experimental diabetes has not been clearly delineated. Diabetes was induced by injection of streptozotocin (STZ) after uninephrectomy. Diabetic animals received no treatment or treatment with MMF (10 mg/kg once daily by gastric gavage) for 8 weeks, non-diabetic rats served as controls. Level of malondialdehyde (MDA) in renal tissue and urine as well as the activity of antioxidant enzymes (AOE) in renal tissue was determined. Renal injury was evaluated. Immunohistochemistry for ED-1 macrophages marker, intercellular adhesion molecule-1 (ICAM-1) and monocyte chemoattractant protein-1 (MCP-1) was performed. Expression of transforming growth factor (TGF)-beta1 protein was determined by Western blotting analysis. Treatment with MMF had no effect on blood glucose level, but did prevent increased urinary albumin excretion and glomerular damage in diabetic rats. Oxidative stress was reduced by MMF treatment, as indicated by a reduction in MDA level in renal tissue and urine. Activity of AOE such as glutathione peroxidase (GSH-PX) was markedly elevated while superoxide dismutase (SOD) and catalase (CAT) were not changed by MMF treatment. In diabetic animals receiving no treatment, there were increases in ED-1-positive cells, ICAM-1 expression and MCP-1 expression in glomeruli and tubulointerstitium, which were effectively suppressed by MMF treatment. Western blotting analysis showed that the expression of TGF-beta1 protein was increased by 1.92-fold in renal tissue in diabetic rats, and MMF treatment significantly reduced the increased expression of TGF-beta1 protein by 45%. Our data suggest that MMF treatment ameliorates early renal injury via the inhibition of oxidative stress and overexpression of ICAM-1, MCP-1 and TGF-beta1 in renal tissue in diabetic rats.

[Determination of iohexol by high performance liquid chromatography].

A fast, sensitive and reproducible method for the determination of iohexol by high performance liquid chromatography with narrow-bore column was developed. The method involved one step deproteinization of plasma by perchloric acid and a simple, isocratic elution system of 7% (v/v) methane solution. The linearity of the method ranged from 0.5 to 500 mg/L of iohexol with a correlation coefficient of more than 0.999 8. Minimum detection limit was 154 pg of the agent in plasma. Retention time for the contrast medium in the elution system was (3.46 +/- 0.02) min and a single run was less than 5 min. The relative standard deviation of iohexol in peak area was less than 4.3% for run to run within a day and 11.4% for day to day operations. The recoveries of iohexol in plasma were over 91.51% at low, middle and high concentrations.

Mycophenolate mofetil treatment for diffuse proliferative lupus nephritis: a multicenter clinical trial in China.

OBJECTIVE: To investigate the efficacy and side effects of mycophenolate mofetil (MMF) in the treatment of diffuse proliferative lupus nephritis( DPLN). METHODS: 75 patients [13 male,62 female; age (31.0 +/- 10.1)y] with biopsy-proven active DPLN from nine hospitals in China from Jan. 1999 to Jan. 2000 were enrolled in this study, of whom 26 patients were refractory to conventional treatment of steroids and cyclophosphamide. 21 patients presented with relapses and 28 patients were newly diagnosed. All the patients were treated with a combined regimen of MMF and steroids. Patients who had severe active renal lesions were initially given intravenous methylprednisolone followed by oral prednisone. The initial dosage of MMF was 0.5 approximately 2.0 g/d and the administrati on maintained at least 6 months. 38 patients had repeat renal biopsy after treatment. SLE disease activity (SLE-DAI) score, renal active index (AI),chronic index(CI) and density of immunoglobulins, complement deposition was assessed before and after MMF treatment. RESULTS: The mean starting dosage of MMF was (1.26 +/- 0.30) g/d, it was reduced to (1.21 +/- 0.30) g/d and (0.95 +/- 0.33) g/d at the end of the 3(rd) and 6(th) month respectively. The post-treatment Hb level increased from (92 +/- 21) g/L to (112 +/- 28) g/L(3 mo) and (116 +/- 21) g/L(6 mo), while proteinuria decreased from (4.24 +/- 2.66) g/d to (2.18 +/- 3.75) g/d (3 mos, P < 0.05) and (1.54 +/- 1.60) g/d( 6 mos,P < 0.01). Renal function impairment present in some of the patients also showed marked improvement. The proportion of patients with positive A-dsDNA antibody and hypocomplementemia was significantly reduced after 6 months of MMF treatment. SLE-DAI score in this group of patients decreased from (16.9 +/- 6.7) to (8.1 +/- 4.8) (P < 0.01) by the end of 3 months. Repeat renal biopsy in 38 patients 3 approximately 6 months after the treatment showed a significant decrement of AI ( 13.30 +/- 5.51) vs (3.38 +/- 1.98),P < 0.01, and an increment of CI (1.62 +/- 1.48) vs (2.62 +/- 1.85), P > 0.05. Immunoglobulins and complement deposition scores decreased from (9.39 +/- 3.51) to(6.71 +/- 3.16 ),P < 0.05. During the study period, 12 episodes of infection (16.0%) were recorded including pneumonia(2.7%), herpes zoster(8.0 %), urinary tract infection(2.7%), and tuberculosis(2.7%). Other side effects included gastrointestinal symptoms (10.7%), hirsutism(6.7%), leukocytopenia (1.3%) and transient elevation of SGPT(2.7%). CONCLUSION: MMF in a dosage of 0.5 approximately 2.0 g/d combined with steroids is effective to control the lupus activity of DPLN and well tolerated by the patients.

[The effects of endothelin blockade on renal expression of angiotensin II type 1 receptor in diabetic hypertensive rats].

OBJECTIVE: To investigate the changes of angiotensin II type 1 (AT1) receptor in the kidney of spontaneously hypertensive rats (SHR) with diabetes and the influence of endothelin receptor antagonist bosentan. METHODS: Streptozotocin-induced diabetic SHR were divided into four groups: groups treated by cilazapril, bosentan + amlodipine, and amlodipine rsspectively, and untreated group, 6 rats in each group. Six SHR rats and six WKY rats were used as hypertensive and normotensive controls respectively. By the end of the 4th week, all rats were anasthetized and catheteization was conducted to their right common carotis arteries to measure the mean arterial blood pressure and collect blood samples to determine the blood sugar and creatinine by biochemical analyzer, and plasma angiotensin II level by radioimmunoassay. Then the rats were killed and their kidneys were taken. The renal angiotensin II (Ang II) receptor and expression of AT1 receptor was determined by RT-PCR and Western blotting. One day before the rats were killed, 24-hour urine was collected to determine the urinary protein and creatinine. RESULTS: In untreated diabetic SHR, enhanced blood pressure and urinary protein excretion, reduced creatinine clearance, as well as significantly increased plasma and renal Ang II levels were observed compared with those in WKY rats. Immunohistochemistry, Western blotting and semi-quantitatively RT-PCR methods showed that the protein and mRNA levels of AT1 receptor in kidney were significantly reduced in untreated diabetic SHR compared to those in WKY rats. All these abnormalities were attenuated by bosentan + amlodipine and cilazapril therapies. CONCLUSION: Bosentan prevents the down-regulation of AT1 receptor in the kidneys of diabetic hypertensive rats.

[Effect of selective cyclooxygenase -2 inhibitor on the renal lesion of streptozotocin-induced diabetic rats and its possible mechanism].

OBJECTIVE: To investigate the effect of selective cyclooxygenase (COX)2 inhibitor-meloxicam on the renal lesion of diabetic rats and its possible mechanism. METHODS: Twenty-nine rats were randomly divided into four groups: normal control rats (n = 6), streptozotocin (STZ)-induced diabetic rats without treatment (n = 8), STZ-induced diabetic rats treated with indomethacin (2 mg x kg(-1) x d(-1)) (n = 6), and STZ-induced diabetic rats treated with meloxicam (2 mg x kg(-1) x d(-1)) (n = 9). Sixteen weeks later, the blood sugar, blood urea nitrogen (BUN), and blood creatinine were examined. Radioimmunoassay was used to examine the prostaglandin(2) (PGE(2)) and Thromboxane B(2) (TXB(2)) in the urine. The expression of transforming growth factor -beta1 (TGF-beta1) and TGF-beta receptor type II (TbetaR2) of the renal cortex were measured by reverse transcription-polymerase chain reaction (RT-PCR) analysis. Immuno-precipitation analysis was carried out to examine the protein level of angiotensin II type 1 (AT1) receptor. Periodic acid-Schiff staining was used to examine the morphological changes by light microscopy. RESULTS: In the STZ-induced diabetic group, the blood levels of sugar, BUN, and creatinine were higher, and the creatinine clearance (Ccr) was remarkably higher in comparison with those in the normal control group (P < 0.05). Ccr was lower in diabetic rats treated with indomethacin and diabetic rats treated with meloxicam than in diabetic rats without treatment (P < 0.05). There was no difference of blood creatinie among these three groups. The blood sugar level in diabetic rats treated with meloxicam was lower than those in the diabetic rats treated with indomethacin and untreated group (all P < 0.05). The renal weight/body weight ratio was significantly higher in the untreated group than that in the control group. The PGE(2), TXB(2) and albumin levels in urine of STZ-induced diabetic rats were 1,641 +/- 288 pg/24 h 5,507 +/- 1,359 pg/24 h, and 46.3 +/- 9.5 microg/24 h respectively, much higher than those in meloxicam group (910 +/- 255 pg/24 h, 3,272 pg/24 h +/- 670 pg/24 h and 17.2 +/- 5.4 microg/24h respectively, all P < 0.01). The urine PGE(2) and albumin was 1,195 +/- 448 pg/24 h and 34.1 +/- 10.2 microg/24 h respectively in the indomethacin group. There was no difference in renal weight/body weight ratio and TXB(2) excretion between the untreated group and indomethacin group. The relative contents of TGF-beta1 and TbetaR2 mRNA expression in renal cortex of STZ-induced diabetic rats were 0.185 +/- 0.037 and 0.194 +/- 0.054, much higher than those in other groups. Glomerular hypertrophy, mesengial expansion, extracellular matrix accumulation, and sclerosis of partial glomeruli were seen in diabetic rats without treatment and those treated with indomethacin. The pathological changes were less in meloxicam group (P < 0.05). CONCLUSION: The selective COX-2 inhibitor meloxicam significantly suppresses TGF-beta1 and TbetaR2 genes expression, elevates the protein level of AT1 receptor and attenuate the renal lesion caused by diabetes. TGF-beta1 and AT1 receptor may be involved in the mechanism concerned.