Objective To evaluate the efficacy and safety of leflunomide (LEF) in induction and maintenance therapy of type Ⅳ and type Ⅴ lupus nephritis. Methods A prospective single-center controlled clinical trial was conducted. Patients with biopsy-confirmed type Ⅳ and type Ⅴ lupus nephritis in one month were recruited. Patients were given either LEF (LEF oral 30 mg/d) or cyclophosphamide (CTX IV 1 g/month) combined with prednisolone (40～60 mg/d, tapered to 10 mg/d). After 6 months of induction therapy, 20 mg/d LEF and 1 g/3months CTX with prednisolone (5~10 mg/d) added respectively. Side effects were recorded and the efficacy and safety were evaluated. Results Forty patients were enrolled (19 in LEF group and 21 in CTX group), and 5 patients withdrew due to adverse events (2 in LEF group and 3 in CTX group) during induction therapy. The total response rate was 88.2％ in LEF group and 72.2％ in CTX group respectively. Complete remission rate was 52.9％ in LEF group and 44.4％ in CTX group after 6 months induction therapy respectively. There was no difference in the change of proteinuria, serum albumin, serum creatinine, C3 and SLEDAI between two groups. Eighteen response patients entered maintenance therapy (7 in LEF group, 11 in CTX group). LEF group was followed up for (18.6±6.5) months, and no patient was found relapse, and 3 patients received repeated renal biopsy, which showed the pathologic type was improved and the activity index decreased, but the chronic index increased slightly. CTX group was followed up for (25.1±9.6) months and 3 patients relapsed. Major adverse event in LEF group was infection, among which herpes zoster was the most common type. Major adverse events in CTX group were infection and amenorrhea . Conclusions LEF combined with steroid appears to be a safe and effective therapy within the induction and maintenance stage of the patients with type Ⅳ and Ⅴ LN. The long-term benefit of LEF needs to be confirmed by additional studies.

Objective To evaluate the relationship of C161T polymorphism in peroxisome proliferator-activated receptor γ (PPARγ) gene with glucocorticoid -induced osteoporosis(GIO) in Chinese population. Methods The C161T genotypes of PPARγ exon 6 were determined by PCR-RFLP in 208 healthy persons(group Ⅰ), 168 patients without GIO(group Ⅱ) and 104 patients of GIO(group Ⅲ) . Bone mineral density(BMD) at lumbar spine, femoral neck， trochanter and Ward’s triangle were measured by dual-energy X-ray absorptiometry(DEXA). Results Hardy-Weinberg equilibrium was evident for PPARγ polymorphism. PPARγ gene CC genotype frequency was lower in group Ⅲ as compared with groupⅠ. CT and TT genotype frequencies were higher in group Ⅲ as compared with groupⅠ. CC, CT and TT genotype frequencies were not significantly different among group Ⅱ, group Ⅱ+Ⅲand groupⅠ. BMD of lumbar spine in group Ⅱand gronp Ⅲpatients with CC genotype was lower as compared to those with CT+TT (P ＜ 0.05). In groupⅡ, the BMD of CC and CT+TT genotypes at lumbar spine area was (1.04±0.17) g/cm2 and (1.02±0.07) g/cm2 respectively. In groupⅢ, the BMD at lumbar spine area was (0.94±0.12) g/cm2, (0.83±0.08) g/cm2 of CC, CT+TT genotypes respectively. The significant difference was also presented after adjusted by age and BMI. Conclusions The frequency of the genotype is not significantly different between groupⅠand groupⅡ+Ⅲ, which indicates that genotype may not be related with the onset of glomerulonephritis. The frequency of the genotype is significantly different between group Ⅰ and group Ⅲ, which indicates that genotype may be related with the onset. Polymorphism of C161T in PPARγ gene may have an effect on the BMD of lumbar spine of the patients receiving glucocorticoid group (Ⅱ+Ⅲ).Allele C may be a protection factor of bone mass.

Objective To investigate the relative frequency and to elucidate the relationship between clinical and pathological features of primary focal segmental glomerulosclerosis(FSGS) in the south of China. Methods The data of 263 adult patients diagnosed as primary FSGS from 1988 to 2005, excluding secondary FSGS by renal biopsies, were reviewed and analyzed retrospectively. Results (1) The incidence of primary FSGS accounted for 7.02% of adult primary glomerular diseases confirmed by renal biopsies, and 6.33% of adult primary nephrotic syndrome(NS) within corresponding period. The predominant patients were young people. The patients presented various degrees of proteinuria, including nephrotic syndrome in 133 cases (about 50.6%). (2) The main pathological characteristics of the 263 FSGS cases were as follows: 48.4% of the patients had a glomerulosclerosis ratio more than 25%, 88.5% of glomerulosclerosis usually accompanied tubulointerstitial lesion, including severe lesions of graded 2 or 3 (about 25.2%). (3) Both glomerulosclerosis and tubulointerstitial lesion were correlated with renal insufficiency. The degree of glomerulosclerosis and tubulointerstitial lesion was negatively correlated with creatinine clearance, but positively with serum creatinine. A positive correlation was also found between glomerulosclerosis and tubulointerstitial lesion (P＜0.01). Tubulointerstitial lesion was a significant influencing factor of renal insufficiency. Conclusions Primary FSGS is one of the major pathological types of adult NS. Glomerulosclerosis and tubulointerstitial lesion, which contribute to renal insufficiency, progress severely when the patients are diagnosed as primary FSGS. Therefore early diagnosis and therapy is recommended to attenuate the disease and is one of the important topics for the nephrologists.

Objective To detect the urinary levels of monocyte chemoattractant protein 1 (MCP-1), β-catenin and cytokeratin 19(CK19) in patients with various renal diseases and to analyze the relationship among these molecular levels in urine and the clinical and pathological features of patients, especially the formation of crescent. Methods Urine specimens of 20 healthy subjects and 124 patients with various renal diseases were collected at the day of renal biopsy and one month after renal biopsy respectively. Urinary levels of MCP-1, β-catenin and CK19 were examined by enzyme linked immunosorbent assay (ELISA). Urinary creatinine, 24-hour urinary protein excretion and serum creatinine were detected at the same time. The indexes of cellular crescent and fibrous crescent were calculated according to the number and the size of crescents respectively. Results All the patients had significantly higher levels of urinary MCP-1, β-catenin and CK19 compared with healthy subjects （P < 0.01）.Levels of urinary MCP-1 and β-catenin were significantly higher in patients with cellular crescent than those in patients without crescent [median 247.54, range 22.17～2335.18 ng/mmol Cr vs median 113.55, range 1.75～1057.77 ng/mmol Cr; median 219.40, range 0.93～3827.50 ng/mmol Cr vs median 82.30，range 4.03～1632.58 ng/mmol Cr, P<0.01]. Urinary levels of MCP-1, β-catenin and CK19 were positively correlated to the indexes of cellular crescent ( r= 0.75，0.21，0.63， P< 0.01). Conclusions Urinary levels of MCP-1, CK19 and β-catenin can be detected By ELISA in healthy subjects and patients with various renal diseases. The levels of urinary MCP-1, β-catenin and CK19 are correlated to the indexes of cellular crescent in the renal biopsy specimens.

Objective To evaluate the efficacy of catheter-restricted filling using the gentamicin-heparin mixed solution in the prevention for catheter-related bacteremia (CRB). Methods Forty-three patients in our center were enrolled from Nov. 2004 to Mar. 2005 in this study, and were randomly assigned to receive either heparin lock solution (H group, heparin 45 g/L, 23 patients) or gentamicin-heparin mixed solution (GH group, gentamicin 4 g/L, heparin 45 g/L, 20 patients) as the catheter lock solution during the interdialytic period. The observative indicators included the incidence of CRB, the concentration of gentamicin in peripheral blood, the residual kidney function (RKF), the side effect of gentamicin, and the change of blood levels of CRP and IL-6. Results Mean CRB-free catheter survival days was 114.13±34.39 (31~200) in GH group and 127.40±32.85 (40~196) in H group. Accumulative catheter day of 2625 day was gained in GH group and 2548 in H group. CRB developed in two patients in the H group whereas none of the patients developed CRB in GH group, however the incidence of CRB between two groups was not significantly different. The trough and peak concentrations of gentamicin in GH group were (0.23±0.12) mg/L and (0.53±0.29) mg/L respectively after two weeks, and(0.26±0.15) mg/L and (0.67±0.32) mg/L after twelve weeks, and no statistical difference was found. The levels of CRP and IL-6 decreased in both groups. At the 16th week, the level of CRP decreased significantly compared to baseline in GH group (P < 0.05), but such difference was not found in H gruop. The RKF decreased in both groups, but the significant differences were not found between two groups. At the 16 weeks, the positive rate of gentamicin-fast E. Coli was 23.6% in GH group and 21.4% in H group respectively, with no significant difference. There were no other antibiotic-resistant bacteria or fungus found in stool culture. Conclusions The gentamicin-heparin mixed solution (4 g/L-45 g/L) is a good catheter lock regimen that has better safety and convenience, and its anti-coagulative efficacy is definite. It may be a beneficial means of reducing the CRB rate in hemodialysis patients with cuff-tunneled catheter.

Objective To investigate the effect of N-terminal fragment(LRR-WSC) fusion protein of polycystin-1 (PC-1NF) on collagen Ⅳ synthesis and degradation in rat glomerular mesangial cells and to explore its mechanism. Methods Rat glomerular mesangial cells were treated with different concentrations of PC-1NF in vitro. The mRNA expression of type Ⅳ collagen, metalloproteinase 2(MMP-2) and tissue inhibitor of metalloproteinase 1(TIMP-1) were detected by real-time fluorescence quantitative PCR. The concentration of type Ⅳ collagen was determined by ELISA. The expression level of c-fos and c-jun was examined by immunocytochemistry. The expression of PKC-α was detected by Western blot. Results After treatment with 4 mg/L PC-1NF for 48 hours, the mRNA level of hype IV collagen was down-regulated [（82±11） copies per million GAPDH] compared with that in the control [（103±16）copies per million GAPDH] （P<0.05）, so was the protein level of type Ⅳ collagen. The mRNA level of TIMP-1 was significantly decreased [（3040±370） copies per million GAPDH] compared with that in the control [（5530±480） copies per million GAPDH] （P<0.01）. However, the mRNA level of MMP-2[（2770±170） copies per million GAPDH] was significantly increased compared with that in the control [（1150±90） copies per million GAPDH] （P<0.01）. Meanwhile, PC-1 NF fusion protein treatment resulted in decreased protein levels of c-jun, c-fos and PKC-α. Conclusions PC-1 NF fusion protein may induce collagen Ⅳ degradation through increasing the ratio of MMP-2 to TIMP-1 in rat mesangial cells, inhibit c-jun and c-fos expression by modulating the PKC-α synthesis, and then decrease the production of collagen Ⅳ in rat glomerular mesanglal cells.

Objective To evaluate the effects of angiotensin II (Ang II) infusion on nephrin expression and podocyte apoptosis in vivo, and investigate the mechanism of proteinuria and glomerulosclerosis induced by Ang II infusion. Methods Thirty-six male Sprague-Dawley rats were randomly assigned to receive either normal saline or Ang II (400 ng&#8226;kg-1&#8226;min-1) by osmotic mini-pump, or to be used as normal control. After the systolic blood pressure was measured, the 24 h urine sample was collected for analysis of proteinuria at day 7, 14, 21, 28. Animals were sacrificed at day 14, 28 respectively. Serum creatinine, renal pathological change, podocyte apoptosis were observed. Expression of nephrin mRNA and protein was examined by RT-PCR, Western blot, and immunofluorescence staining. Results (1) Ang II-infused rats developed significant hypertension associated with marked proteinuria. Significantly positive correlation was found between blood pressure and proteinuria (r = 0.80，P < 0.01). (2)At day 14, Ang II infusion induced the narrowing of slit diaphragm. At day 28, apoptotic podocytes were detected by electron microscopy and TUNEL assay [(2.7±1.6) apoptotic podocytes per glomerular cross section] in Ang II-infused rats. (3) Distribution of nephrin expression was changed from linear to granular pattern in Ang II-infused rats at day 14 and expression of nephrin mRNA and protein increased (P < 0.05). But at day 28, expression of nephrin mRNA and protein was decreased, and significantly negative correlation was found between the expression of nephrin and the number of apoptotic podocytes (r = 0.63，P < 0.01). Conclusion Change in nephrin expression may play a critical role in the pathogenesis of Ang II-induced podocyte apoptosis.

Objective To investigate the effects of advanced glycosylation end products (AGE) on DNA damage in cultured normal rat kidney fibroblasts (NRK-49F) cells, and the potential role of oxidative stress in AGEs-induced genotoxicity. Methods NRK-49F cells were treated with DMEM medium containing AGE-BSA (AGE-bovine serum albumin) at various concentrations (100, 200, 400, 800 mg/L) for 24 h. Cells were treated with serum-free DMEM medium or BSA at various concentration (100, 200, 400, 800 mg/L) for 24 h as control. To evaluate the potential role of oxidative stress in AGE-induced DNA damage, cells were preincubated with or without 10 mmol/L N-acetyl-l-cysteine (NAC) for 24 h and then were treated with AGE (400 mg/L) for another 24 h. Cell proliferation was measured by reduction of AlamarBlue. Single-cell gel electrophoresis (comet assay), a well-established method for quantifying DNA damage, was employed to analyze AGE-induced DNA damage. Results Compared with control medium and BSA, treatment with AGE caused significant inhibition of cell proliferation (P < 0.05)and increase of DNA damage (formation of comet) in a dose-dependent manner. Tail length of comet-formation caused by 200 mg/L AGE (P < 0.05) and 400 mg/L, 800 mg/L AGE (P < 0.01) was significantly longer than those of control group and BSA-treated group. Preincubation with antioxidant NAC attenuated AGE-induced DNA damage. Tail length of NAC pretreated group was significantly shorter than that of non-pretreated group (P < 0.05). However, treatment with BSA for 24 h did not show any increase in tail length (P > 0.05) and had no effect on cell proliferation (P > 0.05), compared with control medium. Conclusions AGE can induce DNA damage and inhibit cell proliferation in NRK-49F cells. Antioxidant can attenuate these effects. The enhanced oxidative stress may be one mechanism of underlying AGE-induced genotoxicity in chronic renal failure.

bjective To establish a method culturing myofibroblasts primarily from cortex of rat kidney. Methods Explanting the pieces of cortex of rat kidney into plastic flask, then cells grew out from the tissue pieces. Cells were identified by observation with inverted phase contrast microscope and electron microscope, as well as by phenotypic protein examination via immunochemistry staining and Western blot analysis. Results The cells showed shuttle shape with single nuclear in each one. Expression of vimentin and α-SMA was positive over all of the cells, which confirmed the myofibroblast phenotype. In contrast, expression of epithelial aminopeptidase P, cytokeratin, desmin, which were known as the markers for both endothelial cells and epithelial cells, was negative in these cells. Cultured cells could be propagated to 5 passage and remained the phenotype of myofibroblsts. Connective tissue growth factor(CTGF) and transforming growth factor β1(TGF-β1) could stimulate cell proliferation, but interferon-gamma had no effect on these cells. Conclusion Cultured myofibroblasts from the cortex of rat kidney can be propagated and have proliferative response to CTGF and TGF-β1 .