Objective To analyze the clinicopathological characteristics of systemic lupus erythematosus (SLE) with secondary antiphospholid syndrome (APS) . Methods Data of 11 cases of SLE with secondary APS (SLE with APS) admitted to Peking Union Medical College Hospital from January 2000 to March 2010 were retrospectively analyzed. Kidney biopsy was performed on all of these patients. Differences of clinicopathology and outcomes between SLE with and without APS were compared. Results Renal involvement was found in all the SLE with APS patients. The prominent clinical manifestations included hypertension (54.5%), nephrotic level of proteinuria （24 h proteinuria ≥3.5 g）(72.7%) and renal insufficiency (45.5%). Diastolic blood pressure, mean arterial pressure and glomerular filtration rate in SLE with APS were significantly higher than those in SLE without APS (all P<0.05). In 8 out of 11 cases (72.7%), APS nephropathy (APSN) in kidney biopsy was found, characterized by small vessel vaso-occlusive lesions. These included thrombotic microangiopathy (TMA), fibrous intimal hyperplasia (FIH), focal cortical atrophy (FCA) and tubular thyroidization. Among those, 5 cases (45.5%) had chronic APSN and 4 (36.4%) had acute APSN (one case had acute and chronic APSN at the same time). The incidences of APSN and acute APSN in the SLE with APS group were significantly higher than those in SLE without APS group (P<0.05). Conclusions The major renal manifestations of SLE with APS are hypertension, nephrotic level of proteinuria and renal insufficiency. Other than lupus nephritis, also a high incidence of APSN is found in SLE with APS patients.

Objective To assess the effect of continuous renal-replacement therapy (CRRT) dose on the outcome of acute renal failure (ARF) patients with meta-analysis of randomized controlled trials (RCTs). Methods Studies were identified by systematic search of peer-reviewed publications in Medline, EMBASE and Cochrane library database through June 2010. All the RCTs that compared the incidence of clinical outcome such as mortality, need for chronic dialysis between standard and low dose CRRT were eligible. The pooled relative risk (RR) for clinical outcome was compiled using a random-effects model. Heterogeneity was evaluated by means of subgroup and sensitivity analysis. Results Six eligible studies were identified. By meta-analysis, standard dose CRRT was associated with non-significant 13% mortality risk reduction （RR 0.87, 95%CI 0.70-1.07, P=0.19）and 13% composite outcome risk reduction of chronic dialysis dependence and mortality (RR 0.87, 95%CI 0.69-1.09, P=0.21), but the trend toward increased chronic dialysis dependence risk among survivors (RR 1.43, 95%CI 0.94-2.18, P=0.09). The overall test for heterogeneity among cohort studies was significant (P=0.001, I2=76.2%). The risk of mortality was significantly lower in some studies of which delivered dose was moer than 35 ml&#8226;kg-1&#8226;min-1, modality was continuous venous-venous hemofiltration (CVVH) and major cause was non-sepsis treated with standard dose CRRT. Conclusions Standard dose CRRT in patients with ARF does not improve survival, renal recovery and composite outcome, but decreases mortality in important subgroups including those with higher delivered dose, CVVH and non-sepsis.

Objective To assess the prognosis and effect on renal function of pediatric urolithiasis caused by melamine-contaminated milk powder (PUMMP) in a long-term follow-up. Methods One hundred and two of 8335 children (≤ 6-year-old) with history of consuming melamine-contaminated milk powder screened in our hospital were followed up for eighteen months after diagnosis. Urinary system ultrasonography, urinalysis, urinary microprotein profiles [microalbumin (ALBU), immunoglobulin G (IgG), and N-acetyl-β-D-glucosidase (NAG)], urinary melamine and cyanuric acid were examined in the first visit and at the end of follow-up. Results Follow-up was completed in 91 children and the stone was excreted in 82 children (90.1%). Stones less than 5 mm in diameter were most vulnerable to discharge, and stones larger than 10 mm could not be expelled without interventions. At the end of follow-up, no melamine or cyanuric acid was found in the urine samples of 74 patients. Urinalysis showed that incidences of proteinuria, microscopic hematuria and leukocyturia were 0%, 5.1% and 2.0%, which were significant different from those in the first visit (Pproteinuria=0.123, Phematuria=0.038 and Pleukocyturia=0.005). Urinary microprotein profiles revealed that some children whose urinalysis was normal still presented glomerular and renal tubular injury and the abnormal rates were 8.8% and 12.1% respectively. The glomerular injury was mainly related to persistent stone, male and younger. Conclusions 90.1% of children with PUMMP passes urinary stones at the end of follow-up. Stone size is the major risk factor of discharge. No melamine or cyanuric acid is found in the urine of children. After eighteen months, glomerular and renal tubular injury is still found in some patients. Further follow-up is necessary.

Objective To explore the effect of uric acid (UA) on the rat glomerular mesangial cells(GMCs) and its possible mechanism in vitro. Methods Cultured rat GMCs were treated with various concentrations of UA (50 μmol/L, 100 μmol/L, 300 μmol/L) in the presence or absence of U0126, apocynin, rotenone. The incorporation of 3H-thymidine (3H-TdR) and cell counting were used to measure GMCs proliferation. GMCs cell-cycle was analyzed by flow cytometry. CyclinD1 and cyclin A2 expression were determined by real-time PCR and Western blotting analysis. ERK1/2 phosphorylation was detected by Western blotting. Reactive oxygen species (ROS) was measured by 2, 7-dichlorofluorescein diacetate (DCFDA) fluorescence. Results (1) Uric acid increased GMCs proliferation in dose-dependent manner compared with the control groups, as assessed by 3H-TdR incorporation and cell counting. GMCs proliferation induced by 300 μmol/L uric acid was increased by more than 1.5-fold assessed by both of the methods. (2) Uric acid decreased cell number in G1/G0 phase and increased cell number in S phase in dose-dependent manner, as assessed by flow cytometry. (3) Uric acid iuduced cyclin D1 and cyclin A2 expression in dose-dependent manner. (4)Uric acid increased pospho-ERK1/2 in dose-dependent manner and ERK1/2 specific inhibitor U0126 could suppress uric acid-induced cell proliferation. The inhibition percentage of U0126 was 22% and 31% assassed by cell counting and 3H-TdR incorporation, respectively. (5) Uric acid increased ROS production in dose-dependent manner. NADPH oxidase inhibitor apocynin could also significantly inhibit uric acid-induced ROS production, ERK phosphorylation and cell proliferation. In contrast, rotenone had no effect on them. Conclusion Uric acid can stimulate rat GMCs proliferation, partially by the activation of ERK pathway via NADPH oxidase-derived ROS generation.

Objective To investigate the role of genistein(Gen) in the expression of connective tissue growth factor (CTGF) induced by parathyroid hormone (PTH) in human renal tubular epithelia cells. Methods Real-time PCR, Western blotting and reporter gene assay were employed to detect the role of Gen in PTH-induced CTGF expression in HK-2 cells. The activity of NF-κB was measured by EMSA to investigate the mechanism by which PTH induced CTGF expression in HK-2 cells. Inhibitors of NF-κB signaling pathway were used to ascertain which signal pathway was involved. Results HK-2 cells had basic amount of CTGF mRNA and protein, which, however, increased significantly after treatment with PTH, and the luciferase activity increased to a higher level as compared with control group after treatment with 10-10 mol/L PTH for 12 h (1.89±0.08 vs 0.99±0.03, P＜0.01). Gen decreased the expressions of CTGF mRNA and protein induced by PTH in dose-dependent manner. The NF-κB of nucleus was inactivation without PTH, while the activity of NF-κB significantly increased after exposed to PTH, with the maximal response of PTH at a concentration of 10-10 mol/L and the best stimulating time at 30 minute. The NF-κB inhibitor PDTC reduced the increase of CTGF transcript levels in response to PTH stimulation. Gen blunted PTH-mediated NF-κB activation. Conclusion Gen inhibits CTGF expression induced by PTH through bloking NF-κB signaling pathway in human renal tubular epithelial cells.

Objective To observe the effect of ulinastatin on the expression of interleukin 15(IL-15), connective tissue growth factor (CTGF) and malondialdehyde (MDA) in rat peritoneal mesothelial cells(RPMCs) induced by high glucose. Methods RPMCs were isolated, cultured and passaged by trypsin, then identified. The third generation of cultured RPMCs were used in the experiment. RPMCs were divided into normal control group, high glucose (1.5%, 2.5%, 4.25%) for 6 hours and 12 hours, high glucose (2.5%) for 3, 6, 12, 24 hours or ulinastatin (160, 320, 640 U/ml) for 12 hours. IL-15 mRNA was detected by real-time PCR. IL-15 and CTGF protein in supernatants was detected by ELISA. MDA protein was detected by TBAS. Results Compared with the control group, the expression of IL-15, CTGF and MDA was significantly increased in the groups stimulated by high glucose (P<0.05) in dose- and time-dependent manner. Ulinastatin could significantly decrease the expression of IL-15, CTGF and MDA induced by high glucose in dose-dependent manner both in protein and gene levels (P<0.05). Conclusions High glucose can up-regulate the expression of IL-15, CTGF and MDA in RPMCs. Ulinastatin can reverse these changes.

Objective To investigate the inhibitory effect and mechanism of rosiglitazone on chemokines secretion in renal tubular epithelial cells(HK-2) stimulated by lipopolysaccharide(LPS). Methods Cells were divided into four groups: control (CON), LPS (1 mg/L), rosiglitazone (10 μmol/L), rosiglitazone (10 μmol/L) +LPS (1 mg/L). MCP-1 and IL-8 expression was measured using real time PCR and ELISA. PPARγ was knockdown by RNAi to investigate whether the inhibitory effect of rosiglitazone was PPARγ-dependent or -independent. The NF-κB in nucleus was detected by Western blotting. The DNA binding activity of NF-κB was determined by electrophoretic mobility shift assay. Results Compared with CON group, the expressions of IL-8 and MCP-1 were increased by (4.30±0.45) and (4.80±1.29) times in mRNA level, (1.39±0.18) and (2.11±0.47) times in protein level, respectively, in LPS-stimulated HK-2 cells (P<0.05). Application of rosiglitazone followed by LPS significantly reduced IL-8 and MCP-1 secretion compared with LPS group (decreasing by 66.37% and 71.88% in mRNA levels, while 41.68% and 47.87% in protein levels) (P<0.05). In pcDNATM 6.2-GW/EmGFP-miPPARγ transfected cells, IL-8 and MCP-1 only were decreased by 18.16% and 16.83% in mRNA level, while 11.39% and 11.86%% in protein level in rosiglitazone pretreated group, showing no significant difference compared with LPS group. Rosiglitazone did not block NF-κB nuclear translocation while significantly inhibiting the DNA binding activity of NF-κB. Conclusions Rosiglitazone inhibits the expressions of MCP-1 and IL-8 via a PPARγ-dependent mechanism in HK-2 cells, resulting from inhibition the DNA binding activity of NF-κB.

Objective To observe the impact of different culture conditions on the growth of the endothelial progenitor cells(EPCs) from rat peripheral blood. Methods Mononuclear cells obtained from rat peripheral blood were isolated by using density gradient centrifugation. According to the culture medium added with vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF) and the dishes were precoated with fibronectin or not, the mononuclear cells were divided into different groups and cultured under different conditions in vitro to compare the differences of the growth conditions. The different growth conditions were recorded and the result was calculated by statistics software. At last, the cells were identified by immunohistochemistry and immunofluorescence. Results The mononuclear cells from rat peripheral blood grew in the manner of keeping close to the wall in vitro. The cells and cell colonies cultured for 7 days hinted: under the same culture conditions, precoating FN was beneficial to the adherent proliferation of EPCs (t=4.43, P<0.05; t=3.70, P<0.05). Excluding the impact of fibronectin, growth factors could promote mononuclear cells differentiation to EPCs, indicating that growth factors enhanced proliferation of EPCs(t=-13.22, P<0.01; t=-10.96, P<0.01). Immunohistochemistry and immunofluorescence showed that, at day 4, 7, 10 of cells cultured, CD34 and CD133 expressions were increased gradually [(35.7±4.2)%, (60.1±3.8)%, (81.8±6.4)%; (3.2±0.9)%, (18.4±7.3)%, (32±3.8)%, respectively]; at day 14, both were decreased [(32.1±5.4)%, (1.9±2.7)%]; but Flk-1 was increased at day 4, 7,10, 14[(31.2±3.5)%, (40.6±5.3)%, (71.2±8.4)%, (81.5±4.1)%]. Conclusions Fibronectin is conducive to adhesion and proliferation of EPCs. VEGF and bFGF play an important role in the differentiation of EPCs. The success culture of EPCs in vitro will provide a sufficient number of seed cells for its application in vascular tissue engineering, and offer new ideas for peripheral blood stem cell transplantation for the treatment of various diseases.

Objective To investigate the protective effect and mechanism of pamidronate disodium on calcification in rat vascular smooth muscle cells (RVSMCs) induced by hyperphosphate. Methods RVSMCs were placed in various culture mediam, including normal phosphate medium(Pi 1.4 mmol/L), high phosphate medium(Pi 4.5 mmol/L), different pamidronate disodium concentrations medium (Pi 4.5 mmol/L+pamidronate disodium 10-5, 10-6, 10-7 mmol/L)． Calcium content and cell protein content were quantified by the O-cresolphthalein complexone method and BCA protein assay respectively． Calcification was visualized by von Kossa staining, and cbfα-1, osteocalcin were quantified by Western blotting． Results After culture for 3 days, calcium content in high phosphate group were much higher than that in control group, and pamidronate disodium groups had a lower calcium content compared with high phosphate group (all P<0.05)． Calcium deposit in RVSMCs was greater in high phosphate group, while pamidronate disodium groups revealed obviously decreased deposit (all P<0.05). Protein expression of cbfα-1 and osteocalcin in pamidronate disodium groups was much lower than that in high phosphate group. Conclusion Pamidronate disodium can protect RVSMCs from phosphate-induced calcification in vitro, which may be associated with the blockage of transformation of RVSMCs into osteoblast.