Objective To investigate the prevalence, characteristics and risk factors of superior vena cava and auxiliary branchs thrombosis in hemodialysis patients with internal jugular venous indwelling catheter. Methods A total of 43 cases on hemodialysis (HD) with indwelling short?-erm catheter in internal jugular vein from June to December in 2007 were enrolled in this study. The clinical data and biochemical indicators were collected to investigate the prevalence, characteristics and risk factors of venous thrombosis around indwelling catheter, such as, superior vena cava and auxiliary branchs in these patients. Results Short-term double lumen internal jugular venous catheter were placed in 43 HD patients. Different degrees of central vein thrombosis were found in 21 of the 43 HD patients (48.8%). The ratio of thrombosis in jugular vein, brachiocephalic vein, subclavical vein and uperior vena cava was 100% (21/21), 28.6％ (6/21), 23.8％(5/21) and 19.0％(4/21), respectively. Ten of the 21 HD patients (47.6%) with central vein thrombosis presented clinical symptoms. Five cases developed edema of the upper extremity, 2 cases had new-onset symptom’s pulmonary embolism, and 3 cases developed blood overflowed from inlet port of circum-catheter. The ratio of diabetes mellitus, malignant tumor, the prevalence of increased level of serum lipoprotein a and plasma homocysteic acid were significantly higher in the HD patients with central vein thrombosis than that in those without central vein thrombosis. The odds ratio of diabetes mellitus, malignant tumor, high serum lipoprotein a and high plasma homocysteic acid was 5.758, 4.750, 6.967 and 8.533, respectively. Conclusions The prevalence of central vein thrombosis in HD patients with short-term indwelling catheter in internal jugular vein is quite high. Its clinical symptom is insidious but dangerous. Diabetes mellitus, malignant tumor, high serum lipoprotein a and high plasma homocysteic acid may be the important risk factors of central vein thrombosis in above HD patients.

Objective To explore the risk factors of invasive fungal infection after kidney transplantation and to evaluate their effect on prognosis. Methods Data of 2573 patients of kidney transplantation in our center from Jan 1994 to May 2008 were analyzed retrospectively. Patients were divided into case group and control group according to fungal infection after operation. Differences of age, preoperative conditions, complications after operation, drainage time, application of broad-spectrum antibiotics, and use of anti-rejection drugs were compared between these two groups to identify the risk factors of postoperative fungal infection. The impact of risk factor amount on the incidence and mortality of invasive fungal infection, as well as on the mortality of patients and graft loss rate was analyzed. Results Compared with control group, the number of aged patients elevated significantly, as well as the incidence of delayed graft function (DGF), acute rejection, CMV infection, liver function impairment, delayed incision healing, and myelosuppression went up significantly in case group. The incidence of long drainage time (>1 week), using broad-spectrum antibiotics (>1 week) and anti-rejection drugs was also increased in case group (P<0.01) . Multivariate Logistic regression showed that aging (≥60 years）, DGF, delayed incision healing, myelosurppression, and using broad-spectrum antibiotics(>1 week) were independent risk factors for invasive fungal infection. With the risk factor number increasing, the incidence and mortality increased significantly (χ2=91.2 and 18.1，respectively, P<0.01), the graft loss rate also increased significantly (χ2=93.0, P<0.01). Conclusion Evaluaton of risk factors and prevention of fungal infection after kidney transplantation are very important for improving the prognosis.

Objective To evaluate the effect of continuous renal replacement therapy (CRRT) on the remove of inflammatory mediators and the function of endothelial cells in patients with multiple organ dysfunction syndrome (MODS). Methods Thirty patients with MODS were enrolled in this study. All of the patients underwent CRRT for at least 24 hours. Peripheral blood levels of IL-1β, IL-4, IL-6, IL-10, TNF-α, E-selectin, sVCAM-1, sICAM-1 and PAF-AH were measured at the beginning and 3, 6, 12, 24 h after initiation of CRRT. Results Nineteen patients survived after 14 days and 17 patients survived after 28 days during therapy. The clinical oxygenation and hemodynamics were improved after 6 h of CRRT. Among inflammatory mediators, the levels of TNF-α, IL-6, IL-10 rose gradually from the beginning [(462.24±331.03) ng/L, (106.39±90.82) ng/L, (124.51±118.39) ng/L), and reached the peak at 12 h [(887.88±975.46) ng/L, (132.01±118.14) ng/L, (167.01±161.66) ng/L], and the levels of IL-1β, IL-4 decreased from initiation of CRRT. But there were no significant differences in the levels of above cytokines between at the beginning and at the end of CRRT. There were significant differences in the levels of cytokines between survival and death group. The level of IL-6 in death group [（145.45±14.28） ng/L] was significantly higher than that in survival group [（106.03±10.86） ng/L]. The level of IL-10 in death group [（94.93±16.09） ng/L] was significantly lower than that in survival group [（143.06±12.24） ng/L]. Levels of E-select, sVCAM-1 and sICAM-1 elevated from the beginning and reached the peak at 12 h, but no significant differences were found between intiation and the end of CRRT. The level of PAF-AH increased after initiation, and there was a significant difference between beginning and the end of CRRT. Levels of cytokines for endothelial cell function were significantly different, such as E-selectin [（287.13±42.70） μg/L vs (266.26±65.26) μg/L], sVCAM-1 [（1697.25±475.24） μg/L vs (1488.10±691.67) μg/L], sICAM-1 [（975.33±142.50） μg/L vs (835.40±332.41) μg/L], and PAF-AH [（9.07±6.38） μg/L vs (16.32±8.95) μg/L]. Conclusions Clinical oxygenation and hemodynamics can be improved, and endothelial cell function can be improved partly by CRRT. There were no significant differences of inflammatory mediator levels between initiation and the end of CRRT. IL-6 and IL-10 can be used as predicators for prognosis of MODS patients.

Objective To assess the safety and efficacy of mycophenolate mofetil （MMF） combined with low dose corticosteroid in the treatment of adults with minimal change nephrotic syndrome and concomitant HBsAg positive (MCNS-HBsAg). Methods Thirty adults with MCNS-HBsAg were enrolled in this prospective study and were assigned to two groups. The Pred group (n=16) received conventional prednisone regimen (prednisone 1 mg&#8226;kg-1&#8226;d-1) and the MMF group (n=14) received low dose of prednisone combined with MMF (MMF 1.0 to 2.0 g/d plus prednisone 0.5 mg&#8226;kg-1&#8226;d-1). Results Hepatitis B virus (HBV) was replicated in 62.5% patients of Pred group versus 35.7% patients of MMF group. 43.8% patients of Pred group versus 21.4% patients of MMF group received lamivudine therapy. Elevation of alanine aminotransferase (ALT) occurred in 50% patients of Pred group and 28.6% patients of MMF group. The complete remission (CR) rate after 24 weeks treatment was 11/14 in Pred group versus 10/12 in MMF group. 6/11 patients of the Pred group and 4/10 patients of the MMF group who achieved CR experienced relapses during follow-up. Conclusions Use of MMF combined with low dose prednisone is as effective as conventional prednisone regimen in treating adults with MCNS-HBsAg. The MMF protocol seems to be superior in HBV reactivation to conventional prednisone protocol.

Objective To elucidate the prevalence and risk factors of cardiovascular disease(CVD) in end-stage renal disease (ESRD) patients on peritoneal dialysis (PD), and to investigate the associated problems in treatment. Methods A total of 254 PD patients in our division were enrolled in this study. CVD history, laboratory measurements, examinations of carotid atherosclerosis and left ventricular hypertrophy by ultrasonography were collected and associated factors were analyzed. The median follow-up time was 49 months. Results The overall prevalence of CVD was 37% (93/254). Diabetes, longer dialysis duration, hypertriglyceridemia, hypoalbuminemia, hypoprealbuminemia were commonly found in the patients with new CVD event. The patients without pre-existing CVD had the higher Ccr, Kt/V, D/Pr, nPCR, serum albumin level. In those with pre-existing CVD, the hypertriglyceridemia and the duration of dialysis were independent predictors of progression of CVD. Differences of LAD, LVST, LVMI and IMT were significant between with and without pre-existing CVD groups. Kaplan-Meier curves showed that the presence of CVD was the independent risk factor of survival. Alb<330 g/L, LAD>39.6 mm and peritonitis were risk factors of CVD. Conclusion The prevalence of CVD in PD patients is quite high. CVD history should be realized, dialysis adequacy should be maintained, and peritonitis should be prevented.

Objective To test the hypothesis that point mutations in the B1 subunit of vacuolar H+-ATPase, which cause inherited type I (or distal) renal tubular acidosis (dRTA), interfere with assembly and proton secretion function of the H+-ATPase. Methods Eight constructs that mimic seven known point mutations in inherited dRTA (M) or wild type (WT) B1 were transfected into a rat inner medullary collecting duct (IMCD) cell line to express GFP-B1 WT or GFP-B1 M fusion proteins. Distributions of GFP-B1 WT or M, and its assembly ability with other subunits (E, H and c), as well as its effect on the ATP hydrolysis activity and proton secretion function of H+-ATPase were tested by immunofluorescent methods, immunoprecipitates, ATP/NADH coupled assay or Na+-independent pHi recovery following acute acid load respectively. Results Immunofluorescence revealed that GFP-B1 WT displayed the same typical vesicular distribution pattern as H+-ATPase, but all GFP-B1 M exhibited more diffused cytoplasm pattern. In co-immunoprecipitation studies, GFP-B1 WT formed complexes with other H+-ATPase subunits (c, H and E) whereas GFP-B1 M did not. Proteins immunoprecipitated with anti-GFP antibody from GFP-B1 WT cells had ATPase activity whereas proteins from GFP-B1 M cells did not. Proton pump-mediated pHi transport was significantly inhibited in GFP-B1 M transfected cells [pHi recovery rate (pH U/min) was 0.007±0.002, 0.004±0.002, 0.002±0.002, 0.003±0.002, 0.006±0.004, 0.009±0.004, 0.015±0.006 in L81P, R124W, M174R, P275R, G316E, P346R, G364S B1 M-transfected IMCD cells, P<0.05, n=5]. However, pHi recovery rates in both GFP-B1 WT cells and the untransfected IMCD cells were similar[(0.040±0.006) pH U/min], and were abolished by 1 μmol/L bafilomycin (specific H+-ATPase inhibitor）. Conclusion B1 point mutations that produce dRTA prevent normal assembly of the H+-ATPase B1 and affect the proton secretion of H+-ATPase in IMCD cells.

Objective To investigate the influence of epigallocatechin gallate（EGCG）on reactive oxygen species(ROS) in mouse podocytes exposed to high glucose. Methods Mouse podocytes cultured in high glucose were exposed to different concentrations of EGCG (0.2, 10, 100 μmol/L) or α-tocopherol(0.2 μmol/L) for 6, 12, 24 hours. The viability of podocytes was detected by MTT. The intracellular formation of ROS was detected by confocal microscopy with fluorescent probe CM-H2DCFDA and was measured by fluorescence microscopy. RT-PCR was used to examine the expression of p22phox, p47phox and p67phox mRNA in cultured podocytes exposed to different concentrations of EGCG. Results Intracellular ROS generation was significantly higher in high glucose than that in control conditions (P<0.01). EGCG could significantly inhibit ROS induced by high glucose significantly(P<0.01). EGCG(100 μmol/L) led to an inhibition of the increased production of NADPH oxidase components of p22phox and p67phox mRNA in high glucose(P<0.05). The expression of p47phox mRNA in high glucose was inhibited by EGCG(0.2 μmol/L) and α-tocopherol(0.2 μmol/L) (P<0.05). Conclusion EGCG can protect cultured mouse podocytes from injury of high glucose by inhibiting ROS formation.

Objective To study the impact and mechanism of continuous venovenous hemofiltration (CVVH) in different ultrafiltration rates on plasma cytokines in porcine endotoxemic shock. Methods Eighteen anesthetized mechanically ventilated pigs weighing 21－34 kg were randomly divided into three groups. In control group (n=6), the pigs received a 15.7 μg/kg endotoxin (E.coli O111:B4) infusion. In CVVH group (n=6) and high volume hemofiltration (HVHF) group (n=6), the pigs received CVVH after the endotoxin infusion for 24 hours with an ultrafiltration rate of 45 ml·kg-1·h-1 and an ultrafiltration rate of 70 ml·kg-1·h-1 respectively. Blood was taken before endotoxin infusion and at 0, 1, 6, 12, 24 h during CVVH. The plasma levels of TNF-α, IL-6, IL-10 and IL-18 were tested by ELISA. Results The survival time in control group was (15.4±5.2) h，CVVH group was (21.4±7.1) h，HVHF group was (22.4±6.7) h. The survival time in CVVH and HVHF group was significantly longer than that of control group（P<0.05）. Heart rate(HR), mean arterial blood pressure(MAP), central venous pressure(CVP) and cardiac output(CO) showed no significant differences among three groups. Plasma BUN and Scr increased gradually after the establishment of porcine endotoxemic shock model. BUN and Scr of CVVH and HVHF group were lower compared to control group（P<0.05）, but there was no significant difference between CVVH and HVHF group（P>0.05）. Plasma TNF-α and IL-6 peaked at T1, IL-10 peaked at T0, then they declined gradually. While IL-18 increased at T0 and did not change after T0. A significant decrease of plasma IL-10 level was observed at T6, T12 and T24 in CVVH group compared with control group (P<0.05). HVHF group accomplished a greater decrease in plasma TNF-α (T6) and IL-10 (T6, T12, T24) levels compared with control group and CVVH group（P< 0.05）. The levels of IL-6 and IL-18 showed no significant differences among three groups. There was a negative correlation between IL-6 and survival time (P<0.05). Conclusions HVHF and CVVH can prolong the survival time of porcine endotoxemic shock. IL-10 can be removed effectively with CVVH and HVHF. HVHF can also remove TNF-α effectively. CVVH and HVHF treatment can both remove BUN and Scr effectively. IL-6 is a powerful independent predictive factor for survival time of porcine endotoxemic shock.

Objective To observe the effects of a new immunosuppressive agent, FTY720, on rat glomerular mesangial cell (GMC) apoptosis and on gene expression profiles of cell cycle regulatory proteins. Methods Rat GMCs were cultured with 20 μmol/L FTY720 for 6 h, 12 h, 24 h and 48 h, and then were evaluated for proliferation through MTT method, and for apoptosis by flow cytometry and fluorescence stainig with Hoechst33258 and PI, and DNA fragmentation analysis. The gene expression profile of cell cycle regulatory proteins was characterized in rat GMCs before and after FTY720 treatment by SuperArray real-time PCR microarray analysis. Results After incubation with FTY720 for 6 h, apoptotic sub-G1 peak was identified in GMC through flow cytometry. After incubation with FTY720 for 12 h, not only apoptosis bodies of GMC were observed by fluorescence staining with Hoechst33258 and PI, but also typical morphological changes of apoptosis were found in GMC. After incubation with FTY720 for 24 h, typical DNA ladder pattern was identified. The percentage of FTY720-induced GMC apoptosis gradually increased with the extension of incubation time. SuperArray real-time PCR microarray analysis revealed that FTY720 could respectively up-regulate the expression of Dnajc2, LOC688900 and RGD1562436_predicted genes to 41.6, 38 and 16 folds. Conclusion FTY720 can induce GMC apoptosis in a time-dependent manner, probably through influencing gene expression of cell cycle regulatory proteins.

Objective To investigate the inhibitory effects of rosiglitazone on the synthesis of reactive oxygen species ( ROS) and the expression of monocyte chemoattractant protein 1 (MCP-1) induced by high glucose in rat mesangial cells. Methods The mesangial cells were divided into six groups: control group( C, 5.6 mmol/L glucose), mannitol group (M, 24.2 mmol/L mannitol+group C), high glucose group( H, 30 mmol/L glucose), R1 group（R1, group H＋10 μmol/L rosiglitazone）, R2 group(R2, group H＋20 μmol/L rosiglitazone), N-acetylcysteine (NAC) group (N, group H＋5 mmol/L NAC, NAC was added 1 h before the stimulation of high glucose). The level of ROS was measured by confocal laser scanning microscopy. The mRNA and the protein expression of MCP-1 were semi-quantitatively determined with reverse transcription-polymerase chain reaction and ELISA respectively. Results No significant differences of ROS and MCP-1 were found between control group and mannitol group. The intracellular ROS induced by high glucose increased by 4.1-fold compared to control group（P<0.01）, which was prevented by rosiglitazone (20 μmol/L) and NAC respectively. The MCP-1 mRNA expression in group R2 and group N was significantly lower than that in group H（P<0.01）. The MCP-1 protein level in group H [(940.9±20.3) ng/L] was higher than that in group C [(403.0±8.1) ng/L] （P<0.01）, and the expression of MCP-1 protein in group R2 [(562.5±15.3) ng/L] and group N [(539.8±8.3) ng/L] was lower than that in group H（P<0.01）. Conclusion Rosiglitazone may suppress high glucose-induced MCP-1 expression by reducing the level of ROS, which may be one of the mechanisms that rosiglitazone plays a direct role in the protection of kidney.

Objective To evaluate the effect of sevelamer hydrochloride on parameters of coronary artery calcification(CAC), mineral metabolism and lipid profile in maintenance hemodialysis (MHD) patients. Methods Medline, CENTRAL and Chinese biomedical database were retrieved by using the key words "sevelamer or Renagel" so as to search the materials about the randomized controlled clinical trials that had compared the effects of sevelamer and calcium-based phosphate binders(CBPB) on cardiovascular calcification in MHD patients. A meta-analysis was conducted. Results Five documents about randomized controlled clinical trials, including 697 patients, from the retrieved 276 documents according to the demand of enrollment. Compared with CBPB, there was a significantly lower coronary artery calcification score in MHD patients treated with sevelamer (weighted mean difference -66.84, 95％CI -126.90 to -6.77). The funnel plot test regarding CAC score did not indicate the existence of publication bias. The multiple mortality of sevelamer group was 4.2％, not significantly different from that of CBPB group 5.6％ (RR=0.76, 95％CI 0.37 to 1.57, P=0.45). However, the hospitalization rate of sevelamer group was lower(RR=0.75, 95％CI 0.59 to 0.95, P=0.02). Sensitive analysis confirmed the nonexistence of differences in CAC score and hospitalization rate between these two groups. Conclusions Sevelamer improves the CAC score of MHD patients compared with CBPB. Treatment with sevelamer does not affect overall mortality, but there is evidence for beneficial effect on multiple all-cause hospitalizations.

Objective To elucidate the prevalence and risk factors of acute kidney injury (AKI) within the post-operative 100 days in adult patients with nonmyeloablative hematopoietic stem cell transplantation (HSCT), and whether AKI influences patients’ survival. Methods Sixty-two adult leukemia patients from three transplant centers in Jiangsu province were treated with similar protocols of nonmyeloablative HSCT. AKI was classified as follows: Grade 0, no AKI; Grade 1, renal dysfunction, Scr increased ≥26.5 μmol/L or increased by 50% to 200% (0.5- to 2-fold) from baseline; Grade 2, Scr increased by 200% to 300% (2- to 3-fold) from baseline; Grade 3, Scr increased >300% (>3-fold) from baseline, or Scr ≥ 353.6 μmol/L with an acute increase of at least 44.2 μmol/L. Results 29% (18/62) of the patients developed AKI within 100 days after nonmyeloablative HSCT. Risk factors of AKI were incomplete HLA-matched transplantation [odds ratio (OR) 3.6, 95% confidence interval (CI) 1.1-13.0]. The complications, including sepsis, veno-occlusive disease of liver and acute graft-versus-host, were also associated with the development of AKI (OR 12.1, 95% CI 2.4-62.4). The overall one-year mortality of the patients was 27.4%. AKI was significantly associated with the mortality (log-rank test, P<0.01). Conclusions AKI is a very common complication in the patients with nonmyeloablative HSCT. It is associated with the incomplete HLA-matched transplantation and complications and has an important impact on the patients’ first year survival.

Objective To investigate the association of transforming growth factor β1(TGF-β1) gene -509C/T polymorphism with the susceptibility and the tubulointerstitial damage (TID) degree of primary nephrotic syndrome (PNS) in Han nationality of Chinese population in Hunan province. Methods Ninety-eight PNS patients and 128 healthy controls were enrolled in the study. The TGF-β1 gene-509C/T polymorphism in the 5’-flanking region was detected with the polymerase chain reaction- restriction fragment length polymorphism (PCR-RFLP) technique, and the serum level of TGF-β1 was determined by enzyme-linked immunosorbent assay (ELISA). The association of TGF-β1 gene －509C/T polymorphism with susceptibility and TID degree of PNS was examined. Results (1) There were 3 genotypes (CC, CT, TT) and 2 allele genes (C, T) of TGF-β1 gene -509 position in PNS and healthy control group. (2) There were no significant differences of genotypes frequency or allele frequency of -509C/T polymorphism in TGF-β1 gene between PNS and healthy control subjects. (3) Significant differences of genotype frequency and allele gene frequency were found among severe TID group of PNS, mild TID group of PNS and healthy control group (all P<0.01）. T allele gene frequency and TT genotype frequency of individuals in severe TID group of PNS were significantly higher than those of other two groups (all P<0.01), while no significant difference was found between mild TID group of PNS and healthy control group. (4) With the development of TID in PNS，the serum TGF-β1 level increased. The serum TGF-β1 level was significantly different among severe TID group of PNS, mild TID group of PNS and healthy control group (all P<0.05), and the serum TGF-β1 level in the individuals with TT genotype was higher than that in those with CC and CT genotype. Conclusions TGF-β1 gene-509C/T polymorphism is not associated with the morbidity of PNS, but associated with the severe degree of TID in PNS. T allele gene may be the important susceptible factor. In addition, the increasing serum TGF-β1 level is associated with the severe degree of TID and TT genotype.

Objective To explore the relationship between the expression of telomerase reverse transcriptase (TERT) and the proliferation of endothelial progenitor cells (EPCs). Methods The bone marrow-derived EPCs form SD rats were cultured in vitro. At the end of week 1, 2, 3 and 4 of culture, MTT assay was used to detect the EPCs proliferation rate in the growth duration; Annexin-V-FITC/PI asaay was applied to examine the apoptosis rate in early stage of EPCs; RT-PCR, immunocytochemistry and Western blotting were employed to detect the TERT mRNA and protein expression. Results The mononuclear cells from rats bone marrow could be induced into endothelial progenitor cells in vitro. The proliferation rate of EPCs from different culture duration took on a singlet curve, with the peak at day 14 (P< 0.01). The apoptosis rate in EPCs was 0.28%, 0.66%, 1.38%, 1.52% respectively at week 1 to 4, increasing along with the growth duration within 28 days. Aging rate of EPCs was 3.04%, 20.28%, 24.36%, 16.52% respectively at the end of 1th, 2th, 3th and 4th weeks, which significantly increased at day 14 (P<0.01) and was highest at day 21, but no significant difference was found. The TERT mRNA and protein expression took on a single curve with its peak at day 14 as well, then reduced with the culture time. Conclusion The down-regulated TERT expression may contribute to the increasing apoptosis rate in early stage and the decreasing proliferaton rate of rat bone marrow-derived EPCs.

Objective To explore the effect of curcumin (Cur) on oxidative stress-induced NRK-52E cells injury. Methods NRK-52E cells were treated with H2O2 at different concentrations as an oxidative stress-induced injury model. Nucleus changes in apoptotic cells were investigated by using Hoechst 33258 staining and photofluorography. Apoptotic rate was evaluated by propidium iodide (PI) staining and flow cytometer (FCM). The expression of Bcl-2 was detected by Western blot assay. Results Apoptosis rate in NRK-52E cells was dose-dependently increased by H2O2 treatment at the concentrations from 100 to 500 μmol/L for 24 h. Expression of Bcl-2 in NRK-52E cells was obviously inhibited by exposure to 500 μmol/L H2O2 (P<0.05). Curcumin, at concentrations of 20 μmol/L and 40 μmol/L, not only decreased an elevated apoptotic rate caused by H2O2[(32.9±8.1)%, (22.23±9.3)% vs (72.7±10.5)%, P<0.05], but also blocked the inhibition of Bcl-2 expression induced by H2O2(P<0.05). Curcumin treatment alone led to an up-regulation of Bcl-2 expression(P<0.05). Conclusions Curcumin significantly protects NRK-52 cells against oxidative stress-induced apoptosis. The cytoprotection may be associated with the inhibition of down-regulation of Bcl-2 expression evoked by H2O2.

Objective To investigate the amelioration mechanism of renal dysfunction by Celecoxib(CXB) through the observation of CXB on Han: SPRD rats with ADPKD. Methods Fifty-seven 3-week-old male Han:SPRD heterozygous(Cy/+) rats were randomly divided into 3 groups(n=19). One group was fed with normal forage(control group), another two groups were fed with low dosage (3 mg&#8226;kg-1&#8226;d-1) and high dosage(10 mg&#8226;kg-1&#8226;d-1) administration of CXB respectively. The BUN and Scr were determined respectively at 3, 5, 7, 9, 12 and 16 weeks. The content of 6-keto-PGF-1α and TXB2 was measured by enzyme-linked immunosorbent assay (ELISA). The expression of TNF-α mRNA was detected by real-time RT-PCR assay. The co-expression of TNF-α and COX-2 was examined by double immunofluorescence labeling technique and laser scanning confocal microscopy. The expression of TNF-α protein was detected by Western blotting. Results BUN and Scr increased continuously in control group and exceeded the normal at 6-week-old and at 8-week-old respectively. At 16-week-old, BUN and Scr were (26.56±9.19) mmol/L and (95.08±67.54) μmol/L. After being treated with CXB, the progression of BUN and Scr was reduced in both low and high dosage group. The content of 6-keto-PGF-1α and TXB2 in low[(1831.68±233.31) ng/L and (156.62±9.29) ng/L] and high dosage group [(1148.57±105.80) ng/L and (157.87±10.16）ng/L] was significantly lower than those in control group [(2792.26±830.48) ng/L and (248.88±93.72) ng/L]. TNF-α mRNA levels in low[(2.52±0.01)×103] and high dosage group[(2.48±0.02)×103] were both decreased as compared to control group[(6.17±0.19)×103]. The co-expression of TNF-α and COX-2 distributed widely in tubulointerstitial area in control group and only few in low dosage group. More TNF-α protein in CXB-treated group was detected than that in control group. Conclusion CXB can slow down disease progression in Han: SPRD rats with renal dysfunction through anti-inflammatory effect, including inhibition of COX-2 activity, blockage of 6-keto-PGF-1α and TXB 2 release, and down-regulation of COX-2 over expression by positive feedback.

Objective To investigate the expression of focal adhesion kinase(FAK) in epithelial-mesenchymal transition(EMT) of TGF-β1-stimulated HK-2 cells and the effect of FAK knockdown by small interfering RNA on EMT. Methods HK-2 cells were grown in DMEM-F12 medium supplemented by 10% fetal bovine serum(FBS). HK-2 cells were cultured in free serum medium for 24 h, then were stimulated by TGF-β1(10 μg/L). The expression of E-cadherin, α-SMA, FAK mRNA and protein were detected by RT-PCR, Western blot and immunofluorescence, respectively. The expression level of phosphorylated FAK-Tyr397 was detected by Western blot. HK-2 cells were transfected with 200 nmol/L FAK-siRNA or negative control siRNA using Lipofectamine 2000. Then the expression of E-cadherin, α-SMA, FAK protein was detected by Western blot. Results The expression of E-cadherin mRNA and protein was markedly decreased in HK-2 cells induced by TGF-β1, and the expression of α-SMA mRNA and protein was dramatically increased. Western blot analysis demonstrated that then protein levels of FAK and p-FAK(Tyr397) were progressively increased in a time-dependent manner in response to TGF-β1 treatment in HK-2 cells. When transfected with FAK-siRNA, the FAK mRNA and protein expression was markedly inhibited with 50% and 41%. Knockdown expression of FAK led to a severe blockage of TGF-β1-induced E-cadherin suppression and α-SMA induction. Conclusions The expression of FAK is up-regulated in HK-2 cells stimulated by TGF-β1. But the EMT induced by TGF-β1 in HK-2 cells is inhibited by FAK knockdown, which suggests that FAK plays an important role in TGF-β1-induced tubular EMT and renal fibrosis.

Objective To investigate the effect of PTH on the expression of CTGF in human renal proximal tubular epithelial cell line HK-2 and the role of nuclear factor kappa-B (NF-κB) signaling pathway. Methods The expression of CTGF mRNA and protein in HK-2 cells was measured by real time PCR and Western blot, respectively. The activity of NF-κB in HK-2 cells was measured by EMSA to investigate the mechanism by which PTH induced CTGF expression. The signaling pathway through which PTH produced biological effect was determined by using NF-κB signaling inhibitor PDTC. To assess the potential role of NF-κB activation in PTH-induced CTGF expression, HK-2 cells were transfected with CTGF luciferase reporter gene in the presence or absence of the PDTC added 12 h before PTH. Results A basal level of CTGF mRNA and protein expression was detected in HK-2 cells, with the maximal response to PTH at concentration of 10－10 mol/L and the best stimulating time at 72 hours. After exposure to PTH (10－10 mol/L) for 12 hours, the maximal level of luciferase activity was 1.96-fold of control. The NF-κB of nucleus was inactivated without PTH, while the activity of NF-κB significantly increased after exposed to PTH, with the maximal response to PTH at 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. Similarly, CTGF mRNA and protein expression were decreased in PDTC-treated cells as compared with the untreated control cells(mRNA 0.33±0.05 vs 3.84±0.68, P＜0.05; protein 0.56±0.23 vs 3.76±0.54, P＜0.01). Conclusions PTH induces the expression of CTGF in HK-2 cells. This induction is dependent on NF-κB signaling pathway.

Objective To observe the expression and distribution of PTEN in renal epithelial-mesenchymal transition(EMT), and to investigate the effect of DJ-1 up-regulation on the expression and distribution of PTEN and the activation of PI3K-Akt signal pathway. Methods Human tubular epithelial cells (HKC cell line) were cultured with 10 μg/L TGF-β1 for 72 h. Protein expressions of PTEN, E-cadherin and α-SMA were measured by Western blot. RT-PCR was used to detect the expression of PTEN mRNA. To over express DJ-1, HKC cells were transfected with pEGFP-N1-DJ-1 via lipofectamine 2000 to induce up-regulation of DJ-1. The efficiency of transfection was examined by fluorescence microscope and Western blot. The expressions of DJ-1, PTEN, p-Akt and Akt in the transfected cells were detected by Western blot, and PTEN mRNA was detected by RT-PCR. The intracellular distribution of PTEN in normal HKC cells, cells stimulated by TGF-β1 and cells transfected with pEGFP-N1-DJ-1 was observed by confocal microscope. Results Normal HKC cells expressed PTEN, E-cadherin, but almost did not express α-SMA. The expressions of PTEN protein, PTEN mRNA and E-cadherin in cells stimulated by TGF-β1 were less as compared to normal cells (P<0.05), while α-SMA expression was increased (P<0.05). The effenciency of green fluorescence was more than 80% in transfected cells. In the DJ-1 transfectants, the expressions of PTEN and PTEN mRNA were suppressed, but p-Akt expression was up-regulated as compared to normal cells. In normal HKC cells, PTEN distributed both in cytoplasm and nucleus. In cells stimulated by TGF-β1, cytoplasmic PTEN was completely lost while nuclear PTEN was increased slightly. In the DJ-1 transfectants, only the nuclear PTEN was observed which was similar to the cells stimulated by TGF-β1. Conclusion In renal interstitial fibrosis, the over-expression of DJ-1 can suppress PTEN expression and activate PI3K-Akt signal pathway.

Objective To observe the effects of megsin gene transfection on mesangial cell proliferation and expression of matrix metalloproteinase-2(MMP-2), tissue inhibitor of metalloproteinase-2（TIMP-2） and type Ⅳ collagen under high glucose concentration , and to investigate the relationship between megsin and mesangial cell proliferation and extracellular matrix metabolism. Methods Mouse glomerular mesangial cells were cultured in high glucose medium, then cell proliferation was measured by MTT at 12, 24 and 48 h respectively after culture. Protein levels of megsin, MMP-2, TIMP-2 in mesangial cells were detected by Western blot. Concentration of type Ⅳcollagen in supernatant of mesangial cells was measured by radioimmunochemistry. Results Under high glucose concentration, megsin and TIMP-2 expression were increased, MMP-2 expression was decreased, the concentration of type Ⅳ collagen in the cellular supernatant was increased, and mesangial cell proliferation was enhanced. After megsin gene transfection, the above changes were more significant. Conclusion Megsin gene induces mesangial cell proliferation and inhibits matrix degradation through TIMP-2 up-regulation and MMP-2 down-regulation, which is probably one of the mechanisms of accelerating glomerulosclerosis.