Objective     To explore the genetic association of C1GALT1 gene polymorphisms with susceptibility and prognosis of IgA nephropathy (IgAN) on Uyghur population in Xinjiang Uyghur Autonomous Region.  Methods    Ninety Uighur patients with IgAN and ninety geographically and age matched healthy controls were recruited. Peripheral blood was collected from recruited individuals for DNA extracting. After amplified by polymerase chain reaction (PCR), genotyping of the four single nucleotide polymorphisms (SNPs) in C1GALT1 gene, which were rs9639031, rs5882115, rs1008898, -527A/G, were detected by direct sequencing analysis. Differences of allele and genotype frequency were analyzed between IgAN and healthy controls. Moreover, the association between these SNPs and the risk and progress in IgAN patients were further analyzed.  Results    (1) The I allele frequency of rs5882115 in C1GALT1 gene was significantly higher in IgAN than that in healthy controls (χ2＝7.788, P＝0.015), no difference in allele frequencies of rs9639031, rs1008898, -527A/G between IgAN and healthy controls was found. Under the dominant mode, the DI+Ⅱ genotype frequencies of rs5882115 was significantly higher in IgAN than that in healthy controls (χ2＝8.563, P＝0.009), no difference in genotype frequencies of rs9639031, rs1008898, -527A/G between IgAN and healthy controls was found. Under the hidden mode, no difference in genotype frequencies of rs5882115, rs9639031, rs1008898, -527A/G between IgAN and healthy controls was found. Logistic single factor regression analysis showed that the risk to IgAN of whom carry I allele of rs5882115 was 2.469 times than the D allele (OR＝2.469), and the risk to IgAN of whom carry DI+Ⅱ genotype was also higher (OR＝2.852). (3) No association was found between these SNPs in C1GALT1 gene and serum creatinine in IgAN.  Conclusion    Association between rs5882115 in C1GALT1 gene and Uighur IgAN susceptibility suggests that there may be variants in C1GALT1 gene or its linked genetic region, which needs further exploration.

Objective     To explore the significance of peptidylarginine deiminase type 4 (PAD4) in the pathogenesis of ANCA-associated vasculitis (AAV) by detecting its level in patients with AAV.  Methods     Sera from 13 patients with AAV, 11 patients with primary chronic kidney disease and 12 healthy controls were collected. Serum PAD4 was detected using commercial ELISA kits. The association between serum PAD4 and BVAS of AAV was further investigated.  Results     (1) The serum level of PAD4 in patients with AAV in active and remission stages were all higher than that in the healthy controls. The serum level of PAD4 in patients with CKD was not found elevated compared with the normal controls. (2) The serum levels of PAD4 in AAV with renal damage were all significantly higher than that in CKD group no matter in active or remission stage. (3) The serum level of PAD4 in AAV with renal damage in active stage was positively correlated with BVAS (r＝0.71, P＝0.02).  Conclusion     PAD4 is involved in the pathogenesis of AAV.

Objective     To analyze the baseline clinical characteristics of the chronic kidney disease (CKD) in aged people in the clinic.  Methods     Patients aged 18 years or older in our CKD clinic from October 2003 to December 2012 were included in this study. According to their age patients were divided into 2 groups: aged CKD group: aged 65 or older and non-aged CKD group: younger than 65. A group of the elderly without CKD from health screening program were selected as aged non-CKD control group. Causes, distributions of stages and complications of CKD in three groups were analyzed.  Results    The major cause of the elderly CKD was hypertension, different from that of younger CKD . The distribution of CKD stage in the elderly was mainly in the G3b stage, different from that in the younger. Anemia and mineral bone disease presented in earlier CKD stage in the aged CKD patients and the prevalence was higher than in the aged non-CKD group. The prevalence of hypertension had no statistical difference between the two CKD groups, but hypertension control rate was lower in aged CKD patients. Conclusions    The clinical characteristics including causes and renal stage are different between the young and aged CKD patients. Complications such as anemia and mineral bone disease presents in earlier renal stage in aged CKD patients which means we should monitor and interfere earlier.

Objective    To explore the effects of low-protein diet supplemented with ketoacids on podocytes as well as local RAS in the kidney of patients with diabetic nephropathy. Methods    A tal of 61 patients with T2DN and CKD stages 3-4 were included. All the patients were randomly divided into two groups: low protein group (0.6 g•kg BW-1•d-1 and 30 kcal•kg BW-1•d-1, LPD) and LPD +Ketoacids (KA) group (0.6 g•kg BW-1•d-1, 30 kcal kg BW-1•d-1, and Ketoacids 100 mg•kg BW-1• d-1). Blood and 24 h urine samples were collected at baseline and every 3 months for routine examination to evaluate the efficacy of LPD + KA diet. Podocytes loss was evaluated by mRNA expression of nephrin, podocin, and synaptopodin in urine at baseline and every 3 months. Urinary angiotensinogen was detected by ELISA at baseline and every 3 months.  Results    After 12 months of follow-up, there were no significant difference statistically in declines of GFR between LPD and LPD+ KA group (P＞0.05). Compared with LPD group, proteinuria in LPD+KA group was decreased [(0.43± 0.35) vs (0.15±0.36) g/24 h, P＜0.01]. Patients in two groups were both in good nutritional condition, and KA independently increased the levels of serum albumin and prealbumin (all P＜0.05). The urinary angiotensinogen/creatinine ratio was correlated with GFR (r＝-0.437, P＝0.001), serum creatinine (r＝-0.733, P＝0.000) and proteinuria (r＝-0.851, P＝0.000); while urinary mRNA expression of podocyte markers (podocin and synaptopodin were correlated with proteinuria (r＝0.340, P＝0.012; r＝0.333, P＝0.014; respectively), but had no correlation with GFR and serum creatinine. After 12 months of follow-up, the urinary angiotensinogen/creatinine ratio in LPD+KA group was lower than in LPD group (P＜0.05). The mRNA expression of podocin and synaptopodin were lower in LPD+ KA group compared with LPD group in urinary sediment (P＜0.05). Further correlation analysis suggested that the change of urinary expression of podocin and synaptopodin had a modest but significant correlation with the change urinary angiotensinogen creatinine ratio change (r＝0.305, P＝ 0.026; r＝0.281, P＝0.04, respectively) after treatment for one year.  Conclusions    Low protein diet supplemented with Ketoacids (LPD + KA) is associated with amelioration of proteinuria, meanwhile nutrition status remains well. The mechanism of these effects may be explained by the role of LPD+KA diet in reducing urine podocyte loss and lowering the angiotensinogen level in the urine.

Objective    To evaluate the value of immune cell functional assay (ImmuKnow CD4+ T cell ATP assay) in monitoring immune status in renal recipients.  Methods    A total of 131 adult renal transplant recipients who received transplantation for the first time were under investigation. According to the dynamic monitoring ATP concentration before operation, 2 week, 1, 3, 6 months after operation and during infect or rejection, samples were divided into the following groups: health control group (HC), pretransplant (Pre-Tx) group, stable (Tx) group, infect group, acute rejection (AR) group, acute kidney injury (AKI) group. Immune cell functions were detected by ImmuKnow CD4+ T cell ATP assay. Lymphocyte subsets (CD4+/CD8+) were analysed and serum concentrations of FK506 were tested. Mixed lymphocyte reaction(MLR) was analysed.  Results    The ATP concentration was no significant difference between Pre-Tx and HC group. The ATP concentration of 2 weeks, 1 months after operation were significantly higher than Pre-Tx group (P＜0.01). After 3 months, 6 months follow-up, the ATP concentration stabilized with time. The ATP concentration of AR group was significantly higher than other three groups (Tx, infect and AKI group, all P＜0.05). The correlation coefficient between the ATP concentration and MLR, CD4+/CD8+, FK506 level were R2＝0.0072, R2＝10-6, R2＝0.004 respectively (all P﹥0.05).  Conclusions    The cell-mediated immunity of recipients is relatively strongger during the first month after transplantation. The ATP concentration is not related to the levels of MLR, CD4+/ CD8+, FK506. ImmuKnow ATP assay is a valuable predictor in acute rejection diagnosis.

Objective     To identify novel biomarker for diabetic nephropathy (DN) by urinary proteomic methods, and to detect the expression of E-cadherin in urine and renal tissue of patients with DN.  Methods    Urine samples were collected from 12 cases of type 1 diabetic nephropathy patients (T1DN), 12 cases of type 2 diabetic nephropathy patients (T2DN), 12 cases of nephritic syndrome patients (NS), and 12 cases of healthy Controls. Comparative proteomic approach of two-dimensional gel electrophoresis (2-DE) and mass spectrometry (MS) were employed to identify DN-related biomarker in urine samples. The differential expression of the identified biomarker in urine samples and renal biopsy specimens were detected by Western blotting and immunohistochemistry method.  Results    E-cadherin was identified by 2DE/MS, which was significantly up-regulated in T1DN and T2DN groups (all P＜ 0.01). Western blotting confirmed the expression of E-cadherin was significantly higher in T1DN and T2DN groups than in NS and Control groups (all P＜0.01). Immunohistochemical stain showed E-cadherin was mainly expressed in the membrane and cytoplasm of renal tubular epithelial cell, and its expression was markedly decreased in DN kidneys compared with healthy Controls (P＜0.05).  Conclusions    E-cadherin is identified as a novel DN-related biomarker, which is specifically increased in urine of DN patients.

Objective     To observe the effects of uric acid (UA) on mitochondrial oxidative damage and apoptosis in renal tubular epithelial cells (HK-2), and investigate the possible mechanism.  Methods     HK-2 cells were exposed to UA (480 μmol/L, 720 μmol/L) for different time (0 h, 24 h, 48 h) in vitro. The mitochondrial ROS production was detected by MitoSOX staining. The mitochondrial membrane potential was measured by JC - 1 staining. The expressions of prohibitin and AIF were examined by Western blotting and immunofluorescence cytochemistry. The cell apoptosis was measured by Annexin V-FITC/PI staining.  Results    The mitochondrial ROS production in HK-2 cells exposed to 480 μmol/L UA was increased than that of control group at 24 h (P﹤0.05), and increased gradually with UA concentration and incubation time increasing, while the mitochondrial membrane potential was reduced at the same time. There were no significant changes in AIF expression and apoptosis rate of HK -2 cells exposed to 480 μmol/L UA for 24 h compared with that of control group (P﹥0.05), while both of them were up-regulated when HK-2 cells were exposed to 480 μmol/L UA for 48 h and 720 μmol/L UA for 24 h and 48 h (P﹤0.05). The prohibitin expression in HK-2 cells exposed to 480 μmol/L UA was reduced than that of control group at 24 h (P ﹤ 0.05), and down - regulated gradually with UA concentration and incubation time increasing.  Conclusion    Uric acid can induce the mitochondrial ROS production increased, the mitochondrial membrane potential reduced, the prohibitin expression down-regulated and the mitochondrial apoptosis pathway activated in HK-2 cells.

Objective     To investigate the role of PI3K/Akt signaling in the regulation of epithelial-mesenchymal transition (EMT) of peritoneal mesothelial cells (PMCs) in peritoneal dialysis in vitro and in vivo. Methods The level of Phosphorylated serine/threonine kinase Akt and the expression of EMT associated gene and protein, including ZO-1, Vimentin and FN, were measured in mice EMT model. In vitro study, phosphorylation level and nuclear translocation of Akt, ZO - 1 and Vimentin expression induced by TGF - β1 in human peritoneal mesothelial cells (HPMCs) were also observed. Moreover, HPMCs were pre-treated by one of PI3K/Akt inhibitor, LY294002, or transfected with dominant-negative Akt plasmid to inhibit PI3K/Akt signaling, then analyzed its effect on Zo-1 and Vimentin expression induced by TGF-β1. Results Compared with the control, thickened parietal peritoneum and remarkable decrease in mRNA and protein of the epithelial marker ZO-1, and notable increased in the expression of mesenchymal markers Vimentin and FN were observed in PD mice (all P＜0.01). Moreover, the phosphorylation of Akt also significantly increased under above condition (P＜ 0.01). In vitro study, with the stimulation of TGF-β1, the expression of Zo-1 was down-regulated, while the expression of Vimentin increased (all P＜0.01). In addition, TGF-β1 remarkably increased pAkt in HPMCs (all P＜0.01) in dose-dependent. However, LY294002 and DN-Akt dramatically inhibited the vimentin expression in HPMCs induced by TGF-β1 after inhibition of pAkt. On the other hand, the expression of ZO - 1 also was restored (P＜0.01). Conclusion PI3K/Akt signaling is involved in EMT of peritoneal mesothelial cells in peritoneal dialysis, and may be a new target for the prevention and treatment of peritoneal fibrosis during PD.

Objective     To investigate the effects and underlying mechanism of the scavenger receptor CD36 in high glucose - induced rat glomerular mesangial cells apoptosis.  Methods     The mesangial cells of rats were divided into 4 groups: control group (5.6 mmol/L glucose), mannitol group (24.2 mmol/L mannitol+5.6 mmol/L glucose), high glucose group (30 mmol/L glucose), CD36 mono- antibody group (30 mmol/L glucose+CD36 mono-antibody). The intracellular ROS level was detected by confocal microscopy with fluorescent probe CM - H2DCFDA. MDA, GSH - PX, 8 - OHDGA in cell supernatant were detected. Apoptosis was determined by flow cytometry followed by Annexin V-FITC/PI double stains. The expression of CD36, Bax and Bcl-2 were detected by RT-PCR and Western blotting.  Results     The expression of CD36 was detected in glomerular mesangial cells. The highest level was found in high glucose group in 24 hours. There was no significant difference found between control group and mannitol group with respect to intracellular ROS generation, MDA, 8-OHDG, GSH-PX level, apoptosis rate, expression of CD36, Bax and Bcl-2 (all P＞0.05). There was no significant difference in the expression of CD36 between CD36 mono - antibody group and high glucose group (P＞0.05）. Compared to control group, the intracellular ROS generation, MDA and 8-OHDG levels, apoptosis rate, the expression of CD36 and Bax were significantly increased, the GSH-PX level and the expression of Bcl-2 were significantly lower in high glucose group (all P＜0.05). Compared to the high glucose group, the intracellular ROS generation, MDA and 8-OHDG levels, apoptosis rate, the expression of Bax were suppressed but the GSH-PX level and the expression of Bcl-2 increased in CD36 mono-antibody group (all P＜0.05). The intracellular ROS level was positively correlated with apoptosis rate, protein expression of CD36 and Bax gene, was negatively correlated with Bcl - 2 protein expression.  Conclusions     CD36 was involved in the high glucose induced apoptosis of mesangial cells which was potentially mediated by an increased level of oxidative stress.

Objective     To observe the changes of foot processes, expression and distribution of transient receptor potential cation channel 6 (TRPC6) in podocytes by puromycin aminonucleoside (PAN) and dexamethasone (DEX) intervention, then to investigate the function of TRPC6 in podocytes and its relation to proteinuria in kidney diseases.     Methods     Podocytes cultured in vitro were divided into three group: control group, PAN stimulation group and DEX intervention group. Mouse podocyte cell line (MPC5) were cultured in 0.02% dimethyl sulfoxide (DMSO) in control group, subjected to PAN (50 μg/ml) treatment alone or with DEX (1 μmol/L) in other two groups for 8 h, 24 h, 48 h. The podocyte morphology was observed and took pictures by phase - contrast microscope, then the differences of morphology and areas were analyzed. The distribution, mRNA expression and protein expression of TRPC6 were detected by indirect immunocyto-fluorescence, real-time quantitative PCR and Western blotting, respectively.  Results    The well - developed podocyte arborization and interconnection was formed in control group, but PAN led to significant shrinkage of podocytes (P＜0.05), together with podocyte foot process retraction, effacement and loss of cell contact. DEX significantly prevented the shrinkage and apoptosis of podocytes. The apoptosis rate was significantly increased after PAN stimulated 48 h (P＜0.05). Real-time quantitative PCR and Western blotting found TRPC6 mRNA and protein expression were prone to increase in PAN group compared with control group (P＜0.05). The distribution of TRPC6 becamed abnormal in PAN group. DEX decreased TRPC6 mRNA and protein expression at 48 h compared with PAN group (P＜0.05). The abnormal distribution of TRPC6 was also alleviated by the protection of DEX.  Conclusion     DEX exerts a direct action to podocyte which retains the integrity of slit diaphragm against podocyte injury, and alleviates proteinuria via stabilizing mRNA, protein expression and distribution of TRPC6.

Objective    To investigate the mechanism of chronic intermittent hypoxia (CIH)- induced renal injury and the protection of metallothionein (MT).  Methods    8-10 weeks old male MT-1 transgenic (MT-TG) mice (n=12) and the wide type (WT) mice (n=12) were randomly divided into two groups respectively, Air mimic control(Ctrl) group (n=6) and CIH group (n=6). The period of chronic intermittent hypoxia was continued for 8 weeks. The CIH paradigm consisted of 20.9% O2 and 8% O2 fraction of inspiration O2 (FiO2) alternation cycles (30 episodes per hour) with 20 seconds at the nadir FiO2 for 12 hours/day during daylight. The nadir hemoglobin oxygen saturations mainly ranged from 60% to 70%. Urine, blood, kidney were collected at the end of study respectively. Histopathology, Western blotting and colorimetric method for related target were performed respectively.  Results     In WT mice, renal fibrosis, the expression of connective tissue growth factor (CTGF), type-1 plasminogen activator inhibitor (PAI - 1), hypoxia - inducible factor 1α (HIF - 1α), transforming growth factor β1 (TGF-β1), phosphorylated Smad2 and the MDA content were significantly increased by CIH (P＜0.01). In WT mice, the expression of MT detected by using Western blotting was significantly decreased by CIH (P＜0.01). However, in MT-TG mice, above-mentioned indicators showed no significant difference between CIH and Ctrl group.  Conclusions     Oxidative stresses is the main mechanism of CIH - induced renal injury. The possible molecular mechanism of CIH - induced renal injury is that CIH increases the expression of HIF - 1α in kidney tissue, then activate the TGF - β1 - Smad2 signaling pathway and lead to the renal fibrosis. The protection of MT on CIH-induced renal injury may be via its antioxidant effect.