Objective To detect the M-type phospholipase A2 receptor (PLA2R), and thrombospondin type-1 domain-containing 7A (THSD7A) expression in renal tissue and the levels of their antibodies in adult idiopathic membranous nephropathy (IMN). Also to determine the value of the two markers in the diagnosis of IMN. Methods One hundred and sixteen patients with biopsy-proven MN at the Second Hospital of Hebei Medical University from December 2014 to August 2015 were enrolled, including 86 patients with IMN, 10 patients with HBV-MN and 10 patients with stage V lupus nephritis (LN-V). Twenty patients with minimal change disease (MCD) were regarded as control group. We conducted immunohistochemical analysis of the presence of THSD7A and PLA2R the Paraffin section and enzyme linked immunosorbent assay (ELISA) detecting serum PLA2R-AB and THSD7A-AB concentration to investigate whether there was a correlation between them and clinical indicators. Results Compared with the SMN and MCD groups, the positive rates of PLA2R and PLA2R-AB were significantly higher in IMN groups. Expression PLA2R was detected in 88.4%, 47.4%, 10% and 0% and PLA2R-AB in 82.6%, 15%, 10%, 0%, respectively, of the patients with IMN, HBV-MN, LN-V and MCD. Expression THSD7A was detected in 2.3% of the patients with IMN while not detected in SMN and MCD. THSD7A-AB antibody was negative in all patients. Compared with serum PLA2R-Ab negative individuals, patients with serum PLA2R-Ab positive had lower serum albumin (P＜0.001), higher urine protein excretion (P=0.01). The sensitivity of PLA2R-AB, PLA2R,THSD7A and PLA2R+THSD7A in the diagnosis of IMN were 82.6%, 88.4%, 2.3%, 88.6%, and the specificity was 92%, 66.7%, 100%, 66.7%, respectively. Conclusions PLA2R in renal tissue and serum PLA2R-AB are specific markers for the diagnosis of IMN, which are closely related with the severity of IMN. Expression of THSD7A is only positive in some of IMN patients with negative PLA2R, which can be used as a supplementary examination of IMN patients with negative PLA2R.

Objective To analyze the clinicopathologic features of proliferative sclerosing IgA nephropathy and the efficacy of prednisone therapy. Methods A retrospective analysis was conducted, enrolling 50 patients with biopsy-proven primary proliferative sclerosing IgA nephropathy who were admitted in the Hospital from January 2005 to June 2015 - 26 males and 24 females, mean age (36.8±10.4) years. Clinicopathologic features and prednisone therapeutic effect were analyzed. Results The clinical manifestations of 50 cases were nephritis syndrome with varying degrees of renal insufficiency, including 32 cases (64.0%) with hypertension, 15 cases (30.0%) with microscopic hematuria. Renal biopsy showed the incidence of glomerular global sclerosis was 17.0%-47.2%, tubular atrophy/ interstitial fibrosis outstanding (T0 50%, T1 32%, T2 18%). After prednisone treatment, compared with sustained remission group and relapse group, invalid patients had higher incidence of hypertension (P＜0.05), relatively lower Hb (P＜0.01) and serum albumin, more significant renal dysfunction (P＜0.01), more severe glomerular global sclerosis, segmental sclerosis, tubular atrophy/interstitial fibrosis, while the lower interstitial inflammatory cell infiltration. During the follow-up, which lasted from 6 to 132 months (median 27.3 months), the effective rate of treatment was 74.0% after sufficient prednisone or half dose prednisone therapy. Repeated recurrence rate was 32.0%. At the end of the follow-up period, 13(26.0%) patients entered the stage of uremia. Conclusions Application of glucocorticoids in the treatment of proliferative sclerosing IgA nephropathy can protect renal function and delay the progression of renal impairment. The efficacy of glucocorticoids therapy is significantly associated with the presence or absence of hypertension, the degree of renal function impairment, and the severity of the onset of renal pathology.

Objective To detect the prevalence of heart valvular calcification (VC) and its related risk factors, and to investigate correlation between serum 25-hydroxyvitamin D3[25(OH)D3] and VC in chronic kidney disease (CKD) stage 3-5 patients. Methods A total of 294 CKD patients stage 3-5 were admitted in The Second Affiliated Hospital of Anhui Medical University. Their clinical and laboratory data were collected, patients were classified into two groups according echocardiography: patients with VC were defined as VC group while others were defined as non-VC group. The differences of 25(OH)D3 level and other data in two group were assessed, and related risk factors of VC were analyzed. Results Among 294 CKD patients, 82 were with VC (27.9%) while 212 were without VC (72.1%); serum 25(OH)D3 level was significantly higher in VC group than in non-VC group [(11.9±9.3) μg/L vs (9.6±7.2) μg/L, P＜0.05]. Age, cystatin C, hypersensitive C-reactive protein, pulmonary artery pressure, proportion of secondary hyperparathyroidism, incidence of abdominal aortic calcification and taking active vitamin D proportion were higher in VC group than in non-VC group (P＜0.05). Two classification logistic regression analyses showed that advanced age, high intact parathyroid hormone (iPTH) and 25(OH)D3, pulmonary arterial hypertension were risk factors for VC in CKD stage 3-5 patients. Conclusions The prevalence of VC is high in CKD stage 3-5 patients. Advanced age, bone metabolic disorder and pulmonary arterial hypertension are associated with VC.

Objective To observe the quantity change of autophagosomes in podocytes and expressions of autophagy-related gene Beclin-1 and microtubule-associated protein 1 light chain 3 (LC3) in different pathological stages of idiopathic membranous nephropathy (IMN), and to explore how autophagy is related to podocyte injury, the occurrence of proteinuria and the disease progression in IMN. Methods Clinical data of 26 patients who were diagnosed as IMN (14 IMN stage 1 and 12 IMN stage 2) admitted to Zhejiang Provincial people's Hospital from January 2013 to December 2014 were retrospectively analyzed. Normal renal tissue from 15 cases of kidney neoplasms with nephrectomy was collected as control. The changes of kidney tissue pathology were detected after PAS and PASM staining by light microscope. The autophagosomes of podocyte were detected by transmission electron microscopy. Expressions of Beclin-1 and LC3 protein were detected by immunohistochemistry. Expressions of LC3 and synaptopodin were detected by immunofluorescence. The correlation of autophagosomes and clinical pathologic factors in IMN patiens was analyzed. Results There were fewer autophagosomes of podocytes and lower expression of Beclin-1 and LC3 protein in IMN group than those in control group (P=0.034, P=0.011, P=0.013, respectively). Moreover, these effects were more obvious with the development of IMN. Compared with those in control group, autophagosomes, Beclin-1 and LC3 protien were reduced in IMN stage 2 group (P=0.009, P=0.030, P=0.015); the number of autophagosomes and the expressions of LC3 and Beclin-1 were decreased in IMN stage 1 group as well, however statistically insignificant (P=0.352, P=0.087, P=0.128); Comparisons between IMN stage 2 patients and IMN stage 1 patients shown significant difference in the number of autophagosomes (P=0.030), but no significant difference in expressions of Beclin-1 and LC3 (P=0.355, P=0.181). Autophagosomes number was not correlated with serum creatinine, serum urea nitrogen, 24-hour urinary protein and eGFR (all P＞0.05). The expressions of synaptopodin and LC3 protein were lower in IMN group than those in control group. Conclusion Autophagy may contribute to podocyte injury and the production of protein urine in IMN, and may be closely related to the progression of disease.

Objective To compare the survival rates of elderly hemodialysis (HD) and peritoneal dialysis (PD) patients and identify their independent prognostic predictors. Methods Patients aging ＞60 years old who initiated dialysis between January 1, 2008 and December 31, 2014 were included. Propensity score method (PSM) was applied to adjust for selection bias. Kaplan-Meier method was used to obtain survival curves and a Cox regression model was used to evaluate risk factors for mortality. Results 447 eligible patients with maintenance dialysis were identified, 236 with hemodialysis and 211 with peritoneal dialysis. 174 pairs of patients were matched, with the baseline data [age, gender, Charlson comorbidity index (CCI) and the primary disease] between two groups showing no significant difference (P＞0.05). Cardiovascular events, cerebrovascular events and infection were major causes of death in both groups and there was no significant difference in the causes of death between two groups (P＞0.05). The overall survival rates at 1 and 5 year were 93.6% and 63.4% respectively in HD group, 91.9% and 61.5% in PD group. The differences of total survival rates between HD and PD patients were not significant (P＞0.05). Cox regression analysis showed age(≥80 year) (P＜0.001, HR=1.058, 95%CI 1.028-1.088), diabetic nephropathy (P=0.001, HR=2.161, 95%CI 1.384-3.373), CCI≥5 (P=0.007, HR=1.935, 95%CI 1.201-3.117) were independent prognostic risk predictors in HD patients; age(≥80 year) (P=0.022, HR=1.043, 95%CI 1.006-1.081), serum albumin level ＜ 35 g/L (P=0.025, HR=1.776, 95%CI 1.075-2.934), and prealbumin (P=0.012, HR=0.968, 95%CI 0.944-0.993) were independent prognostic predictors in PD patients. Conclusions The differences of total survival rates between aged HD and PD patients are not significant. Age, diabetic nephropathy, CCI≥5 and age, serum albumin＜35 g/L, prealbumin＞30 g/L respectively influence the survival of elderly HD and PD patients.

Objective To retrospectively analyze the characteristics of age distribution and clinical nutritional parameters in secondary hyperparathyroidism (SHPT) patients undergoing parathyroidectomy (PTX). Methods Clinical data of 496 SHPT patients undergoing PTX from 2011 to 2015 in the First Affiliated Hospital with Nanjing Medical University were collected and recorded. Age stratification of SHPT patients was observed. The levels of nutritional parameters in different age groups were compared using ANOVA analysis. The relationship between intact parathyroid hormone (iPTH) and nutritional parameters was explored using Spearman's correlation. Results There were 274 males in 496 SHPT patients who were aged (46.0±11.4) years. Chronic glomerulonephritis was the major primary cause of patients (92.1%). Their dialysis vintage was (7.7±3.6) years. The proportion of SHPT patients receiving hemodialysis was 92.9%. In SHPT patients serum levels of calcium, phosphorus, iPTH and alkaline phosphatase (ALP) were (2.6±0.2) mmol/L, (2.2±0.5) mmol/L, (2290.0±1294.2) ng/L, and (564.7±537.8) U/L, respectively. Levels of serum albumin (Alb) in all age groups were lower than the reference range. Serum calcium, ALP, and iPTH levels among age groups were different with statistical significance, while serum phosphorus levels among age groups shown no statistically significant difference. Compared with patients aged ≤18 years old and 19~30 years old, the level of ln[ALP] was lower in patients aged 61-70 years old (P＜0.05). Conclusions Severe SHPT patients are mainly receiving hemodialysis and aged between 30 and 60 years old. Chronic glomerulonephritis is a primary cause of SHPT patients. In order to increase the patients' endurance of operations and reduce the occurrence of postoperative complications, malnutrition in SHPT patients is to be alleviated before PTX.

Objective To investigate the effect and mechanism of chitosan on vascular smooth muscle cell proliferation of uremia patients with arteriovenous fistula. Methods Primarily culturing the VSMCs of uremia patients with arteriovenous fistula and patients without uremia by explants adherent method, and taking the second generation. VSMCs from patients without uremia cultured with 20% FBS medium were non-uremia group, VSMCs of uremia patients cultured with 20% FBS medium were uremia group, VSMCs of uremia patients with 100 μg/ml chitosan were uremia+chitosan group. The expression of α-SMA was detected by immunohistochemistry. The changes of migration and invasion of VSMCs were detected by scratches and transwell migration assays. The mRNA expressions of TLR4 and PCNA were measured by real-time PCR. VSMCs of uremia patients with arteriovenous fistula were intervened with different doses of chitosan (0, 100 and 500 μg/ml), and the protein expressions of TLR4, MyD88 and NF-κB were detected by Western blotting. Results Compared with those in non-uremia group, in uremia group and uremia+chitosan group α-SMA was up-regulated, migration and invasion of VSMCs were enhanced, and mRNA expressions of TLR4 and PCNA were increased (all P＜0.05). Compared with those in uremia group, the level of α-SMA was significantly decreased, the ability of migration and invasion of VSMCs were decreased, and the mRNA expressions of TLR4 and PCNA were decreased (all P＜0.05). TLR4, MyD88 and NF-κB protein expressions were reduced in concentration-dependent manner by 100 and 500 μg/ml chitosan. Conclusions (1) In vitro, chitosan decreases the ability of migration and invasion of VSMCs of uremia patients with arteriovenous fistula. (2) Chitosan inhibits the proliferation of VSMCs, which may be relevant in the decreased expressions of TLR4, MyD88 and NF-κB.

Objective To investigate the regulation of melatonin (MT) on Toll-like receptor 4 (TLR4) signaling in diabetic db/db mice kidneys. Methods The 48 10-week-old male db/db mice were randomly divided into db/db group, db/db+MT 50 μg/kg group, db/db+MT 100 μg/kg group and db/db+MT 200 μg/kg group, each consisting of 12 mice. These mice received i.p. injections of MT These mice received i.p. injections of MT [dissoved in phosphate buffer solution (PBS)/ dimethylsulfoxide (DMSO) solution, given every day]. Alternatively, 12 db/m mice served as the control group. db/m and db/db group were injected i.p. with the same volume of PBS/DMSO solution. The animals were sacrificed after 12 weeks of dosage administration. Blood glucose (BG), body weight (BW), kidney weight (KW) and 24 h urinary albumin excretion rate (UAER) were determined; Kidney pathological lesions were evaluated by renal pathological staining. Immunohistochemistry of renal TLR4, NF-κB p65, and ED-1 was performed to determine the immunoreactivity. Western blotting was used to detect the expression of renal TLR4, myeloid differentiation factor 88 (MyD88), TIR-domain-containing adaptor inducing interferon-β (TRIF), interferon regulatory factor 3 (IRF-3) and NF-κB p65, while the mRNA expressions of renal tumor necrosis factor -α (TNF-α) and monocyte chemotactic protein-1 (MCP-1) were evaluated by real-time PCR. Results Compared with control group, the levels of BG, BW, KW and UAER were much higher in db/db mice group (P＜0.01), while KW in db/db+MT (100, 200 μg/kg) groups and UAER level in db/db+MT (50, 100, 200 μg/kg) groups were distinctly decreased compared with those in db/db group (P＜0.01). In week 12 db/db mice, the glomerular mesangial expansion index and tubulointerstitial injury index were increased compared with those in db/m mice (P＜0.01). The above kidney histopathologic lesions were distinctly ameliorated by 50, 100, 200 μg/kg MT (P＜0.05). Immunohistochemistry intensity of renal TLR4, NF-κB p65 and ED-1 displayed obvious differences between db/m mice and db/db mice (P＜0.01), and that were remarkably decreased in db/db+MT (50, 100, 200 μg/kg) mice compared with db/db mice (P＜0.05). Western blotting showed that the protein expression of renal TLR4, MyD88, TRIF, IRF-3 and NF-κB p65 were stronger in db/db group compared with those in db/m group (P＜0.05) and weaker in db/db+MT (50, 100, 200 μg/kg) groups compared with those in db/db group (P＜0.05). Futhermore, the mRNA expressions of renal MCP-1 and TNF-α were higher in db/db group compared with those in db/m group (P＜0.01) and lower in db/db+MT (50, 100, 200 μg/kg) groups compared with those in db/db group (P＜0.01). Conclusion Melatonin may partly down-regulate TLR4 signaling pathway to inhibit Inflammatory reaction and alleviate kidney injury in diabetic db/db mice.

Objective To observe the effect of JLP on transdifferentiation of human renal proximal tubular epithelial cells (HK-2), and to investigate the role of p38 MAPK signaling pathway in this process. Methods The knock-down plasmids of JLP were constructed. HK-2 cells were randomly divided into four groups: negative control cells (Ctrl-shRNA group), knock-down jlp cells (jlp-shRNA group), negative control cells with FGF-2 treatment (FGF-2 group) and knock-down jlp cells with FGF-2 treatment(jlp-shRNA+FGF-2 group). The expressions of JLP, E-cadherin, TGF-β1, α-SMA, p-p38 MAPK protein were detected by Western blotting．After the induction of FGF-2 for 24 hours, the expressions of α-SMA, COL-I, FN were detected by immunocytochemistry. Results Compared with Ctrl-shRNA group, the expression of JLP protein was significantly down-regulated in FGF-2 group. Compared with FGF-2 group, the expressions of TGF-β1, α-SMA, p-p38 MAPK protein were significantly up-regulated, while E-cadherin protein was significantly down-regulated (P＜0.05). Compared with FGF-2 group, the expressions of α-SMA, COL-I, FN immunostaining increased markedly in jlp-shRNA+FGF-2 group. Conclusion Scaffolding protein JLP is critical in preventing EMT in the course of fibrosis through the inhibition of p-p38 activation in HK-2 cells.

Objective To establish adriamycin-induced focal segmental glomerular sclerosis (FSGS) mice model, and observe the expressions of and relation between oxidative stress and p38 MAPK signal pathway in renal injury. Methods Eight-week-old male Balb/c mice were randomly divided into FSGS group (n=20) and control group (n=20). In FSGS group mice were intravenously injected with a single dose of adriamycin (0.01 mg/g), and mice in control group were received saline with the same dose. At day 3, 7, 14, 22 and 32, urine protein-to-urine creatinine ratio (P/C) was detected. At day 22 and 32, serum creatinine, blood urea nitrogen, nitric oxide (NO) and reactive oxygen species (ROS) in blood and urine, and ROS in kidney tissues were detected; changes of pathological morphology in renal tissue were analyzed by HE stain; the expressions of NF-κB, CD36, IL-13, BAX and Bcl-2 mRNA were detected by real time quantitative PCR; the expressions of NF-κB, p-p38 and p-ERK1/2 protein were detected by Western blotting. Results Compared with that in control group, P/C was gradually increasing in FSGS group, and peaked at day 22 (P＜0.05). At day 22 and 32, mice had higher creatinine, serum creatinine, urea nitrogen, ROS and NO in FSGS group than those in control group (all P＜0.05). There were mild hyperplasia of mesangial cells and mesangial matrix, segment with moderate exacerbations, podocytes with significant proliferation, and the capillary loops of the stenosed in the glomerular in FSGS group at day 32. Compared with those in control group, the mRNA expression of NF-κB, BAX, IL-13 and CD36, and the protein expressions of NF-κB and p-p38 MAPK were gradually increased in FSGS group, all showed statistical differences at day 32 (all P＜0.05); the expression of p-ERK1/2 was increased at day 22 (P＜0.05) but was reduced at day 32 (P＜0.05). Conclusions Adriamycin has induced FSGS in mice successfully, which may through oxidative stress activating p38, up-regulating NF-κB, increasing the inflammatory cytokines and inducing apoptosis pathways.