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The aim of the study was to characterize by molecular profiling two glomerular diseases: IgA nephropathy (IgAN) and focal segmental glomerulosclerosis (FSGS) and to identify potential molecular markers of IgAN and FSGS progression. The expressions of 90 immune-related genes were compared in biopsies of patients with IgAN (n=33), FSGS (n=17) and in controls (n=11) using RT-qPCR. To identify markers of disease progression, gene expression was compared between progressors and non-progressors in 1 year follow-up. The results were verified on validation cohort of patients with IgAN (n=8) and in controls (n=6) using laser-capture microdissection, that enables to analyze gene expression separately for glomeruli and interstitium. In comparison to controls, patients with both IgAN and FSGS, had lower expression of BAX (apoptotic molecule BCL2-associated protein) and HMOX-1 (heme oxygenase 1) and higher expression of SELP (selectin P). Furthermore, in IgAN higher expression of PTPRC (protein-tyrosine phosphatase, receptor-type C) and in FSGS higher expression of BCL2L1 (regulator of apoptosis BCL2-like 1) and IL18 compared to control was observed. Validation of differentially expressed genes between IgAN and controls on another cohort using laser-capture microdissection confirmed higher expression of PTPRC in glomeruli of patients with IgAN. The risk of progression in IgAN was associated with higher expression EDN1 (endothelin 1) (AUC=0.77) and FASLG (Fas ligand) (AUC=0.82) and lower expression of VEGF (vascular endothelial growth factor) (AUC=0.8) and in FSGS with lower expression of CCL19 (chemokine (C-C motif) ligand 19) (AUC=0.86). Higher expression of EDN1 and FASLG along with lower expression of VEGF in IgAN and lower expression of CCL19 in FSGS at the time of biopsy can help to identify patients at risk of future disease progression.

The present study was performed to examine in two-kidney, one-clip (2K1C) Goldblatt hypertensive mice: first, the relative contribution of angiotensin II receptor subtypes 1A (AT(1A)) and 1B (AT(1B)); second, the role of angiotensin II type 2 (AT(2)) receptors in the development of hypertension in wild-type (AT(1A)+/+) and AT(1A) receptor knockout (AT(1A)-/-) mice; and third, the role of increased nitric oxide synthase activity in counteracting the hypertensinogenic action of angiotensin II in this model.AT(1A)+/+ and AT(1A)-/- mice underwent clipping of one renal artery and were infused with either saline vehicle or selective AT(2) receptor agonist CGP-42112A (CGP). Blood pressure was monitored by radiotelemetry. Blood pressure responses to the nitric oxide synthase inhibitor nitro-L-arginine-methyl-ester were evaluated.AT(1A)+/+ mice responded to clipping by a rise in blood pressure that was not modified by CGP infusion. Clip placement caused a slight increase in blood pressure in AT(1A)-/- mice that remained significantly lower than in AT(1A)+/+ mice. Acute nitric oxide synthase inhibition caused greater increase in blood pressure in 2K1C/AT(1A)+/+ than in AT(1A)+/+ mice.The present data support the critical role of AT(1A) receptors in the development of 2K1C hypertension, whereas AT(1B) receptors play only a minor role in blood pressure regulation in this model of angiotensin II-dependent hypertension. Activation of AT(2) receptors does not play an antagonistic role in the AT(1) receptor-mediated hypertensinogenic actions of angiotensin II in this model. Finally, enhanced nitric oxide synthase activity plays a protective role by counteracting the vasoconstrictor influences of angiotensin II in 2K1C hypertensive mice.

The reason of variability of clinical course and progression to end-stage renal failure (ESRF) of two widespread chronic nephropathies--autosomal dominant polycystic kidney disease (ADPKD) and IgA nephropathy (IGAN) is not clear. The endothelial dysfunction is considered in the number of factors possibly influencing the prognosis of these nephropathies. Our study tried to verify the hypothesis that endothelial nitric oxide synthase (ecNOS) gene polymorphisms in intron 4 could have some relevance to the progression of ADPKD and/or IgA nephropathy.We examined 128 Czech patients with ADPKD (62 males, 66 females) and 93 patients with IGAN (51 males, 42 females). As a control group, we used 100 genetically unrelated healthy subjects (50 men, 50 women, mean age 51.2 +/- 8.2). The genomic DNA was amplified by polymerase chain reaction (PCR) and the products were separated on 1.5% agarose gel and visualized by ultraviolet transillumination. We compared homozygous subjects for ecNOSb allele with homozygous and heterozygous subjects for ecNOSa allele.The frequencies of ecNOSa/b + a/a and ecNOSb/b genotypes were 19% (19/100) and 81% (81/100) in the control group. The frequencies of ecNOSa/b + a/a and ecNOSb/b genotypes in ADPKD patients were: 26.6% (8/30) and 73.4% (22/30) in ADPKD patients with normal renal function, 30% (9/30) and 70% (21/30) in ADPKD with ESRF, 35.2% (18/51) and 64.8% (33/51) in young ADPKD patients, 60% (12/20) and 40% (8/20) in ADPKD patients with ESRF later than in 62 years. In IGAN, the frequencies of ecNOSa/b + a/a and ecNOSb/b genotypes were 24% (12/50) and 76% (38/50) in IgA with normal renal function and 20.9% (9/43) and 79.1% (38/43) in IgA with ESRF.Both in ADPKD and IGAN groups, there was no significant difference in the frequencies of ecNOS genotypes between patients with normal renal function and age matched patients with ESRF and between patients with normal renal function and control group. The frequency of ecNOSa allele was significantly higher in a number limited group ADPKD patients with ESRF later than in 62 years (Chi-square test p0.05). This higher frequency of a allele among ADPKD patients with later onset of ESRF could suggest the trend of positive influence of a allele in ADPKD patients.

Background: Significant phenotypical variability is observed in autosomal dominant polycystic kidney disease (ADPKD). Dysregulation of vascular endothelial growth factor (VEGF) expression in the kidney has been demonstrated in a wide range of renal diseases. The aim of the present study was to assess the influence of the –2578 C/A and the –1154 G/A polymorphisms in the regulatory region of the VEGF gene upon the progression of ADPKD toward end-stage renal disease (ESRD). Methods: The study was performed on 283 ADPKD patients (145 males, 138 females, mean age 51.7 ± 10.3 years) who had reached ESRD. Patients were divided into three groups: (1) ESRD development later than in 63 years (slow progressors, n = 47), (2) ESRD development before 45 years (rapid progressors, n = 69), and (3) ESRD development between 45 and 63 years (intermediate progressors, n = 167). Genetically unrelated healthy Czech individuals were analyzed as a control group (n = 311, 153 males, 158 females, mean age 44.6 ± 9.2 years). DNA samples were genotyped for the –2578 C/A and for the –1154 G/A polymorphisms of the VEGF gene promoter. The serum levels of VEGF were established in 111 healthy Czech individuals from the control group. Results: The VEGF –2578 C/A and –1154 G/A genotype distribution showed no differences among the groups of slow, rapid and intermediate progressors. The age of ESRD with regard to different genotypes was not significantly different in all ADPKD patients. However, the AA genotype of the –2578 C/A polymorphism was associated with a significantly higher age of ESRD than other genotypes in rapid progressors (42.7 vs. 40.5 years, p = 0.01). The CG haplotype was found significantly more frequent in ADPKD rapid progressors than in slow progressors (p = 0.047). Serum levels of VEGF did not significantly differ in the control group, according to different genotypes of both polymorphisms. Conclusion: To conclude, AA genotype of the –2578 C/A polymorphism was related to better prognosis of the disease in a limited group of ADPKD patients. Classical genetic recessive and dominant model did not find significant influence of separate VEGF polymorphisms on the progression of ADPKD. Accordingly, CG haplotype was associated with earlier onset of ESRD in ADPKD patients.

BACKGROUND; A significant phenotypical variability is observed in autosomal dominant polycystic kidney disease (ADPKD), the most common renal hereditary disease. Endothelin-1 (ET-1) has been suggested to be an important disease-promoting factor of the kidney. Endothelin-converting enzyme-1 (ECE-1) is the main protease responsible for ET-1 generation by cleavage of its functionally inactive precursor. We examined the influence of the ECE-1b C-338A polymorphism on the progression of ADPKD toward end-stage renal disease (ESRD). The A allele was suggested to be associated with higher plasma level of ET-1.200 ADPKD patients (107 males, 93 females) who had reached ESRD were analyzed. Patients were divided into three groups: (1) 47 patients (23 males, 24 females) with ESRD later than in 63 yr (slow progressors); (2) 71 patients (38 males, 33 females) with ESRD before 45 yr (rapid progressors); and (3) 82 patients (46 males, 36 females) with ESRD between 45-63 yr. Moreover, we analyzed 160 genetically unrelated healthy Czech subjects as the control group (82 males, 78 females, mean age 51.4 +/- 8.2 yr). DNA samples from collected blood were genotyped for ECE-1b C-338A polymorphism using described polymerase chain reaction (PCR) followed by restriction enzyme digestion. We compared the frequencies of different genotypes between the groups of slow and rapid progressors and the ages of ESRD with regard to different genotypes.The ECE-1b C-338A genotype distribution showed no differences among the groups of slow progressors, rapid progressors, ADPKD group with ESRD between 45-63 yr and control group. Comparing the ages of ESRD of all patients, we did not find significant differences in the ages with regard to different genotypes: CC (51.5 +/- 10.1 yr), AC (51.6 +/- 11.4 yr), AA (48.2 +/- 5.9 yr). There was a tendency to lower age of ESRD in AA homozygotes in comparison with other genotypes (t-test, p = 0.12). We found no influence of gender.We excluded the effect of ECE-1b C-338A polymorphism on the progression of ADPKD. We could observe a mild tendency toward faster decline of renal function in AA homozygous individuals.

Background: A significant phenotypical variability is observed in autosomal dominant polycystic kidney disease (ADPKD). The variability can not be fully explained by the genetic heterogenity of the disease. We examined the influence of the ACE I/D polymorphism, adducin Trp460Gly polymorphism and the association of both polymorphisms on the progression of ADPKD towards end-stage renal failure (ESRF). Methods: 320 ADPKD patients (pts) were analyzed, 220 pts (113 males, 107 females) with ESRF before 63 years of age, with a subgroup (rapid progressors) of 20 pts (12 males, 8 females) with ESRF before 40 years of age, 52 pts (23 males, 29 females) with ESRF later than 63 years of age (slow progressors), 48 ADPKD pts (18 males, 30 females) with mean age ±50 years with serum creatinine Results: The α-adducin genotypes showed no differences among the groups of slow progressors (74% Gly/Gly, 22.9% Gly/Trp and 3.1% Trp/Trp), pts with ESRF before 63 years of age (67.7% Gly/Gly, 30.5% Gly/Trp and 1.8% Trp/Trp) and rapid progressors (75% Gly/Gly, 25% Gly/Trp). The ACE genotypes did not differ among the groups of slow progressors (27.1% I/I, 44.8% I/D and 28.1% D/D), pts with ESRF before 63 years of age (23.6% I/I, 51.4% I/D and 25% D/D) and rapid progressors (20% I/I, 55% I/D and 25% D/D). The distribution did not differ from the control group. The ages of ESRF according to different genotypes did not significantly differ. We observed a significant tendency to better prognosis in Trp allele carriers for I/I genotype in comparison with Gly/Gly homozygous subjects. Conclusion: The ACE and α-adducin polymorphisms do not play a significant role in the progression of ADPKD to ESRF.

Background: Kidney transplant recipients (KTRs) with “operational tolerance” (OT) maintain a functioning graft without immunosuppressive (IS) drugs, thus avoiding treatment complications. Nevertheless, IS drugs can influence gene-expression signatures aiming to identify OT among treated KTRs. Methods: We compared five published signatures of OT in peripheral blood samples from 18 tolerant, 183 stable, and 34 chronic rejector KTRs, using gene-expression levels with and without adjustment for IS drugs and regularised logistic regression. Findings: IS drugs explained up to 50% of the variability in gene-expression and 20–30% of the variability in the probability of OT predicted by signatures without drug adjustment. We present a parsimonious consensus gene-set to identify OT, derived from joint analysis of IS-drug-adjusted expression of five published signature gene-sets. This signature, including CD40, CTLA4, HSD11B1, IGKV4–1, MZB1, NR3C2, and RAB40C genes, showed an area under the curve 0⋅92 (95% confidence interval 0⋅88–0⋅94) in cross-validation and 0⋅97 (0⋅93–1⋅00) in six months follow-up samples. Interpretation: We advocate including adjustment for IS drug therapy in the development stage of gene-expression signatures of OT to reduce the risk of capturing features of treatment, which could be lost following IS drug minimisation or withdrawal. Our signature, however, would require further validation in an independent dataset and a biomarker-led trial. Funding: FP7-HEALTH-2012-INNOVATION-1 [305147:BIO-DrIM] (SC,IR-M,PM,DSt); MRC [G0801537/ID:88245] (MPH-F); MRC [MR/J006742/1] (IR-M); Guy's&StThomas’ Charity [R080530]&[R090782]; CONICYT-Bicentennial-Becas-Chile (EN-L); EU:FP7/2007–2013 [HEALTH-F5–2010–260687: The ONE Study] (MPH-F); Czech Ministry of Health [NV19–06–00031] (OV); NIHR-BRC Guy's&StThomas' NHS Foundation Trust and KCL (SC); UK Clinical Research Networks [portfolio:7521].

Contains fulltext : 234032.pdf (Publisher’s version ) (Closed access) Recurrence of primary membranous nephropathy after transplantation occurs in up to 44% of patients and is driven by PLA2R antibody. Here, we asked whether genetic determinants could improve risk prediction. First, we sequenced PLA2R1 and HLA-D loci in 248 patients with primary membranous nephropathy and identified two independent single nucleotide polymorphisms (SNPs) at risk for primary membranous nephropathy at each locus. These were rs9271188 (intergenic between HLA-DRB1 and HLA-DQA1,) and rs9275086 (intergenic between HLA-DQB1 and HLA-DQA2) at the HLA-D locus along with rs6726925 and rs13018963 at the PLA2R1 locus. Then we investigated whether primary membranous nephropathy at-risk variants were associated with recurrence in a retrospective cohort of 105 donor-recipient pairs and a replication cohort of 40 pairs. Seven SNPs located between HLA-DRB1 and HLA-DQA1 in linkage disequilibrium with rs9271188, and three SNPs in the PLA2R1 region predicted recurrence when presented by the donor, but not when presented by the recipient. The two SNPs in the HLA-D region most strongly associated with recurrence (rs9271705 and rs9271550) were confirmed in the replication cohort. A genetic risk score based on the two best predictors at each locus (rs9271705, rs9271550, rs17830558, and rs3828323) identified a group of patients with high risk of recurrence. Thus, our results suggest that the graft contributes to recurrence of primary membranous nephropathy through the disease susceptibility HLA-D and PLA2R1 SNPs in an autoimmune milieu. Further studies are needed before implementation of genetic testing for these in donor selection.