Immunogenicity, Immunological Memory and Monitoring of Disease Activity Following an Anamnestic Immunization With the 13-Valent Pneumococcal Conjugate Vaccine in Children With Idiopathic Nephrotic Syndrome.

Anamnestic 13-valent pneumococcal conjugate vaccine immunization did not affect the relapse risk in pediatric idiopathic nephrotic syndrome. Pneumococcal serotype (PS)-specific antibody titers increased significantly in all groups. Children receiving immunomodulatory treatments (IMTs) displayed significantly lower levels of PS-specific antibodies for 3/8 serotypes tested. PS-specific B-cell counts significantly increased only in healthy controls and patients receiving corticosteroids.

Effect of Leflunomide on Treatment of Pediatric Renal Transplant Recipients With BK Virus Infection.

OBJECTIVES: Infection with the BK virus is a significant complication after renal transplant and can progress to BK virus nephropathy and graft dysfunction. There is no consensus on the management of BK virus infection in pediatric renal transplant recipients. The most common therapeutic option is immunosuppression reduction, which can increase rejection risk. We aimed to examine the effect of leflunomide, an agent with antiviral and immunosuppressive actions, in a case series of pediatric renal transplant recipients with BK virus infection. MATERIALS AND METHODS: Routine screening with blood BK virus DNA polymerase chain reaction was performed regularly in all of our renal transplant patients. When BK virus was detected, we reduced tacrolimus levels, discontinued mycophenolate mofetil, and started active treatment with leflunomide. Treatment with leflunomide was continued until BK virus was undetectable by polymerase chain reaction in at least 2 blood samples 2 weeks apart. RESULTS: All pediatric patients developed BK virus infection in a mean period of 3.9 months after transplant. Graft dysfunction was evident in all patients with 20% to 100% elevation of creatinine from baseline. Afterleflunomide initiation, all patients had undetectable levels of BK virus by plasma polymerase chain reaction in at least 2 different samples within a mean period of 3.4 months, and renal function had normalized back to the baseline. None of our patients had evidence of hepatotoxicity or anemia on regular monitoring, with no other adverse events. Renal function remained stable in the follow-up period with no reoccurrence of BK viremia up to the date of this writing. CONCLUSIONS: Treatment with leflunomide resulted in rapid BK virus clearance and preservation of renal function with no adverse effects.

Bone Mineral Density and Vascular Calcification in Children and Young Adults With CKD 4 to 5 or on Dialysis.

Introduction: Older adults with chronic kidney disease (CKD) can have low bone mineral density (BMD) with concurrent vascular calcification. Mineral accrual by the growing skeleton may protect young people with CKD from extraosseous calcification. Our hypothesis was that children and young adults with increasing BMD do not develop vascular calcification. Methods: This was a multicenter longitudinal study in children and young people (5-30 years) with CKD stages 4 to 5 or on dialysis. BMD was assessed by tibial peripheral quantitative computed tomography (pQCT) and lumbar spine dual-energy X-ray absorptiometry (DXA). The following cardiovascular imaging tests were undertaken: cardiac computed tomography for coronary artery calcification (CAC), ultrasound for carotid intima media thickness z-score (cIMTz), pulse wave velocity z-score (PWVz), and carotid distensibility for arterial stiffness. All measures are presented as age-adjusted and sex-adjusted z-scores. Results: One hundred participants (median age 13.82 years) were assessed at baseline and 57 followed up after a median of 1.45 years. Trabecular BMD z-score (TrabBMDz) decreased (P = 0.01), and there was a nonsignificant decrease in cortical BMD z-score (CortBMDz) (P = 0.09). Median cIMTz and PWVz showed nonsignificant increase (P = 0.23 and P = 0.19, respectively). The annualized increase in TrabBMDz (ΔTrabBMDz) was an independent predictor of cIMTz increase (R 2 = 0.48, ß = 0.40, P = 0.03). Young people who demonstrated statural growth (n = 33) had lower ΔTrabBMDz and also attenuated vascular changes compared with those with static growth (n = 24). Conclusion: This hypothesis-generating study suggests that children and young adults with CKD or on dialysis may develop vascular calcification even as their BMD increases. A presumed buffering capacity of the growing skeleton may offer some protection against extraosseous calcification.

Novel and Founder Pathogenic Variants in X-Linked Alport Syndrome Families in Greece.

Alport syndrome (AS) is the most frequent monogenic inherited glomerulopathy and is also genetically and clinically heterogeneous. It is caused by semi-dominant pathogenic variants in the X-linked COL4A5 (NM_000495.5) gene or recessive variants in the COL4A3/COL4A4 (NM_000091.4/NM_000092.4) genes. The disease manifests in early childhood with persistent microhematuria and can progress to proteinuria and kidney failure in adolescence or early adulthood if left untreated. On biopsy, pathognomonic features include alternate thinning, thickening and lamellation of the glomerular basement membrane (GBM), in the presence of podocyte foot process effacement. Although previous studies indicate a prevalence of AS of about 1/50,000, a recent publication reported a predicted rate of pathogenic COL4A5 variants of 1/2320. We herewith present 98 patients (40 M/58 F) from 26 Greek families. We are selectively presenting the families segregating the X-linked form of AS with pathogenic variants in the COL4A5 gene. We found 21 different pathogenic variants, 12 novel: eight glycine and one proline substitutions in the collagenous domain, one cysteine substitution in the NC1 domain, two premature termination of translation codons, three splicing variants, one 5-bp insertion/frameshift variant, one indel-frameshift variant and four gross deletions. Notably, patients in six families we describe here and three families we reported previously, carried the COL4A5-p.G624D substitution, a founder defect encountered all over Europe which is hypomorphic with mostly milder symptomatology. Importantly, on several occasions, the correct genetic diagnosis reclassified patients as patients with AS, leading to termination of previous immunosuppressive/cyclosporine A therapy and a switch to angiotensin converting enzyme inhibitors (ACEi). With the understanding that all 98 patients span a wide range of ages from infancy to late adulthood, 15 patients (11 M/4 F) reached kidney failure and 11 (10 M/1 F) received a transplant. The prospects of avoiding lengthy diagnostic investigations and erroneous medications, and the advantage of delaying kidney failure with very early administration of renin-angiotensin-aldosterone system (RAAS) blockade, highlights the importance of timely documentation of AS by genetic diagnosis.

Plasma Brain Natriuretic Peptide Levels in Children with Chronic Kidney Disease and Renal Transplant Recipients: A Single Center Study.

Pediatric chronic kidney disease (CKD) patients, as well as kidney transplant patients, are at an increased risk of developing cardiovascular disease. BNP measurement, as a biomarker of cardiovascular risk, has been recommended to this high-risk population. Plasma BNP levels were measured in 56 CKD children in either pre-dialysis stage, hemodialysis (HD) or renal transplant recipients (RTRs) and in 76 sex- and age-matched healthy controls. BNP levels were investigated in HD children, before and after the completion of their HD session. BNP levels in total CKD population, in pre-dialysis stage patients and on HD were significantly higher, compared to the respective controls. HD children had higher BNP levels compared to CKD patients in the pre-dialysis stage. Moreover, post-HD BNP concentration was slightly higher than pre-HD, with the difference being marginally statistically significant. BNP was positively correlated with eGFR, creatinine, cystatin-C and parathormone and negatively with albumin and 25-hydroxyvitamin D. A positive correlation between BNP concentration and the ratio of E/A in pulse-wave Doppler echocardiography was also observed. In conclusion, CKD pediatric patients, mainly those undergoing HD, have high plasma BNP levels which do not decrease after the HD session. This is indicative of a greater risk for future cardiovascular disease.

The burden of subclinical cardiovascular disease in children and young adults with chronic kidney disease and on dialysis.

BACKGROUND: Cardiovascular disease (CVD) is a common cause of morbidity and mortality even in young people with chronic kidney disease (CKD). We examined structural and functional CV changes in patients ˂30 years of age with CKD Stages 4 and 5 and on dialysis. METHODS: A total of 79 children and 21 young adults underwent cardiac computed tomography for coronary artery calcification (CAC), ultrasound for carotid intima-media thickness (cIMT), carotid-femoral pulse wave velocity (cfPWV) and echocardiography. Differences in structural (CAC, cIMT z-score, left ventricular mass index) and functional (carotid distensibility z-score and cfPWV z-score) measures were examined between CKD Stages 4 and 5 and dialysis patients. RESULTS: Overall, the cIMT z-score was elevated [median 2.17 (interquartile range 1.14-2.86)] and 10 (10%) had CAC. A total of 16/23 (69.5%) patients with CKD Stages 4 and 5 and 68/77 (88.3%) on dialysis had at least one structural or functional CV abnormality. There was no difference in the prevalence of structural abnormalities in CKD or dialysis cohorts, but functional abnormalities were more prevalent in patients on dialysis (P < 0.05). The presence of more than one structural abnormality was associated with a 4.5-fold increased odds of more than one functional abnormality (95% confidence interval 1.3-16.6; P < 0.05). Patients with structural and functional abnormalities [cIMT z-score >2 standard deviation (SD) or distensibility <-2 SD) had less carotid dilatation (lumen:wall cross-sectional area ratio) compared with those with normal cIMT and distensibility. CONCLUSIONS: There is a high burden of subclinical CVD in young CKD patients, with a greater prevalence of functional abnormalities in dialysis compared with CKD patients. Longitudinal studies are required to test these hypothesis-generating data and define the trajectory of CV changes in CKD.

The immunopathogenesis of idiopathic nephrotic syndrome: a narrative review of the literature.

Idiopathic nephrotic syndrome (INS) is a common glomerular disease in childhood, and the immunological involvement in the pathogenesis of non-genetic INS, although not fully elucidated, is evident. This narrative review aims to offer a concise and in-depth view of the current knowledge on the immunological mechanisms of the development of INS as well as the role of the immunological components of the disease in the responsiveness to treatment. T cell immunity appears to play a major role in the INS immunopathogenesis and has been the first to be linked to the disease. Various T cell immunophenotypes are implicated in INS, including T-helper-1, T-helper-2, T-helper-17, and T regulatory cells, and various cytokines have been proposed as surrogate biomarkers of the disease; however, no distinct T helper or cytokine profile has been conclusively linked to the disease. More recently, the recognition of the role of B cell mediated immunity and the various B cell subsets that are dysregulated in patients with INS have led to new hypotheses on the underlying immunological causes of INS. Finally, the disambiguation of the exact mechanisms of the INS development in the future may be the key to the development of more targeted personalized approaches in managing INS. CONCLUSIONS: INS demonstrates particularly interesting immunopathogenetic pathways, in which multiple interactions between T cell and B cell immunity and the podocyte are involved. The disambiguation of these pathways will provide promising novel therapeutic targets in INS. WHAT IS KNOWN: • INS is the most common glomerular disease in the paediatric population, and its onset and relapses have been linked to various immunological triggers. • Multiple immunological mechanisms have been implicated in the pathogenesis of INS; however, no single distinct immunological profile has been recognized. WHAT IS NEW: • Th17 cells and Treg cells play an important role in the immune dysregulation in INS. • Transitional B cell levels as well as the transitional/memory B cell ratio have been correlated to nephrotic relapses and have been proposed as biomarkers of INS relapses in SSNS patients.

Routine serum biomarkers, but not dual-energy X-ray absorptiometry, correlate with cortical bone mineral density in children and young adults with chronic kidney disease.

BACKGROUND: Biomarkers and dual-energy X-ray absorptiometry (DXA) are thought to be poor predictors of bone mineral density (BMD). The Kidney Disease: Improving Global Outcomes guidelines suggest using DXA if the results will affect patient management, but this has not been studied in children or young adults in whom bone mineral accretion continues to 30 years of age. We studied the clinical utility of DXA and serum biomarkers against tibial cortical BMD (CortBMD) measured by peripheral quantitative computed tomography, expressed as Z-score CortBMD, which predicts fracture risk. METHODS: This was a cross-sectional multicentre study in 26 patients with CKD4 and 5 and 77 on dialysis. RESULTS: Significant bone pain that hindered activities of daily living was present in 58%, and 10% had at least one low-trauma fracture. CortBMD and cortical mineral content Z-scores were lower in dialysis compared with CKD patients (P = 0.004 and P = 0.02). DXA BMD hip and lumbar spine Z-scores did not correlate with CortBMD or biomarkers. CortBMD was negatively associated with parathyroid hormone (PTH; r = -0.44, P < 0.0001) and alkaline phosphatase (ALP; r = -0.22, P = 0.03) and positively with calcium (Ca; r = 0.33, P = 0.001). At PTH <3 times upper limit of normal, none of the patients had a CortBMD below -2 SD (odds ratio 95% confidence interval 7.331 to infinity). On multivariable linear regression PTH (ß = -0.43 , P < 0.0001), ALP (ß = -0.36, P < 0.0001) and Ca (ß = 0.21, P = 0.005) together predicted 57% of variability in CortBMD. DXA measures did not improve this model. CONCLUSIONS: Taken together, routinely used biomarkers, PTH, ALP and Ca, but not DXA, are moderate predictors of cortical BMD. DXA is not clinically useful and should not be routinely performed in children and young adults with CKD 4-5D.

Results in the ESPN/ERA-EDTA Registry suggest disparities in access to kidney transplantation but little variation in graft survival of children across Europe.

One of the main objectives of the European health policy framework is to ensure equitable access to high-quality health services across Europe. Here we examined country-specific kidney transplantation and graft failure rates in children and explore their country- and patient-level determinants. Patients under 20 years of age initiating kidney replacement therapy from January 2007 through December 2015 in 37 European countries participating in the ESPN/ERA-EDTA Registry were included in the analyses. Countries were categorized as low-, middle-, and high-income based on gross domestic product. At five years of follow-up, 4326 of 6909 children on kidney replacement therapy received their first kidney transplant. Overall median time from kidney replacement therapy start to first kidney transplantation was 1.4 (inter quartile range 0.3-4.3) years. The five-year kidney transplantation probability was 48.8% (95% confidence interval: 45.9-51.7%) in low-income, 76.3% (72.8-79.5%) in middle-income and 92.3% (91.0-93.4%) in high-income countries and was strongly associated with macro-economic factors. Gross domestic product alone explained 67% of the international variation in transplantation rates. Compared with high-income countries, kidney transplantation was 76% less likely to be performed in low-income and 58% less likely in middle-income countries. Overall five-year graft survival in Europe was 88% and showed little variation across countries. Thus, despite large disparities transplantation access across Europe, graft failure rates were relatively similar. Hence, graft survival in low-risk transplant recipients from lower-income countries seems as good as graft survival among all (low-, medium-, and high-risk) graft recipients from high-income countries.

Plasma Urotensin II levels in children and adolescents with chronic kidney disease: a single-centre study.

BACKGROUND: Increased plasma Urotensin II (UII) levels have been found in adults with renal diseases. Studies in children are scarce. The objective of the study is to estimate plasma UII levels in subjects with chronic kidney disease (CKD) stages 3 to 5 and renal transplant recipients (RTR). In addition, the correlation of UII with anthropometric features and biochemical parameters was assessed. METHODS: Fifty-four subjects, aged 3 to 20 years old, 23 with CKD, 13 with end-stage kidney disease (ESKD) undergoing hemodialysis (HD) and 18 RTR were enrolled. A detailed clinical evaluation was performed. Biochemical parameters of renal and liver function were measured. Plasma UII levels were measured in all patients and in 117 healthy controls, using a high sensitive enzyme immunoassay (EIA) kit. All data were analyzed using STATA™ (Version 10.1). RESULTS: Median UII and mean log-transformed UII levels were significantly higher in CKD and RTR patients compared to healthy subjects (p < 0.001). HD patients had higher but not statistically significant UII and log-UII levels than controls. UII levels increased significantly at the end of the HD session and were higher than controls and in line to those of other patients. The geometric scores of UII in HD (before dialysis), CKD and RTR patients increased respectively by 42, 136 and 164% in comparison with controls. Metabolic acidosis was associated with statistical significant change in log-UII levels (p = 0.001). Patients with metabolic acidosis had an increase in UII concentration by 76% compared to those without acidosis. CONCLUSIONS: Children and adolescents with CKD, particularly those who are not on HD and RTR, have significantly higher levels of UII than healthy subjects. UII levels increase significantly at the end of the HD session. The presence of metabolic acidosis affects significantly plasma UII levels.

Long-term outcome of pediatric kidney transplantation: A single-center experience from Greece.

Pediatric kidney Tx has critically altered the outcome in ESRD pediatric patients. The aims of this study were to determine long-term graft and patient survival in a homogeneous ethnic population. We reviewed the medical charts of pediatric kidney Tx performed between 1990 and 2012 in Greece. Seventy-five kidney Txs were performed from LRD and 62 from DD. The 10- and 20-yr graft survival was higher in LRD Tx compared with DD Tx. Both patient and graft survival at 10 and 20 yr after Tx were similar in LRD Tx from grandparents compared with parents (92.9% vs. 93.4% 20-yr patient survival, 71.4% vs. 78.7% and 57.1% vs. 72.1%, 10- and 20-yr graft survival, respectively). However, there was a decreasing tendency in LRD Tx rates in period 2001-2012 compared with period 1990-2000 (47.1% vs. 62.7%). Risk factors for poor five-yr graft survival were DD Tx, and induction treatment with ALG compared with basiliximab, but their effect attenuated at 10 yr after Tx. In conclusion, Tx from LRD may offer efficient survival outcomes irrespective of donor age, suggesting that even older LRD could be an excellent option for the 1st kidney Tx in children and adolescents.

Safety and immunogenicity of booster immunization with 7-valent pneumococcal conjugate vaccine in children with idiopathic nephrotic syndrome.

Safety and immunogenicity of a booster dose of 7-valent pneumococcal conjugate vaccine (PCV7) were evaluated in 29 patients with idiopathic nephrotic syndrome (INS), who had been primed 12 months earlier with one dose of PCV7. PCV7 was not associated with increased risk of INS relapse (RR=0.77, p=0.8) and serotype-specific antibodies increased in all subjects at 1 month (p<0.01). The quantitative characteristics of immune response and the effect of treatment with mycophenolate mofetil and/or cyclosporine A following booster PCV7 were similar with primary response. Additional PCV7 doses could be safely given in children with INS to increase circulating antibodies above the protective threshold.

Acute renal failure in a child with thrombocytopenic purpura caused by acute Epstein-Barr virus infection after treatment with anti-D immunoglobulin.

Immune thrombocytopenia (ITP) in children is usually a benign, self-limiting disorder. An acute Epstein-Barr virus (EBV) infection usually causes atypical lymphocytosis and mild decrease in platelets. Severe thrombocytopenia is an extremely rare complication. Anti-D immunoglobulin has been used for treatment of ITP in Rh(D)-positive nonsplenectomized patients. Severe hemolysis and acute renal failure are extremely rare complications that may be aggravated by the presence of an acute EBV infection. It is believed that anti-D immunoglobulin triggers an unusual virus-induced immune response causing hemolysis. We present a 4-year-old girl with ITP caused by an acute EBV infection that developed acute kidney injury following treatment with anti-D immunoglobulin. The patient recovered completely from thrombocytopenia and renal dysfunction. Intravascular hemolysis and acute kidney injury are consistent with anti-D immunoglobulin mechanism of action. Pediatric patients treated with anti-D immunoglobulin for ITP should be closely monitored for signs and symptoms of hemolysis that may be aggravated by the presence of EBV infection leading to impaired renal function.

Safety, immunogenicity and kinetics of immune response to 7-valent pneumococcal conjugate vaccine in children with idiopathic nephrotic syndrome.

Safety, immunogenicity and kinetics of 7-valent pneumococcal-conjugate vaccine (PCV7) immune response were evaluated in 33 children with idiopathic nephrotic syndrome (INS) and 16 controls. PCV7 was not associated with increased risk of relapse (RR=0.80, p=0.61). Serotype (PS)-specific antibodies increased in all subjects at 1 month (p<0.01) with inferior immunogenicity for 3/7 PS in patients on immunomodulators (p<0.02). Most patients retained protective antibodies at 12-14 months although at lower levels for 4/7 PS (p<0.08). Different PS kinetics in INS suggests antibody monitoring and revaccination when necessary for protection from pneumococcal infections.

Serum osteoprotegerin, RANKL and fibroblast growth factor-23 in children with chronic kidney disease.

Osteoprotegerin (OPG), receptor activator of the nuclear factor κB ligand (RANKL) and fibroblast growth factor-23 (FGF-23) play a central role in renal osteodystrophy. We evaluated OPG/RANKL and FGF-23 levels in 51 children with chronic kidney disease (CKD) [n = 26 stage 3 or 4 (CKD3-4) and n = 25 stage 5 (CKD5)] and 61 controls. Any possible association with intact parathyroid hormone (iPTH) and bone turnover markers was also investigated. The OPG levels were lower in the CKD3-4 group (p < 0.001) and higher in the CKD5 group (p < 0.01) than in the controls, while RANKL levels did not differ. The FGF-23 levels were higher in both patient groups (p < 0.0001), while the levels of phosphate and iPTH were higher only in the CKD5 group (p < 0.0001). There were independent positive correlations between OPG and RANKL (ß = 0.297, p < 0.01) and FGF-23 (ß = 0.352, p < 0.05) and a negative correlation with the bone resorption marker TRAP5b (ß = -0.519, p < 0.001). OPG was positively correlated with iPTH (R = 0.391, p < 0.01). An independent positive correlation between FGF-23 and phosphate (ß = 0.368, p < 0.05) or iPTH (ß = 0.812, p < 0.0001) was noted. In conclusion, we found that higher OPG levels in patients with CKD stage 5 correlated with the levels of RANKL, FGF-23, iPTH, and TRAP5b. These findings may reflect a compensatory mechanism to the negative balance of bone turnover. High FGF-23 levels in early CKD stages may indicate the need for intervention to manage serum phosphate (Pi) levels.

Broad and unexpected phenotypic expression in Greek children with steroid-resistant nephrotic syndrome due to mutations in the Wilms' tumor 1 (WT1) gene.

Mutations in the Wilms' tumor suppressor gene 1 (WT1), most commonly within exons 8 or 9 or intron 9, are found in cases with the overlapping conditions of Denys-Drash and Frasier syndromes, as well as in patients with steroid-resistant nephrotic syndrome (SRNS). This study investigated the presence of WT1 gene mutations in cases with childhood SRNS, along with an evaluation of their clinical outcome. Twenty-seven Greek children with sporadic (19 cases) and familial (8 cases) SRNS were tested. Four phenotypically female patients with sporadic SRNS were found to carry de novo WT1 mutations, including two cases with p.R394W, and one case each with p.R366H, or n.1228+5G>A. Karyotype analysis found 46XX in three cases, but 46XY in one. No phenotype-genotype correlations were apparent in the WT1 gene positive cases since their clinical presentation varied broadly. Interestingly, one patient with a pathological WT1 nucleotide variation responded fully to combined therapy with cyclosporine A and corticosteroids. This study further illustrates that investigation of WT1 gene mutations is clinically useful to support definitive diagnosis in children presenting with SRNS in order to direct the most appropriate clinical management.

Successful treatment of steroid-resistant nephrotic syndrome associated with WT1 mutations.

The Wilms' tumor suppressor gene 1 (WT1) encodes a transcription factor involved in kidney and gonadal development. WT1 is also a key regulator of podocyte functions and mutations have been found in a small percentage of children with isolated or syndromal steroid-resistant nephrotic syndrome. It is commonly assumed that the nephrotic syndrome (NS) in patients with WT1 mutations is unresponsive to therapy and characterized by rapid progression to end-stage renal disease. We report long-term observations in 3 children with focal-segmental glomerulosclerosis associated with WT1 mutations and NS (2 cases) or nephrotic range proteinuria (1 case). All patients showed a favorable response to an intensified therapy consisting of cyclosporin A (CyA) in combination with induction therapy with intravenous and oral prednisone. Treatment with angiotensin-converting enzyme inhibitors and angiotensin receptor blockers was added to the regimen at various times. As shown both by the short-term response and during long-term follow-up, this treatment resulted in clinical remission of the NS and/or significant reduction of proteinuria, while normal renal function could be maintained over many years. Thus, glomerular diseases in selected patients with mutations in genes regulating renal development and podocyte function may respond to combination therapy with CyA and corticosteroids.

Nucleotide variations in the NPHS2 gene in Greek children with steroid-resistant nephrotic syndrome.

Mutations in the NPHS2 gene, encoding podocin, are a major cause of autosomal-recessive steroid-resistant nephrotic syndrome (SRNS) in childhood, accounting for up to 30% of sporadic and 20-40% of familial cases. Among 22 Greek children with a clinical diagnosis of SRNS, mutation analysis was performed in all eight NPHS2 gene exons, using denaturing gradient gel electrophoresis and DNA sequencing. The frequency of all nucleotide variations found in patients was also evaluated in 100 unrelated samples (18-30 years) with no known history of nephrotic disease. Three pathogenic genotypes (R138Q/R138Q, R229Q/A295T, and R168H/R168H) accounted for 3/14 (21%) of sporadic patients; the A295T mutation in exon 8 (c.883G>A) is novel and predicted in silico to be pathogenic. Among the familial cases, a single patient was heterozygous for R229Q. Several known polymorphisms were found, including the in cis variants IVS3-46C>T plus IVS3-21C>T, IVS7+7A>G A and exonic variants S96S (c.288C>T), A318A (c.954T>C), and L346L (c.1038A>G), with allele frequencies comparable to those in other populations. A novel substitution (IVS3-17C>T) was found in two related patients, but in no controls. In conclusion, podocin mutations do not appear to be a major cause of SRNS in Greek children, although the study cohort was small. However, NPHS2 gene analysis could still be considered in Greek SRNS patients to support appropriate management. The present study also contributes potentially useful observations for the clinical management of SRNS patients.

Absence of mutations in the HOXA11 and HOXD11 genes in children with congenital renal malformations.

Experimental studies have shown that homeobox genes are essential for the development of the kidney and urinary tract. Hoxa11/Hoxd11 double mutant mice demonstrate renal agenesis or hypoplasia. Since, to our knowledge, these genes have never been examined for alterations in humans with congenital anomalies of the kidney and urinary tract (CAKUT), we investigated whether mutations of HOXA11/HOXD11 genes are associated with non-syndromal congenital renal parenchymal malformations. DNA samples from 26 unrelated children with unilateral renal agenesis (URA), 20 with renal hypodysplasia (RHD) and 13 with multicystic dysplastic kidney (MCDK) were included in the study. Exons 1 and 2 of the HOXA11/HOXD11 genes were amplified individually by polymerase chain reaction (PCR) using 12 unique oligonucleotide primers. Single-strand conformation polymorphism (SSCP) analysis of overlapping polymerase chain reaction products was performed. SSCP analysis revealed no variant band shifts in the samples of the amplified segments of the 59 patients, suggesting lack of either mutation or polymorphisms. Our findings do not support the hypothesis that mutations in the HOXA11/HOXD11 coding regions are involved in the pathogenesis of human non-syndromal congenital renal parenchymal malformations. Further studies are necessary, since other genes known to affect nephrogenesis, as well as genetic and environmental factors, may be involved.