Ankana Daga

An international multicenter study comparing arrhythmia prevalence between the intracardiac lateral tunnel and the extracardiac conduit type of Fontan operations

OBJECTIVEnThe study objective was to determine whether the extracardiac conduit Fontan confers an arrhythmia advantage over the intracardiac lateral tunnel Fontan.nnnMETHODSnThis multicenter study of 1271 patients compared bradyarrhythmia (defined as need for pacing) and tachyarrhythmia (defined as needing antiarrhythmic therapy) between 602 patients undergoing the intracardiac Fontan and 669 patients undergoing the extracardiac Fontan. The median age at the time of the Fontan procedure was 2.1 years (interquartile range, 1.6-3.2 years) for the intracardiac group and 3.0 years (interquartile range, 2.4-3.9) for the extracardiac group (Pxa0<xa0.0001). The median follow-up was 9.2 years (interquartile range, 5-12.8) for the intracardiac group and 4.7 years (interquartile range, 2.8-7.7) for the extracardiac group (Pxa0<xa0.0001).nnnRESULTSnEarly postoperative (<30 days) bradyarrhythmia occurred in 24 patients (4%) in the intracardiac group and 73 patients (11%) in the extracardiac group (Pxa0<xa0.0001). Early postoperative (<30 days) tachyarrhythmia occurred in 32 patients (5%) in the intracardiac group and 53 patients (8%) in the extracardiac group (Pxa0=xa0not significant). Late (>30 days) bradyarrhythmia occurred in 105 patients (18%) in the intracardiac group and 63 patients (9%) in the extracardiac group (Pxa0<xa0.0001). Late (>30 days) tachyarrhythmia occurred in 58 patients (10%) in the intracardiac group and 23 patients (3%) in the extracardiac group (Pxa0<xa0.0001). By multivariate analysis factoring time since surgery, more patients in the extracardiac group had early bradycardia (odds ratio, 2.9; 95%xa0confidence interval, 1.8-4.6), with no difference in early tachycardia, late bradycardia, or late tachycardia.nnnCONCLUSIONSnOverall arrhythmia burden is similar between the 2 groups, but the extracardiac Fontan group had a higher incidence of early bradyarrhythmias. There was no difference in the incidence of late tachyarrhythmias over time between the 2 operations. Therefore, the type of Fontan performed should be based on factors other than an anticipated reduction in arrhythmia burden from the extracardiac conduit.

Prevalence of Arrhythmias Late After the Fontan Operation

The extracardiac conduit (ECC) modification of the Fontan procedure has been theorized to reduce the risk of sinus node dysfunction and atrial arrhythmia compared with the intra-atrial lateral tunnel (ILT) Fontan. This study aimed to compare the prevalence of early and late arrhythmias in patients who underwent ECC and ILT Fontan from a similar era with long-term follow-up at a single institution. A retrospective cohort study was conducted of all patients who underwent ECC or ILT Fontan from 1995 to 2005 at The Childrens Hospital of Philadelphia. Bradyarrhythmias (including sinus node dysfunction), tachyarrhythmias, and pacemaker burden prevalence was determined throughout early (<30 days) and late (>30 days) postoperative periods. Of 434 patients undergoing the Fontan procedure during the study period, a total of 87 and 106 patients who underwent ECC and ILT Fontan, respectively, met the inclusion criteria. There were no significant differences in risk of sinus node dysfunction or tachyarrhythmia in both early and late postoperative periods. Although the overall risk of late postoperative pacemaker therapy was lower for the ECC cohort (4.9% vs 15.7%, p=0.03), when adjusting for follow-up time, no significant difference was observed (odds ratio 3.1, 95% confidence interval 0.6 to 15.2, p=0.16). In conclusion, the overall prevalence of late postoperative arrhythmias observed after contemporary Fontan modifications is low. Intra-atrial reentrant tachycardia, a potentially fatal complication of the atriopulmonary Fontan operation was infrequently encountered in both ECC and ILT Fontan cohorts. Pacemaker use was higher in the ILT group, although this difference may be explained by differences in follow-up time. Despite the low prevalence of arrhythmias after contemporary Fontan modifications, ongoing surveillance is warranted as the onset of arrhythmias may emerge after longer follow-up time.

Whole exome sequencing frequently detects a monogenic cause in early onset nephrolithiasis and nephrocalcinosis

The incidence of nephrolithiasis continues to rise. Previously, we showed that a monogenic cause could be detected in 11.4% of individuals with adult-onset nephrolithiasis or nephrocalcinosis and in 16.7-20.8% of individuals with onset before 18 years of age, using gene panel sequencing of 30 genes known to cause nephrolithiasis/nephrocalcinosis. To overcome the limitations of panel sequencing, we utilized whole exome sequencing in 51 families, who presented before age 25 years with at least one renal stone or with a renal ultrasound finding of nephrocalcinosis to identify the underlying molecular genetic cause of disease. In 15 of 51 families, we detected a monogenic causative mutation by whole exome sequencing. A mutation in seven recessive genes (AGXT, ATP6V1B1, CLDN16, CLDN19, GRHPR, SLC3A1, SLC12A1), in one dominant gene (SLC9A3R1), and in one gene (SLC34A1) with both recessive and dominant inheritance was detected. Seven of the 19 different mutations were not previously described as disease-causing. In one family, a causative mutation in one of 117 genes that may represent phenocopies of nephrolithiasis-causing genes was detected. In nine of 15 families, the genetic diagnosis may have specific implications for stone management and prevention. Several factors that correlated with the higher detection rate in our cohort were younger age at onset of nephrolithiasis/nephrocalcinosis, presence of multiple affected members in a family, and presence of consanguinity. Thus, we established whole exome sequencing as an efficient approach toward a molecular genetic diagnosis in individuals with nephrolithiasis/nephrocalcinosis who manifest before age 25 years.

Whole Exome Sequencing of Patients with Steroid-Resistant Nephrotic Syndrome

BACKGROUND AND OBJECTIVESnSteroid-resistant nephrotic syndrome overwhelmingly progresses to ESRD. More than 30 monogenic genes have been identified to cause steroid-resistant nephrotic syndrome. We previously detected causative mutations using targeted panel sequencing in 30% of patients with steroid-resistant nephrotic syndrome. Panel sequencing has a number of limitations when compared with whole exome sequencing. We employed whole exome sequencing to detect monogenic causes of steroid-resistant nephrotic syndrome in an international cohort of 300 families.nnnDESIGN, SETTING, PARTICIPANTS, & MEASUREMENTSnThree hundred thirty-five individuals with steroid-resistant nephrotic syndrome from 300 families were recruited from April of 1998 to June of 2016. Age of onset was restricted to <25 years of age. Exome data were evaluated for 33 known monogenic steroid-resistant nephrotic syndrome genes.nnnRESULTSnIn 74 of 300 families (25%), we identified a causative mutation in one of 20 genes known to cause steroid-resistant nephrotic syndrome. In 11 families (3.7%), we detected a mutation in a gene that causes a phenocopy of steroid-resistant nephrotic syndrome. This is consistent with our previously published identification of mutations using a panel approach. We detected a causative mutation in a known steroid-resistant nephrotic syndrome gene in 38% of consanguineous families and in 13% of nonconsanguineous families, and 48% of children with congenital nephrotic syndrome. A total of 68 different mutations were detected in 20 of 33 steroid-resistant nephrotic syndrome genes. Fifteen of these mutations were novel. NPHS1, PLCE1, NPHS2, and SMARCAL1 were the most common genes in which we detected a mutation. In another 28% of families, we detected mutations in one or more candidate genes for steroid-resistant nephrotic syndrome.nnnCONCLUSIONSnWhole exome sequencing is a sensitive approach toward diagnosis of monogenic causes of steroid-resistant nephrotic syndrome. A molecular genetic diagnosis of steroid-resistant nephrotic syndrome may have important consequences for the management of treatment and kidney transplantation in steroid-resistant nephrotic syndrome.

Mutations in six nephrosis genes delineate a pathogenic pathway amenable to treatment

No efficient treatment exists for nephrotic syndrome (NS), a frequent cause of chronic kidney disease. Here we show mutations in six different genes (MAGI2, TNS2, DLC1, CDK20, ITSN1, ITSN2) as causing NS in 17 families with partially treatment-sensitive NS (pTSNS). These proteins interact and we delineate their roles in Rho-like small GTPase (RLSG) activity, and demonstrate deficiency for mutants of pTSNS patients. We find that CDK20 regulates DLC1. Knockdown of MAGI2, DLC1, or CDK20 in cultured podocytes reduces migration rate. Treatment with dexamethasone abolishes RhoA activation by knockdown of DLC1 or CDK20 indicating that steroid treatment in patients with pTSNS and mutations in these genes is mediated by this RLSG module. Furthermore, we discover ITSN1 and ITSN2 as podocytic guanine nucleotide exchange factors for Cdc42. We generate Itsn2-L knockout mice that recapitulate the mild NS phenotype. We, thus, define a functional network of RhoA regulation, thereby revealing potential therapeutic targets.Nephrotic syndrome is the second most common chronic kidney disease but there are no targeted treatment strategies available. Here the authors identify mutations of six genes codifying for proteins involved in cytoskeleton remodelling and modulation of small GTPases in 17 families with nephrotic syndrome.

Diagnosis and Risk Factors of Acute Kidney Injury in Very Low Birth Weight Infants

BACKGROUNDnAcute kidney injury (AKI) is common in critically ill premature infants. There is a lack of consensus on the diagnostic definition of AKI in very low birth weight (VLBW) infants. The primary aim of this study was to determine the incidence and risk factors for AKI in VLBW infants using the AKI network (AKIN) and pRIFLE (pediatric Risk, Injury, Failure, Loss, End-Stage) criteria and to evaluate whether Clinical Risk Index for Babies (CRIB II) score is a predictor of AKI. The secondary objective was to determine the extent of agreement between the AKIN and pRIFLE criteria in the diagnosis of AKI in VLBW infants.nnnMETHODSnThis was a retrospective chart review of 115 VLBW (< 1500xa0g) infants born in an academic center with a Level 3B neonatal intensive care unit. Multiple congenital anomalies, transfer to other centers, or death within the first 2xa0weeks were the exclusion criteria. Relevant data were collected and analyzed in the first 2xa0weeks postnatally.nnnRESULTSnAKI incidence, according to AKIN and pRIFLE criteria, was 20.1% and 22.6%, respectively. As per the interrater reliability analysis, there was a fair agreement between the two criteria (kappaxa0=xa00.217). AKI was nonoliguric. The length of stay was significantly longer in the AKI group. Prenatal nonsteroidal anti-inflammatory drug exposure, lower gestational age, lower birth weight, respiratory distress syndrome, mechanical ventilation, patent ductus arteriosus, hypotension, late onset sepsis, and higher CRIB II scores were significantly associated with AKI. Our regression analysis found CRIB II scores to be an independent risk factor for AKI (odds ratioxa0=xa01.621; 95% confidence interval, 1.230-2.167; pxa0=xa00.001).nnnCONCLUSIONnThe determination of AKI using the pRIFLE and AKIN criteria yielded different results. pRIFLE appears to be more sensitive in VLBW infants. A high CRIB II score was recorded for AKI. Future studies are necessary to develop a uniform definition and identify the risk factors to improve the outcomes in this population.

Mutations in multiple components of the nuclear pore complex cause nephrotic syndrome

Steroid-resistant nephrotic syndrome (SRNS) almost invariably progresses to end-stage renal disease. Although more than 50 monogenic causes of SRNS have been described, a large proportion of SRNS remains unexplained. Recently, it was discovered that mutations of NUP93 and NUP205, encoding 2 proteins of the inner ring subunit of the nuclear pore complex (NPC), cause SRNS. Here, we describe mutations in genes encoding 4 components of the outer rings of the NPC, namely NUP107, NUP85, NUP133, and NUP160, in 13 families with SRNS. Using coimmunoprecipitation experiments, we showed that certain pathogenic alleles weakened the interaction between neighboring NPC subunits. We demonstrated that morpholino knockdown of nup107, nup85, or nup133 in Xenopus disrupted glomerulogenesis. Re-expression of WT mRNA, but not of mRNA reflecting mutations from SRNS patients, mitigated this phenotype. We furthermore found that CRISPR/Cas9 knockout of NUP107, NUP85, or NUP133 in podocytes activated Cdc42, an important effector of SRNS pathogenesis. CRISPR/Cas9 knockout of nup107 or nup85 in zebrafish caused developmental anomalies and early lethality. In contrast, an in-frame mutation of nup107 did not affect survival, thus mimicking the allelic effects seen in humans. In conclusion, we discovered here that mutations in 4 genes encoding components of the outer ring subunits of the NPC cause SRNS and thereby provide further evidence that specific hypomorphic mutations in these essential genes cause a distinct, organ-specific phenotype.

Advillin acts upstream of phospholipase C ϵ1 in steroid-resistant nephrotic syndrome

Steroid-resistant nephrotic syndrome (SRNS) is a frequent cause of chronic kidney disease. Here, we identified recessive mutations in the gene encoding the actin-binding protein advillin (AVIL) in 3 unrelated families with SRNS. While all AVIL mutations resulted in a marked loss of its actin-bundling ability, truncation of AVIL also disrupted colocalization with F-actin, thereby leading to impaired actin binding and severing. Additionally, AVIL colocalized and interacted with the phospholipase enzyme PLCE1 and with the ARP2/3 actin-modulating complex. Knockdown of AVIL in human podocytes reduced actin stress fibers at the cell periphery, prevented recruitment of PLCE1 to the ARP3-rich lamellipodia, blocked EGF-induced generation of diacylglycerol (DAG) by PLCE1, and attenuated the podocyte migration rate (PMR). These effects were reversed by overexpression of WT AVIL but not by overexpression of any of the 3 patient-derived AVIL mutants. The PMR was increased by overexpression of WT Avil or PLCE1, or by EGF stimulation; however, this increased PMR was ameliorated by inhibition of the ARP2/3 complex, indicating that ARP-dependent lamellipodia formation occurs downstream of AVIL and PLCE1 function. Together, these results delineate a comprehensive pathogenic axis of SRNS that integrates loss of AVIL function with alterations in the action of PLCE1, an established SRNS protein.

Genetic variants in the LAMA5 gene in pediatric nephrotic syndrome

BACKGROUNDnNephrotic syndrome (NS), a chronic kidney disease, is characterized by significant loss of protein in the urine causing hypoalbuminemia and edema. In general, ∼15% of childhood-onset cases do not respond to steroid therapy and are classified as steroid-resistant NS (SRNS). In ∼30% of cases with SRNS, a causative mutation can be detected in one of 44 monogenic SRNS genes. The gene LAMA5 encodes laminin-α5, an essential component of the glomerular basement membrane. Mice with a hypomorphic mutation in the orthologous gene Lama5 develop proteinuria and hematuria.nnnMETHODSnTo identify additional monogenic causes of NS, we performed whole exome sequencing in 300 families with pediatric NS. In consanguineous families we applied homozygosity mapping to identify genomic candidate loci for the underlying recessive mutation.nnnRESULTSnIn three families, in whom mutations in known NS genes were excluded, but in whom a recessive, monogenic cause of NS was strongly suspected based on pedigree information, we identified homozygous variants of unknown significance (VUS) in the gene LAMA5. While all affected individuals had nonsyndromic NS with an early onset of disease, their clinical outcome and response to immunosuppressive therapy differed notably.nnnCONCLUSIONnWe here identify recessive VUS in the gene LAMA5 in patients with partially treatment-responsive NS. More data will be needed to determine the impact of these VUS in disease management. However, familial occurrence of disease, data from genetic mapping and a mouse model that recapitulates the NS phenotypes suggest that these genetic variants may be inherited factors that contribute to the development of NS in pediatric patients.

Panel sequencing distinguishes monogenic forms of nephritis from nephrosis in children

BACKGROUNDnAlport syndrome (AS) and atypical hemolytic-uremic syndrome (aHUS) are rare forms of chronic kidney disease (CKD) that can lead to a severe decline of renal function. Steroid-resistant nephrotic syndrome (SRNS) is more common than AS and aHUS and causes 10% of childhood-onset CKD. In recent years, multiple monogenic causes of AS, aHUS and SRNS have been identified, but their relative prevalence has yet to be studied together in a typical pediatric cohort of children with proteinuria and hematuria. We hypothesized that identification of causative mutations by whole exome sequencing (WES) in known monogenic nephritis and nephrosis genes would allow distinguishing nephritis from nephrosis in a typical pediatric group of patients with both proteinuria and hematuria at any level.nnnMETHODSnWe therefore conducted an exon sequencing (WES) analysis for 11 AS, aHUS and thrombotic thrombocytopenic purpura-causing genes in an international cohort of 371 patients from 362 families presenting with both proteinuria and hematuria before age 25 years. In parallel, we conducted either WES or high-throughput exon sequencing for 23 SRNS-causing genes in all patients.nnnRESULTSnWe detected pathogenic mutations in 18 of the 34 genes analyzed, leading to a molecular diagnosis in 14.1% of families (51 of 362). Disease-causing mutations were detected in 3 AS-causing genes (4.7%), 3 aHUS-causing genes (1.4%) and 12 NS-causing genes (8.0%). We observed a much higher mutation detection rate for monogenic forms of CKD in consanguineous families (35.7% versus 10.1%).nnnCONCLUSIONSnWe present the first estimate of relative frequency of inherited AS, aHUS and NS in a typical pediatric cohort with proteinuria and hematuria. Important therapeutic and preventative measures may result from mutational analysis in individuals with proteinuria and hematuria.