Whole-Genome Linkage and Association Scan in Primary,Nonsyndromic Vesicoureteric Reflux

Primary vesicoureteric reflux accounts for approximately 10% of kidney failure requiring dialysis or transplantation, and sibling studies suggest a large genetic component. Here, we report a whole-genome linkage and association scan in primary, nonsyndromic vesicoureteric reflux and reflux nephropathy. We used linkage and family-based association approaches to analyze 320 white families (661 affected individuals, generally from families with two affected siblings) from two populations (United Kingdom and Slovenian). We found modest evidence of linkage but no clear overlap with previous studies. We tested for but did not detect association with six candidate genes (AGTR2, HNF1B, PAX2, RET, ROBO2, and UPK3A). Family-based analysis detected associations with one single-nucleotide polymorphism (SNP) in the UK families, with three SNPs in the Slovenian families, and with three SNPs in the combined families. A case-control analysis detected associations with three additional SNPs. The results of this study, which is the largest to date investigating the genetics of reflux, suggest that major loci may not exist for this common renal tract malformation within European populations.Vesicoureteric reflux (VUR) is abnormal movement of urine from the bladder retrogradely through the vesicoureteric junctions into the upper urinary tract. This is a study of primary VUR, i.e., VUR that is not secondary to bladder outflow obstruction caused by neurogenic damage or urethral valves or part of a multiorgan syndrome. VUR is usually a benign condition but can be associated with transient kidney damage, acute inflammation from ascending pyelonephritis, or permanent damage as a consequence of scarring after infection and/or congenital kidney defects histologically comprising renal hypoplasia (too few nephrons) and/or renal dysplasia (incomplete differentiation).^1–3 These renal defects are grouped under the term reflux nephropathy (RN). In the United Kingdom, RN accounts for 12% of the approximately 40,000 adults and 7% of the 768 children who require renal transplantation and/or life-long dialysis.⁴Traditionally, the diagnosis of VUR has been based on cystography with radiodense or radioisotopic materials to visualize retrograde passage of urine. Williams et al.³ reviewed 15 cystography studies in well children: The largest study reported no VUR in 722 children, whereas some of the smaller studies reported much higher percentages of affected individuals. The true prevalence of (primary) VUR in children remains uncertain: 1% is probably conservative, and 10 to 20% is possible.³ Screening studies of first-degree relatives of individuals with VUR identifies reflux in one third to one half of siblings^5,6 and 65% of offspring.⁷ Futhermore, there is a high concordance of primary VUR in identical twins,⁸ and families have been identified with multiple generations affected by primary VUR and RN.⁹ Collectively, these studies suggest that there is a substantial genetic component to VUR.The first genome-wide linkage analysis for VUR, based on seven kindreds,⁹ provided preliminary evidence for a locus on chromosome 1 and also for genetic heterogeneity. In this study, multipoint parametric and nonparametric linkage analysis was undertaken; however, one of the markers defining the interval on chromosome 1, GATA176C01, was subsequently found to be on chromosome 2 (Ensembl release 55, July 2009), so this localization should be treated with caution. Subsequent studies using similar kindreds^10–12 have supported the notion that the condition is genetically heterogeneous. In the largest linkage study of VUR before this report, Kelly et al.¹³ performed a linkage genome scan of 609 individuals (283 affected individuals in 129 families) and detected six to seven regions with suggestive evidence of linkage,¹⁴ one of which at chromosome 2q37 attained genome-wide significance when analyzed in a phenotypically derived subset of the data. The high incidence in offspring of affected individuals and the large number of pedigrees consistent with autosomal dominant inheritance, albeit with reduced penetrance, is in keeping with a dominant model; however, recently, a locus was identified on 12p11-q13 using a recessive model.¹⁵Here we report on linkage and association analysis in affected sibling pairs from two populations. We used the Affymetrix NspI array to generate genome-wide data, adding in haplotype-tagging single-nucleotide polymorphisms (SNPs) to obtain full coverage for six candidate genes: AGTR2, HNF1B, PAX2, RET, ROBO2, and UPK3A.¹⁶