The position of ureteric budding and its relationship to vesico-ureteric reflux and kidney defects in mice

Vesico-ureteric reflux (VUR) is a congenital urinary tract defect that results in the retrograde flow of urine from the bladder to the kidneys. It is caused by a defect in the formation of the uretero-vesical junction such that the ureter does not insert properly into the bladder. VUR affects up to 1% of the population and patients are at increased risk of developing recurrent urinary tract infections, hypertension, and end-stage renal disease. Patients with VUR frequently have malformed kidneys and short intravesical ureters, one component of the uretero-vesical junction. The co-occurrence of these defects suggests that they arise from a common developmental mechanism, notably, abnormal formation of the ureteric bud. The ureteric bud is an outgrowth of the mesonephric duct and develops into both the kidney and the urinary tract. We hypothesize that proper ureteric bud development is critical for the formation of a normal kidney and an intact uretero-vesical junction. We further hypothesize that VUR is associated with mutations in genes that direct early formation of the ureteric bud. Our objective is to identify mouse models of VUR to determine if an abnormal ureteric bud precedes the development of small or malformed kidneys and a refluxing urinary tract.;Since VUR is associated with a malformed kidney in Pax2 1Neu+/- mice, but a normal kidney in C3H/HeJ mice, we characterized the kidney phenotype in C3H/HeJ mice by assessing for VUR and kidney size at adulthood. Surprisingly, C3H/HeJ mice continued to reflux, but now had larger kidneys than C57BL/6J mice. We hypothesized that this change in kidney size was caused by a reduction in nephron number at birth and subsequent renal hypertrophy by adulthood. Nephrons are the functional units of the kidney and reduced nephron number leads to reduced renal function and can predispose to hypertension, renal disease, and compensatory renal hypertrophy. We therefore counted the number of nephrons in newborn and adult C3H/HeJ and C57BL/6J kidneys. Our results demonstrated that C3H/HeJ mice had the same number of nephrons as C57BL/6J at birth and at adulthood. Furthermore, nephron number did not correlate with kidney size or body weight at birth or during adulthood in either mouse strain. These results suggest that in these inbred mice, nephron number appears to be independent of kidney size. Our results also demonstrate that C3H/HeJ mice are actually a model of VUR and normal kidneys.;From human studies, VUR can be a genetic disorder. To identify the gene(s) that cause(s) VUR in the C3H/HeJ mouse, we performed genetic studies on backcross and intercross progeny derived from a cross between C3H/HeJ and C57BL/6J mice. We identified a 24Mb genomic region on chromosome 12, VUR1, which is linked to the VUR phenotype (LOD=7.8, p=0.001). We hypothesize that the candidate gene(s) within VUR1 will be expressed either by the ureteric bud or by the surrounding mesenchyme and will be critical for establishing proper kidney and urinary tract development. We believe that the candidate gene(s) responsible for VUR in the C3H/HeJ mouse will be important to consider in human studies of VUR.;We identified VUR in two mouse lines at birth: the Pax2 1Neu+/- mouse and the inbred C3H/HeJ mouse. At postnatal day 1, Pax21Neu+/- and C3H/HeJ mice had a 32% and 100% incidence of VUR, respectively. Control mice (CD1 and C57BL/6J) had a 6.25% and 0% incidence of VUR, respectively. We measured kidney size in all mice tested for VUR and Pax21Neu+/- mice had small malformed kidneys while C3H/HeJ mice had small but no malformed kidneys at birth when compared to controls. To identify a specific urinary tract abnormality, we measured the length of the intravesical ureter. Both refluxing mouse models had significantly shorter intravesical ureters compared to controls, providing evidence for a defective uretero-vesical junction. Lastly, to understand the embryonic origin of VUR, we characterized the position of the ureteric bud along the mesonephric duct. At embryonic day 10.5, both Pax21Neu+/- and C3H/HeJ embryos had ureteric buds that exited from a more caudal position along the mesonephric duct compared to controls. Furthermore, detailed analysis of the kidney and urinary tracts throughout development revealed that both Pax21Neu+/- and C3H/HeJ embryos had urinary tracts that were severely delayed in their development such that the ureters achieved a independent insertion into the bladder later than in controls. Our data suggest that a caudal origin of the ureteric bud is associated with a delay in urinary tract development, a short intravesical ureter, and the development of VUR.