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Altered Anoctamin-1 (ANO 1) Tyrosine Phosphorylation In Congenital Ureteropelvic Junction Obstruction

Saturday, October 20, 2012
Grand Ballroom A/B (Hilton Riverside)
Manuela Hunziker1, Jan-H. Gosemann1, Anne-Marie O'Donnell1, Nicolae Corcionivoschi1, Luis A. Alvarez2 and Prem Puri1, (1)Our Lady's Children's Hospital, National Children's Research Centre, Dublin, Ireland, (2)University College Dublin, Conway Institute of Biomedical Research, Dublin, Ireland

Purpose: Ureteropelvic junction (UPJ) obstruction is the most common cause of congenital hydronephrosis in children. The pathophysiology of UPJ obstruction and the exact mechanism of pelviureteral peristalsis are poorly understood. Coordinated contractions of smooth muscle cells (SMCs) produce the motor patterns for transmission of peristaltic waves across the UPJ. ANO1, a Ca2+ activated chloride channel, has been shown to play a key role in muscle wall contractions in the gastrointestinal tract. Expression of ANO1 has also recently been described in SMCs of the murine renal pelvis and in the urethra of mice, rats and sheep. Furthermore, phosphorylation has been shown to inhibit Ca2+ activated chloride channels in SMCs. We designed this study to investigate the hypothesis that ANO1 is expressed in SMCs of the human UPJ and that ANO1 tyrosine phosphorylation is altered in UPJ obstruction.

Methods: Fresh frozen specimens of ureteropelvic junction obstruction (n=10) and control specimens from patients who underwent Wilms' tumor nephrectomy (n=5) were prepared. Western blot was performed to evaluate ANO1 expression and tyrosine phosphorylation patterns. In addition analysis of ANO1 and α-smooth muscle actin using confocal-immunofluoresence-double staining technique and 3D reconstruction was carried out.

Results: Western blot revealed markedly increased tyrosine phosphorylation in UPJ obstruction compared to controls (Fig. 1). Western blotting showed decreased ANO1 expression in UPJ obstruction compared to controls (Fig 2). ANO1 immunoreactivity was decreased in SMCs of UPJ obstruction compared to controls (Fig. 3).

Conclusion: We provide evidence, for the first time, of the presence of ANO1 expression in the human UPJ. Altered ANO 1 tyrosine phosphorylation observed in UPJ obstruction may inhibit Ca2+ activated chloride channels in SMCs leading to failure of transmission of peristaltic waves in ureteropelvic junction obstruction.