National Children's Research Centre, Our Lady's Children's Hospital, Dublin, Ireland; School of Medicine and Medical Science and Conway Institute of Biomolecular and Biomedical Research, University College Dublin, Ireland.
National Children's Research Centre, Our Lady's Children's Hospital, Dublin, Ireland.
J Pediatr Surg. 2020 Aug;55(8):1621-1625. doi: 10.1016/j.jpedsurg.2020.02.001. Epub 2020 Feb 11.
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. Anoctamin-1 (ANO1), a Ca-activated chloride channel, has been shown to play a key role in muscle wall contractions in the gastrointestinal tract. We designed this study to investigate the hypothesis that ANO1 is expressed in smooth muscle cells (SMCs) of the human UPJ and that tyrosine phosphorylation is altered in UPJ obstruction.
Fresh frozen specimens of UPJ obstruction (n = 28) and control specimens from patients who underwent Wilms' tumor nephrectomy (n = 20) were prepared. Western blot (WB) was performed to evaluate levels of ANO1 protein expression and changes in tyrosine phosphorylation. In addition analysis of ANO1 and phalloidin using confocal-immunofluoresence-double staining and 3D reconstruction were carried out.
Our WB results revealed increased tyrosine phosphorylation in UPJ obstruction samples compared to controls, and decreased ANO1 expression in UPJ obstruction. Confocal microscopy showed that ANO1 immunoreactivity was decreased in SMCs of UPJ obstruction compared to controls.
We provide evidence, for the first time, of the presence of ANO1 expression in the human UPJ. We speculate that altered tyrosine phosphorylation, observed in UPJ obstruction, may lead to a failure of transmission of peristaltic waves in UPJ obstruction by inhibiting Ca-activated chloride channels in SMCs.
肾盂输尿管连接部（UPJ）梗阻是儿童先天性肾积水的最常见原因。UPJ 梗阻的病理生理学和肾盂输尿管蠕动的确切机制尚未完全清楚。Anoctamin-1（ANO1），一种钙激活的氯离子通道，已被证明在胃肠道的肌壁收缩中发挥关键作用。我们设计本研究旨在验证以下假说，即 ANO1 在人 UPJ 的平滑肌细胞（SMCs）中表达，并且 UPJ 梗阻时酪氨酸磷酸化发生改变。
制备 UPJ 梗阻（n=28）和因 Wilms 瘤行肾切除术（n=20）的患者的 UPJ 梗阻和对照标本。通过 Western blot（WB）评估 ANO1 蛋白表达水平和酪氨酸磷酸化的变化。此外，还使用共聚焦免疫荧光双重染色和 3D 重建分析 ANO1 和鬼笔环肽。
我们的 WB 结果显示，与对照组相比，UPJ 梗阻样本中的酪氨酸磷酸化增加，而 ANO1 表达减少。共聚焦显微镜显示，与对照组相比，SMCs 中 ANO1 免疫反应性在 UPJ 梗阻中降低。
我们首次提供了 ANO1 在人 UPJ 中表达的证据。我们推测，在 UPJ 梗阻中观察到的改变的酪氨酸磷酸化可能通过抑制 SMC 中的钙激活氯离子通道，导致 UPJ 梗阻中蠕动波的传递失败。
