OBJECTIVE Ureteropelvic junction obstruction (UPJO) is a common renal obstructive disorder, but its pathogenic mechanisms remain largely unclear. We aimed to investigate the potential involvement of the renin-angiotensin system in congenital UPJO pathogenesis. METHODS Differentially expressed proteins in exosomes isolated from amniotic fluid of patients with congenital UPJO were characterized using iTRAQ (isobaric tags for relative and absolute quantification)-based proteomics. The expressions of angiotensin-converting enzyme (ACE) and aminopeptidase N (AP-N) in HK2 cells were inhibited by quinapril and siRNA, respectively. Cell proliferation and reactive oxygen species were measured by EdU staining and flow cytometry, respectively. Gene expression was detected by Western blot or qRT-PCR. The inflammatory factors were measured through ELISA. Mice that underwent unilateral ureteral obstruction were used as the animal model. RESULTS The identity of exosomes from amniotic fluids was confirmed by the expression of CD9 and CD26. In total, 633 differentially expressed proteins were identified in the amniotic fluid-derived exosomes from patients with UPJO, including 376 up- and 257 down-regulated proteins associated with multiple biological processes. Of them, ACE and AP-N were significantly decreased in the amniotic fluid exosomes. Inhibition of ACE and AP-N resulted in suppressed cell proliferation; repressed IARP, AT1R, and MAS1 expression; elevated ROS production; and increased IL-1β, TNF-α, and IL-6 levels in HK2 cells. Decreased ACE expression and elevated IL-1β levels were also observed in the mouse model. CONCLUSION Suppression of ACE and AP-N expression mediates congenital UPJO pathogenesis by repressing renal tubular epithelial proliferation, promoting ROS production, and enhancing inflammatory factor expression.

中文翻译：

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基于iTRAQ的蛋白质组学和体外实验揭示了ACE和AP-N在肾素-血管紧张素系统介导的先天性输尿管盆腔连接阻塞中的重要作用。

目的输尿管骨盆连接处梗阻（UPJO）是一种常见的肾梗阻性疾病，但其致病机制仍不清楚。我们旨在研究肾素-血管紧张素系统在先天性UPJO发病机理中的潜在作用。方法使用基于iTRAQ（相对和绝对定量的等压标记）的蛋白质组学对先天性UPJO患者羊水分离的外泌体中差异表达的蛋白质进行表征。奎那普利和siRNA分别抑制HK2细胞中血管紧张素转化酶（ACE）和氨肽酶N（AP-N）的表达。通过EdU染色和流式细胞术分别测量细胞增殖和活性氧种类。通过蛋白质印迹或qRT-PCR检测基因表达。通过ELISA测量炎性因子。将经历了单侧输尿管阻塞的小鼠用作动物模型。结果通过CD9和CD26的表达证实了羊水中外泌体的身份。在来自UPJO患者的羊水来源的外泌体中，总共鉴定出633种差异表达的蛋白质，包括376种与多种生物学过程相关的上调蛋白和257种下调蛋白。其中，羊水外泌体中ACE和AP-N显着降低。ACE和AP-N的抑制导致细胞增殖受到抑制；抑制了IARP，AT1R和MAS1的表达；ROS产生增加；HK2细胞中的IL-1β，TNF-α和IL-6水平升高。在小鼠模型中还观察到ACE表达降低和IL-1β水平升高。