Transforming growth factor-β (TGFβ) has been reported to be dysregulated in malformed ureters. There exists, however, little information on whether altered TGFβ levels actually perturb ureter development. We therefore hypothesised that TGFβ has functional effects on ureter morphogenesis. Tgfb1, Tgfb2 and Tgfb3 transcripts coding for TGFβ ligands, as well as Tgfbr1 and Tgfbr2 coding for TGFβ receptors, were detected by quantitative polymerase chain reaction in embryonic mouse ureters collected over a wide range of stages. As assessed by in situ hybridisation and immunohistochemistry, the two receptors were detected in embryonic urothelia. Next, TGFβ1 was added to serum-free cultures of embryonic day 15 mouse ureters. These organs contain immature smooth muscle and urothelial layers and their in vivo potential to grow and acquire peristaltic function can be replicated in serum-free organ culture. Such organs therefore constitute a suitable developmental stage with which to define roles of factors that affect ureter growth and functional differentiation. Exogenous TGFβ1 inhibited growth of the ureter tube and generated cocoon-like dysmorphogenesis. RNA sequencing suggested that altered levels of transcripts encoding certain fibroblast growth factors (FGFs) followed exposure to TGFβ. In serum-free organ culture exogenous FGF10 but not FGF18 abrogated certain dysmorphic effects mediated by exogenous TGFβ1. To assess whether an endogenous TGFβ axis functions in developing ureters, embryonic day 15 explants were exposed to TGFβ receptor chemical blockade; growth of the ureter was enhanced, and aberrant bud-like structures arose from the urothelial tube. The muscle layer was attenuated around these buds, and peristalsis was compromised. To determine whether TGFβ effects were limited to one stage, explants of mouse embryonic day 13 ureters, more primitive organs, were exposed to exogenous TGFβ1, again generating cocoon-like structures, and to TGFβ receptor blockade, again generating ectopic buds. As for the mouse studies, immunostaining of normal embryonic human ureters detected TGFβRI and TGFβRII in urothelia. Collectively, these observations reveal unsuspected regulatory roles for endogenous TGFβ in embryonic ureters, fine-tuning morphogenesis and functional differentiation. Our results also support the hypothesis that the TGFβ up-regulation reported in ureter malformations impacts on pathobiology. Further experiments are needed to unravel the intracellular signalling mechanisms involved in these dysmorphic responses. © 2019 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.

中文翻译：

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过度活跃或对转化生长因子-β的阻滞均会导致特定的输尿管畸形。

据报道，在畸形的输尿管中转化生长因子-β（TGFβ）失调。然而，关于改变的TGFβ水平是否实际上干扰输尿管发育的信息很少。因此，我们假设TGFβ对输尿管形态发生有功能作用。通过定量聚合酶链反应，在收集了广泛阶段的胚胎小鼠输尿管中检测到了编码TGFβ配体的Tgfb1，Tgfb2和Tgfb3转录本，以及编码TGFβ受体的Tgfbr1和Tgfbr2。如通过原位杂交和免疫组织化学评估，在胚胎尿路上皮中检测到两种受体。接下来，将TGFβ1添加至胚胎第15天小鼠输尿管的无血清培养物中。这些器官包含未成熟的平滑肌和尿道上皮层，它们在体内生长和获得蠕动功能的潜力可以在无血清器官培养物中复制。因此，这些器官构成了合适的发育阶段，通过该阶段来定义影响输尿管生长和功能分化的因素的作用。外源性TGFβ1抑制输尿管的生长并产生茧状畸形。RNA测序表明，暴露于TGFβ后，编码某些成纤维细胞生长因子（FGFs）的转录本水平发生了变化。在无血清器官培养物中，外源性FGF10而不是FGF18消除了由外源性TGFβ1介导的某些畸形作用。为了评估内源性TGFβ轴是否在发育中的输尿管中起作用，将胚胎第15天的外植体暴露于TGFβ受体的化学阻断作用；输尿管的生长得到增强，并且尿路上皮管出现了异常的芽状结构。这些芽周围的肌肉层减弱，蠕动受到损害。为了确定TGFβ的作用是否仅限于一个阶段，将小鼠胚胎第13天输尿管的外植体（更原始的器官）暴露于外源性TGFβ1，再次产生茧样结构，并暴露于TGFβ受体，再次产生异位芽。至于小鼠研究，正常胚胎人输尿管的免疫染色在尿路上皮中检测到了TGFβRI和TGFβRII。总的来说，这些观察结果揭示了在胚胎输尿管中内源性TGFβ的意外调节作用，微调的形态发生和功能分化。我们的结果也支持以下假设：输尿管畸形中报告的TGFβ上调会影响病理生物学。需要进一步的实验来揭示参与这些畸形反应的细胞内信号传导机制。©2019作者。John Wiley＆Sons Ltd代表大不列颠及爱尔兰病理学会出版的《病理学杂志》。