Shentu Yangping, Jiang Huanchang, Liu Xiaoyuan, Chen Hao, Yang Dicheng, Zhang Jinqi, Cheng Chen, Zheng Yulin, Zhang Yang, Chen Chaosheng, Zheng Chenfei, Zhou Ying
Department of Pathology, The First Affiliated Hospital of Wenzhou Medical University, Wenzhou, China.
Department of Internal Medicine, The First Affiliated Hospital of Wenzhou Medical University, Wenzhou, China.
Front Physiol. 2020 Dec 16;11:517912. doi: 10.3389/fphys.2020.517912. eCollection 2020.
Peritoneal dialysis (PD) is a treatment for end stage renal disease patients, but it can also cause peritoneal fibrosis. Nestin is known as a neural stem cell marker and it has many functions. The hypoxia induced factor (HIF) signaling pathway can be activated under hypoxia conditions, leading to the overexpression of some angiogenesis related genes. The aim of our study is to demonstrate Nestin's role in the development of peritoneal fibrosis (PF), and to provide a new target (Nestin) to treat PF.
PD mice models were constructed by an intraperitoneal administration of PDS at 10 ml/100g/d for 4 weeks. Nestin-positive cells were isolated from peritonea of Nestin-GFP mice by flow cytometry. The relationship of Nestin and HIF1-α-VEGFA pathway was detected by Nestin knockdown, Co-immunoprecipitation and immunofluorescence. Also, proteasomal activity was demonstrated by CHX and MG132 application, followed by Western blotting and Co-immunoprecipitation.
In our experiments, we found that Nestin expression resulted in PF. Also, HIF1-α/VEGFA pathway was activated in PF. Nestin knockdown reduced the level of HIF1-α. Nestin directly bound to HIF1-α and protected HIF1-α from proteasomal degradation. Overexpression of HIF1-α reverts the fibrosis levels in Nestin-knockdown cells. In brief, Nestin inhibited the degradation of HIF1-α by mitigating its ubiquitination level, leading to the activation of HIF1-α signaling pathway, and eventually promoted PF.
We found a novel mechanism of PF that Nestin promotes by protecting HIF1-α from proteasomal degradation. Taken together, our key findings highlight a novel mechanism by which the silencing of Nestin hinders HIF1- α -induced PF.
腹膜透析（PD）是终末期肾病患者的一种治疗方法，但它也会导致腹膜纤维化。巢蛋白是一种神经干细胞标志物，具有多种功能。缺氧诱导因子（HIF）信号通路在缺氧条件下可被激活，导致一些血管生成相关基因的过表达。本研究的目的是证明巢蛋白在腹膜纤维化（PF）发展中的作用，并为治疗PF提供一个新靶点（巢蛋白）。
通过腹腔注射10 ml/100g/d的PDS构建PD小鼠模型，持续4周。通过流式细胞术从巢蛋白绿色荧光蛋白（Nestin-GFP）小鼠的腹膜中分离出巢蛋白阳性细胞。通过巢蛋白敲低、免疫共沉淀和免疫荧光检测巢蛋白与HIF1-α-VEGFA通路的关系。此外，通过应用环己酰亚胺（CHX）和MG132证明蛋白酶体活性，随后进行蛋白质印迹和免疫共沉淀。
在我们的实验中，我们发现巢蛋白表达导致PF。此外，HIF1-α/VEGFA通路在PF中被激活。巢蛋白敲低降低了HIF1-α的水平。巢蛋白直接与HIF1-α结合，并保护HIF1-α不被蛋白酶体降解。HIF1-α的过表达逆转了巢蛋白敲低细胞中的纤维化水平。简而言之，巢蛋白通过降低其泛素化水平抑制HIF1-α的降解，导致HIF1-α信号通路的激活，并最终促进PF。
我们发现了PF的一种新机制，即巢蛋白通过保护HIF1-α不被蛋白酶体降解来促进PF。综上所述，我们的主要发现突出了一种新机制，即巢蛋白的沉默阻碍HIF1-α诱导的PF。
