基于EMT的肾小管上皮-树突状细胞转分化及其Runx1-PU.1调控研究

In recent years, it becomes the focus of researches that EMT endows cells with properties of stem cells. Based on the previous studies, we found that renal tubular epithelial cells (RTEC) could transdifferentiate to dendritic -like cells exhibiting some phenotype and functionality of dendritic cells（DC） , further enhance inflammation in renal tubulointerstitium. However, the relationship between DC-like cells transdifferentiation and epithelial-mesenchymal transitions（EMT） is unclear. Recently, we further found that both RTEC stimulated with TGF-β and DC-like cells could express Runx1, a master regulator of haematopoiesis, and PU.1 which is required for differentiation of DC, as well as, a key downstream target of Runx1 in adult mouse haematopoiesis. It has been demonstrated that EMT derived cells exhibit differentiation potential of stem-like cells. Therefore, we speculate that this DC-like cells transdifferentiation may be another form of EMT in inflammation. Based on the relationship between EMT and stem cell-like properties, we make a hypothesis that RTEC may acquire the haematopoietic lineage differentiation potential through expressing Runx1 in the process of EMT, and may futher be drived differentiation to DC-like cells by induction of PU.1. Therefore, in this study, we will investigate the relationship between this DC-like cells transdifferentiation and EMT, as well as, the mechanisms of transdifferentiation through the regulation of Runx1 and PU.1 based on EMT and stem cell-like properties. It may futher elucidate the diversity role of RTEC and provide a novel therapeutic strategy in renal tubulointerstitial lesions and fibrosis

近年上皮间充质转化（EMT）尤其与干性关系受关注。我们发现肾小管上皮细胞（RTEC）也可以树突状细胞（DC）转化形式参与肾间质炎症调节，该转分化机制及与EMT关系不明。继而发现转分化后DC样细胞或经TGF-β刺激RTEC，均表达血系细胞分化转录因子Runx1及下游靶点PU.1。现知上皮细胞经历EMT可获得干细胞样分化潜能，且Runx1和PU.1分别参与调控早期胚胎造血干细胞产生以及DC分化发育。推测上述DC样转分化，可能系RTEC随炎症微环境改变经EMT获干性后另一分化形式，而Runx1和PU.1可能共同参与调控EMT中RTEC向DC转化。为此本项目从EMT与干性角度，拟通过观察DC样转分化与EMT关系，EMT及其干性对Runx1、PU.1表达影响，以及两者在DC样转分化中调控作用。以期结合EMT探讨DC样转分化的可能机制，为进一步阐释肾小管间质损伤及纤维化机制以及临床早期干预提供新策略。

本项目在前期研究基础上，基于EMT并结合RUNX1等调控，从分子调控机制层面，进一步探讨了肾小管上皮细胞DC样转分化以及与EMT两者转分化的相互关系。本项目首先继续从DC-SIGN免疫调节作用及机制角度，结合急性肾损伤患者观察，进一步验证了DC-SIGN可通过串话调控TLR4信号通路中Myd88非依赖途径，介导肾小管上皮细胞参与肾脏炎症损伤。继而从炎症性疾病角度，进一步验证该DC-SIGN/TLR4信号串话通路，亦与调控巨噬细胞参与动脉粥样硬化斑块形成密切相关。上述结果有助于进一步阐释炎症状态下DC-SIGN免疫调节作用机制，且提示抑制DC-SIGN与TLR4两者相互作用，可成为肾脏等炎症性疾病新的防治策略。在上述基础上，进一步从肾脏炎症机制角度，发现证实参与DC-SIGN表达调控的RUNX1，其不仅可通过与NF-κB p50相互作用并籍TLR4下游通路调控M1型巨噬细胞极化，亦可通过线粒体脂肪酸氧化途径调抑M2型巨噬细胞极化，介导肾脏炎症反应及肾损伤。表明RUNX1作为炎症反应关键调控分子，其亦可成为肾脏等炎症性疾病新的干预靶点。随后从肾小管上皮细胞DC样转分化与EMT关系角度，进一步证实RUNX1可协同PU.1调控DC-SIGN表达以及肾小管上皮细胞DC样转分化，同时证实RUNX1也参与调控肾小管上皮细胞EMT过程以及此过程中的DC样转分化。提示RUNX1上述双向调控作用，可能与炎症微环境中不同炎性刺激下肾小管上皮细胞出现上述两类转分化，以及两者转分化间存在紧密联系或互相影响有关。进一步从EMT及其干性角度，发现RUNX1调控作用与肾小管上皮细胞EMT中干性形成密切关联，且后者获得干性后可明显促进RUNX1高表达。结合上述结果表明，肾小管上皮细胞发生DC样转分化，可能与其EMT过程中获得干性后出现另一分化形式有关，而RUNX1及其下游PU.1可能是上述转分化的关键分子调控基础。在此基础上，继续从RUNX1调控EMT作用机制角度，结合体内体外实验，进一步证实RUNX1可籍PI3Kp110δ/Akt信号通路，调控EMT发生以及肾间质纤维化形成。表明RUNX1可能亦是参与肾纤维化形成的关键调控分子，且提示RUNX1-p110δ通路可成为肾纤维化防治的新靶标。上述结果有助于进一步阐述肾小管间质损伤及纤维化机制，并为临床防治提供新的研究思路及干预策略，均有待进一步研究。