Epithelial Cells as Active Player In Fibrosis: Findings from an <em>In Vitro</em> Model

Kidney fibrosis, a scarring of the tubulo-interstitial space, is due to activation of interstitial myofibroblasts recruited locally or systemically with consecutive extracellular matrix deposition. Newly published clinical studies correlating acute kidney injury (AKI) to chronic kidney disease (CKD) challenge this pathological concept putting tubular epithelial cells into the spotlight. In this work we investigated the role of epithelial cells in fibrosis using a simple controlled in vitro system. An epithelial/mesenchymal 3D cell culture model composed of human proximal renal tubular cells and fibroblasts was challenged with toxic doses of Cisplatin, thus injuring epithelial cells. RT-PCR for classical fibrotic markers was performed on fibroblasts to assess their modulation toward an activated myofibroblast phenotype in presence or absence of that stimulus. Epithelial cell lesion triggered a phenotypical modulation of fibroblasts toward activated myofibroblasts as assessed by main fibrotic marker analysis. Uninjured 3D cell culture as well as fibroblasts alone treated with toxic stimulus in the absence of epithelial cells were used as control. Our results, with the caveats due to the limited, but highly controllable and reproducible in vitro approach, suggest that epithelial cells can control and regulate fibroblast phenotype. Therefore they emerge as relevant target cells for the development of new preventive anti-fibrotic therapeutic approaches.肾纤维化（Kidney fibrosis）是指肾小管间质间隙（tubulo-interstitial space）发生瘢痕化，其发病与局部或全身募集的间质肌成纤维细胞（interstitial myofibroblasts）活化及后续细胞外基质沉积（extracellular matrix deposition）密切相关。此前主流病理学说秉持上述机制，但最新发表的将急性肾损伤（acute kidney injury, AKI）与慢性肾病（chronic kidney disease, CKD）相关联的临床研究对此提出了挑战，使肾小管上皮细胞成为研究焦点。本研究借助简易可控的体外（in vitro）实验体系，探究了上皮细胞在纤维化进程中的作用。我们构建了由人近端肾小管上皮细胞（human proximal renal tubular cells）与成纤维细胞（fibroblasts）组成的上皮-间质三维细胞培养模型，通过给予毒性剂量的顺铂（Cisplatin）损伤上皮细胞。随后，分别在存在或缺失该损伤刺激的条件下，采用逆转录聚合酶链反应（RT-PCR）检测成纤维细胞的经典纤维化标志物（classical fibrotic markers）表达水平，以评估其向活化肌成纤维细胞表型（activated myofibroblast phenotype）的转化情况。实验结果显示，上皮细胞损伤可触发成纤维细胞向活化肌成纤维细胞的表型转化，这一结论通过主要纤维化标志物的检测得到验证。本研究设置了两组对照：未受损伤的三维细胞培养体系，以及无上皮细胞存在、仅用毒性刺激处理的单纯成纤维细胞培养体系。尽管本研究所用的体外模型存在一定局限，但该体系具备高度可控性与可重复性，我们的结果提示上皮细胞能够调控成纤维细胞的表型。因此，上皮细胞有望成为开发新型预防性抗纤维化治疗手段的关键靶细胞。