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Renal fibrosis is thought to be the final common pathway leading to chronic kidney disease (CKD) and end-stage renal failure. Except for renal replacement therapy, no adequate treatment regimen is available; therefore studies on the treatment of renal fibrosis have attracted significant interest. In recent years, studies have shown that traditional Chinese medicine (TCM) may represent an attractive source to produce drugs with antifibrosis effects. The aim of this study was to establish a robust cell-based high-content screening (HCS) approach to identify TCM compounds with antifibrosis effects in NRK49F cells following TGF-β1 exposure. When designing the model, one of the most important steps involved the stability and reproducibility of this cell-based model. Therefore, we initially optimized the experimental parameters. Then, our HCS model was validated using SB525334, an inhibitor of the TGF-β1 receptor, and curcumin and emodin, two TCM compounds with well-documented anti-renal fibrosis activity. Subsequently, the proven reliable HCS model was used to screen a standard TCM compound library, which included 344 TCM molecules. Based on our HCS algorithm, a total of 16 compounds were identified to have prospective inhibitory activity. These compounds were further validated by verification experiments. Strikingly, eight compounds have been shown to inhibit renal fibrosis; six of them had rarely been described in the literature, namely, Ligustrazine, Glycyrrhizic acid, Astragaloside iv, Hydroxysafflor Yellow A, Crocin, and Gypenosides. To the best of our knowledge, this is the first study in which a HCS assay was performed to identify TCM compounds with anti-renal fibrosis effects. The HCS approach was successfully applied to screen active compounds and will be propitious to further anti-renal fibrosis drugs discovery research. Meanwhile, it may offer possibilities for identifying lead compounds for treating other diseases from registered Chinese herbal medicines.

Establishing a Cell-Based High-Content Screening Assay for TCM Compounds with Anti-Renal Fibrosis Effects