Abstract
Chronic kidney disease (CKD) is a worldwide health problem with growing prevalence in developing countries. Renal tubular epithelial-mesenchymal transition (EMT) is a critical step and key factor in the development of this condition. Renal tubulointerstitial fibrosis is a basic pathological change at the later stages of the disease. Therefore, blocking the development of EMT could be a critical factor in curing CKD. We have established a cell-based high-content screening (HCS) method to identify inhibitors of EMT in human proximal tubular epithelial (HK-2) cells by automatic acquisition and processing of dual-fluorescent labeled images. With the aid of chromatographic separation and mass spectrometry, we achieved the rapid and reliable screening of active compounds from the Chinese herbal medicine Tong-Mai-Yang-Xin-Wan (TMYX) for treating EMT. Five fractions were found to exert anti-EMT activity and were further identified by liquid chromatography coupled with tandem mass spectrometry. Glycyrrhizic acid, glyasperin A, and licorisoflavan A were found to inhibit EMT. The proposed approach was successfully applied to screen active compounds from TMYX on TGF-β1-stimulated HK-2 cells and may offer a new means for identifying lead compounds for treating EMT from registered Chinese herbal medicines.
Highlights
Chronic kidney disease (CKD) is a worldwide health problem, and its incidence and mortality have increased in recent years
Several active compounds isolated from Chinese herbal medicines, such as salvianolic acid B [13,14,15], curcumin [16], and norcantharidin [17] were reported to prevent TGF-β1-induced proliferation in human proximal tubule epithelial (HK-2) cells
After the compounds stimulate the cells for an appropriate time, high-content images can be obtained by detecting of fluorescent markers with automated microscopy
Summary
Chronic kidney disease (CKD) is a worldwide health problem, and its incidence and mortality have increased in recent years. Identifying lead compounds with anti-EMT effects from Chinese herbal medicines is of extremely high relevance. Several active compounds isolated from Chinese herbal medicines, such as salvianolic acid B [13,14,15], curcumin [16], and norcantharidin [17] were reported to prevent TGF-β1-induced proliferation in human proximal tubule epithelial (HK-2) cells. A high-content whole-well imaging approach has been established to screen compounds against drug-induced cardiotoxicity [19]; and drug-induced human hepatotoxicity [20]. This kind of screening includes the identification of novel nuclear export inhibitors [21], as well as compounds that detect hepatotoxicity in HepG2 cells [22]. The anti-EMT effects of glycyrrhizic acid, glyasperin A, and licorisoflavan A were further validated by in vitro assays
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