Inhibitory effect of Sanguinarine on TGF-β1-induced endothelial-mesenchymal transition of HUVEC

Abstract

Objective To investigate the effect of Sanguinarine (SAN) on TGF-β1-induced endothelial-mesenchymal transition (EMT) of human umbilical vein endothelial cells (HUVECs) and its underlying mechanism. Methods TGF-β1 (10 ng/mL) was used to stimulate HUVECs to observe the effect of SAN at various concentrations on TGF-β1-induced EMT of HUVEC. CCK8 assay was employed to detect the cell viability of HUVECs stimulated by SAN. The morphology of HUVEC was observed under inverted phase-contrast microscope. The expression levels of collagen-1, fibronectin, N-cadherin in HUVEC were determined by real-time semi-quantitative reverse transcription polymerase chain reaction (RT-PCR). Western blot was conducted to detect the expressions of p-Smad2, Smad2, p-Smad3 and Smad3 in HUVECs. Results CCK8 assay showed that the cell viability of HUVEC did not change obviously under the stimulation of SAN at various concentrations for 72 h. Optical microscopy indicated that HUVEC exhibited regular ' cobblestone' morphology under normal circumstances but switched to fusiform cells after treated with TGF-β1. This phenomenon was apparently regressed under the costimulation of TGF-β1 and SAN (0.25 μM). The results of RT-PCR showed that the expression levels of collagen-1, fibronectin, N-cadherin were upregulated with the stimulation of TGF-β1. Western blot showed that the expressions of p-Smad2 and p-Smad3 increased under the stimulation of TGF-β1, but was regressed by addition of SAN. Conclusion SAN could inhibit the development of EMT of HUVEC induced by TGF-β1, suggesting its potentially important role in the treatment of myocardial fibrosis. Key words: Sanguinarine; TGF-β1; Human umbilical vein endothelial cells; Endothelial-mesenchymal transition