Liuwei Dihuang Decoction Drug-containing Serum Attenuates Transforming Growth Factor-beta 1-induced Epithelial-mesenchymal Transition in HK-2 Cells by Inhibiting NF-kappa B/Snail Signaling Pathway

Objectives: The nuclear factor-kappa B (NF-kappa B) signaling pathway plays an important role in regulating tubular epithelial-mesenchymal transition (EMT), an indispensable cellular programme for driving organ fibrosis and tumor progression. Liuwei Dihuang Decoction (LWD) is an effective Chinese formula for treating chronic renal failure. Methods: First, by using morphological examination, immunofluorescence staining assay, RT-qPCR, and Western blot analysis, in vitro experiments were designed to analyze NF-kappa B and EMT markers (including Snail, alpha-SMA, and E-cadherin) in transforming growth factor-beta 1 (TGF-beta 1) induced renal tubular epithelial cells (HK-2) and to detect the expression levels of LWD-CS co-treatment. Then, the recombinant lentiviral vector was overexpressed and knocked down by NF kappa B and transfected into HK-2 cells. Cells were treated with TGF-beta 1 (10 ng/ml) with blank serum or LWD-containing serum, respectively, and the expression of these molecules in the NF-kappa B/Snail signaling pathway was further evaluated. Results: Our results confirmed that TGF-beta 1 could induce EMT, nuclear translocation of NF-kappa B p65, and activate the NF-kappa B/Snail signaling pathway in HK-2 cells. Furthermore, NF-kappa B knocked-down dramatically increases the TGF-beta 1-induced mRNA and protein expression level of E-cadherin and reduces the level of Snail and alpha-SMA; this is reversed by NF-kappa B overexpression. LWD can decrease the EMT levels through the NF-kappa B/Snail signaling activation in TGF-beta 1-induced EMT of HK-2 cells. Conclusion: The present study provides evidence suggesting a novel mechanism that LWD exerts anti-fibrosis effects through inhibiting activation of the NF-kappa B/Snail signaling pathway and consequently downregulating the TGF-beta 1-induced EMT in renal tubular epithelial cells.