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KMID : 0363320160370010065
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Journal of Korean Oriental Internal Medicine
2016 Volume.37 No. 1 p.65 ~ p.76
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Inhibition of Migration and Invasion of Human Bladder Cancer 5637 cells by Hwangheuk-san
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Shim Won-Suk
Kim Min-Serh
Park Sang-Eun
Choi Yung-Hyun
Hong Sang-Hoon
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Abstract
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Objectives: The purpose of this study was to identify the inhibitory effects of Hwangheuk-san (HHS), a Korean multi-herb
formula comprising four medicinal herbs, on cell migration and invasion, two critical cellular processes that are often deregulated
during metastasis, using the human bladder cancer 5637 cell line.
Methods: Cell viability, motility, and invasion were assessed by 3-(4,5-dimethyl-2 thiazolyl)-2,5-diphnyl-2H-tetrazolium
bromide (MTT), wound healing migration, and Transwell assays, respectively. Gene expression was detected by Western blot
analysis. In addition, the activities of matrix metalloproteinases (MMPs) and the values for transepithelial electrical resistance
(TER) were analyzed using a Gelatinase Activity Assay Kit and an Epithelial Tissue Voltohmmeter, respectively.
Results: Our data indicated that within the concentration range that was not cytotoxic, HHS effectively inhibited the cell
motility and invasiveness of 5637 cells. HHS markedly decreased the expression and activity of MMP-2 and MMP-9, which
was associated with unregulation of tissue inhibitors of metalloproteinase (TIMP)-1 and TIMP-2. Further investigation revealed
that phosphorylation of phosphatidylinositol 3-kinase (PI3K) and AKT was decreased in HHS-treated 5637 cells, and a
PI3K/AKT inhibitor synergistically reduced the inhibition of migration and invasion and also inactivated MMP-2 and MMP-9.
Moreover, HHS increased the tightening of tight junctions (TJs), which was demonstrated by an increase in the TER, and reduced
the expression the levels of claudin family members (claudin-3 and -4), which are major components involved in the tightening of TJs.
Conclusions: The present findings demonstrated that HHS attenuated the migration and invasion of bladder cancer 5637
cells by modulating the activity of the PI3K/Akt signaling pathway and also through TJ tightening.
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KEYWORD
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Hwangheuk-san, invasion, MMP, PI3K/Akt, tight junction
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