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Overexpressed sFRP3 exerts an inhibitory effect on hepatocellular carcinoma via inactivation of the Wnt/β-catenin signaling pathway

Abstract

Hepatocellular carcinoma (HCC) is recognized as the most common malignancy of the liver in adults. Many human cancers have been associated with the oncogenic activation of the Wnt/β-catenin signaling pathway. The secreted frizzled-related proteins (sFRPs) function as negative regulators of the Wnt signaling and have important implications in carcinogenesis. This study aims to investigate the possible regulatory effects of sFRP3 on the Wnt/β-catenin signaling pathway and their interactions in HCC occurrence. Firstly, sFRP3 expression was quantified in the collected cancer and adjacent normal tissue samples from HCC patients. The lowly expressed sFRP3 in HCC tissues was found to be correlated with HCC development. The expression of sFRP3 was regulated by a lentivirus-based packaging system, and the Wnt/β-catenin signaling pathway was inactivated by DDK-1 in HepG2 cells. The expressions of Wnt1, β-catenin and the nuclear translocation of β-catenin were determined, both of which were down-regulated by sFRP3 overexpression. CCK8 assay, EdU staining, Colony formation assay, flow cytometry, scratch test and Transwell assay were employed to test cell viability, proliferation, cell cycle, apoptosis, migration and invasion, respectively. Overexpressed levels of sFRP3 were found to produce a reduction in MMP-2, MMP-7, MMP-9, PCNA, Ki67, and Bcl-2 expressions but an increase in the expressions of caspase-3 and Bax. In addition, overexpression of sFRP3 inhibited cell proliferation, migration, invasion, and colony formation, but promoted cell cycle arrest and cell apoptosis in HCC cells. The addition of the Wnt/β-catenin signaling pathway inhibitor, DKK-1, reversed the contributory effect of sFRP3 silencing on HCC development. Lastly, in vivo tumor formation was inhibited by enforced sFRP3 expressions. The obtained results suggested that sFRP3 acts as an anti-oncogene in HCC by inhibiting the activation of the Wnt/β-catenin signaling pathway.

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Fig. 1: sFRP3 is poorly expressed in HCC tissues and cell lines.
Fig. 2: Overexpression of sFRP3 blocks HepG2 cell proliferation but promotes apoptosis.
Fig. 3: Overexpressed sFRP3 suppresses migration and invasion of HepG2 cells.
Fig. 4: Overexpressed sFRP3 suppresses the activation of Wnt1/β-catenin signaling pathway in HepG2 cells.
Fig. 5: sFRP3 suppresses HepG2 cell proliferation and promotes cell apoptosis via inhibition of the Wnt1/β-catenin signaling pathway.
Fig. 6: sFRP3 suppresses HepG2 cell migration and invasion via inhibition of the Wnt1/β-catenin signaling pathway.
Fig. 7: Overexpressed sFRP3 suppresses tumor formation and growth in nude mice.

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Acknowledgements

This study was supported by Beijing Natural Science Foundation (No. 7202181), China-Japan Friendship Hospital Youth Science and Technology Excellence Project (No. 2014-QNYC-B-01), the Research Fund of the China-Japan Friendship Hospital (No. 2014-2-MS-9) and the National Natural Science Foundation (No. 91729301; No. 81630103).

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Correspondence to Shi-Yu Du.

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Fang, L., Gao, C., Bai, RX. et al. Overexpressed sFRP3 exerts an inhibitory effect on hepatocellular carcinoma via inactivation of the Wnt/β-catenin signaling pathway. Cancer Gene Ther 28, 875–891 (2021). https://doi.org/10.1038/s41417-020-0201-z

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