Huaier aqueous extract inhibits colorectal cancer stem cell growth partially via downregulation of the Wnt/β-catenin pathway

Tao Zhang, Ke Wang, Junshu Zhang, Xiaolei Wang, Zhigang Chen, Chao Ni, Fuming Qiu, Jian Huang, Tao Zhang, Ke Wang, Junshu Zhang, Xiaolei Wang, Zhigang Chen, Chao Ni, Fuming Qiu, Jian Huang

Abstract

The existence of cancer stem cells (CSCs) is central to the pathogenesis and therapy resistance of colorectal cancer. The aim of this study was to evaluate whether Huaier aqueous extract, a Chinese medicine, has efficacy against CSCs and to investigate the mechanisms of its anticancer effects. It was observed that the Huaier extract significantly inhibited the spheroid formation potential (P<0.05) and decreased the aldehyde dehydrogenase (ALDH)-positive cell population in colorectal primary cancer cells (P<0.05). Western blotting analysis and Wnt/β-catenin reporter assays revealed that the Huaier extract downregulated the Wnt/β-catenin self-renewal pathway. This is the first study to demonstrate that Huaier aqueous extract acts as an effective agent for eradicating colorectal CSCs and identifies the Wnt/β-catenin pathway as its potential target, which may be a new approach for colorectal cancer therapy.

Keywords: Huaier aqueous extract; antitumor activity; cancer stem cells; colorectal cancer.

Figures

Figure 1
Figure 1
Huaier aqueous extract suppresses cell viability in colorectal cancer cells. (A) Dose-response curves of primary colorectal cancer cells (T1 and T2 cells) treated with Huaier extract for 48 h. The antiproliferative effect of the Huaier aqueous extract was measured using the MTS assay. Bars denote the standard deviation. (B) Huaier extract caused morphological changes in the colorectal cancer cells. The images of colorectal cancer cells were obtained 48 h after the addition of Huaier extract.
Figure 2
Figure 2
Huaier aqueous extract inhibits the formation of spheroids. (A) Colorectal cancer cells (T1 and T2 cells) were incubated with the Huaier extract for 14 days under spheroid-forming conditions. The size of the spheroids was then evaluated under a microscope. (B) Quantification of spheroid-formation with colorectal cancer cells treated as in (A). Bars denote the standard deviation. *P<0.05 vs. untreated cells.
Figure 3
Figure 3
Huaier aqueous extract decreases the ALDH-positive cell population. Colorectal cancer cells (T1 and T2 cells) were treated with Huaier extract for seven days and subjected to the Aldefluor assay. (A) Proportion of the ALDH-positive cell subpopulation in total cancer cells. Bars denote the standard deviation.*P<0.05 vs. untreated cells. (B) Representative flow cytometry. ALDH1, aldehyde dehydrogenase.
Figure 4
Figure 4
Huaier aqueous extract downregulates the Wnt/β-catenin pathway. (A) Expression of β-catenin and cyclin D1 was detected by western blot analysis in colorectal cancer cells either untreated or treated with Huaier extract for 48 h. (B) TOP/FOPflash reporter transiently infected colorectal cancer cells were treated with Huaier extract for 24 h. The Huaier extract decreased the percentage of TOP reporter. Bars denote the standard deviation (*P<0.05).

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