| Background and object:Colorectal cancer (CRC) is one of the most common tumor diseases in China [1]. Several factors have been shown to contribute to colon cancer, such as inflammatory bowel disease, obesity, high consumption of meat or alcohol, smoking and diabetes. The combination of surgery, locational radiotherapy, chemotherapy and immunological therapy has been the main treatment of CRC. Even so, a poor prognosis has been showed in the advanced CRC patients. Tetrastigma hemsleyanum Diels et Gilg (Sanyeqing), a traditional Chinese medicinal herb, exhibits a tumor suppressing effects in several malignant diseases. And it is used in the folk therapy of pneumonia, hepatitis, scrofula, infantile febrile convulsion, high fever, menstrual disorders, sore throat, asthma and rheumatism. The rhizomes of tetrastigma hemsleyanum are rich in flavonoids, including rutin, quercitrin, isoquercitrin, astragalin, kaempferol-3-O-rutinoside, procyanidin B1/B2 and catechin. The mixture is called total flavonoids of tetrastigma hemsleyanum (TFTH) and several of them have been reported to inhibit the proliferation of tumor cells. In addition, TFTH also has been reported to elicit a strong anti-tumorigenic activity against several types of carcinoma.In this study, we treated CRC with different TFTH in vitro, and evaluated if TFTH could prevent inflammation-associated CRC in mice. Besides, we also investigated the underlying mechanism of TFTH-mediated tumor suppression.Materials and Methods:Human colon cancer cell lines HT29 and SW620 were used in this research. The proliferation of colon cancer cells was evaluated through CCK8 assay and plate colony formation assay. Cell invasion and migration were evaluated by Cell wound scratch assay and Transwell invasion experiments. Cell cycle was measured by flow cytometry. Furthermore, western blot and quantitative reverse transcription PCR (RT-qPCR) were performed to evaluate the expression of genes, such as Leucine-rich repeat-containing G-protein-coupled receptor 5 (Lgr5), Cyclin Dl, c-Myc, E-Cadherin and Vimentin. Mice were induced to form CRC with azoxymethane (AOM) and dextran sulfate sodium (DSS), and treated with either daily TFTH gavage or vehicle. Tumor volume and number were compared between the groups.Results:The results suggested that TFTH treatment could decreased cell proliferation, clonal forming capacity, invasion and migration ability. Cell cycle was trapped in G0/G1-phase by TFTH in a dose-dependent manner. EMT process was inhibited after TFTH treatment. Meanwhile, TFTH down-regulated the activation of Wnt/β-catenin pathway, increased protein level of Vimentin and reduced the expression of Lgr5, Cyclin D1, c-Myc and E-Cadherin. AOM/DSS models, Furthermore, we analyzed the effect of TFTH on inflammation-associated colorectal cancer in mice. Compared with control groups, mice given a treatment of TFTH via gavage had substantially fewer tumors and reduced tumor volume. More importantly, mice that received TFTH treatment displayed significantly fewer colorectal tumors. Colorectal sections demonstrated that TFTH-treated groups showed minor heteromorphism, highlighting the strong anti-cancer activity of TFTH in vivo. Additionally, Wnt/p-catenin pathway expression were examined in colorectal sections. The treatment groups showed decreased the levels of both Lgr5 and Cyclin D1 expression. Thus, TFTH may act as an anti-proliferation drug of CRC by inhibiting Wnt pathway activation, suppressing Lgr5 and Cyclin D1 and reducing the sternness of tumor cells and blocking the mitotic cycle.Conclusions:In conclusion, the results suggested that TFTH inhibits the cell proliferation, invasive and migratory ability of CRC in vitro and in vivo by supressing activity of the Wnt/β-catenin pathway and modulating the expression of associated genes, Lgr5, Cyclin D1, c-myc, E-cadherin and Vimentin expression. In this study, we verified the effective components of Tetrastigma hemsleyanum through modern pharmacological theory, providing a new direction for CRC therapy. |
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