Effect of Asperuloside on the malignant progression of colorectal cancer cells by regulating the SIRT1/AMPK signaling pathway
XU Ming-yue,FU Wen-zheng
Department of Colorectal Surgery, Tianjin Union Medical Center; Tianjin Coloproctology Institute, Tianjin300071, China
Abstract:
Objective To investigate the effect of Asperuloside on the malignant progression of colorectal cancer cells by regulating the silent mating type information regulation 2 homolog-1 (SIRT1)/adenosine monophosphate activated protein kinase (AMPK) signaling pathway. Methods Human colorectal cancer cell line HCT116 was cultured in vitro and its nude mouse transplantation model was constructed. The viability of HCT116 cells under different concentrations of Asperuloside treatment and the tumor growth in nude mice under different doses of Asperuloside treatment were detected to screen for the optimal in vitro cell action concentration and in vivo nude mouse action dose of Asperuloside. HCT116 cells and their nude mouse models were randomly separated into a control group, a Asperuloside group, an Ex-527 (SIRT1 inhibitor) group, and a Asperuloside+Ex-527 group. After grouping, CCK-8 assay, flow cytometry, Transwell assay, and cell scratch assay were applied to detect the cell viability, apoptosis, migration, and invasion of HCT116 cells in each group. The tumor volume of nude mice was detected, and immunohistochemical staining was applied to detect the growth of tumor cells (Ki67 positive rate) and microvascular density (CD34 positive rate) of nude mice in each group. Immunoblotting was applied to detect the expression of proteins related to the proliferation (PCNA), apoptosis (Bax, Cleaved caspase-3, caspase-3), epithelial mesenchymal transition (EMT) (E-cadherin, Vimentin), and SIRT1/AMPK signaling pathway in HCT116 cells and nude mice in each group. Results Compared with the control group, the apoptosis rate of HCT116 cells, Bax and Cleaved caspase-3, caspase-3, E-cadherin, SIRT1 protein expression, and p-AMPK/AMPK were increased in the Asperuloside group (P <0.05), the cell viability, invasion number, migration rate, tumor volume, Ki67 and CD34 positive rates, PCNA and Vimentin protein expression were reduced (P <0.05); the trend of changes in various indicators of HCT116 cells in the Ex-527 group was opposite to that in the Asperuloside group. Ex-527 can eliminate the inhibitory effect of coumarin on the malignant progression of HCT116 cells. Conclusion Asperuloside inhibits the in vitro migration, invasion, and proliferation of colorectal cancer cells by activating the SIRT1/AMPK signaling pathway, promotes apoptosis, inhibits their growth in nude mice, and ultimately delays the malignant progression of colorectal cancer.
