ABSTRACT
Metastasis is one of the main reasons causing death in cancer patients. It was reported that chemotherapy might induce metastasis. In order to uncover the mechanism of chemotherapy-induced metastasis and find solutions to inhibit treatment-induced metastasis, the relationship between epithelial-mesenchymal transition (EMT) and doxorubicin (DOX) treatment was investigated and a redox-sensitive small interfering RNA (siRNA) delivery system was designed. DOX-related reactive oxygen species (ROS) were found to be responsible for the invasiveness of tumor cells in vitro, causing enhanced EMT and cytoskeleton reconstruction regulated by Ras-related C3 botulinum toxin substrate 1 (RAC1). In order to decrease RAC1, a redox-sensitive glycolipid drug delivery system (chitosan-ss-stearylamine conjugate (CSO-ss-SA)) was designed to carry siRNA, forming a gene delivery system (CSO-ss-SA/siRNA) down-regulating RAC1. CSO-ss-SA/siRNA exhibited an enhanced redox sensitivity compared to nonresponsive complexes in 10 mmol/L glutathione (GSH) and showed a significant safety. CSO-ss-SA/siRNA could effectively transmit siRNA into tumor cells, reducing the expression of RAC1 protein by 38.2% and decreasing the number of tumor-induced invasion cells by 42.5%. When combined with DOX, CSO-ss-SA/siRNA remarkably inhibited the chemotherapy-induced EMT in vivo and enhanced therapeutic efficiency. The present study indicates that RAC1 protein is a key regulator of chemotherapy-induced EMT and CSO-ss-SA/siRNA silencing RAC1 could efficiently decrease the tumor metastasis risk after chemotherapy.
ABSTRACT
@#[Abstract] Objective: To investigate the effects of miR-223-3p on the proliferation and apoptosis of hepatocellular carcinoma (HCC) cells by regulating Ras-related C3 botulinum toxin substrate 1 (RAC1) and its possible mechanism. Methods: Thirty pairs of HCC and corresponding para-cancer tissues resected in Jilin Central Hospital from August 2016 to August 2018 were collected for this study; in addition, human HCC cell lines SMMC-7721, BEL-7402, HepG2 and human normal hepatocyte QSG-7701 were also collected. The expression level of miR-223-3p in HCC tissue and cell lines was detected by qPCR. miR-223-3p mimics, miR-223-3p inhibitor and siRAC1 were transfected into SMMC-7221 cells, respectively. CCK-8 assay, Colony formation assay and Annexin V-FITC/PI staining Flow cytometry were used to detect the proliferation, clone formation and apoptosis of SMMC-7721 cells, respectively. The relationship between miR-223-3p and RAC1 was confirmed by Dual luciferase reporter gene assay. The protein level of RAC1 in SMMC-7721 cells was detected by Western blotting. Results: The expression of miR-223-3p in HCC tissues was significantly lower than that in paracaner tissues (P<0.01), and had significant correlation with pathological characteristics, such as tumor size, TNM stage, EdmondsonSteiner grade (all P<0.05 or P<0.01). miR-223-3p expression in HCC cell lines was significantly lower than that in QSG-7701 cells with the lowest expression in SMMC-7721 cells. Dual luciferase reporter gene assay confirmed that RAC1 was a target gene of miR-223-3p, and miR-223-3p negatively regulated RAC1 expression. Over-expression of miR-223-3p significantly inhibited the proliferation and colony formation (P<0.05 or P<0.01) of SMMC-7721 cells and promoted cell apoptosis (P<0.01). Contrarily, knockdown of miR-223-3p reversed the inhibitory effect of miR-223-3p mimics on cells. Conclusion: miR-223-3p over-expression inhibits proliferation and colony formation and promotes apoptosis of HCC cells, the mechanism of which may be related with its targeted down-regulation of RAC1.