Astragalus polysaccharides suppressed cisplatin-resistance of colorectal cancer TH-29/DDPcells via regulating miR-20a/TGFBR2 axis / 中国肿瘤生物治疗杂志

ABSTRACT

@# Objective: To investigate the effect of astragalus polysaccharides (APS) on proliferation, invasion, apoptosis and drugresistance of cisplatin-resistant colorectal cancer (CRC) HT-29/DDP cells through regulating miR-20a/TGFBR2 axis, and to explore the possible mechanism. Methods: Human CRC HT-29 cells and HT-29/DDP cells were used as non-drug resistant and resistant cell models, respectively; HT-29/DDP cells were randomly divided into four groups, including untreated (HT-29/DDP) group, APS treatment group, miR-20a mimics + APS group, and si-TGFBR2 + APS group. qPCR and Western blotting were applied to detect the expressions of miR-20a and TGFBR2 in HT-29/DDP cells treated with different concentrations ofAPS (0, 0.5, 1.0, 1.5 and 2.0 mg/ml). Subsequently, dual luciferase reporter gene assay was used to verify whether TGFBR2 was a target gene of miR-20a. In addition, CCK-8, Transwell andAnnexin V-FITC/PI double staining were applied to examine the effect ofAPS on proliferation, invasion and apoptosis of HT29/DDP cells. Furthermore, subcutaneous HT-29/DDP cell xenograft model was established on nude mice, and the effect ofAPS on the growth of transplanted tumor was observed. Results: APS significantly inhibited the proliferation of HT-29/DDP cells in a dose-dependent manner (P<0.01). Meanwhile, the expression of miR-20a was down-regulated in HT-29/DDP cells treated with APS, while the expression of TGFBR2 was significantly up-regulated (all P<0.01). Additionally, dual luciferase reporter gene assay result showed that TGFBR2 was a direct target of miR-20a in HT-29/DDP cells and its expression was suppressed. Furthermore, APS could enhance the drug sensitivity of HT-29/DDP cells through downregulating the inhibitory effect of miR-20a on TGFBR2 expression, thereby suppressed proliferation and invasion, and induced apoptosis of HT-29/DDP cells in vitro and in vivo. It was also found that this effect was related with the suppression of PCNA and Bcl-2 proteins and promotion of Bax and Caspase-3 proteins. Conclusion: APS reverses the resistance of HT-29/DDPcells to cisplatin by down-regulating the inhibitory effect of miR-20a on TGFBR2 expression.