黄芪多糖通过miR-10b-5p/AGPAT3分子轴调控结直肠癌细胞顺铂耐药性

Astragalus polysaccharide regulates cisplatin resistance in colorectal cancer cells through miR-10b-5p/AGPAT3 molecular axis

目的探讨黄芪多糖(astragalus polysaccharide,APS)通过miR-10b-5p/(1-acylglycerol-3-phosphate-O-acyltransferase 3;AGPAT3)分子轴调控结直肠癌(colorectal cancer,CRC)细胞顺铂(cisdiammine dichloroplatinum;CDDP)耐药性的分子机制。方法采用RT-qPCR检测SW620及SW620/CDDP细胞miR-10b-5p表达水平;使用MTT法或Transwell法检测不同浓度的APS或不同浓度的顺铂处理SW620及SW620/CDDP细胞后的细胞活力;通过双荧光素酶报告基因系统检测miR-10b-5p与AGPAT3的靶向结合关系;使用Western blotting检测AGPAT3、E-cadherin、Vimentin、Ki-67及zinc finger E-box binding homeobox 1(Zeb1)的表达情况。结果miR-10b-5p在SW620/CDDP细胞中表达水平显著上调(P<0.01);经不同浓度的APS处理后,SW620/CDDP细胞增殖(P<0.05)及迁移(P<0.05)水平受到显著抑制,APS与顺铂联合使用,显著增加了SW620/CDDP细胞对顺铂的敏感性(P<0.05);双荧光素酶报告基因验证AGPAT3是miR-10b-5p的靶基因;Western blotting检测结果显示,过表达miR-10b-5p或敲降AGPAT3可显著抑制E-cadherin的表达(P<0.01),并上调Vimentin、Ki-67及Zeb1的表达水平(P<0.01)。结论APS通过miR-10b-5p/AGPAT3分子轴调控CRC细胞顺铂耐药性,APS可作为CRC的化疗辅助药物,调节细胞对化疗药物的敏感性。

Objective To investigate the regulation of astragalus polysaccharide(APS) through miR-10 b-5 p/(1-acylglycerol-3-phosphate-O-acyltransferase 3;AGPAT3) molecular axis to regulate colorectal cancer(CRC) cells with cisplatin(Cisdiammine dichloroplatinum;CDDP) molecular mechanism of drug resistance.Methods RT-qPCR was used to detect the expression level of miR-10 b-5 p in SW620 and SW620/CDDP cells;the MTT method or Transwell method was used to detect the cell viability of SW620 and SW620/CDDP cells treated with different concentrations of APS or different concentrations of cisplatin;The dual luciferase reporter gene system detects the targeted binding relationship between miR-10 b-5 p and AGPAT3;Western blotting is used to detect the expression of AGPAT3,E-cadherin, Vimentin, Ki-67 and zinc finger E-box binding homeobox 1(Zeb1) Happening.Results The expression level of miR-10 b-5 p in SW620/CDDP cells was significantly up-regulated(P<0.01);after treatment with different concentrations of APS,the proliferation(P<0.05) and migration(P<0.05) levels of SW620/CDDP cells were significantly affected Inhibition, the combination of APS and cisplatin significantly increased the sensitivity of SW620/CDDP cells to cisplatin(P<0.05);dual luciferase reporter gene verified that AGPAT3 is the target gene of miR-10 b-5 p;Western blotting results It is shown that overexpression of miR-10 b-5 p or knockdown of AGPAT3 can significantly inhibit the expression of E-cadherin(P<0.01),and up-regulate the expression levels of Vimentin, Ki-67 and Zeb1(P<0.01).Conclusion APS regulates the cisplatin resistance of CRC cells through the miR-10 b-5 p/AGPAT3 molecular axis.APS can be used as a chemotherapy adjuvant for CRC to regulate the sensitivity of cells to chemotherapeutic drugs.