Wei Weitian, Wang Liang, Xu Liwei, Liang Jinxiao, Teng Lisong
Department of Surgical Oncology, Zhejiang University School of Medicine First Affiliated Hospital, No. 79 Qingchun Road, Shangcheng District, Hangzhou 310009, China.
Department of Surgical Oncology, Zhejiang Cancer Hospital, No. 1, East Banshan Road, Gongshu District, Hangzhou 310022, China.
ACS Omega. 2021 Nov 17;6(47):31435-31446. doi: 10.1021/acsomega.1c02945. eCollection 2021 Nov 30.
the present study aims to investigate the function of miR-199 on gemcitabine (GEM)-resistance in pancreatic cancer, as well as the underlying mechanism.
the GEM-resistant SW1990 cell line (SW1990/SZ) was established. The CCK-8 assay was used to detect the cell viability. The self-renewal of SW1990/SZ cells was evaluated by sphere formation and the colony formation assay. The apoptosis was detected by flow cytometry and the migration ability was measured by the transwell assay. The dual-luciferase gene reporter assay was utilized to confirm the binding between miR-199 and Snail. The expression level of CD44, ALDH1, Nanog, E-cadherin, Vimentin, β-catenin, and Snail was determined by the Western blotting assay.
the cell sphere formation rate, number of spheres, and expression level of CD44, ALDH1, and Nanog in GEM-treated SW1990/SZ cells were significantly suppressed by miR-199, accompanied by declined proliferation ability, an increased apoptotic rate, inhibited migration ability, and suppressed EMT progression. The binding site between miR-199 and 3'-UTR of Snail was predicted and confirmed. The inhibitory effect of miR-199 on self-renewal of SW1990/GZ cells and the faciliating property of miR-199 on the inhibitory effect of GEM against the proliferation ability, migration ability, and EMT progression were abolished by overexpressing Snail.
MiR-199 reversed the resistance to GEM in pancreatic cancer by suppressing stemness through regulating the EMT.
本研究旨在探讨miR-199在胰腺癌吉西他滨（GEM）耐药中的作用及其潜在机制。
建立GEM耐药的SW1990细胞系（SW1990/SZ）。采用CCK-8法检测细胞活力。通过成球实验和集落形成实验评估SW1990/SZ细胞的自我更新能力。采用流式细胞术检测细胞凋亡，采用Transwell实验检测细胞迁移能力。利用双荧光素酶基因报告实验证实miR-199与Snail之间的结合。采用蛋白质印迹法检测CD44、ALDH1、Nanog、E-钙黏蛋白、波形蛋白、β-连环蛋白和Snail的表达水平。
miR-199显著抑制GEM处理的SW1990/SZ细胞的细胞成球率、球状体数量以及CD44、ALDH1和Nanog的表达水平，同时伴随着增殖能力下降、凋亡率增加、迁移能力抑制和上皮-间质转化（EMT）进程受抑制。预测并证实了miR-199与Snail的3'-UTR之间的结合位点。过表达Snail消除了miR-199对SW1990/GZ细胞自我更新的抑制作用以及miR-199对GEM抑制增殖能力、迁移能力和EMT进程的促进作用。
MiR-199通过调节EMT抑制干性，从而逆转胰腺癌对GEM的耐药性。
