葛根素通过调控PI3K/AKT信号通路影响食管鳞状细胞癌体内外生长和转移

Puerarin Affects the Growth and Metastasis of Esophageal Squamous Cell Carcinoma in Vitro and in Vivo by Regulating PI3K/AKT Signaling Pathway

目的:探究葛根素对食管鳞状细胞癌体内外生长和转移的影响及机制。方法:MTT实验确定葛根素给药浓度为20、40、80μmol/L,取TE-13细胞,设置空白对照组、葛根素20、40、80μmol/L给药组,CCK-8法检测细胞增殖能力、细胞划痕实验检测细胞迁移能力、细胞侵袭实验检测细胞侵袭能力、软琼脂克隆形成实验检测细胞生长能力、流式细胞术检测细胞凋亡及细胞周期、Western blot检测细胞磷脂酰肌醇-3激酶(phosphatidylinositol 3-kinase,PI3K)和蛋白激酶B(Protein Kinase B,PKB,又称Akt)磷酸化水平;构建TE-13细胞裸鼠皮下移植瘤模型,并设置模型对照组、葛根素25mg/kg、50 mg/kg、100 mg/kg灌胃给药组,检测给药期间小鼠瘤体积变化,给药结束后取肿瘤组织称重、Western blot检测瘤组织中PI3K和Akt磷酸化水平,取肺组织拍照,苏木精-伊红染色法(hematoxylin-eosin staining,HE staining)检测肺组织肿瘤转移情况。结果:相较于空白对照组,葛根素20、40、80μmol/L组细胞增殖、迁移、侵袭能力显著下降、克隆形成数目显著减少、细胞凋亡显著增加、细胞周期显著阻滞、PI3K和Akt磷酸化水平明显降低(P <0. 05);相较于模型对照组,葛根素25、50、100μmol/L灌胃给药组小鼠瘤体积生长缓慢、肺组织转移节点显著减少、肿瘤组织PI3K和Akt磷酸化水平明显降低(P <0. 05)。结论:葛根素能够抑制食管鳞状细胞癌的体内外生长和转移,其机制可能为抑制PI3K/AKT信号通路。

Objective: To investigate the effect and mechanism of puerarin on the growth and metastasis of esophageal squamous cell carcinoma in vitro and in vivo. Methods: TE-13 cell were divided into the control group and puerarin treatment groups( 20、40、80 μmol/L). CCK-8 method was used to detect the cell proliferation ability,cell scratch test was used to detect the cell migration ability,cell invasion experiment was used to detect the cell invasion ability in each group,soft agar clone formation test was used to detect the cell growth ability,flow cytometry was used to detect the cell apoptosis and the cell cycle,Western blot was used to detect the levels of PI3 K and Akt phosphorylation in each group. TE-13 A nude mouse subcutaneously transplanted tumor model was established,mice were randomly divided into the model group,25 mg/kg,50 mg/kg and 100 mg/kg puerarin groups. The tumor volume changes in each group during the administration period were measured. After administration,tumor tissues were collected and weighed,the levels of PI3 K and Akt phosphorylation were detected in the tumor tissues by Western blot. Lung tissues were photographed and HE staining was used to detect the tumor metastasis. Results: Compared with the control group,the cell proliferation,migration,and invasion ability in puerarin 20,40,and 80 μmol/L groups were significantly reduced,the number of colony formation was significantly reduced,and the apoptosis was significantly increased,the cell cycle was blocked,the levels of PI3 K and Akt phosphorylation were significantly reduced( P < 0. 05). Compared with the model group,The tumor volume growths of mice in puerarin 25 mg/kg,50 mg/kg,100 mg/groups were slow,the lung tissue metastasis nodes were significantly reduced,and the levels of PI3 K and Akt phosphorylation in tumor tissues were significantly reduced( P < 0. 05). Conclusion: Puerarin can inhibit the growth and metastasis of esophageal squamous cell carcinoma in vitro and in vivo,and its mechanism may be the inhibition of PI3 K/AKT signaling pathway.