白桦脂酸上调DHX15表达抑制食管鳞癌细胞增殖和迁移的研究

Inhibitory Effect of Betulinic Acid on the Proliferation and Migration of Esophageal Squamous Cell Carcinoma Cells by Up-Regulating DHX15 Expression

目的:探究白桦脂酸对食管鳞癌细胞Eca109的增殖、迁移的影响及其可能的作用机制。方法:在含有0,20,40,80μg·ml^(-1)白桦脂酸培养液中培养Eca109细胞。体外转染siRNA-DHX15至Eca109细胞后在白桦脂酸培养液培养,细胞分为NC-siRNA组、DHX15 siRNA组、NC-siRNA+白桦脂酸组、DHX15 siRNA+白桦脂酸组,CCK-8法检测细胞增殖,平板细胞克隆形成实验检测菌落形成,免疫印迹(WB)法检测(天冬-谷-丙-组氨酸)盒多肽(DHX15)、vimentin、MMP-2、MMP-9蛋白表达; Transwell小室系统检测细胞迁移、侵袭能力。结果:随着白桦脂酸剂量的增加,细胞抑制率逐渐升高(P<0.05),当白桦脂酸为80μg·ml^(-1)时,细胞抑制率最高。与空白组相比,白桦脂酸各组DHX15蛋白表达均升高;随着处理剂量的增加,蛋白表达逐渐升高(P<0.05)。与空白组、NC-siRNA组相比,DHX15 siRNA组DHX15表达降低,菌落数数量、侵袭、迁移数量增多,NC-siRNA+白桦脂酸组DHX15表达升高,菌落数数量、侵袭、迁移数量减少(P<0.05)。与DHX15 siRNA+白桦脂酸组相比,DHX15 siRNA组DHX15表达降低、菌落数数量、侵袭、迁移数量增多,NC-siRNA+白桦脂酸组DHX15表达升高,菌落数数量、侵袭、迁移数量增多(P<0.05)。结论:白桦脂酸可通过上调DHX15抑制食管鳞癌细胞Eca109的增殖、迁移,其机制可能与抑制迁移有关蛋白表达有关。

Objective: To explore the influences of betulinic acid on the proliferation and migration of esophageal squamous cell carcinoma cell Eca109 and the possible mechanism. Methods: Eca109 cells were cultured in the culture medium respectively containing 0,20,40 and 80 μg·ml-1 betulinic acid. After transfection of siRNA-DHX15 to Eca109 cells in vitro,the cells were cultured in betulinic acid medium and divided into NC-siRNA group,DHX15 siRNA group,NC-siRNA+betulinic acid group,and DHX15 siRNA+betulinic acid group. CCK-8 assay was used to detect the cell proliferation,colony formation assay was used to detect the colony formation,Western blotting (WB) was used to detect the expressions of (aspartic acid-glutamic acid-alanine-histidine) box polypeptide (DHX15),vimentin,MMP-2 and MMP-9 proteins;Transwell cell system was used to detect the cell migration and invasion. Results:With the dose increase of betulinic acid,the cell inhibition rate gradually increased (P< 0.05),when the concentration of betulinic acid was 80 μg·ml-1,the cell inhibition rate was the highest. Compared with those in the control group,the expressions of DHX15 protein in betulinic acid groups increased;the protein expression increased gradually with the dose increase of betulinic acid (P <0.05). Compared with that in the control group and NC-siRNA group,the expression of DHX15 in siRNA group decreased,the number of colonies,the number of invasion and the number of migration increased,the expression of DHX15 in NC-siRNA + betulinic acid group was elevated,and the numbers of colonies,invasion and migration decreased (P<0.05). Compared with that in DHX15 siRNA+betulinic acid group,DHX15 expression in DHX15 siRNA group decreased,the number of colonies,the number of invasion and the number of migration increased,the expression of DHX15 in NC-siRNA+betulinic acid group was elevated,and the numbers of colonies,invasion and migration increased (P<0.05). Conclusion: Betulinic acid can inhibit the proliferation and migration of Eca109 in esophageal squamous cell carcinoma cells by up-regulating DHX15,which may be related to the inhibition of migration related protein expression.