Li Shuai, Guo Lianyi
Graduate School, First Affiliated Hospital, Jinzhou Medical University, Jinzhou 121000, China.
Department of Gastroenterology, First Affiliated Hospital, Jinzhou Medical University, Jinzhou 121000, China. *Corresponding author, E-mail:
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi. 2018 Jan;34(1):59-64.
Objective To investigate the mechanisms of pseudolaric acid B (PAB) blocks cell cycle and inhibits invasion and migration in human hepatoma HepG2 cells. Methods The proliferation effect of PAB on HepG2 cells was evaluated by MTT assay. The effect of PAB on the cell cycle of HepG2 cells was analyzed by flow cytometry. Immunofluorescence cytochemical staining was applied to observe the effect of PAB on the α-tubulin polymerization and expression in HepG2 cells. Transwell chamber invasion assay and wound healing assay were performed to detect the influence of PAB on the migration and invasion ability of HepG2 cells. Western blotting was used to determine the expressions of α-tubulin, E-cadherin and MMP-9 in HepG2 cells after treated with PAB. Results PAB inhibited the proliferation of HepG2 cells in a dose-dependent manner and blocked the cell cycle in G2/M phase. PAB significantly changed the polymerization and decreased the expression of α-tubulin. The capacities of invasion and migration of HepG2 cells treated by PAB were significantly depressed. The protein levels of α-tubulin and MMP-9 decreased while the E-cadherin protein level increased. Conclusion PAB can inhibits the proliferation of HepG2 cells by down-regulating the expression of α-tubulin and influencing its polymerization, arresting HepG2 cells in G2/M phase. Meanwhile, PAB also can inhibit the invasion and migration of HepG2 cells by lowering cytoskeleton α-tubulin and MMP-9, and increasing E-cadherin.
目的 探讨土槿皮酸B（PAB）阻断人肝癌HepG2细胞周期并抑制其侵袭和迁移的机制。方法 采用MTT法评估PAB对HepG2细胞的增殖作用。通过流式细胞术分析PAB对HepG2细胞周期的影响。应用免疫荧光细胞化学染色观察PAB对HepG2细胞中α-微管蛋白聚合及表达的影响。进行Transwell小室侵袭实验和划痕愈合实验，检测PAB对HepG2细胞迁移和侵袭能力的影响。采用蛋白质印迹法检测PAB处理后HepG2细胞中α-微管蛋白、E-钙黏蛋白和基质金属蛋白酶-9（MMP-9）的表达。结果 PAB以剂量依赖性方式抑制HepG2细胞增殖，并将细胞周期阻断在G2/M期。PAB显著改变α-微管蛋白的聚合状态并降低其表达。PAB处理后的HepG2细胞侵袭和迁移能力明显降低。α-微管蛋白和MMP-9的蛋白水平降低，而E-钙黏蛋白的蛋白水平升高。结论 PAB可通过下调α-微管蛋白表达并影响其聚合，使HepG2细胞阻滞于G2/M期，从而抑制HepG2细胞增殖。同时，PAB还可通过降低细胞骨架α-微管蛋白和MMP-9以及增加E-钙黏蛋白，抑制HepG2细胞的侵袭和迁移。
