砷诱导肝母细胞瘤HepG2细胞凋亡机理研究

目的 研究三氧化二砷(As203)诱导肝母细胞瘤细胞系HePG2细胞凋亡的机理及其对细胞核基质相关蛋白——前髓白血病蛋白(PML)表达的影响。方法 Hep2细胞培养于MEM培养基中,用不同浓度As2O3分别处理24或96h。运用原位末端脱氧核苷转换酶(TdT)标记法(TUNEL)和DNA阶梯法检测细胞凋亡。以激光共振聚交显微镜及Western杂交观察PML的表达。结果 TUNEL阳性的凋亡细胞及DNA阶梯片段可见于As203处理组。用2μmol/L As203处理的Hep2细胞核基质蛋白内PML表达减弱。激光共振聚交图像显示,经2μmol/L As203处理的Hep2细胞核中,PML蛋白的表达明显减弱;用5μmol/L As203处理后,PML在Hep2细胞核中的微小点状表达几乎消失。结论 As203在体外可明显抑制Hep2细胞生长,其诱导HePG2细胞凋亡的作用依时间及剂量不同而异。As203可降解HepG2细胞核中PML蛋白,且PML的表达降低与细胞凋亡密切相关。核基质相关蛋白PML可能是As203作用的靶蛋白。

Mechanism of arsenic trioxide-induced apoptosis in hepatic blastoma cells HepG2

OBJECTIVE: To investigate the mechanism of arsenic trioxide (As(2)O(3)) induced apoptosis in hepatic blastoma cells HepG2 and its effects on cell nuclear matrix related protein promyelocytic leukaemia (PML). METHODS: HepG2 cells were cultured in MEM medium and treated with different concentrations of As(2)O(3) for either 24 h or 96 h. In situ terminal deoxynucleotidyl transferase (TdT) labeling (TUNEL) and DNA ladder were applied to detect apoptosis. Confocal microscopy and western blot were performed to observe the expression of PML. RESULTS: TUNEL positive apoptotic cells and DNA ladder could be detected in As(2)O(3) treated groups. The expression of PML decreased in HepG2 cells with 2 micro mol/L As(2)O(3), and micropunctates characteristic of PML protein in HepG2 cell nuclei almost disappeared after the treatment of 5 micro mol/L As(2)O(3). CONCLUSION: As(2)O(3) induces HepG2 tumor cell apoptosis in a time- and concentration-dependent manner. As(2)O(3) may degradate the PML protein in HepG2 cell nuclei. The decreased expression of PML is closely correlated with apoptosis. Nuclear matrix associated protein PML could be the target of As(2)O(3) therapy.