论文部分内容阅读
目的:研究7-(4-甲氧基苯基)-5,8a-二苯基-1,2,3,7,8,8a-六氢咪唑[1,2-a]吡啶(TIP-6)对培养的人肝癌HepG2细胞和人正常肝细胞L02增殖的影响。方法:用MTT法和台盼蓝染色法检测TIP-6对细胞增殖的影响;相差显微镜观察细胞形态学的变化;流式细胞术(FCM)分析细胞周期改变;吖啶橙荧光染色观察自噬泡的变化;Annexin V/7-AAD和DAPI染色检测细胞凋亡;DNA电泳检测凋亡梯形带的产生。细胞免疫化学法测定NF-κB的表达。结果:TIP-6浓度为90～200μmol/L时,细胞增殖受到抑制,且与浓度、时间有关;当浓度为200μmol/L、处理72h时,两种细胞增殖数分别只有对照的12.10%和18.75%,空泡化也随剂量和时间的增加而加剧,空泡数目越来越多、体积越来越大;FCM结果显示,细胞被阻滞在G2/M期,且HepG2比L02细胞敏感。吖啶橙荧光染色证实自噬及自噬性细胞死亡会随着化合物浓度和时间增加而增多;AnnexinV/7-AAD、DAPI染色及DNA电泳均证实凋亡并非TIP-6诱导HepG2及L02细胞增殖抑制的主要形式;细胞免疫化学法的结果显示,随着核转录因子NF-κB表达量的上调,自噬泡会随之增加,细胞增殖却相应减少。结论:TIP-6可能诱导自噬抑制培养的肝癌细胞和肝细胞增殖。自噬性细胞死亡可能与NF-κB蛋白的活化有关。
AIM: To investigate the effect of 7- (4-methoxyphenyl) -5,8a-diphenyl-1,2,3,7,8,8a-hexahydroimidazo [1,2-a] ) On the proliferation of cultured human hepatoma HepG2 cells and human normal liver cells L02. Methods: The effects of TIP-6 on cell proliferation were detected by MTT assay and trypan blue staining. The morphological changes of cells were observed by phase contrast microscopy. Cell cycle was analyzed by flow cytometry (FCM). Acridine orange fluorescence staining was used to detect autophagy Apoptosis was detected by Annexin V / 7-AAD and DAPI staining. Apoptotic ladder was detected by DNA electrophoresis. Immunocytochemistry was used to determine the expression of NF-κB. Results: When the concentration of TIP-6 was 90 ~ 200μmol / L, the cell proliferation was inhibited and the concentration and time were related. When the concentration of 200μmol / L and 72h, the proliferation of the two cells were only 12.10% and 18.75 %, Vacuolization also aggravated with dose and time, the number of cavities was more and more and the volume was larger and larger. FCM results showed that cells were arrested in G2 / M phase and HepG2 was more sensitive than L02 cells. Acridine orange fluorescence staining confirmed that autophagy and autophagic cell death increased with the concentration and time of compound. AnnexinV / 7-AAD, DAPI staining and DNA electrophoresis confirmed that apoptosis was not induced by TIP-6 in HepG2 and L02 cells The main form of inhibition; results of Cellular Immunochemistry showed that as the expression of nuclear transcription factor NF-κB increased, autophagic vacuoles increased, but the cell proliferation decreased accordingly. Conclusion: TIP-6 may induce autophagy to inhibit the proliferation of hepatoma cells and hepatocytes. Autophagic cell death may be related to the activation of NF-κB protein.