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分别用含10、20、40、60μmol/L的菹草类胡萝卜素提取物(CEPC)培养液处理人肝癌细胞(QGY-7703)48h、96h和144h,在这三个处理时间各剂量组对肝癌细胞的抑制率平均值范围分别为0.14%-23.07%、39.59%-70.61%和71.65%-87.01%。经10、20和40μmol/L的CEPC培养液处理肝癌细胞24h、48h和72h,用激光扫描共聚焦显微术(LSCM)观察细胞形态,出现了肝癌细胞数量明显减少,细胞体积缩小、皱缩变形,细胞核呈现“新月状”、条状甚至碎片状,细胞核中呈黄色的DNA面积较明显地减小等典型的凋亡细胞形态特征。以流式细胞术分析用CEPC处理肝癌细胞后各时相细胞的百分比,与对照组比较,用10μmol/L和20μmol/L浓度的CEPC处理肝癌细胞48h后,使细胞周期中的G_0/G_1期的细胞比例极显著增加(P<0.01),分别增加了23.8%和35.6%,而在G_2/M期没有明显的变化,在S期则相应减少。用LSCM测定了肝癌细胞内的Ca~(2+)浓度,与对照组比较,经20μmol/LCEPC处理48h后能引起细胞内Ca~(2+)浓度极显著上升(P<0.01),剂量组细胞内Ca~(2+)荧光强度为对照组的1.5倍。以上结果表明CEPC对人肝癌细胞QGY-7703的增殖具有明显的抑制作用,且在一定程度上呈时间-效应和剂量-效应依赖关系。在较短的时间内及使用较小的CEPC剂量能有效地诱导肝癌细胞凋亡,CEPC使肝癌细胞阻滞于G_0/G_1期发生凋亡。CEPC能极显著提高肝癌细胞内的Ca~(2+)浓度,提示Ca~(2+)浓度升高可能是CEPC诱导肝癌细胞发生调亡的重要原因。本项研究结果为进一步研究和开发菹草类胡萝卜素的功能和价值打下了重要基础。
Human liver cancer cells (QGY-7703) were treated with CEPC (10, 20, 40 and 60μmol / L) for 48h, 96h and 144h respectively. The average inhibition rates of hepatoma cells ranged from 0.14% -23.07%, 39.59% -70.61% and 71.65% -87.01%, respectively. The hepatocellular carcinoma cells were treated with 10, 20 and 40 μmol / L CEPC for 24 h, 48 h and 72 h, respectively. The morphology of the cells was observed by laser scanning confocal microscopy (LSCM). The number of HCC cells was significantly decreased, Deformation, the nucleus showed “crescent”, strip or even fragments, DNA nuclei in the yellow area is more obvious to reduce the morphological characteristics of typical apoptotic cells. Flow cytometry was used to analyze the percentage of cells in each phase after treatment with CEPC. Compared with the control group, the hepatocellular carcinoma cells were treated with CEPC at concentrations of 10μmol / L and 20μmol / L for 48h, the G 0 / G 1 phase (P <0.01), increased by 23.8% and 35.6%, respectively. However, there was no significant change in G 2 / M phase and a corresponding decrease in S phase. The concentration of Ca2 + in hepatocellular carcinoma cells was determined by LSCM. Compared with the control group, the concentration of Ca2 + in cells treated with 20μmol / L CEPC for 48h significantly increased (P <0.01) Intracellular Ca 2+ fluorescence intensity was 1.5 times that of the control group. The above results showed that CEPC significantly inhibited the proliferation of human hepatoma cells QGY-7703, and to a certain extent, the time-effect and dose-effect dependence. In a short period of time and using a small dose of CEPC can effectively induce apoptosis of liver cancer cells, CEPC so that liver cancer cells arrested in G_0 / G_1 phase of apoptosis. CEPC can significantly increase the concentration of Ca ~ (2+) in hepatoma cells, suggesting that elevated Ca ~ (2+) may be an important reason for the apoptosis induced by CEPC. The results of this study lay an important foundation for further researching and developing the function and value of carotenoids.