不同方法炮制甘遂对LO2细胞周期与凋亡的影响

Effect of Kansui Radix prepared by different processes on LO2 cell cycle and apoptosis

目的:探讨不同方法炮制所得甘遂各样品对人正常肝细胞LO2细胞周期与凋亡的影响。方法:以人正常肝细胞LO2为研究对象,采用MTT法检测甘遂醋制前后各炮制品(生品、清炒品、醋润品、醋制品)对LO2细胞活性的影响,采用倒置显微镜观察细胞形态,采用流式细胞术研究甘遂醋制前后各炮制品对LO2细胞周期和凋亡的影响。结果:与阴性对照组比较,甘遂生品可明显降低LO2细胞的活性(P<0.01)和形态,显著升高S期细胞百分率(P<0.05),显著降低G2/M期细胞百分率(P<0.01),显著增加人正常肝细胞LO2细胞的早期凋亡率、晚期凋亡坏死率、总凋亡率(P<0.01);与甘遂生品组比较,甘遂各炮制品均能降低对人正常肝细胞LO2的增殖抑制作用(P<0.01)和形态变差的趋势,其降低增殖抑制作用的顺序为醋制品>醋润品>清炒品,且呈一定的量-效关系(r醋制=0.999,r醋润=0.913,r清炒=0.914,r生品=0.984);且均能显著增加G2/M期细胞百分率(P<0.05,P<0.05,P<0.01),显著减少人正常肝细胞LO2的早期凋亡率、晚期凋亡坏死率、总凋亡率(P<0.01),其增加G2/M期细胞百分率的顺序和降低凋亡率的顺序均为醋制品>醋润品>清炒品。结论:甘遂炮制后可通过影响LO2细胞周期与凋亡降低其毒性,且甘遂醋制工艺中的炒与醋润2种操作在醋制降低甘遂的肝毒性中能够起到协同作用,从而为揭示上述2种操作在甘遂醋制减毒工艺中的合理性和醋制工艺优化提供了一定的依据。

Objective： To discuss the effect of Kansui Radix prepared by different processes on cell cycle and apoptosis of nor real human liver cell lines LO2. Method： With normal human liver cell lines LO2 as the study object, the MqT method was adopted to study the effect of Kansui Radix prepared by different processes, including Kansui Radix, stirbaking Kansui Radix, Kansui Radix moistening with vinegar and Kansui prepared by different processes, on LO2 cell activity. The cellular morphological changes were ob served by inverted microscope. The effect of Kansui Radix stirbaked with vinegar on LO2 cell cycle and apoptosis was observed by flow cytometry. Result： Compared with the negative control group, Kansui could obviously inhibit the activity of human normal liver cell lines LO2 （P 〈 0. 01 ） , and significantly increase the percentage of LO2 cells in S phase （P 〈 0. 05 ）, notably decrease the percentage of LO2 cells in GE/M phase （P 〈 0. 01 ）, significantly increase the early apoptosis rate, late apoptosis rate and necrosis rate and total apoptosis rate of human normal liver cell lines LO2 （P 〈 0. 01 ）. Compared with the Kansui group, all of the other processed Kansui samples could significantly decrease the eel1 proliferation inhibition （ P 〈 0. 01 ） , and the trend of morphological degradation. Besides, they could significantly increase the percentage of LO2 ceils in G2/M phase （P 〈 0. 05, P 〈 0. 05, P 〈 0. 01 ） , significantly decrease the early apoptosis rate, late apoptosis rate and necrosis rate, and total apoptosis rate of human normal liver cell lines LO2 （P 〈 0. 01 ）. The order of the increase in the percentage of cells in G2/M phase and the decrease in apoptosis rate was Kansui Radix stir baked with vinegar 〉 Kansui Radix moistening with vinegar 〉 stirbaking Kansui Radix. Conclusion ： The toxicity of processed Kan sui could be reduced by affecting LO2 cell cycle and apoptosis. The processes of stirbaking and moistening with vinegar can play a synergistic effect in the detoxication of human normal liver cell lines LO2, which provides a basis for unveiling the rationality of stir baking with vinegar of Kansui in the detoxieation, as well as the optimizing the process.