蔓荆子黄素对小鼠单核巨噬细胞增殖和凋亡的影响

Effect of Vitexicarpin on proliferation and apoptosis of monocyte-macrophages in mice

目的通过蔓荆子黄素对小鼠腹腔巨噬细胞活性及RAW264.7细胞增殖和凋亡作用的研究,探讨蔓荆子黄素对单核巨噬细胞的影响。方法分别用含有不同浓度蔓荆子黄素作用于小鼠腹腔巨噬细胞与小鼠单核巨噬细胞RAW264.7,于24、48及72 h后采用CCK-8检测细胞活性。使用AO/EB染色观察RAW264.7细胞凋亡变化;应用流式细胞仪检测RAW264.7细胞凋亡率。通过荧光探针DCFH-DA检测RAW264.7细胞活性氧水平,使用JC-1染色标记线粒体膜电位的变化。结果 CCK-8法检测结果显示,蔓荆子黄素对小鼠腹腔巨噬细胞的细胞活性及RAW264.7细胞的增殖都有抑制作用,呈剂量与时间依赖关系,尤其对具有增殖能力的RAW264.7细胞活性抑制作用更强。AO/EB染色结果显示,RAW264.7细胞经浓度为5及10μmol/L的蔓荆子黄素处理后,细胞表现为凋亡形态;流式细胞仪检测结果显示,蔓荆子黄素诱导RAW264.7细胞Sub-G1期代表凋亡细胞的百分比增加(P<0.05)。蔓荆子黄素作用于RAW264.7细胞后,细胞内活性氧水平提高;JC-1染色结果显示,蔓荆子黄素能使RAW264.7细胞的线粒体膜电位下降。结论蔓荆子黄素能够抑制小鼠腹腔巨噬细胞与小鼠单核巨噬细胞RAW264.7活性,尤其对具有增殖能力的RAW264.7细胞活性抑制作用更强,可以促进其凋亡,其作用机制与增加细胞内活性氧水平和降低线粒体膜电位有关。

Objective To investigate the effect of Vitexicarpin on proliferation and apoptosis of monocytemacrophage.Method Mouse peritoneal macrophages and monocyte-macrophage cell line RAW264.7 cells were treated with different concentrations of Vitexicarpin.Cell activity was tested by CCK8 24,48,and 72 hours post treatment.Apoptosis rate was determined by AO/EB staining and flow cytometry.Levels of reactive oxygen species in RAW264.7 cells and mitochondrial membrane potential were obtained by fluorescence probe DCFH-DA and JC-1 staining,respectively.Results CCK8 assay showed thatVitexicarpin inhibited cellular proliferation of peritoneal macrophages and RAW264.7 cells in a dose-and time-dependent manner.AO/EB staining data suggested that RAW264.7 cells treated with Vitexicarpin(5μmol/L and 10μmol/L)manifested apoptotic morphology.Flow cytometry indicated that Vitexicarpin increased percentage of Sub-G1 phase in RAW264.7 cells(P<0.05).The activity of reactive oxygen species in RAW264.7 cells was increased with introduction of Vitexicarpin.JC-1 staining showed that the mitochondrial membrane potential of RAW264.7 cells was decreased by Vitexicarpin.Conclusion Vitexicarpin inhibits the cellular proliferation and promotes apoptosis of macrophages through increase of intracellular reactive oxygen species and decrease of mitochondrial membrane potential.