山萘酚对人肝癌Bel-7402细胞增殖、迁移、侵袭及凋亡的影响

Effect of kaempferol on the proliferation,migration,invasion,and apoptosis of human hepatoma Bel-7402 cells

目的探讨山萘酚对人肝癌Bel-7402细胞增殖、迁移、侵袭及凋亡的影响,并探讨其相关分子机制。方法将体外培养的肝癌Bel-7402细胞随机分为对照组和低、中、高浓度实验组,实验组分别用低、中、高浓度山萘酚(25、50、100μmol/L)处理,对照组予以等量二甲基亚砜处理。用CCK-8法检测山萘酚对Bel-7402细胞活力的影响;平板克隆检测山萘酚对细胞克隆形成能力的影响;细胞划痕和Transwell小室侵袭实验检测山萘酚对细胞迁移和侵袭的影响;Western Blot检测凋亡及周期相关蛋白表达情况。多组间比较采用单因素方差分析,进一步两两比较采用LSD-t检验。结果对照组和低、中、高浓度实验组的细胞活力在处理24 h后分别为(100.00±2.72)%、(75.70±2.42)%、(62.79±2.45)%、(43.41±2.11)%,与对照组比较,各实验组细胞活力均明显下降(P值均<0.05)。各组细胞克隆形成数分别为923.3±35.2、682.7±24.4、464.0±22.0、327.3±14.0,与对照组比较,各实验组细胞克隆形成能力均明显下降(P值均<0.05)。处理24 h后,各组的相对迁移率分别为(100.00±1.11)%、(63.33±1.16)%、(51.72±3.23)%、(37.18±2.71)%,穿膜细胞数目分别为212.0±3.0、134.0±2.0、71.0±2.0、34.0±1.0,与对照组比较,各实验组相对迁移率和穿膜细胞数均明显下降(P值均<0.05)。处理48 h后,与对照组比较,低、中、高浓度实验组抗凋亡蛋白Bcl-2的表达水平均明显降低(P值均<0.05),促凋亡蛋白Bax表达水平均明显升高(P值均<0.05),周期蛋白CyclinD1表达水平均明显降低(P值均<0.05)。结论山萘酚可抑制人肝癌Bel-7402细胞的增殖、迁移和侵袭能力,促进其凋亡,作用机制可能与调控凋亡蛋白Bax/Bcl-2以及下调周期蛋白CyclinD1的表达有关。

Objective To investigate the effect of kaempferol on the proliferation,migration,invasion,and apoptosis of human hepatoma Bel-7402 cells and related molecular mechanism.Methods Hepatoma Bel-7402 cells cultured in vitro were randomly divided into control group and low-,middle-,and high-concentration experimental groups.The experimental groups were treated with low-,middle-,and high-concentration kaempferol(25,50,and 100μmol/L),and the control group was treated with an equal volume of dimethyl sulfoxide.CCK-8 assay was used to observe the effect of kaempferol on the viability of Bel-7402 cells;plate colony formation assay was used to evaluate the effect of kaempferol on cell colony formation ability;wound healing assay and Transwell chamber were used to observe the effect of kaempferol on cell migration and invasion;Western blot was used to measure the expression of apoptosis-and cycle-related proteins.A one-way analysis of variance was used for comparison between multiple groups,and the least significant difference t-test was used for further comparison between two groups.Results After 24 hours of treatment,the cell viability was 100.00%±2.72%in the control group and 75.70%±2.42%,62.79%±2.45%,and 43.41%±2.11%,respectively,in the low-,middle-,and high-concentration experimental groups,and compared with the control group,the experimental groups had a significant reduction in cell viability(all P<0.05).The number of cell colonies was 923.3±35.2 in the control group and 682.7±24.4,464.0±22.0,and 327.3±14.0,respectively,in the low-,middle-,and high-concentration experimental groups,and compared with the control group,the experimental groups had a significant reduction in cell colony formation ability(all P<0.05).After 24 hours of treatment,the relative migration rate was 100.00%±1.11%in the control group and 63.33%±1.16%,51.72%±3.23%,and 37.18%±2.71%,respectively,in the low-,middle-,and high-concentration experimental groups,and the number of transmembrane cells was 212.0±3.0 in the control group and 134.0±2.0,71.0±2.0,and 34.0±1.0,respectively,in the low-,middle-,and high-concentration experimental groups;compared with the control group,the experimental groups had significant reductions in relative migration rate and number of transmembrane cells(all P<0.05).After 48 hours of treatment,compared with the control group,the low-,middle-,and high-concentration experimental groups had a significant reduction in the expression of the anti-apoptotic protein Bcl-2(all P<0.05),a significant increase in the expression of the pro-apoptotic protein Bax(all P<0.05),and a significant reduction in the expression of C<italic/>yclinD1(all P<0.05).Conclusion Kaempferol can inhibit the proliferation,migration,and invasion of human hepatoma Bel-7402 cells and promote the apoptosis of such cells,possibly by regulating the apoptosis proteins Bax and Bcl-2 and downregulating the expression of CyclinD1.