低氘水抑制宫颈癌Hela细胞增殖和侵袭能力的研究

Research of inhibition of deuterium-depleted water for cervical cancer Hela cell proliferation and invasiveness

目的探讨培养基中氘浓度的下降对宫颈癌Hela细胞增殖、侵袭能力的影响。方法用含不同氘浓度的培养基培养Hela细胞,分为150×10-6、100×10-6、75×10-6和50×10-6共4组,其中,氘浓度为150×10-6的培养基为对照组,其他为实验组;用四甲基噻唑蓝(MTT)法、划痕实验检测随着培养基中氘浓度下降,Hela细胞增殖和迁移侵袭力的变化;流式细胞术检测含不同氘浓度的培养基对Hela细胞周期的影响;蛋白免疫印迹和免疫组化实验检测含不同氘浓度的培养基对Hela细胞肿瘤相关蛋白p21、Na+/K+-ATPase表达的影响。结果 MTT实验结果显示,随着培养基中氘浓度的降低Hela细胞增殖得到明显的抑制,其中氘浓度为50×10-6的培养基抑制效果最明显,相同条件下与对照组相比在72 h抑制率达到39.54%(P<0.01);划痕实验可观察到Hela细胞在氘浓度为75×10-6、50×10-6的培养基中细胞迁移能力得到明显抑制(P<0.05);Hela细胞在氘浓度为75×10-6、50×10-6的培养基中培养24 h后,流式细胞术检测S期明显低于对照组(P<0.01);Hela细胞肿瘤相关蛋白p21在氘浓度为100×10-6、75×10-6和50×10-6的培养基中的表达与对照组比较,差异均有高度统计学意义(P<0.01)。结论培养基中氘浓度的下降对宫颈癌Hela细胞增殖、侵袭能力有明显的抑制作用。

Objective To examine the effects of the proliferation and invasiveness in Hela cell lines by the deuteriumdepleted water （DDW）.Methods The Hela cell lines were cultured in the presence of media containing different concentrations of deuterium （150×10-6,100×10-6,75×10-6 and 50×10-6）,while the 150×10-6 concentration of deuterium was taken as the control group,the other concentrations of deuterium were taken as the experimental group.MTT and wound scratch assay were used to examine the effects of DDW on the proliferation and migration of Hela cells.More over,the flow cytometry was used to detect the effects of DDW on the cell cycle of Hela cell lines.Finally,the expression levels of p21 and Na＋/K＋-ATPase were assessed by using Western blot and immunohistochemistry.Results The proliferation of Hela cells was significantly suppressed when culturing in three concentrations of DDW （100×10-6,75×10-6and 50×10-6） medium,the inhibition effect was obvious in 50×10-6 concentration of DDW,the 72 h inhibition ratio was 39.54％ （P ＜ 0.01）.The result of wound scratch assay showed that the invasiveness ability of Hela cells was significantly decreased in 75×10-6 and 50×10-6 concentration of DDW （P ＜ 0.05）.Cell cycle analysis revealed that DDW caused cell cycle arrest and decreased the amounts of cells in the S phase after 24 h cultivation in 75×10-6 and 50×10-6concentration of DDW （P ＜ 0.01）.Western blot suggested that compared with the control group,DDW （100×10-6,75×10-6and 50×10-6） could up-regulate the expression of p21 （P ＜ 0.01）.Conclusion The decreased of deuterium concentration in the medium can significant inhibit the proliferation and invasiveness of Hela cells.