可降解的Mg-1%Ca合金抑制宫颈癌细胞体外增殖、迁移及侵袭

Degradable Mg-1%Ca alloy inhibits cervical cancer cell proliferation, migration and invasion in vitro

目的研究Mg-1%Ca合金的降解性能及对宫颈癌细胞体外增殖、细胞周期及迁移、侵袭的作用。方法检测Mg-1%Ca合金在DMEM培养基中pH值及Mg^(2+)、Ca^(2+)浓度的变化。根据GB/T 16886.5标准制备浸提液,取宫颈癌细胞(SiHa/HeLa),与Mg-1%Ca合金浸提液共培养,CCK-8法检测细胞存活率,平板克隆实验检测细胞增殖,流式细胞术分析细胞周期,划痕和Transwell侵袭实验检测细胞迁移和侵袭能力。取人宫颈上皮永生化细胞H8,与Mg-1%Ca合金浸提液共培养,CCK-8法检测细胞存活率。结果随着浸泡时间的延长,Mg-1%Ca合金浸提液和纯Mg浸提液的pH值均逐渐升高。Mg-1%Ca合金浸提液在24、72、168 h的pH值分别为(7.95±0.04)、(8.15±0.04)和(8.24±0.06),均低于纯Mg浸提液的pH值(P<0.001)。浸提24 h时,Mg-1%Ca合金浸提液中Mg^(2+)浓度增加至(21.642±0.348)mmol/L,但低于纯Mg浸提液中Mg^(2+)浓度(P<0.001)。浸提24 h时,Mg-1%Ca合金浸提液和纯Mg浸提液中的Ca^(2+)浓度都是降低的,分别为(0.431±0.065)mmol/L和(0.403±0.055)mmol/L。Mg-1%Ca合金可明显抑制宫颈癌细胞(SiHa/HeLa)的增殖,宫颈癌细胞SiHa/HeLa存活率均<70%。Mg-1%Ca合金浸提液对H8细胞有一定抑制作用,但H8细胞存活率>70%。Mg-1%Ca合金明显减少SiHa/HeLa细胞的克隆形成数(P<0.001),增加SiHa/HeLa细胞处于G0/G1期的细胞数(P<0.01),减少SiHa/HeLa细胞迁移距离(P<0.001)和侵袭细胞数(P<0.05)。Mg-1%Ca合金浸提液与纯Mg浸提液相比,对两种宫颈癌细胞的抑制作用无明显差别。结论Mg-1%Ca合金降解速率更慢,具有良好的生物相容性。Mg-1%Ca合金可抑制宫颈癌细胞(SiHa/HeLa)的体外增殖,调控G0/G1期细胞周期阻滞,并能抑制其发生迁移和侵袭。这种抑制的机制可能与微环境中Mg^(2+)浓度和pH值升高有关。

Objective To study the degradation properties of Mg-1%Ca alloy and its effect on proliferation,cell cycle and migration and invasion of cervical cancer cells in vitro.Methods The changes of pH and Mg^(2+)and Ca^(2+)concentration of Mg-1%Ca alloy in DMEM medium were detected.The extract was prepared according to GB/T 16886.5 standard,cervical cancer cells(SiHa/HeLa)were taken and co-cultured with Mg-1%Ca alloy extract,cell viability was detected by CCK-8 assay,cell proliferation was detected by plate cloning assay,cell cycle was analyzed by flow cytometry,and cell migration and invasion ability was detected by scratch and Transwell invasion assay.Human cervical epithelial immortalized cells H8 were taken and co-cultured with Mg-1%Ca alloy immersion solution,and cell viability was detected by CCK-8 method.Results The pH values of both Mg-1%Ca alloy extract and pure Mg extract gradually were increased with the prolongation of the immersion time.pH values of Mg-1%Ca alloy extract at 24,72 and 168 h were(7.95±0.04),(8.15±0.04)and(8.24±0.06),respectively,which were lower than those of pure Mg extract(P<0.001).The Mg^(2+)concentration in the Mg-1%Ca alloy leachate was increased to(21.642±0.348)mmol/L at 24 h of leaching,but it was lower than that in the pure Mg leachate(P<0.001).The Ca^(2+)concentration in both Mg-1%Ca alloy extract and pure Mg extract was decreased to(0.431±0.065)mmol/L and(0.403±0.055)mmol/L,respectively,at 24 h.Mg-1%Ca alloy significantly inhibited the proliferation of cervical cancer cells(SiHa/HeLa),and both the cervical cancer cells SiHa/HeLa survival rate were<70%.Mg-1%Ca alloy extract had some inhibitory effect on H8 cells,but the survival rate of H8 cells was>70%.Mg-1%Ca alloy significantly reduced the number of clone formation of SiHa/HeLa cells(P<0.001),it increased the number of cells in G0/G1 phase of SiHa/HeLa cells(P<0.01),and it reduced the cell migration distance(P<0.001)and the number of invasive cells of SiHa/HeLa(P<0.05).The inhibitory effect of Mg-1%Ca alloy extract on the 2 types of cervical cancer cells did not show any significant difference when compared with the pure Mg extract.Conclusion Mg-1%Ca alloy has a slower degradation rate and good biocompatibility.Mg-1%Ca alloy inhibited the proliferation of cervical cancer cells(SiHa/HeLa)in vitro,modulated G0/G1 phase cell cycle block and inhibited the occurrence of migration and invasion.The mechanism of the inhibition may be related to the elevated Mg^(2+)concentration and pH in the microenvironment.