镉通过类雌激素样作用促进人乳腺癌MCF-7细胞增殖

Estrogen-like effect of cadmium on promoting proliferation of human breast cancer MCF-7 cells

[目的]研究镉对人乳腺癌MCF-7细胞增殖及雌激素受体(estrogen receptor,ER)蛋白表达水平的影响及其可能机制。[方法]用1×10^(-6)、1×10^(-7)、1×10^(-8)、1×10^(-9)、1×10^(-10)、1×10^(-11) mol/L的17β-雌二醇(后称雌激素)培养不同来源的A、B两株MCF-7细胞4 d,应用CCK8法筛选敏感细胞株并确定雌激素促细胞增殖作用最强的浓度。用1×10^(-6)、1×10^(-7)、1×10^(-8)、1×10^(-9)、10^(-10)、1×10^(-11) mol/L的镉溶液染毒敏感细胞株,确定镉促进细胞增殖的最大浓度。设置阴性对照组(去雌激素培养液)、实验组(1×10^(-8) mol/L镉)、阳性对照组(1×10^(-9) mol/L雌激素),通过克隆形成实验检测镉对MCF-7细胞集落形成能力的影响。利用ER抑制剂(ICI182780)初步探讨镉致细胞增殖的可能机制,增设实验抑制剂组(1×10^(-8) mol/L镉+1×10^(-7) mol/L ER抑制剂)、阳性抑制剂组(1×10^(-9) mol/L雌激素+1×10^(-7) mol/L ER抑制剂),通过CCK8法、流式细胞术和Western blot分别检测各组的细胞增殖率、细胞周期S期比例和ER表达水平。[结果]实验所用B株MCF-7细胞为雌激素敏感细胞株,且当雌激素浓度为1×10^(-9) mol/L时,对MCF-7细胞的促进增殖作用[(203.55±36.65)%]最大(P<0.001)。与阴性对照组(100%)相比,1×10^(-8)~1×10^(-10) mol/L镉溶液可促进MCF-7细胞增殖[(163.78±31.90)%^(176.88±10.06)%](P<0.001)。1×10^(-8) mol/L镉可促进细胞集落形成(P<0.05),增加细胞S期比例(P<0.001),提高细胞ER蛋白表达水平(P<0.001)。与实验组相比,加入ER抑制剂能够抑制镉对MCF-7细胞的促增殖作用[由(135.17±23.96)%降至(107.66±7.64)%](P<0.01),抑制镉诱导的细胞S期比例增加[由(24.17±0.53)%降至(12.36±0.43)%](P<0.001),拮抗镉诱导的ER表达增加[由(56.19±3.67)%降至(38.84±1.04)%](P<0.05)。[结论]镉可以促进人乳腺癌细胞MCF-7增殖和ER表达,这种作用可被ER抑制剂所抑制,提示镉可能具有雌激素样作用。

[Objective] To investigate the effect and potential mechanism of cadmium（Cd） on the proliferation of human breast cancer MCF-7 cells and the expression of estrogen receptor（ER）. [Methods] A and B strains of MCF-7 cells were treated with 17β-estradiol（hereinafter referred to as estrogen） at 1×10-6, 1×10-7, 1×10-8, 1×10-9, 1×10-10, and 1×10-11 mol/L, respectively, for 4 d. CCK8 assay was employed to detect a susceptible MCF-7 cell strain and to identify the concentration of estrogen promoting maximum cell proliferation. Then, the susceptible MCF-7 cell strain was treated with 1×10-6, 1×10-7, 1×10-8, 1×10-9, 1×10-10, and 1×10-11 mol/L cadmium solution, respectively, to identifythe concentration of cadmium promoting maximum cell proliferation. We set a negative control group（medium without estrogen）, an experimental group（1×10-8 mol/L cadmium）, and a positive control group（1×10-9 mol/L estrogen） to evaluate colony formation ability by clonogenicity assay. Additionally, we set an experimental inhibitor group [1×10-8 mol/L Cd＋1×10-7 mol/L ER antagonist（ICI182780）] and a positive inhibitor group（1×10-9 mol/L estrogen＋1×10-7 mol/L ER antagonist） to detect cell proliferation, the ratio of S phase cells, and the expression level of ER by CCK8, flow cytometry, and Western blot, respectively. [Results] The MCF-7 cells of strain B were estrogen sensitive cells, and the 1×10-9 mol/L estrogen treatment induced the maximum cell proliferation [（203.55±36.65）% ]. Compared with the negative control group（100% ）, 1×10-8-1×10-10 mol/L cadmium significantly promoted proliferation of MCF-7 cells [（163.78±31.90）% -（176.88±10.06）% ]（P 〈 0.001）. Besides, 1×10-8 mol/L cadmium treatment promoted colony formation of MCF-7 cells（P 〈 0.05）, increased the ratio of S phase cells（P 〈 0.001）, and elevated ER protein expression level（P 〈 0.001）. Compare with the experimental group, ER antagonist suppressed the effect of cadmium on proliferation of MCF-7 cells [from（135.17±23.96）% to（107.66±7.64）% ]（P 〈 0.01）, reduced the ratio of S phase cells [from（24.17±0.53）% to（12.36±0.43）% ]（P 〈 0.001）, and down-regulated ER protein expression level [from（56.19±3.67）% to（38.84±1.04）% ]（P 〈 0.05）.[Conclusion] Cadmium can promote proliferation and ER expression of breast cancer MCF-7 cells, and the effect can be inhibited by ER antagonist, indicating estrogen-like effect of cadmium.