125I粒子联合AZD1152对三阴性乳腺癌细胞增殖和凋亡的影响

目的：研究¹²⁵I粒子联合Aurora激酶抑制剂AZD1152对三阴性乳腺癌(TNBC)细胞MDA-MB-231增殖和凋亡的影响。方法：试验设对照组(常规培养)、¹²⁵I粒子照射组(照射组)、1 μmol/L AZD1152作用MDA-MB-231细胞48 h组(抑制组)、¹²⁵I粒子照射与1 μmol/L AZD1152联合作用MDA-MB-231细胞48 h组(联合组)。采用免疫荧光检测细胞多核形成；四甲基噻唑蓝(MTT)法检测细胞增殖抑制率；CCK-8检测细胞活力；流式细胞术PI单染检测细胞周期；流式细胞术Annexin V/PI双染观察各组细胞凋亡情况；Western blot检测各组细胞中Cyclin B1、组蛋白H3的表达及其磷酸化水平的改变，以及凋亡相关蛋白Bcl-2、Bax、PARP表达的变化。结果：免疫荧光检测发现1 μmol/L AZD1152作用MDA-MB-231细胞48 h时可见到多核细胞形成；MTT试验结果显示对照组、照射组、抑制组和联合组的细胞增殖抑制率分别为(0.61±0.32)%、(17.62±1.41)%、(29.67±0.41)%、(53.17±1.26)%；CCK-8检测显示细胞存活率分别为(94.88±0.22)%、(59.21±0.14)%、(42.05±0.17)%、(32.12±0.36)%；细胞周期检测发现G2/M期占比分别为(18.99±0.15)%、(38.05±0.23)%、(49.80±0.32)%、(75.52±0.45)%，与对照组比较差异均具有统计学意义(P<0.05)。照射组、抑制组和联合组的细胞凋亡率分别为(17.48±0.24)%、(29.23±0.02)%、(63.11±0.27)%，均较对照组(0.31±0.03)%升高(P<0.05)。流式细胞术发现抑制组细胞出现多核及多倍体细胞，易形成非整倍体。Western blot检测结果显示联合组较其他3组，抗凋亡蛋白Bcl-2、组蛋白H3的磷酸化和Cyclin B1蛋白表达减少(P<0.05)，而促凋亡蛋白Bax和PARP蛋白剪切明显增加(P<0.05)。结论：¹²⁵I粒子对AZD1152有增敏作用，¹²⁵I联合AZD1152可显著抑制MDA-MB-231细胞组蛋白H3磷酸化及Cyclin B1水平，从而抑制细胞增殖，诱导细胞凋亡。

The combination of 125I particles and AZD1152 efficiently inhibited proliferation and promoted apoptosis of a triple negative breast cancer cell line

OBJECTIVE: To study effects of AZD1152 combined with ¹²⁵I particles on proliferations and apoptoses of triple negative breast cancer cells MDA-MB-231. METHODS: Cultured cells were divided into several groups; control, ¹²⁵I particle irradiation, 1 μmol/L AZD1152 treatment with 48 h, treated with ¹²⁵I particle irradiation and 1 μmol/L AZD1152 treated with 48 h. Cell proliferation inhibitory rates were detected using the MTT and cell viability using the CCK-8 assays. Cell ploidies,cell cycles and apoptoses were detected using flow cytometry with cells separately stained by propidium iodide (PI) and by Annexin V/PI double-staining. Expressions of apoptosis-related proteins (Bcl-XL, Bcl-2 and PARP), CyclinB1 and Phosphorylation levels of Histone H3 were analyzed using Western blotting. RESULTS: After treatments for 48 h, cell proliferation inhibitory rates were (0.61±0.32)%,(17.62±1.41)%,(29.67±0.41)%,(53.17±1.26)%,respectively. Cell viability rates were (94.88±0.22)%,(59.21±0.14)%,(42.05±0.17)% (32.12±0.36)%,respectively. The proportions of G2/M phase were (18.99±0.15)%,(38.05±0.23)%,(49.80±0.32)%,(75.52±0.45)%,respectively. The apoptosis rates of MDA-MB-231 cells were (17.48±0.24)%,(29.23±0.02)%,(63.11±0.27)%,with significantly differences in the different groups (P<0.05). Compared with (0.31 ±0.03)% in the control group, the observed increases were significant (P<0.05). Immunofluorescence and flow cytometry analyses showed that multinucleated and polyploid cells appeared in the 1μmol/L AZD1152 group in 48 hours,which were easy to form aneuploid cells. Compared with the irradiated,inhibitor and control groups,the combined irradiation with 1 μmol/L AZD1152 for 48 h effectively down-regulated the expressions of Bcl-2,p-Histone H3,CyclinB1 and promoted Bax and dissection of PARP (P<0.05). CONCLUSION: The combination of ¹²⁵I particles and AZD1152 treatments efficiently inhibited proliferation and promoted apoptosis in breast cancer cell line MDA-MB-231.