双硫仑螯合铜对MCF-7和MCF-10A细胞超微结构和力学特性的影响

目的:基于原子力显微镜(atomic force microscope,AFM)在纳米级分辨率和皮牛级力学测量探讨双硫仑(disulfiram,DSF)螯合氯化铜复合物(DSF/Cu)对人乳腺癌MCF-7和正常乳腺上皮MCF-10A细胞膜超微结构及力学特性的影响。方法:采用流式细胞术比较DSF/Cu作用于2种细胞后凋亡率和细胞周期的改变。AFM探测细胞表面形貌、超微结构、细胞高度、宽度及粗糙度的变化。利用AFM纳米压痕技术检测DSF/Cu对2种细胞硬度(杨氏模量)的影响。结果:流式细胞术结果显示:DSF/Cu可显著诱导MCF-7细胞凋亡,且呈浓度依赖性;而对MCF-10A细胞影响不大。细胞周期检测显示DSF/Cu对2种细胞的细胞周期影响不同。AFM成像观察发现,400和800 nmol/L的DSF/Cu作用于MCF-7细胞6 h后,细胞形态明显皱缩,体积变小,表面变得平整,丝状伪足明显回缩、变短,有些变成了板状伪足,甚至完全消失。进一步定量分析发现细胞宽度变小,高度增高,均方根粗糙度和平均粗糙度均减小,且呈浓度依赖性。而对MCF-10A细胞则影响甚微。最后,单细胞水平生物力学检测显示:DSF/Cu作用于MCF-7和MCF-10A细胞6 h后,2种细胞的杨氏模量随药物浓度的增加均有所增加,且呈浓度依赖性,但MCF-7细胞增加的比例要远远高于MCF-10A细胞。结论:DSF/Cu可能通过特异性影响MCF-7细胞的生物力学特性发挥高效低毒的抗乳腺肿瘤作用。

Effects of DSF/Cu on surface ultrastructures and mechanical properties of human breast cancer and normal breast epithelial cells

AIM: To study the effects of disulfiram/copper complex (DSF/Cu) on ultrastructures and mechanical properties of human breast cancer and normal breast epithelial cells by atomic force microscopy (AFM) based on the nanoscale resolution and piconewton force measurement level. METHODS: The change of cell cycle and apoptotic rate of MCF-7 cells and MCF-10A cells induced by DSF/Cu were compared by flow cytometry. The cell surface morphology, ultrastructure, height, width and roughness were detected by AFM. The effects of DSF/Cu on the hardness (Young's modulus) of the 2 kinds of cells were determined by AFM with indentation technique. RESULTS: DSF/Cu significantly induced apoptosis of the MCF-7 cells in a dose-dependent manner, whereas had little effect on the MCF-10A cells. The cell cycle analysis showed that DSF/Cu induced G₂/M arrest in the MCF-7 cells, but led to G₀/G₁ arrest in the MCF-10A cells. The AFM images showed that the MCF-7 cells shrank and showed smaller and smoother morphology, and the filopodia were retracted obviously, even some became into lamellipodia, or disappeared completely after treated with DSF/Cu at concentrations of 400 and 800 nmol/L. The quantitative analysis indicated that the MCF-7 cells showed smaller width and larger height, and the root mean square roughness and average roughness were decreased significantly in a dose-dependent manner after treated with DSF/Cu at concentrations of 400 and 800 nmol/L. However, little effect in the MCF-10A cells was observed. The biomechanics test at a single cell level demonstrated that the Young's modulus of the MCF-7 cells and MCF-10A cells were both increased, yet the proportion increased in the MCF-7 cells was much higher than that in the MCF-10A cells after treated with DSF/Cu at concentrations of 400 and 800 nmol/L. CONCLUSION: DSF/Cu has strong antitumor effect on breast cancer with high efficiency and low toxicity by changing the properties of the biomechanics specifically.