基于原子力显微镜的溶液流动环境下癌细胞力学特性测量研究  

Measuring The Mechanical Properties of Cancerous Cells in Fluidic Environments by Atomic Force Microscopy

作　　者：魏佳佳 李密[1,2,3] 冯雅琦 刘连庆 WEI Jia-Jia;LI Mi;FENG Ya-Qi;LIU Lian-Qing(State Key Laboratory of Robotics,Shenyang Institute of Automation,Chinese Academy of Sciences,Shenyang 110016,China;Institutes for Robotics and Intelligent Manufacturing,Chinese Academy of Sciences,Shenyang 110169,China;University of Chinese Academy of Sciences,Beijing 100049,China)

机构地区：[1]中国科学院沈阳自动化研究所,机器人学国家重点实验室,沈阳110016 [2]中国科学院机器人与智能制造创新研究院,沈阳110169 [3]中国科学院大学,北京100049

出　　处：《生物化学与生物物理进展》2022年第10期2041-2053,共13页Progress In Biochemistry and Biophysics

基　　金：国家自然科学基金(62273330,61922081,61873258);中国科学院前沿科学重点研究计划(ZDBS-LY-JSC043);辽宁省“兴辽英才计划”(XLYC1907072)资助项目。

摘　　要：目的细胞力学特性在细胞生理病理活动过程中起着重要的调控作用,开展细胞力学特性研究对于揭示生命活动内在规律具有重要意义。原子力显微镜(AFM)的发明为单细胞力学特性表征提供了新的强大工具,基于AFM压痕分析的细胞力学特性探测已成为生命科学领域的重要研究方法,为单细胞行为研究带来了大量新认识。然而,现有基于AFM的细胞力学特性研究主要集中在静态溶液环境,而癌细胞在体内转移过程中处于脉管系统的动态液流环境下,因此现有的测量结果无法完全反映溶液流动环境下的癌细胞真实生理行为,特别是目前对于肿瘤转移过程中液流环境与癌细胞之间相互作用的力学机制的认知还十分有限。本文通过将AFM与液流控制技术结合建立了溶液流动环境下的细胞力学特性测量方法。方法基于两侧开口培养皿并结合注射泵/抽取泵液流控制搭建细胞培养基动态液流系统,并将其分别与AFM及光学倒置显微镜进行集成。选取MCF-7(人乳腺癌细胞)和HGC-27(人未分化胃癌细胞)两种癌细胞进行实验。利用细胞培养基动态液流系统培养细胞并分析溶液流速以及流动时间对细胞生长及力学特性的影响。在光学显微镜导引下控制AFM对培养基静态/流动环境下生长的细胞进行压痕实验以获取力曲线,并利用Hertz-Sneddon模型对力曲线进行分析得到细胞杨氏模量。利用荧光染色试剂分析溶液流动环境对细胞活性及细胞骨架蛋白的影响。结果首先分析了溶液流动环境对细胞生长的影响,实验结果表明,与静态培养环境相比,培养基动态液流环境可更好地促进细胞生长。随后分别对静态和流动环境下生长的癌细胞力学特性进行了测量,结果表明当癌细胞生长环境由静态变为动态后细胞的杨氏模量显著减小,且溶液流动环境会导致细胞骨架结构的变化,显示了溶液流动环境对癌Objective Cell mechanics plays an important role in regulating cellular physiological and pathological processes and investigating cell mechanics is meaningful for revealing the underlying mechanisms guiding life activities.The advent of atomic force microscopy(AFM)provides a novel powerful tool for singlecell studies and AFM-based indentation assay has become an important method for characterizing cell mechanics in the field of life sciences,yielding numerous new insights into single-cell behaviors.However,current studies of cell mechanics by AFM are commonly performed in static environment,while cancerous cells are in flow environment of vascular fluids during tumor metastasis,and thus the results obtained in static environment cannot completely reflect the real behaviors of cancerous cells in fluidic condition.Particularly,so far knowledge of the mechanical mechanisms guiding the interactions between fluidic microenvironment and cancerous cells in the process of tumor metastasis is still limited.Here,based on AFM,a method allowing measuring the mechanical properties of single cells in fluidic environments is developed.Methods A fluidic cell culture medium device was established on a petri dish with openings on both sides with the use of an injection pump and an extraction pump.The fluidic cell culture medium device was integrated with AFM to measure the mechanical properties of cells in fluidic environments.The fluidic cell culture medium device was also integrated with inverted optical microscope which had a heating plate to observe the growth states of cells in fluidic environments.MCF-7 cells(human breast cancer cell)and HGC-27 cells(human gastric undifferentiated carcinoma cell)were used for the experiments.The fluidic cell culture medium device was used to grow cells to examine the effects of medium flow rate and flow time on cell proliferation and cell mechanics.Under the guidance of optical microscopy,AFM probe was moved to cells grown in static culture medium or fluidic culture medium to perform indent

关 键 词：原子力显微镜 溶液流动环境 癌症转移 细胞 力学特性 杨氏模量 

分 类 号：Q27[生物学—细胞生物学] Q66