Cell Stiffness Analysis And Cell Classification Based On Quantitative Characterization Of Cytoskeleton Structure

Cell stiffness is one of the important physical properties of cells,which can effectively characterize the physiological state of cells and has a complex relationship with human diseases.A large number of studies have shown that the stiffness of a single cell is closely related to the adhesion,movement and metastasis of cancer cells,and there is a strong correlation between the stiffness and cellular malignancy.Many researchers use cell stiffness as a cellular marker and diagnostic parameter to diagnose diseases(including cancer)and to determine the metastatic potential of cancer cells.At present,there are many methods to detect the stiffness of cells,such as AFM,micropipette and microfluidic.These methods have been used successfully to characterize cell stiffness by Young's modulus.However,it can be found that the stiffness values of the same cell lines measured by different detection methods differ greatly and are uncertain.Therefore,it is necessary to study and compare cell stiffness with new methods.Cytoskeleton,as the structural framework of cells,determines the stiffness of cells and can effectively reflect a variety of physiological characteristics and states of cells.When a healthy cell lesion becomes cancerous,the cytoskeleton degrades from a fairly ordered structure to an irregular structure with reduced stiffness.During this process,the structural changes of the microfilaments in the cytoskeleton played a major role in the change of cell stiffness.The analysis of the microfilament cytoskeleton structure can obtain the stiffness information of cells and even distinguish the pathological changes of cells by the characteristics of the microfilament cytoskeleton structure.Based on the bladder cell microfilament skeleton structure,this paper puts forward to quantify bladder cell microfilament skeleton structure parameters of microfilament length and the standard deviation of sinusoidal microfilament angle,and study correlation between cytoskeleton structure and stiffness of bladder cancer.In this paper,the microfilament cytoskeleton was used to characterize the bladder cell stiffness and to classify normal bladder cells and bladder cancer cells.The research of this paper is as follows:1.The parameters of microfilament length and the standard deviation of sinusoidal microfilament angle are proposed to quantify the microfilament structure of normal bladder cells and bladder cancer cells respectively.Quantitative data showed that the average length of microfilaments in normal bladder cells was longer than that in cancer bladder cells.The standard deviation of sinusoidal microfilament angle of normal bladder cells is smaller than that of bladder cancer cells,suggesting that the microfilaments of normal bladder cells are more orderly arranged.2.The stiffness of single bladder cell corresponds to the skeleton structure of the microfilaments by modification of the substrate of the culture dish.In this paper,a cell culture dish was designed and modified to facilitate cell localization.AFM was used to obtain the stiffness of normal bladder cells and bladder cancer cells,and then the cell stiffness was corresponding to the fluorescence image of the microfilament skeleton through the localization of the substrate of the culture dish.The results showed that bladder cancer cells were softer than normal bladder cells,with a lower Young's modulus and a shorter,more disorganized microfilament skeleton to match.After that,this paper carried out AFM morphology imaging of normal bladder cells and bladder cancer cells,and the surface morphology showed that parallel arrangement of actin filaments could be seen in normal bladder cells,while fewer actin filaments were found in bladder cancer cells,which was consistent with the results of fluorescence staining of cell microfilament skeleton.3.In this paper,we analyzed the correlation between the bladder cell stiffness and the structural parameters of the microfilament(microfilament length,the standard deviation of sinusoidal microfilament angle),and classified the normal bladder cell and bladder cancer cells by the stiffness and the microfilament parameters respectively.The results showed that the bladder cell stiffness was positively correlated with the microfilament length,and the bladder cell stiffness was negatively correlated with the standard deviation of sinusoidal microfilament angle,and the microfilament length had a dominant effect on the bladder cell stiffness.The support vector machine(SVM)was used to classify normal bladder cells and bladder cancer cells.The results showed that the accuracy of bladder cell classification by stiffness alone was 84.2%,and the accuracy of bladder cell classification by microfilament cytoskeleton parameter was 94.7%,indicating that the structure of microfilament cytoskeleton could be used to distinguish normal bladder cells from bladder cancer cells.