Physics Theses and Dissertations
Permanent URI for this collectionhttp://hdl.handle.net/1903/2800
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Item High Resolution Mapping of Intracellular Mechanical Properties during Key Stages of Cancer Progression(2022) Nikolic, Milos; Scarcelli, Giuliano; Tanner, Kandice; Biophysics (BIPH); Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)The mechanical phenotype of the living cell is critical for survival following deformations due to confinement and fluid flow. Furthermore, in recent years mechanical interaction between cells and the cellular environment has been implicated as one of the key regulators of cancer progression and malignant transformation. Due to the need to better understand the mechanical properties of invasive cells and how the mechanical phenotype plays a role in cancer progression, several microrheology techniques have been applied to study cell mechanics in a range of in vitro environments. However, many of these techniques have been limited either to studying cells in only one type of environment (e.g. 2D), with limited resolution, or with invasive probes. To begin to address this question, in this dissertation we aim to quantify the mechanical state of cells in a broader range of different contexts and geometries. To do this we use Brillouin microscopy, a non-contact, label free, non-invasive technique which enables us to probe the mechanical response of cells in a wide range of complex microenvironments. Here we introduce an improved Brillouin microscope with improved signal and acquisition speed which enables us to perform biological studies at the single cell level. Using the improved Brillouin microscopy, we find that individual cells can be softer as function of the invasive potential, but that cells are able to dynamically change their mechanical properties across many different contexts. We validate our results using complementary microrheology methods such as atomic force microscopy and broadband optical tweezer microrheology. We directly observe changes in cell mechanics in key processes relevant for metastatic migration, as well as a function of external and internal parameters like morphology, ECM properties, intracellular factors, and cell-cell cooperativity during multicellular tissue organization. These results support the paradigm that the mechanical state of a cell is a dynamic parameter that varies as a consequence of the microenvironmental and functional context, in addition to the observable changes in cell’s mechanical properties due to malignant transformation.Item UNDERSTANDING CELL DIFFERENTIATION AND MIGRATION WITH MULTIVARIATE CELL SHAPE QUANTIFICATION(2018) Chen, Desu; Losert, Wolfgang; Biophysics (BIPH); Digital Repository at the University of Maryland; University of Maryland (College Park, Md.)This thesis focuses on developing multivariate quantification methods of cell shape to facilitate understanding of physiological processes such as cell differentiation and migration. Cell shape reflects complex intracellular and extracellular factors affecting cell function. However, analyses associating cell shape and cell function need to account for challenges of multivariate interpretation, single-cell heterogeneity and reproducibility. Specifically, Human Bone Marrow Stromal Cells (hBMSCs) population in nanofiber scaffolds can develop osteogenic differentiation without chemical cues. I developed a method based on Support Vector Machine (SVM) to train classifiers as boundaries in the shape metric spaces to identify the day 1 cell shape phenotype of hBMSCs population in nanofiber scaffolds. To reduce the effect of single-cell heterogeneity in the population phenotyping, the “supercell” method was introduced to generate average measurements of small groups of cells for SVM training. To overcome the multivariate complexity in biological interpretation, a feature selection process was implemented to select the most significant cell shape metrics. The predictive potential of the achieved classifiers was validated by subsampling. It was found that in nanofiber scaffolds, hBMSCs were narrower with more elongated and dendritic shape and rougher cell boundary. Further, I found that increase in nanofiber density enhanced hBMSCs osteogenic differentiation potential. The pre-trained classifiers successfully predict the modulation of nanofiber density on hBMSCs fates and single-cell shape. While much can be learned from cell shapes alone, it is important to note that shapes can change with time, especially for migrating cells. The second part of my thesis focuses on analysis of shape dynamics. Quantification for cell shape dynamics at the subcellular level was developed to understand the coordination of the subcellular myosin localization and the cell boundary dynamics in neutrophil migration in vivo. The correlation of myosin localization and positive cell boundary curvature was identified as a unique in vivo neutrophil migration phenotype. Correlations of myosin localization and local cell boundary dynamics in vivo were found to be affected by cell motility and polarization. This analysis framework shown here can also be used to study the link between other subcellular features and neutrophil migration and shape dynamics.