Changes in the Mechanical Properties of Cells undergoing Neoplastic Transformation

Abstract

Macrophages are an important component of the innate immune response and their ability to migrate toward pathogens is critical to their success as first responders. Several proteins are known to contribute to the migration of macrophages and their roles have been studied using Boyden chambers, Dunn chambers, and micropipette pointsources. However, visualizing macrophage migration has been difficult due to their inherently strong adhesion. We have been able to study macrophage migration in two dimensions using a novel surface preparation in which PDMS coated coverslips are stamped with the extracellular matrix protein fibronectin. LR5 macrophages were individually tracked on these surfaces in the presence of CSF-1, a physiologically relevant chemokine, using time-lapse microscopy. Analysis of the trajectories of motion has allowed us to describe motility in terms of speed, persistence time, and the random motility coefficient. LR5 cells in which endogenous Cdc42 or WASp was reduced using small interfering RNA were also examined to determine the role of these proteins in migration. Our results indicate that reduction of WASp levels leads to a significant reduction in motility. The knockdown of Cdc42 leads to a reduction in the random motility coefficient and sensitivity to ligand chemistry. Our results are in contrast to other studies which have linked Cdc42 and WASp to directional sensing but have found no defect in random motility. The roles of PI3K in macrophage migration are currently being investigated. We are also beginning to study the forces macrophages produce while migrating using microfabricated post array detectors. The cell lines with reduced protein levels will also be studied on posts to determine the role of each protein in force production. 1639-Pos Board B531The Nanomechanical Signature of Breast Cancer Cancer initiation and progression follow complex molecular and structural changes in the extracellular matrix and in the cellular architecture of living tissue. Yet, it remains poorly understood how the transformation from health to malignancy alters the mechanical properties of cells within the tumour microenvironment. Here we show using an indentation-type atomic force microscope (IT-AFM) that unadulterated human breast biopsies display distinct stiffness profiles. Correlative stiffness maps obtained on normal and benign tissues show uniform stiffness profiles that are characterized by a single distinct peak. In contrast, malignant tissues have a broad distribution resulting from tissue heterogeneity with a prominent low-stiffness peak representative of cancer cells. Similar findings are seen in specific stages of breast cancer in MMTV-PyMT transgenic mice. Further evidence obtained from the lungs of mice with late-stage tumours shows that migration and metastatic spreading is correlated to the low-stiffness of hypoxia-associated cancer cells. Overall, nanomechanical profiling by IT-AFM provides quantitative indicators in the clinical diagnostics of breast cancer with translational significance. ...