Mechano-chemical regulation of cell migration on nanofabricated scaffolds mimicking in vivo substrata

Migration;Cancer;Microenvironment;Biomedical Engineering

Park, JinSeokLevchenko, Andre ;

Johns Hopkins University

关键词: Migration;    Cancer;    Microenvironment;    Biomedical Engineering;   

Others  :  https://jscholarship.library.jhu.edu/bitstream/handle/1774.2/60284/PARK-DISSERTATION-2014.pdf?sequence=1&isAllowed=y

瑞士|英语

来源: JOHNS HOPKINS DSpace Repository

【 摘 要 】

In cell and tissue development and physiology, cell migration is of paramount significance. It is essential in such diverse processes as immune surveillance, tissue repair, embryonic development, and cancer metastasis. Cell migration is a tightly orchestrated phenomenon, involving cyclic adhesion, membrane protrusion and contraction regulated by a complicated network of intra/intercellular signaling pathways and cytoskeletal reorganization. Although commonly studied in the lab using flat and hard cell adhesion substrata, in vivo migration of mammalian cells usually depends on successful navigation of topographically, mechanically, and chemically complex and diverse environments. Furthermore, rapidly accumulating information suggests that such cell properties as cell polarity, migration, and proliferation can be strongly affected by the mechanical characteristics of the microenvironment, particularly the topography of the extracellular matrix (ECM). Therefore, it is of interest to explore the mechanisms underlying the ability of various cell types, especially metastatic cancer cells, to detect and respond to changes in their mechanical environment. We aim to understand broadly not only how cells sense external cues from nanofabricated substrata that mimic in vivo structure but also, in turn, how they regulate migratory behavior.Supporting ECM structures such as collagen are generally inhomogeneous and variable on the scale of a single cell. We thus applied to design graded post density arrays (GPDA) as a simplified model of the inhomogeneous topographical surface of in vitro tissue. With this designed substratum, we focused on analyzing 1205Lu melanoma cell line which has a high degree of aggressiveness in terms of tissue invasion and metastasis because of a lack of functional PTEN, a negative regulator of phosphoinositide 3-kinase (PI3K) signaling, a common mutation observed in many cancers and Scbl2 benign melanoma cell line as a control. We investigated whether the direction of cell migration is determined by topographical density combined with intrinsic characteristics of cells: 1205Lu cells whose PI3K activity is dominant over Rho kinase (ROCK) activity exhibited directional migration from dense to sparse post arrays in contrast to Scbl2 on high ECM coated GPDA whose ROCK is dominant over PI3K showing the opposite directional migration. Furthermore, we examined the persistence of cell migration depending on the dynamics of PI3K signaling in response to local post density arrays, i.e. the progressively enhanced persistent migration with increased anisotropic cue as increasing the fractions of persistent PI3K dynamics vs. oscillatory and random dynamics. Also, the computational model to describe PI3K dynamics-regulating machinery composed of Rac, Rho and ECM-cell contacts was proposed. Moreover, cancer cells remodel surrounding ECM and this remodeling creates aligned ECM fiber structure, which can serve as a strong mechanical cue. The role of this topographical cue in single cell migration has been reported; its involvement in collective epithelial migration is still unexplored. We investigated the role of this topographical cue in collective cell migratory behavior and, particularly, epithelial-mesenchymal transition (EMT), on a collagen type I pre-coated grooved substratum, 800 nm of width, height and depth, that mimics reorganized fibrous structures of extracellular matrix. In contrast to observations on the flat substratum, healthy epithelial monolayers on nano-structued substrata formed finger-like structures, which marginal cells migrate significantly faster migration than sub-marginal cells along the direction of overall guided migration. Cells at the tips of these structures frequently lost contact with neighboring cells and disseminate from the original collective cell sheet, which was accompanied by the expression of EMT markers. Additionally we found that YAP (Yes-associated protein) is activated in these finger-like projections on structured substrata and that a knockdown of YAP expression induces higher expression of E-cadherin and activation of ROCK followed by suppression of cell migration of tip cells. These data shown here argue that topographical cues can strongly modify collective cell migration and induce EMT, even in healthy cells. We also suggest that YAP as a critical regulator of collective cell migration in the microenvironment that has pronounced topographical cues, e.g., aligned fibers of extracellular matrix.