Cell-Matrix Interactions and Migration

National Institute Of Dental & Craniofacial Research

Investigators

Linked publications & trials

Abstract

Extracellular matrix molecules, integrin receptors, cytoskeletal proteins, proteases, physical forces, and regulators of these molecular systems combine to mediate and regulate cell migration, signaling, and cancer cell invasion by complex, integrated mechanisms. We are addressing the following specific questions: 1. How are the functions of integrin receptors, the extracellular matrix, and the cytoskeleton integrated, and how are they coordinated to produce effective cell migration or invasion? 2. What biophysical properties and signaling mechanisms are important for efficient cell migration or for cancer cell invasion? We are using a variety of cell and molecular biology approaches to address these questions, including New Approach Methodologies (NAMs) with live-cell wide-field, confocal, and two-photon time-lapse microscopy with fluorescent protein chimeras, biochemical and signal transduction analyses, as well as methods for evaluating local matrix deformations in response to forces impinging upon, or generated by, migrating or invading mammalian cells. We use a variety of fluorescent molecular chimeras and mutants of cytoskeletal proteins, various configurations of extracellular matrix molecules, and experimental alterations in microenvironmental physical properties as part of a long-term program to analyze their functions in integrin-mediated processes in various 3D matrix environments. We are also testing an ex vivo model system for analyzing cell interactions and migration in a physiological 3D environment to complement our studies on cell-matrix interactions in vitro. This latter approach will be using mouse fascia as the source of native extracellular matrix. The role of the major alpha5-beta1 human integrin receptor in the pathogenesis of osteoarthritis was previously examined collaboratively. A fragment of the extracellular matrix protein fibronectin containing the FN 7-10 domains with its RGD and synergy binding sites activates the alpha5-beta1 integrin in primary human chondrocytes. This activation triggers downstream signaling to produce the matrix metalloproteinase MMP-13. This sequence suggests a pathophysiological role for degradation fragments in a feedback loop in which matrix fragments induce production of this protease, which in turn further degrades endogenous fibronectin to produce more pathological fragments. We collaborated further to write a review for Nature Reviews regarding these and multiple other studies further implicating integrins – including alpha5-beta1 – in not only the pathogenesis of osteoarthritis, but also in mediating normal joint development and homeostasis. Our combined approach involving mechanistic characterization of the cell adhesive, protrusive, and migratory interactions with various types of extracellular matrix should provide novel approaches to understanding and potentially ameliorating migratory or invasive processes used by cells during abnormal embryonic development and cancer invasion. An in-depth understanding of the precise ways in which cells interact with their extracellular matrix environment should also facilitate tissue engineering studies for future regenerative medicine. In all our research projects, our Section emphasizes rigorous and responsible conduct of research to conduct investigations that are reproducible and transparent, e.g., by describing in detail our specific methods, reagents, and the instrument settings we use, to be able to ensure that our work can be replicated. We perform at least 3 independent biological repeats (vs. technical repeats) for each experiment we report, as well as using orthogonal approaches with skepticism to test any important conclusion. A key component of rigorous and reproducible research, we feel, is the continuous conscientious use of electronic lab notebooks that provide forensic tracking and backups (we use the Federal version of LabArchives), as well as retaining two backups of all primary data, of which one copy is retained at NIH after a lab member leaves. We evaluate and communicate error and uncertainty by using quantification, unbiased data analysis approaches, and appropriate statistical tests. In our publications, we discuss reservations, caveats, and limitations of our work and the assumptions on which it is based. We aim to construct experimental tests to obtain yes/no and/or quantitative answers in order to rigorously test our hypotheses. We welcome negative results as opening new avenues of exploration into areas that had not been predicted. We avoid conflicts of interest in our research, as well as in our reviewing for journals and grant funding agencies, and we take care to submit our research to journals with unbiased peer review, as well as avoiding “predatory” journals. We encourage collaboration and interdisciplinary research, which enriches and broadens our approaches. This approach includes sharing our underlying primary data as much as practical, e.g., by deposition in public repositories after publication of our research. Public availability of the raw data underlying research allows others to evaluate and replicate our findings. We also continue to emphasize the training and mentoring of the next generation of biomedical researchers.