Three-dimensional matrix fiber alignment modulates cell migration and MT1-MMP utility by spatially and temporally directing protrusions

Multiple attributes of the three-dimensional (3D) extracellular matrix (ECM) have been independently implicated as regulators of cell motility, including pore size, crosslink density, structural organization, and stiffness. However, these parameters cannot be independently varied within a complex 3D ECM protein network. We present an integrated, quantitative study of these parameters across a broad range of complex matrix configurations using self-assembling 3D collagen and show how each parameter relates to the others and to cell motility. Increasing collagen density resulted in a decrease and then an increase in both pore size and fiber alignment, which both correlated significantly with cell motility but not bulk matrix stiffness within the range tested. However, using the crosslinking enzyme Transglutaminase II to alter microstructure independently of density revealed that motility is most significantly predicted by fiber alignment. Cellular protrusion rate, protrusion orientation, speed of migration, and invasion distance showed coupled biphasic responses to increasing collagen density not predicted by 2D models or by stiffness, but instead by fiber alignment. The requirement of matrix metalloproteinase (MMP) activity was also observed to depend on microstructure, and a threshold of MMP utility was identified. Our results suggest that fiber topography guides protrusions and thereby MMP activity and motility.

[1]  Kevin W Eliceiri,et al.  Contact guidance mediated three-dimensional cell migration is regulated by Rho/ROCK-dependent matrix reorganization. , 2008, Biophysical journal.

[2]  Denis Wirtz,et al.  Hypoxia and the extracellular matrix: drivers of tumour metastasis , 2014, Nature Reviews Cancer.

[3]  Steven C George,et al.  Noninvasive assessment of collagen gel microstructure and mechanics using multiphoton microscopy. , 2007, Biophysical journal.

[4]  Alan Wells,et al.  2D protrusion but not motility predicts growth factor–induced cancer cell migration in 3D collagen , 2012, The Journal of cell biology.

[5]  Kenneth M. Yamada,et al.  Direct comparisons of the morphology, migration, cell adhesions, and actin cytoskeleton of fibroblasts in four different three-dimensional extracellular matrices. , 2011, Tissue engineering. Part A.

[6]  E. Thompson,et al.  Collagen induced MMP-2 activation in human breast cancer , 2004, Breast Cancer Research and Treatment.

[7]  Amit Pathak,et al.  Biophysical regulation of tumor cell invasion: moving beyond matrix stiffness. , 2011, Integrative biology : quantitative biosciences from nano to macro.

[8]  Yunfeng Feng,et al.  Dimensional and temporal controls of three-dimensional cell migration by zyxin and binding partners , 2012, Nature Communications.

[9]  D. Lauffenburger,et al.  Migration of tumor cells in 3D matrices is governed by matrix stiffness along with cell-matrix adhesion and proteolysis. , 2006, Proceedings of the National Academy of Sciences of the United States of America.

[10]  M. Dembo,et al.  Traction force microscopy of migrating normal and H-ras transformed 3T3 fibroblasts. , 2001, Biophysical journal.

[11]  B. R. Bass,et al.  3D collagen alignment limits protrusions to enhance breast cancer cell persistence. , 2014, Biophysical journal.

[12]  Ben Fabry,et al.  3D Traction Forces in Cancer Cell Invasion , 2012, PloS one.

[13]  F. Lanni,et al.  Cell traction forces on soft biomaterials. I. Microrheology of type I collagen gels. , 2001, Biophysical journal.

[14]  Micah Dembo,et al.  Influence of type I collagen surface density on fibroblast spreading, motility, and contractility. , 2003, Biophysical journal.

[15]  Stephanie I. Fraley,et al.  A distinctive role for focal adhesion proteins in three-dimensional cell motility , 2010, Nature Cell Biology.

[16]  Denis Wirtz,et al.  Three-dimensional cell migration does not follow a random walk , 2014, Proceedings of the National Academy of Sciences.

[17]  Stephanie I. Fraley,et al.  Reply: reducing background fluorescence reveals adhesions in 3D matrices , 2010, Nature Cell Biology.

[18]  L. Kaufman,et al.  Flow and magnetic field induced collagen alignment. , 2007, Biomaterials.

[19]  L. Kaufman,et al.  Pore size variable type I collagen gels and their interaction with glioma cells. , 2010, Biomaterials.

[20]  C. Rueden,et al.  Bmc Medicine Collagen Density Promotes Mammary Tumor Initiation and Progression , 2022 .

[21]  D. Wirtz,et al.  The Arp2/3 complex mediates multigeneration dendritic protrusions for efficient 3‐dimensional cancer cell migration , 2013, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[22]  Paul Matsudaira,et al.  Computational model for cell migration in three-dimensional matrices. , 2005, Biophysical journal.

[23]  Robert M. Hoffman,et al.  Physical limits of cell migration: Control by ECM space and nuclear deformation and tuning by proteolysis and traction force , 2013, The Journal of cell biology.

[24]  G. Giannelli,et al.  Role of Cell Surface Metalloprotease Mt1-Mmp in Epithelial Cell Migration over Laminin-5 , 2000, The Journal of cell biology.

[25]  Wilhelm Burger,et al.  Digital Image Processing - An Algorithmic Introduction using Java , 2008, Texts in Computer Science.

[26]  M. McNiven,et al.  Invasive matrix degradation at focal adhesions occurs via protease recruitment by a FAK–p130Cas complex , 2012, The Journal of cell biology.

[27]  D A Lauffenburger,et al.  Mathematical model for the effects of adhesion and mechanics on cell migration speed. , 1991, Biophysical journal.

[28]  Qin Su,et al.  A Multifunctional Lentiviral-Based Gene Knockdown with Concurrent Rescue that Controls for Off-Target Effects of RNAi , 2010, Genom. Proteom. Bioinform..

[29]  F H Silver,et al.  Self-assembly of collagen fibers. Influence of fibrillar alignment and decorin on mechanical properties. , 1997, Biophysical journal.

[30]  Denis Wirtz,et al.  Focal adhesion size uniquely predicts cell migration , 2013, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[31]  H. Birkedal‐Hansen,et al.  Gelatinase A (MMP-2) activation by skin fibroblasts: dependence on MT1-MMP expression and fibrillar collagen form. , 2001, Matrix biology : journal of the International Society for Matrix Biology.

[32]  Alexandra Jilkine,et al.  Membrane Tension Maintains Cell Polarity by Confining Signals to the Leading Edge during Neutrophil Migration , 2012, Cell.

[33]  Stephen J. Weiss,et al.  Protease-dependent versus -independent cancer cell invasion programs: three-dimensional amoeboid movement revisited , 2009, The Journal of cell biology.

[34]  Pei-Hsun Wu,et al.  Statistical analysis of cell migration in 3D using the anisotropic persistent random walk model , 2015, Nature Protocols.

[35]  Jian Zhang,et al.  Microtubules stabilize cell polarity by localizing rear signals , 2014, Proceedings of the National Academy of Sciences.

[36]  Colin Ng,et al.  Shape-dependent cell migration and focal adhesion organization on suspended and aligned nanofiber scaffolds. , 2013, Acta biomaterialia.

[37]  Paolo P. Provenzano,et al.  Collagen reorganization at the tumor-stromal interface facilitates local invasion , 2006, BMC medicine.

[38]  Jiaxi Zhou,et al.  Mechanical force affects expression of an in vitro metastasis-like phenotype in HCT-8 cells. , 2010, Biophysical journal.

[39]  M. d’Ortho,et al.  The activation of ProMMP-2 (gelatinase A) by HT1080 fibrosarcoma cells is promoted by culture on a fibronectin substrate and is concomitant with an increase in processing of MT1-MMP (MMP-14) to a 45 kDa form. , 1998, Journal of cell science.

[40]  V. Barocas,et al.  Comparison of 2D fiber network orientation measurement methods. , 2009, Journal of biomedical materials research. Part A.

[41]  E. Thompson,et al.  Collagen-induced activation of the M(r) 72,000 type IV collagenase in normal and malignant human fibroblastoid cells. , 1992, Cancer research.

[42]  D. Weitz,et al.  A blind spot in confocal reflection microscopy: the dependence of fiber brightness on fiber orientation in imaging biopolymer networks. , 2010, Biophysical journal.

[43]  Cynthia A. Reinhart-King,et al.  Tensional homeostasis and the malignant phenotype. , 2005, Cancer cell.

[44]  Mikala Egeblad,et al.  Matrix Crosslinking Forces Tumor Progression by Enhancing Integrin Signaling , 2009, Cell.

[45]  C. Jackson,et al.  Three-dimensional collagen matrices induce delayed but sustained activation of gelatinase A in human endothelial cells via MT1-MMP. , 2000, The international journal of biochemistry & cell biology.

[46]  Muhammad H. Zaman,et al.  Modeling Persistence in Mesenchymal Cell Motility Using Explicit Fibers , 2014, Langmuir : the ACS journal of surfaces and colloids.

[47]  Yu-Li Wang,et al.  The regulation of traction force in relation to cell shape and focal adhesions. , 2011, Biomaterials.

[48]  J. Urbach,et al.  Size-dependent rheology of type-I collagen networks. , 2010, Biophysical journal.

[49]  Kenneth M. Yamada,et al.  Direct visualization of protease activity on cells migrating in three-dimensions. , 2009, Matrix Biology.