Fibronectin in Layer-by-Layer Assembled Films Switches Tumor Cells between 2D and 3D Morphology.

Tumor cells showing a 3D morphology and in coculture with endothelial cells are a valuable in vitro model for studying cell-cell interactions and for the development of pharmaceuticals. Here, we found that HepG2 cells, unlike endothelial cells, show differences in adhesion to fibronectin alone, or in combination with poly(allylamine hydrochloride). This response allowed us to engineer micropatterned heterotypic cultures of the two cell types using microfluidics to pattern cell adhesion. The resulting cocultures exhibit spatially encoded and physiologically relevant cell function. Further, we found that the protrusive, migratory and 3D morphological responses of HepG2 are synergistically modulated by the constituents of the hybrid extracellular matrix. Treating the hybrid material with the cross-linking enzyme transglutaminase inhibited 3D morphogenesis of tumor cells. Our results extend previous work on the role of fibronectin in layer-by-layer assembled films, and demonstrate that cell-specific differences in adhesion to fibronectin can be used to engineer tumor cell cocultures.

[1]  Roger D. Kamm,et al.  Identification of drugs as single agents or in combination to prevent carcinoma dissemination in a microfluidic 3D environment , 2015, Oncotarget.

[2]  P Bajcsy,et al.  Empirical gradient threshold technique for automated segmentation across image modalities and cell lines , 2015, Journal of microscopy.

[3]  David J. Barry,et al.  Open source software for quantification of cell migration, protrusions, and fluorescence intensities , 2015, The Journal of cell biology.

[4]  Xian Xu,et al.  Three-dimensional in vitro tumor models for cancer research and drug evaluation. , 2014, Biotechnology advances.

[5]  R Core Team,et al.  R: A language and environment for statistical computing. , 2014 .

[6]  Hybrid cell adhesive material for instant dielectrophoretic cell trapping and long-term cell function assessment. , 2011, Langmuir : the ACS journal of surfaces and colloids.

[7]  P. Carmeliet,et al.  Molecular mechanisms and clinical applications of angiogenesis , 2011, Nature.

[8]  Shuichi Takayama,et al.  High-throughput 3D spheroid culture and drug testing using a 384 hanging drop array. , 2011, The Analyst.

[9]  Cedric E. Ginestet.  ggplot2: Elegant Graphics for Data Analysis , 2011 .

[10]  Kristopher E Kubow,et al.  Fibronectin forms the most extensible biological fibers displaying switchable force-exposed cryptic binding sites , 2009, Proceedings of the National Academy of Sciences.

[11]  Anne L Plant,et al.  The treatment of collagen fibrils by tissue transglutaminase to promote vascular smooth muscle cell contractile signaling. , 2009, Biomaterials.

[12]  Paul R Van Tassel,et al.  Multilayer nanofilms as substrates for hepatocellular applications. , 2008, Biomaterials.

[13]  John T Elliott,et al.  Quantifying myosin light chain phosphorylation in single adherent cells with automated fluorescence microscopy , 2007, BMC Cell Biology.

[14]  L. Addadi,et al.  Dynamic study of the transition from hyaluronan‐ to integrin‐mediated adhesion in chondrocytes , 2006, The EMBO journal.

[15]  Micah Dembo,et al.  The dynamics and mechanics of endothelial cell spreading. , 2005, Biophysical journal.

[16]  J. Hubbell,et al.  Synthetic biomaterials as instructive extracellular microenvironments for morphogenesis in tissue engineering , 2005, Nature Biotechnology.

[17]  James P. Freyer,et al.  The Use of 3-D Cultures for High-Throughput Screening: The Multicellular Spheroid Model , 2004, Journal of biomolecular screening.

[18]  Mauro Piacentini,et al.  Transglutaminase 2: an enigmatic enzyme with diverse functions. , 2002, Trends in biochemical sciences.

[19]  Christopher S. Chen,et al.  Engineering cellular microenvironments to improve cell-based drug testing. , 2002, Drug discovery today.

[20]  M. Mrksich,et al.  Using electroactive substrates to pattern the attachment of two different cell populations , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[21]  G. Whitesides,et al.  Patterning Mammalian Cells Using Elastomeric Membranes , 2000 .

[22]  S. Akimov,et al.  Tissue Transglutaminase Is an Integrin-Binding Adhesion Coreceptor for Fibronectin , 2000, The Journal of cell biology.

[23]  M L Yarmush,et al.  Effect of cell–cell interactions in preservation of cellular phenotype: cocultivation of hepatocytes and nonparenchymal cells , 1999, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[24]  D. Lauffenburger,et al.  Cell Migration: A Physically Integrated Molecular Process , 1996, Cell.

[25]  M. Schwartz,et al.  The extracellular matrix as a cell survival factor. , 1993, Molecular biology of the cell.

[26]  D. Scudiero,et al.  Feasibility of a high-flux anticancer drug screen using a diverse panel of cultured human tumor cell lines. , 1991, Journal of the National Cancer Institute.

[27]  J. Vacanti,et al.  Control of tumor growth in animals by infusion of an angiogenesis inhibitor. , 1980, Proceedings of the National Academy of Sciences of the United States of America.

[28]  G. Schatten,et al.  Adhesion of cells to surfaces coated with polylysine. Applications to electron microscopy , 1975, The Journal of cell biology.

[29]  R. Hynes.  Alteration of cell-surface proteins by viral transformation and by proteolysis. , 1973, Proceedings of the National Academy of Sciences of the United States of America.

[30]  J. Tukey.  Comparing individual means in the analysis of variance. , 1949, Biometrics.