title: Three-Dimensional High-Throughput Cell Encapsulation Platform to Study Changes in Cell-Matrix Interactions.
abstract: In their native extracellular microenvironment, cells respond to a complex array of biochemical and mechanical cues that can vary in both time and space. High-throughput methods that allow characterization of cell-laden matrices are valuable tools to screen through many combinations of variables, ultimately helping to evolve and test hypotheses related to cell-ECM signaling. Here, we developed a platform for high-throughput encapsulation of cells in peptide-functionalized poly(ethylene glycol) hydrogels. Hydrogels were synthesized using a thiol-ene, photoclick reaction, which allowed the cell matrix environment to be modified in real time. Matrix signals were dynamically altered by in situ tethering of RGDS (0-1.5 mM), a fibronectin-derived adhesive peptide that induced more elongation than RLD or IKVAV, and/or by increasing the matrix modulus (1 to 6 kPa). This method was demonstrated with aortic valvular interstitial cells (VICs), a population of cells responsible for the pathological fibrosis and matrix remodeling that leads to aortic stenosis. VIC response to cell-matrix interactions was characterized by quantifying cell morphology and the fraction of cells exhibiting α-smooth muscle actin (αSMA) stress fibers, a hallmark of the myofibroblast phenotype. VICs elongated in response to RGDS addition, with a dramatic change in morphology within 24 h. Myofibroblast activation was also dependent on RGDS addition, with VICs exhibiting high activation (16-24%) in 1 kPa gels with RGDS. Response to RGDS was path-dependent, with the amount of time exposed to the adhesive ligand important in determining VIC morphology and activation. Although VIC aspect ratios were dependent on the amount of time spent in a stiff vs soft gel, low levels of VIC activation (≤4%) were observed in any gels cultured in higher modulus (6 kPa vs 1 kPa) microenvironments.
authors.affiliation: Department of Chemical and Biological Engineering, ‡Department of Materials Science, and §Howard Hughes Medical Institute and the BioFrontiers Institute, University of Colorado at Boulder , Boulder, Colorado 80303, United States.
authors.forename: Kelly M
authors.initials: KM
authors.lastname: Mabry
authors.affiliation: Department of Chemical and Biological Engineering, ‡Department of Materials Science, and §Howard Hughes Medical Institute and the BioFrontiers Institute, University of Colorado at Boulder , Boulder, Colorado 80303, United States.
authors.forename: Megan E
authors.initials: ME
authors.lastname: Schroeder
authors.affiliation: Department of Chemical and Biological Engineering, ‡Department of Materials Science, and §Howard Hughes Medical Institute and the BioFrontiers Institute, University of Colorado at Boulder , Boulder, Colorado 80303, United States.
authors.forename: Samuel Z
authors.initials: SZ
authors.lastname: Payne
authors.affiliation: Department of Chemical and Biological Engineering, ‡Department of Materials Science, and §Howard Hughes Medical Institute and the BioFrontiers Institute, University of Colorado at Boulder , Boulder, Colorado 80303, United States.
authors.forename: Kristi S
authors.initials: KS
authors.lastname: Anseth
mesh_terms: D001021:Aortic Valve; D002450:Cell Communication; D002478:Cells, Cultured; D005109:Extracellular Matrix / Q000378:metabolism*; D020100:Hydrogels
chemical_list: D020100:Hydrogels
keywords: aortic stenosis; biomaterial; extracellular matrix; fibroblast; high-throughput; hydrogel