title: Multilayer three-dimensional filter paper constructs for the culture and analysis of aortic valvular interstitial cells.
abstract: Culturing aortic valvular interstitial cells in an environment that models the aortic valve is an essential step towards understanding the progression of calcific aortic valve disease. Here the adaption of a three-dimensional (3-D) stacked paper-based culture system is presented for analyzing valve cells in a thick collagen gel matrix. Filter paper layers, modeled after a 96-well plate design, were printed with a wax well-plate template and then seeded with valve cell and collagen mixtures that quickly gelled into 3-D cultures. Stacking these layers permitted extensive customization of culture thickness and cell density profiles to model the full thickness of native valve tissue. Aortic valvular interstitial cells seeded into the paper-based constructs consistently demonstrated high survival up to 14 days of culture with significant increases in cell number through the first 3 days of culture. After 4 days following seeding, valve cells in single layer cultures showed reduced smooth muscle α-actin expression with a stabilized cell density, suggesting a transition from an activated phenotype to a more quiescent state. Valve cells in multilayer cultures demonstrated the ability to migrate from layer to layer and had the highest smooth muscle α-actin expression in areas with predicted low oxygen tensions. These results establish the filter-paper-based method as a viable culture system for analyzing valve cells in an in vitro 3-D model of the aortic valve.
authors.affiliation: Department of Bioengineering, Rice University, Houston, TX 77005, USA.
authors.forename: Matthew C
authors.initials: MC
authors.lastname: Sapp
authors.affiliation: Department of Bioengineering, Rice University, Houston, TX 77005, USA.
authors.forename: Hannelle J
authors.initials: HJ
authors.lastname: Fares
authors.affiliation: Department of Bioengineering, Rice University, Houston, TX 77005, USA.
authors.forename: Ana C
authors.initials: AC
authors.lastname: Estrada
authors.affiliation: Department of Bioengineering, Rice University, Houston, TX 77005, USA. Electronic address: grande@rice.edu.
authors.forename: K Jane
authors.initials: KJ
authors.lastname: Grande-Allen
mesh_terms: D000199:Actins / Q000096:biosynthesis; D000818:Animals; D001021:Aortic Valve / Q000166:cytology / Q000378:metabolism*; D002465:Cell Movement; D002470:Cell Survival; D002478:Cells, Cultured; D005786:Gene Expression Regulation; D032389:Myocytes, Smooth Muscle / Q000166:cytology / Q000378:metabolism*; D010209:Paper; D013552:Swine; D023822:Tissue Engineering
chemical_list: D000199:Actins
keywords: Cell activation; Collagen; ECM (extracellular matrix); Heart valve