title: Active tissue stiffness modulation controls valve interstitial cell phenotype and osteogenic potential in 3D culture.
abstract: Calcific aortic valve disease (CAVD) progression is a highly dynamic process whereby normally fibroblastic valve interstitial cells (VIC) undergo osteogenic differentiation, maladaptive extracellular matrix (ECM) composition, structural remodeling, and tissue matrix stiffening. However, how VIC with different phenotypes dynamically affect matrix properties and how the altered matrix further affects VIC phenotypes in response to physiological and pathological conditions have not yet been determined. In this study, we develop 3D hydrogels with tunable matrix stiffness to investigate the dynamic interplay between VIC phenotypes and matrix biomechanics. We find that VIC populated within hydrogels with valve leaflet like stiffness differentiate towards myofibroblasts in osteogenic media, but surprisingly undergo osteogenic differentiation when cultured within lower initial stiffness hydrogels. VIC differentiation progressively stiffens the hydrogel microenvironment, which further upregulates both early and late osteogenic markers. These findings identify a dynamic positive feedback loop that governs acceleration of VIC calcification. Temporal stiffening of pathologically lower stiffness matrix back to normal level, or blocking the mechanosensitive RhoA/ROCK signaling pathway, delays the osteogenic differentiation process. Therefore, direct ECM biomechanical modulation can affect VIC phenotypes towards and against osteogenic differentiation in 3D culture. These findings highlight the importance of the homeostatic maintenance of matrix stiffness to restrict pathological VIC differentiation.
  
  
  
  We implement 3D hydrogels with tunable matrix stiffness to investigate the dynamic interaction between valve interstitial cells (VIC, major cell population in heart valve) and matrix biomechanics. This work focuses on how human VIC responses to changing 3D culture environments. Our findings identify a dynamic positive feedback loop that governs acceleration of VIC calcification, which is the hallmark of calcific aortic valve disease. Temporal stiffening of pathologically lower stiffness matrix back to normal level, or blocking the mechanosensitive signaling pathway, delays VIC osteogenic differentiation. Our findings provide an improved understanding of VIC-matrix interactions to aid in interpretation of VIC calcification studies in vitro and suggest that ECM disruption resulting in local tissue stiffness decreases may promote calcific aortic valve disease.
authors.affiliation: Department of Biomedical Engineering, Cornell University, Ithaca, NY, USA.
authors.forename: Bin
authors.initials: B
authors.lastname: Duan
authors.affiliation: Department of Bioengineering, University of Illinois at Chicago, Chicago, IL, USA.
authors.forename: Ziying
authors.initials: Z
authors.lastname: Yin
authors.affiliation: Department of Biomedical Engineering, Cornell University, Ithaca, NY, USA.
authors.forename: Laura
authors.initials: L
authors.lastname: Hockaday Kang
authors.affiliation: Department of Bioengineering, University of Illinois at Chicago, Chicago, IL, USA.
authors.forename: Richard L
authors.initials: RL
authors.lastname: Magin
authors.affiliation: Department of Biomedical Engineering, Cornell University, Ithaca, NY, USA. Electronic address: jtb47@cornell.edu.
authors.forename: Jonathan T
authors.initials: JT
authors.lastname: Butcher
mesh_terms: D000469:Alkaline Phosphatase / Q000378:metabolism; D001021:Aortic Valve / Q000166:cytology*; D001696:Biomechanical Phenomena; D018929:Cell Culture Techniques / Q000379:methods*; D002454:Cell Differentiation; D002478:Cells, Cultured; D002648:Child; D003432:Cross-Linking Reagents / Q000737:chemistry; D005109:Extracellular Matrix / Q000378:metabolism; D006801:Humans; D020136:Hydrogel, Polyethylene Glycol Dimethacrylate / Q000494:pharmacology; D020100:Hydrogels / Q000737:chemistry; D056885:Interstitial Cells of Cajal / Q000166:cytology* / Q000187:drug effects / Q000201:enzymology; D058628:Myofibroblasts / Q000166:cytology / Q000187:drug effects; D010012:Osteogenesis / Q000187:drug effects*; D010641:Phenotype; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D013997:Time Factors; D054460:rho-Associated Kinases / Q000378:metabolism; D020742:rhoA GTP-Binding Protein / Q000378:metabolism
chemical_list: D003432:Cross-Linking Reagents; D020100:Hydrogels; D047428:Protein Kinase Inhibitors; D020136:Hydrogel, Polyethylene Glycol Dimethacrylate; D054460:rho-Associated Kinases; D000469:Alkaline Phosphatase; D020742:rhoA GTP-Binding Protein
keywords: Calcific aortic valve disease; Elastography; Hydrogel; Magnetic resonance imaging; Myofibroblast; Stiffness