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Kristi S Anseth - One of the best experts on this subject based on the ideXlab platform.
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collagen networks within 3d peg hydrogels support valvular Interstitial Cell matrix mineralization
Acta Biomaterialia, 2021Co-Authors: Megan E Schroeder, Andrea Gonzalez Rodriguez, Kelly F Speckl, Cierra J Walker, Firaol S Midekssa, Joseph C Grim, Robert M Weiss, Kristi S AnsethAbstract:Enzymatically degradable hydrogels were designed for the 3D culture of valvular Interstitial Cells (VICs), and through the incorporation of various functionalities, we aimed to investigate the role of the tissue microenvironment in promoting the osteogenic properties of VICs and matrix mineralization. Specifically, porcine VICs were encapsulated in a poly(ethylene glycol) hydrogel crosslinked with a matrix metalloproteinase (MMP)-degradable crosslinker (KCGPQG↓IWGQCK) and formed via a thiol-ene photoclick reaction in the presence or absence of collagen type I to promote matrix mineralization. VIC-laden hydrogels were treated with osteogenic medium for up to 15 days, and the osteogenic response was characterized by the expression of RUNX2 as an early marker of an osteoblast-like phenotype, osteocalcin (OCN) as a marker of a mature osteoblast-like phenotype, and vimentin (VIM) as a marker of the fibroblast phenotype. In addition, matrix mineralization was characterized histologically with Von Kossa stain for calcium phosphate. Osteogenic response was further characterized biochemically with calcium assays, and physically via optical density measurements. When the osteogenic medium was supplemented with calcium chloride, OCN expression was upregulated and mineralization was discernable at 12 days of culture. Finally, this platform was used to screen various drug therapeutics that were assessed for their efficacy in preventing mineralization using optical density as a higher throughput readout. Collectively, these results suggest that matrix composition has a key role in supporting mineralization deposition within diseased valve tissue.
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collagen networks within 3d peg hydrogels support valvular Interstitial Cell matrix mineralization
Social Science Research Network, 2020Co-Authors: Megan E Schroeder, Andrea Gonzalez Rodriguez, Kelly F Speckl, Cierra J Walker, Firaol S Midekssa, Joseph C Grim, Kristi S AnsethAbstract:Enzymatically degradable hydrogels were designed for the 3D culture of valvular Interstitial Cells (VICs), and through the incorporation of various functionalities, we aimed to investigate the role of the tissue microenvironment in promoting the osteogenic properties of VICs and matrix mineralization. Specifically, VICs were encapsulated in a poly(ethylene glycol) hydrogel crosslinked with a matrix metalloproteinase (MMP)-degradable crosslinker (KCGPQG↓IWGQCK) and formed via a thiol-ene photoclick reaction in the presence or absence of collagen type I to promote matrix mineralization. VIC-laden hydrogels were treated with osteogenic medium for up to 15 days, and the osteogenic response was characterized by the expression of genetic markers, RUNX2 as an early marker of an osteoblast-like phenotype, and OCN, as a marker of a mature osteoblast-like phenotype. In addition, matrix mineralization was characterized histologically with a Von Kossa stain for calcium phosphate. Osteogenic response was further characterized biochemically with calcium assays, and physically via optical density measurements. When the osteogenic medium was supplemented with calcium chloride, OCN gene expression and mineralization were discernable within 12 days of culture. Finally, this platform was used to screen various drug therapeutics that were assessed for their efficacy in preventing mineralization using optical density as a higher throughput read out. Collectively, these results suggest that matrix composition has a key role in supporting mineralization deposition within diseased valve tissue.
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bioapplications of networks based on photo cross linked hyperbranched polymers
Macromolecular Symposia, 2010Co-Authors: Sara Pedron, Kristi S Anseth, Julie A Benton, Paula Bosch, C PeinadoAbstract:This article deals with some of the most recent developments in the use of hyperbranched polymers in biomedical applications. Some examples have been selected to show their potential in drug delivery, tissue engineering, imaging technologies and molecular imprinting. Moreover, the preparation of methacrylic networks using chemically-modified hyperbranched polymers as multifunctional macromonomers by photopolymerization is described. The capability to support valve Interstitial Cell culture was demonstrated and the adhesion and functionality of the Cells was related to the mechanical properties of these materials.
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characterization of valvular Interstitial Cell function in three dimensional matrix metalloproteinase degradable peg hydrogels
Biomaterials, 2009Co-Authors: Julie A Benton, Kristi S Anseth, Benjamin D FairbanksAbstract:Abstract Valvular Interstitial Cells (VICs) maintain functional heart valve structure and display transient fibroblast and myofibroblast properties. Most Cell characterization studies have been performed on plastic dishes; while insightful, these systems are limited. Thus, a matrix metalloproteinase (MMP) degradable poly(ethylene glycol) (PEG) hydrogel system is proposed in this communication as a useful tool for characterizing VIC function in 3D. When encapsulated, VICs attained spread morphology, and proliferated and migrated as shown through real-time Cell microscopy. Additionally, fibronectin derived pendant RGD was incorporated into the system to promote integrin binding. As RGD concentration increased from 0 to 2000 μ m , VIC process extension and integrin αvβ3 binding increased within two days. By day 10, integrin binding was equalized between conditions. VIC morphology and rate of process extension were also increased through decreasing the hydrogel matrix density presented to the Cells. VIC differentiation in response to exogenously delivered transforming growth factor-beta1 (TGF-β1) was also examined within the hydrogel networks. TGF-β1 increased expression of alpha smooth muscle actin (αSMA) and collagen-1 at both the mRNA and protein level by day 2 of culture, indicating myofibroblast differentiation, and was sustained over the course of the study (2 weeks). These studies demonstrate the utility, flexibility, and biological activity of this MMP-degradable system for the characterization of VICs, an important Cell population for tissue engineering viable valve replacements and understanding valvular pathobiology.
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characterization of valvular Interstitial Cell function in three dimensional matrix metalloproteinase degradable peg hydrogels
Biomaterials, 2009Co-Authors: Julie A Benton, Kristi S Anseth, Benjamin D FairbanksAbstract:Valvular Interstitial Cells (VICs) maintain functional heart valve structure and display transient fibroblast and myofibroblast properties. Most Cell characterization studies have been performed on plastic dishes; while insightful, these systems are limited. Thus, a matrix metalloproteinase (MMP) degradable poly(ethylene glycol) (PEG) hydrogel system is proposed in this communication as a useful tool for characterizing VIC function in 3D. When encapsulated, VICs attained spread morphology, and proliferated and migrated as shown through real-time Cell microscopy. Additionally, fibronectin derived pendant RGD was incorporated into the system to promote integrin binding. As RGD concentration increased from 0 to 2000 microM, VIC process extension and integrin alpha(v)beta(3) binding increased within two days. By day 10, integrin binding was equalized between conditions. VIC morphology and rate of process extension were also increased through decreasing the hydrogel matrix density presented to the Cells. VIC differentiation in response to exogenously delivered transforming growth factor-beta1 (TGF-beta1) was also examined within the hydrogel networks. TGF-beta1 increased expression of alpha smooth muscle actin (alphaSMA) and collagen-1 at both the mRNA and protein level by day 2 of culture, indicating myofibroblast differentiation, and was sustained over the course of the study (2 weeks). These studies demonstrate the utility, flexibility, and biological activity of this MMP-degradable system for the characterization of VICs, an important Cell population for tissue engineering viable valve replacements and understanding valvular pathobiology.
Avrum I Gotlieb - One of the best experts on this subject based on the ideXlab platform.
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the progression of calcific aortic valve disease through injury Cell dysfunction and disruptive biologic and physical force feedback loops
Cardiovascular Pathology, 2013Co-Authors: Avrum I GotliebAbstract:Calcific aortic valve disease (CAVD) is the most common form of heart valve disease in Western society and results in the second most common cardiovascular surgery performed. Despite its prevalence, high morbidity, and high mortality, the pathogenesis of CAVD still eludes our understanding. This review article brings together experimental in vivo and in vitro as well as human in vivo research in Cell and molecular pathobiology to construct an overarching hypothesis regarding the development and progression of CAVD. We focus on injury, Cell dysfunction, and disruptive biologic and physical forces, and how they function in positive feedback loops that result in the eventual calcification of the valve. We propose that injury, inflammation, matrix remodeling, and physical forces are all processes that influence each other and alter the normal physiologic functions of a key player in the pathogenesis of CAVD: the valve Interstitial Cell. We propose that the different phenotypes of the valve Interstitial Cell play essential roles in the pathogenesis of CAVD. We describe important physiologic processes which become dysfunctional including proliferation, migration, secretion of growth factors, chemokines and cytokines, and matrix remodeling. We also describe the emergence of chondrogenesis and osteogenesis in the fibrotic valve that lead to the severe clinical conditions of CAVD. CAVD appears to have a complex pathogenesis which fortunately can be studied in vitro and in vivo to identify ways to detect, treat, and prevent CAVD.
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fibroblast growth factor 2 promotes in vitro heart valve Interstitial Cell repair through the akt1 pathway
Cardiovascular Pathology, 2012Co-Authors: Avrum I GotliebAbstract:Abstract Background Fibroblast growth factor-2 promotes in vitro heart valve Interstitial Cell repair. Fibroblast growth factor-2 acts through betaglycan which is known to bind both transforming growth factor-β and fibroblast growth factor-2 at different locations on the molecule. When fibroblast growth factor-2 binds to betaglycan, transforming growth factor-β binding to betaglycan is reduced, allowing for more transforming growth factor-β to be available to activate pSmad2/3 which then promotes repair. This study investigates another pathway through which fibroblast growth factor-2 regulates valve Interstitial Cell repair. Methods We used an in vitro model of Cell culture disruption. Confluent valve Interstitial Cell monolayers were disrupted, creating an experimental wound in the confluent monolayer, and incubated in treatments of exogenous fibroblast growth factor-2, anti-fibroblast growth factor receptor antibody, active Akt1, and Akt inhibitor. Valve Interstitial Cell monolayers were immunohistochemically stained and quantified for nuclear pSmad2/3 at the wound edge. The extent of closure was measured up to 96 h after disruption. Results Anti-fibroblast growth factor receptor antibody significantly increased both nuclear pSmad2/3 staining at the wound edge and wound closure compared to nontreated control. This increase was less than that seen when valve Interstitial Cells were treated with fibroblast growth factor-2 and combined treatments of fibroblast growth factor-2 and anti-fibroblast growth factor receptor antibody did not further increase nuclear pSmad2/3 staining compared to fibroblast growth factor-2 alone. This suggested that the regulation of wound closure by fibroblast growth factor-2 also involved pathways other than transforming growth factor-β/Smad signaling. Treatment with Akt1 significantly increased wound closure, while Akt inhibitor reduced closure as compared to nontreated valve Interstitial Cells. Fibroblast growth factor-2 and fibroblast growth factor-2 neutralizing antibody up-regulated and down-regulated phosphorylated Akt1 expression in valve Interstitial Cells, respectively. Conclusion Fibroblast growth factor-2 promotes valve Interstitial Cell repair in two ways: the fibroblast growth factor-2/fibroblast growth factor-2 receptor interaction through the activation of Akt1 independent of the transforming growth factor-β/Smad2/3 signaling pathway and the previously described transforming growth factor-β/Smad signaling.
Michael S. Sacks - One of the best experts on this subject based on the ideXlab platform.
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serotonin receptor 2b signaling with Interstitial Cell activation and leaflet remodeling in degenerative mitral regurgitation
Journal of Molecular and Cellular Cardiology, 2018Co-Authors: Kathryn H Driesbaugh, Michael S. Sacks, Salma Ayoub, Mark A Oyama, Emanuela Branchetti, Juan B Grau, Samuel Keeney, Kimberly Glass, Nancy Rioux, John QuackenbushAbstract:Abstract Aims Mitral valve Interstitial Cells (MVIC) play an important role in the pathogenesis of degenerative mitral regurgitation (MR) due to mitral valve prolapse (MVP). Numerous clinical studies have observed serotonin (5HT) dysregulation in cardiac valvulopathies; however, the impact of 5HT-mediated signaling on MVIC activation and leaflet remodeling in MVP have been investigated to a limited extent. Here we test the hypothesis that 5HT receptors (5HTRs) signaling contributes to MVP pathophysiology. Methods and results Diseased human MV leaflets were obtained during cardiac surgery for MVP; normal MV leaflets were obtained from heart transplants. MV RNA was used for microarray analysis of MVP patients versus control, highlighting genes that indicate the involvement of 5HTR pathways and extraCellular matrix remodeling in MVP. Human MV leaflets were also studied in vitro and ex vivo with biomechanical testing to assess remodeling in the presence of a 5HTR2B antagonist (LY272015). MVP leaflets from Cavalier King Charles Spaniels were used as a naturally acquired in vivo model of MVP. These canine MVP leaflets (N = 5/group) showed 5HTR2B upregulation. This study also utilized CB57.1ML/6 mice in order to determine the effect of Angiotensin II infusion on MV remodeling. Histological analysis showed that MV thickening due to chronic Angiotensin II remodeling is mitigated by a 5HTR2B antagonist (LY272015) but not by 5HTR2A inhibitors. Conclusion In humans, MVP is associated with an upregulation in 5HTR2B expression and increased 5HT receptor signaling in the leaflets. Antagonism of 5HTR2B mitigates MVIC activation in vitro and MV remodeling in vivo . These observations support the view that 5HTR signaling is involved not only in previously reported 5HT-related valvulopathies, but it is also involved in the pathological remodeling of MVP.
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regulation of valve Interstitial Cell homeostasis by mechanical deformation implications for heart valve disease and surgical repair
Journal of the Royal Society Interface, 2017Co-Authors: Salma Ayoub, Chunghao Lee, Kathryn H Driesbaugh, Wanda Anselmo, Connor T Hughes, Giovanni Ferrari, Robert C Gorman, Joseph H Gorman, Michael S. SacksAbstract:Mechanical stress is one of the major aetiological factors underlying soft-tissue remodelling, especially for the mitral valve (MV). It has been hypothesized that altered MV tissue stress states lead to deviations from Cellular homeostasis, resulting in subsequent Cellular activation and extraCellular matrix (ECM) remodelling. However, a quantitative link between alterations in the organ-level in vivo state and in vitro-based mechanobiology studies has yet to be made. We thus developed an integrated experimental-computational approach to elucidate MV tissue and Interstitial Cell responses to varying tissue strain levels. Comprehensive results at different length scales revealed that normal responses are observed only within a defined range of tissue deformations, whereas deformations outside of this range lead to hypo- and hyper-synthetic responses, evidenced by changes in α-smooth muscle actin, type I collagen, and other ECM and Cell adhesion molecule regulation. We identified MV Interstitial Cell deformation as a key player in leaflet tissue homeostatic regulation and, as such, used it as the metric that makes the critical link between in vitro responses to simulated equivalent in vivo behaviour. Results indicated that Cell responses have a delimited range of in vivo deformations that maintain a homeostatic response, suggesting that deviations from this range may lead to deleterious tissue remodelling and failure.
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in situ deformation of the aortic valve Interstitial Cell nucleus under diastolic loading
Journal of Biomechanical Engineering-transactions of The Asme, 2007Co-Authors: Hsiaoying Shadow Huang, Jun Liao, Michael S. SacksAbstract:Within the aortic valve (AV) leaflet resides a population of Interstitial Cells (AVICs), which serve to maintain tissue structural integrity via protein synthesis and enzymatic degradation. AVICs are typically characterized as myofibroblasts, exhibit phenotypic plasticity, and may play an important role in valve pathophysiology. While it is known that AVICs can respond to mechanical stimuli in vitro, the level of in vivo AVIC deformation and its relation to local collagen fiber reorientation during the cardiac cycle remain unknown. In the present study, the deformation of AVICs was investigated using porcine AV glutaraldehyde fixed under 0–9 0 mm Hgtransvalvular pressures. The resulting change in nuclear aspect ratio (NAR) was used as an index of overall Cellular strain, and dependencies on spatial location and pressure loading levels quantified. Local collagen fiber alignment in the same valves was also quantified using small angle light scattering. A tissue-level finite element (FE) model of an AVIC embedded in the AV extraCellular matrix was also used explore the relation between AV tissue- and Cellularlevel deformations. Results indicated large, consistent increases in AVIC NAR with transvalvular pressure (e.g., from mean of 1.8 at 0m m Hgto a mean of 4.8 at 90 mm Hg), as well as pronounced layer specific dependencies. Associated changes in collagen fiber alignment indicated that little AVIC deformation occurs with the large amount of fiber straightening for pressures below 1m m Hg, followed by substantial increases in AVIC NAR from 4m m Hg to 90 mm Hg. While the tissue-level FE model was able to capture the qualitative response, it also underpredicted the extent of AVIC deformation. This result suggested that additional micromechanical and fiber-compaction effects occur at high pressure levels. The results of this study form the basis of understanding transvalvular pressure-mediated mechanotransduction within the native AV and first time quantitative data correlating AVIC nuclei deformation with AV tissue microstructure and deformation. DOI: 10.1115/1.2801670
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correlation between heart valve Interstitial Cell stiffness and transvalvular pressure implications for collagen biosynthesis
American Journal of Physiology-heart and Circulatory Physiology, 2006Co-Authors: David W Merryman, Farshid Guilak, Inchan Youn, Howard Lukoff, Paula M Krueger, Richard A Hopkins, Michael S. SacksAbstract:It has been speculated that heart valve Interstitial Cells (VICs) maintain valvular tissue homeostasis through regulated extraCellular matrix (primarily collagen) biosynthesis. VICs appear to be ph...
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the effects of Cellular contraction on aortic valve leaflet flexural stiffness
Journal of Biomechanics, 2006Co-Authors: David W Merryman, Hsiaoying Shadow Huang, Frederick J Schoen, Michael S. SacksAbstract:Abstract The aortic valve (AV) leaflet contains a heterogeneous Interstitial Cell population composed predominantly of myofibroblasts, which contain both fibroblast and smooth muscle Cell characteristics. The focus of the present study was to examine aortic valve Interstitial Cell (AVIC) contractile behavior within the intact leaflet tissue. Circumferential strips of porcine AV leaflets were mechanically tested under flexure, with the AVIC maintained in the normal, contracted, and contraction-inhibited states. Leaflets were flexed both with (WC) and against (AC) the natural leaflet curvature, both before and after the addition of 90 mM KCl to elicit Cellular contraction. In addition, a natural basal tonus was also demonstrated by treating the leaflets with 10 μ M thapsigargin to completely inhibit AVIC contraction. Results revealed a 48% increase in leaflet stiffness with AVIC contraction (from 703 to 1040 kPa, respectively) when bent in the AC direction ( p = 0.004 ) , while the WC direction resulted only in 5% increase (from 491 to 516.5 kPa, respectively—not significant) in leaflet stiffness in the active state. Also, the loss of basal tonus of the AVIC population with thapsigargin treatment resulted in 76% (AC, p = 0.001 ) and 54% (WC, p = 0.036 ) decreases in leaflet stiffness at 5 mM KCl levels, while preventing contraction with the addition of 90 mM KCl as expected. We speculate that the observed layer dependent effects of AVIC contraction are primarily due to varying ECM mechanical properties in the ventricularis and fibrosa layers. Moreover, while we have demonstrated that AVIC contractile ability is a significant contributor to AV leaflet bending stiffness, it most likely serves a role in maintaining AV leaflet tissue homeostasis that has yet to be elucidated.
K M J Menon - One of the best experts on this subject based on the ideXlab platform.
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metformin direct inhibition of rat ovarian theca Interstitial Cell proliferation
Fertility and Sterility, 2012Co-Authors: M Will, Murugesan Palaniappan, Helle Peegel, Pradeep P Kayampilly, K M J MenonAbstract:Objective To determine whether metformin has direct effects on ovarian theca-Interstitial (T-I) Cell proliferation through activation of adenosine monophosphate–activated protein kinase (AMPK). Design In vitro experimental study. Setting Academic medical center laboratory. Animal(s) Immature Sprague-Dawley female rats. Intervention(s) Ovarian T-I Cells were isolated, purified, and cultured in the absence (control) or presence of insulin (1 μg/mL) with or without metformin or other activators/inhibitors of AMPK (AICAR, compound C). Main Outcome Measure(s) Proliferation assessed by determination of expression levels of proteins involved in Cell cycle progression, cyclin D3, and cyclin-dependent kinase 4 (CDK4) with Western blot analysis, and determination of DNA synthesis with bromodeoxyuridine (BrdU) incorporation assay; activation of AMPK, Erk1/2, and S6K1 determined by Western blot analysis with the use of antibodies specific for the phosphorylated (activated) forms. Result(s) Metformin inhibited insulin-induced ovarian T-I Cell proliferation and the up-regulation of the Cell cycle regulatory proteins, cyclin D3 and CDK4. Metformin independently activated AMPK in a dose-dependent manner. Treatment with metformin inhibited insulin-induced activation of Erk1/2 and S6K1. This effect was reversed with the addition of compound C, a known AMPK inhibitor. Conclusion(s) Metformin directly inhibits proliferation of ovarian T-I Cells via an AMPK-dependent mechanism. These findings further validate the potential benefits of metformin in the treatment of conditions associated with hyperinsulinemia and excessive growth of ovarian T-I Cells (such as polycystic ovary syndrome).
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lipoprotein enhancement of ovarian theca Interstitial Cell steroidogenesis relative contribution of scavenger receptor class b type i and adenosine 5 triphosphate binding cassette type a1 transporter in high density lipoprotein cholesterol transport and androgen synthesis
Endocrinology, 2003Co-Authors: Susan Sucheta, Salman Azhar, K M J MenonAbstract:The theca-Interstitial Cells take up plasma high-density lipoprotein (HDL)- and low-density-lipoprotein-derived cholesterol to convert into steroid hormones. The uptake of HDL-derived cholesterol is mediated by the scavenger receptor, class B, type I (SR-BI). In nonsteroidogenic Cells, HDL-stimulated efflux of cholesterol has been shown to be mediated by the ATP-binding cassette A1 (ABCA1) transporter. Its expression has not been documented in steroidogenic Cells. The goal of the present study was to determine: 1) the role of SR-BI in theca-Interstitial Cell androgen production; 2) whether theca-Interstitial Cells express ABCA1 transporter mRNA; and 3) the relative roles of SR-BI and ABCA1 transporter in androgen production. The ABCA1 transporter mRNA expression in rat theca-Interstitial Cells was shown using RT-PCR and Northern blot analyses. The role of SR-BI and ABCA1 in androstenedione production was also examined by treating Cells with anti-SR-BI and 2-hydroxypropyl-beta-cyclodextrin in the presence and absence of human chorionic gonadotropin and/or human HDL(3). The treatment of theca-Interstitial Cells with anti-SR-BI antibody blocked more than 90% of HDL plus human chorionic gonadotropin-stimulated androstenedione production, and selective HDL-CE uptake. On the other hand, the use of inhibitors of ABCA1 transporter function had no discernible effect on HDL-supported androgen production. These data demonstrate that, although theca-Interstitial Cells express both SR-BI and ABCA1 transporter mRNA, the SR-BI pathway supplies the majority of the cholesterol required for androgen production. Furthermore, the present study presents evidence for a crucial role for SR-BI in HDL-mediated androgen production.
Jonathan T Butcher - One of the best experts on this subject based on the ideXlab platform.
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active tissue stiffness modulation controls valve Interstitial Cell phenotype and osteogenic potential in 3d culture
Acta Biomaterialia, 2016Co-Authors: Bin Duan, Ziying Yin, Laura Hockaday Kang, Richard L Magin, Jonathan T ButcherAbstract: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. Statement of Significance 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.
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stiffness and adhesivity control aortic valve Interstitial Cell behavior within hyaluronic acid based hydrogels
Acta Biomaterialia, 2013Co-Authors: Bin Duan, Laura A Hockaday, Edi Kapetanovic, Kevin H Kang, Jonathan T ButcherAbstract:Bioactive and biodegradable hydrogels that mimic the extraCellular matrix and regulate valve Interstitial Cells (VIC) behavior are of great interest as three-dimensional (3-D) model systems for understanding mechanisms of valvular heart disease pathogenesis in vitro and the basis for regenerative templates for tissue engineering. However, the role of stiffness and adhesivity of hydrogels in VIC behavior remains poorly understood. This study reports the synthesis of methacrylated hyaluronic acid (Me-HA) and oxidized and methacrylated hyaluronic acid, and the subsequent development of hybrid hydrogels based on modified HA and methacrylated gelatin (Me-Gel) for VIC encapsulation. The mechanical stiffness and swelling ratio of the hydrogels were tunable with the molecular weight of the HA and the concentration/composition of the precursor solution. The encapsulated VIC in pure HA hydrogels with lower mechanical stiffness showed a more spreading morphology compared to their stiffer counterparts and dramatically up-regulated alpha smooth muscle actin expression, indicating more activated myofibroblast properties. The addition of Me-Gel in Me-HA facilitated Cell spreading, proliferation and VIC migration from encapsulated spheroids and better maintained the VIC fibroblastic phenotype. The VIC phenotype transition during migration from encapsulated spheroids in both Me-HA and Me-HA/Me-Gel hydrogel matrixes was also observed. These findings are important for the rational design of hydrogels for controlling the VIC morphology, and for regulating the VIC phenotype and function. The Me-HA/Me-Gel hybrid hydrogels accommodated with VIC are promising as valve tissue engineering scaffolds and 3-D models for understanding valvular pathobiology.
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cyclic strain anisotropy regulates valvular Interstitial Cell phenotype and tissue remodeling in three dimensional culture
Acta Biomaterialia, 2012Co-Authors: Russell A Gould, Karen Chin, Thom P Santisakultarm, Amanda Dropkin, Jennifer M Richards, Chris B Schaffer, Jonathan T ButcherAbstract:Many planar connective tissues exhibit complex anisotropic matrix fiber arrangements that are critical to their biomechanical function. This organized structure is created and modified by resident fibroblasts in response to mechanical forces in their environment. The directionality of applied strain fields changes dramatically during development, aging, and disease, but the specific effect of strain direction on matrix remodeling is less clear. Current mechanobiological inquiry of planar tissues is limited to equibiaxial or uniaxial stretch, which inadequately simulates many in vivo environments. In this study, we implement a novel bioreactor system to demonstrate the unique effect of controlled anisotropic strain on fibroblast behavior in three-dimensional (3-D) engineered tissue environments, using aortic valve Interstitial fibroblast Cells as a model system. Cell seeded 3-D collagen hydrogels were subjected to cyclic anisotropic strain profiles maintained at constant areal strain magnitude for up to 96 h at 1 Hz. Increasing anisotropy of biaxial strain resulted in increased Cellular orientation and collagen fiber alignment along the principal directions of strain and Cell orientation was found to precede fiber reorganization. Cellular proliferation and apoptosis were both significantly enhanced under increasing biaxial strain anisotropy (P < 0.05). While cyclic strain reduced both vimentin and alpha-smooth muscle actin compared to unstrained controls, vimentin and alpha-smooth muscle actin expression increased with strain anisotropy and correlated with direction (P < 0.05). Collectively, these results suggest that strain field anisotropy is an independent regulator of fibroblast Cell phenotype, turnover, and matrix reorganization, which may inform normal and pathological remodeling in soft tissues.
