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Microarray analyses to quantify advantages of 2D and 3D hydrogel culture systems in maintaining the native valvular interstitial cell phenotype.

ABSTRACT: Valvular interstitial cells (VICs) actively maintain and repair heart valve tissue; however, persistent activation of VICs to a myofibroblast phenotype can lead to aortic stenosis. To better understand and quantify how microenvironmental cues influence VIC phenotype and myofibroblast activation, we compared expression profiles of VICs cultured on poly(ethylene glycol) (PEG) gels to those cultured on tissue culture polystyrene (TCPS), as well as fresh isolates. In general, VICs cultured in hydrogel matrices had lower levels of activation (<10%), similar to levels seen in healthy valve tissue, while VICs cultured on TCPS were ?75% activated myofibroblasts. VICs cultured on TCPS also exhibited a higher magnitude of perturbations in gene expression than soft hydrogel cultures when compared to the native phenotype. Using peptide-modified PEG gels, VICs were seeded on (2D), as well as encapsulated in (3D), matrices of the same composition and modulus. Despite similar levels of activation, VICs cultured in 2D had distinct variations in transcriptional profiles compared to those in 3D hydrogels. Genes related to cell structure and motility were particularly affected by the dimensionality of the culture platform, with higher expression levels in 2D than in 3D. These results indicate that dimensionality may play a significant role in dictating cell phenotype (e.g., through differences in polarity, diffusion of soluble signals), and emphasize the importance of using multiple metrics when characterizing cell phenotype.

SUBMITTER: Mabry KM

PROVIDER: S-EPMC4661067 | biostudies-literature | 2016 Jan

REPOSITORIES: biostudies-literature

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Microarray analyses to quantify advantages of 2D and 3D hydrogel culture systems in maintaining the native valvular interstitial cell phenotype.

Mabry Kelly M KM Payne Samuel Z SZ Anseth Kristi S KS

Biomaterials 20150928

Valvular interstitial cells (VICs) actively maintain and repair heart valve tissue; however, persistent activation of VICs to a myofibroblast phenotype can lead to aortic stenosis. To better understand and quantify how microenvironmental cues influence VIC phenotype and myofibroblast activation, we compared expression profiles of VICs cultured on poly(ethylene glycol) (PEG) gels to those cultured on tissue culture polystyrene (TCPS), as well as fresh isolates. In general, VICs cultured in hydrog ...[more]

PMID: 26433490

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Project description:IntroductionLack of effective pharmacological treatment makes valvular calcification a significant clinical problem in patients with valvular disease and bioprosthetic/mechanical valve replacement therapies. Elevated levels of reactive oxygen species (ROS) in valve tissue have been identified as a prominent hallmark and driving factor for valvular calcification. However, the therapeutic value of ROS-modulating agents for valvular calcification remains elusive. We hypothesized that ROS-modulating shape-specific cerium oxide nanoparticles (CNPs) will inhibit oxidative stress-induced valvular calcification. CNPs are a class of self-regenerative ROS-modulating agents, which can switch between Ce3+ and Ce4+ in response to oxidative microen-vironment. In this work, we developed oxidative stress-induced valve calcification model using two patient-derived stenotic valve interstitial cells (hVICs) and investigated the therapeutic effect of shape-specific CNPs to inhibit hVIC calcification.MethodsHuman valvular interstitial cells (hVICs) were obtained from a normal healthy donor and two patients with calcified aortic valves. hVICs were characterized for their phenotypic (mesenchymal, myofibroblast and osteoblast) marker expression by qRT-PCR and antioxidant enzymes activity before and after exposure to hydrogen peroxide (H2O2)-induced oxidative stress. Four shape-specific CNPs (sphere, short rod, long rod, and cube) were synthesized via hydrothermal or ultra-sonication method and characterized for their biocompatibility in hVICs by alamarBlue® assay, and ROS scavenging ability by DCFH-DA assay. H2O2 and inorganic phosphate (Pi) were co-administrated to induce hVIC calcification in vitro as demonstrated by Alizarin Red S staining and calcium quantification. The effect of CNPs on inhibiting H2O2-induced hVIC calcification was evaluated.ResultshVICs isolated from calcified valves exhibited elevated osteoblast marker expression and decreased antioxidant enzyme activities compared to the normal hVICs. Due to the impaired antioxidant enzyme activities, acute H2O2-induced oxidative stress resulted in higher ROS levels and osteoblast marker expression in both diseased hVICs when compared to the normal hVICs. Shape-specific CNPs exhibited shape-dependent abiotic ROS scavenging ability, and excellent cytocompatibility. Rod and sphere CNPs scavenged H2O2-induced oxidative stress in hVICs in a shape- and dose-dependent manner by lowering intracellular ROS levels and osteoblast marker expression. Further, CNPs also enhanced activity of antioxidant enzymes in hVICs to combat oxidative stress. Cube CNPs were not effective ROS scavengers. The addition of H2O2 in the Pi-induced calcification model further increased calcium deposition in vitro in a time-dependent manner. Co-administration of rod CNPs with Pi and H2O2 mitigated calcification in the diseased hVICs.ConclusionsWe demonstrated that hVICs derived from calcified valves exhibited impaired antioxidant defense mechanisms and were more susceptible to oxidative stress than normal hVICs. CNPs scavenged H2O2-induced oxidative stress in hVICs in a shape-dependent manner. The intrinsic ROS scavenging ability of CNPs and their ability to induce cellular antioxidant enzyme activities may confer protection from oxidative stress-exacerbated calcification. CNPs represent promising antioxidant therapy for treating valvular calcification and deserve further investigation.

Dataset Information

Microarray analyses to quantify advantages of 2D and 3D hydrogel culture systems in maintaining the native valvular interstitial cell phenotype.

Publications

Microarray analyses to quantify advantages of 2D and 3D hydrogel culture systems in maintaining the native valvular interstitial cell phenotype.

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