Dataset Information

Differential effects of cell adhesion, modulus and VEGFR-2 inhibition on capillary network formation in synthetic hydrogel arrays.

ABSTRACT: Efficient biomaterial screening platforms can test a wide range of extracellular environments that modulate vascular growth. Here, we used synthetic hydrogel arrays to probe the combined effects of Cys-Arg-Gly-Asp-Ser (CRGDS) cell adhesion peptide concentration, shear modulus and vascular endothelial growth factor receptor 2 (VEGFR2) inhibition on human umbilical vein endothelial cell (HUVEC) viability, proliferation and tubulogenesis. HUVECs were encapsulated in degradable poly(ethylene glycol) (PEG) hydrogels with defined CRGDS concentration and shear modulus. VEGFR2 activity was modulated using the VEGFR2 inhibitor SU5416. We demonstrate that synergy exists between VEGFR2 activity and CRGDS ligand presentation in the context of maintaining HUVEC viability. However, excessive CRGDS disrupts this synergy. HUVEC proliferation significantly decreased with VEGFR2 inhibition and increased modulus, but did not vary monotonically with CRGDS concentration. Capillary-like structure (CLS) formation was highly modulated by CRGDS concentration and modulus, but was largely unaffected by VEGFR2 inhibition. We conclude that the characteristics of the ECM surrounding encapsulated HUVECs significantly influence cell viability, proliferation and CLS formation. Additionally, the ECM modulates the effects of VEGFR2 signaling, ranging from changing the effectiveness of synergistic interactions between integrins and VEGFR2 to determining whether VEGFR2 upregulates, downregulates or has no effect on proliferation and CLS formation.

SUBMITTER: Nguyen EH

PROVIDER: S-EPMC3970236 | biostudies-literature | 2014 Feb

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Differential effects of cell adhesion, modulus and VEGFR-2 inhibition on capillary network formation in synthetic hydrogel arrays.

Nguyen Eric H EH Zanotelli Matthew R MR Schwartz Michael P MP Murphy William L WL

Biomaterials 20131211 7

Efficient biomaterial screening platforms can test a wide range of extracellular environments that modulate vascular growth. Here, we used synthetic hydrogel arrays to probe the combined effects of Cys-Arg-Gly-Asp-Ser (CRGDS) cell adhesion peptide concentration, shear modulus and vascular endothelial growth factor receptor 2 (VEGFR2) inhibition on human umbilical vein endothelial cell (HUVEC) viability, proliferation and tubulogenesis. HUVECs were encapsulated in degradable poly(ethylene glycol) ...[more]

PMID: 24332391

Similar Datasets

Project description:Activation of vascular endothelial cells (ECs) by growth factors initiates a cascade of events during angiogenesis in vivo consisting of EC tip cell selection, sprout formation, EC stalk cell proliferation, and ultimately vascular stabilization by support cells. Although EC functional assays can recapitulate one or more aspects of angiogenesis in vitro, they are often limited by undefined substrates and lack of dependence on key angiogenic signaling axes. Here, we designed and characterized a chemically-defined model of endothelial sprouting behavior in vitro using human induced pluripotent stem cell-derived endothelial cells (iPSC-ECs). We rapidly encapsulated iPSC-ECs at high density in poly(ethylene glycol) (PEG) hydrogel spheres using thiol-ene chemistry and subsequently encapsulated cell-dense hydrogel spheres in a cell-free hydrogel layer. The hydrogel sprouting array supported pro-angiogenic phenotype of iPSC-ECs and supported growth factor-dependent proliferation and sprouting behavior. iPSC-ECs in the sprouting model responded appropriately to several reference pharmacological angiogenesis inhibitors of vascular endothelial growth factor, NF-?B, matrix metalloproteinase-2/9, protein kinase activity, and ?-tubulin, which confirms their functional role in endothelial sprouting. A blinded screen of 38 putative vascular disrupting compounds from the US Environmental Protection Agency's ToxCast library identified six compounds that inhibited iPSC-EC sprouting and five compounds that were overtly cytotoxic to iPSC-ECs at a single concentration. The chemically-defined iPSC-EC sprouting model (iSM) is thus amenable to enhanced-throughput screening of small molecular libraries for effects on angiogenic sprouting and iPSC-EC toxicity assessment.Angiogenesis assays that are commonly used for drug screening and toxicity assessment applications typically utilize natural substrates like Matrigel(TM) that are difficult to spatially pattern, costly, ill-defined, and may exhibit lot-to-lot variability. Herein, we describe a novel angiogenic sprouting assay using chemically-defined, bioinert poly(ethylene glycol) hydrogels functionalized with biomimetic peptides to promote cell attachment and degradation in a reproducible format that may mitigate the need for natural substrates. The quantitative assay of angiogenic sprouting here enables precise control over the initial conditions and can be formulated into arrays for screening. The sprouting assay here was dependent on key angiogenic signaling axes in a screen of angiogenesis inhibitors and a blinded screen of putative vascular disrupting compounds from the US-EPA.

Project description:Here, we have developed a novel method for forming hydrogel arrays using surfaces patterned with differential wettability. Our method for benchtop array formation is suitable for enhanced-throughput, combinatorial screening of biochemical and biophysical cues from chemically defined cell culture substrates. We demonstrated the ability to generate these arrays without the need for liquid handling systems and screened the combinatorial effects of substrate stiffness and immobilized cell adhesion peptide concentration on human mesenchymal stem cell (hMSC) behavior during short-term 2-dimensional cell culture. Regardless of substrate stiffness, hMSC initial cell attachment, spreading, and proliferation were linearly correlated with immobilized CRGDS peptide concentration. Increasing substrate stiffness also resulted in increased hMSC initial cell attachment, spreading, and proliferation; however, examination of the combinatorial effects of CRGDS peptide concentration and substrate stiffness revealed potential interplay between these distinct substrate signals. Maximal hMSC proliferation seen on substrates with either high stiffness or high CRGDS peptide concentration suggests that some baseline level of cytoskeletal tension was required for hMSC proliferation on hydrogel substrates and that multiple substrate signals could be engineered to work in synergy to promote mechanosensing and regulate cell behavior.Statement of significanceOur novel array formation method using surfaces patterned with differential wettability offers the advantages of benchtop array formation for 2-dimensional cell cultures and enhanced-throughput screening without the need for liquid handling systems. Hydrogel arrays formed via our method are suitable for screening the influence of chemical (e.g. cell adhesive ligands) and physical (stiffness, size, shape, and thickness) substrate properties on stem cell behavior. The arrays are also fully compatible with commercially available micro-array add-on systems, which allows for simultaneous control of the insoluble and soluble cell culture environment. This study used hydrogel arrays to demonstrate that synergy between cell adhesion and mechanosensing can be used to regulate hMSC behavior.

Dataset Information

Differential effects of cell adhesion, modulus and VEGFR-2 inhibition on capillary network formation in synthetic hydrogel arrays.

Publications

Differential effects of cell adhesion, modulus and VEGFR-2 inhibition on capillary network formation in synthetic hydrogel arrays.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets