Biomimetic Hydrogels with Immobilized EphrinA1 for Therapeutic Angiogenesis

Abstract

The formation of a microvasculature is regulated in large part by cell-cell interactions. Ephrins and their Eph receptors mediate cell adhesion, repulsion, and migration, all critical processes in angiogenesis. 1 Here, we use a covalently immobilized ephrinA1, conjugated to poly(ethylene glycol), to induce vessel formation both in vitro and in vivo in poly(ethylene glycol) diacrylate (PEGDA) hydrogels. Human umbilical vein endothelial cell (HUVEC) tubulogenesis in matrix metalloproteinase- sensitive hydrogels was visualized from 6 hours to 7 days in response to three different concentrations of PEG-ephrinA1. The deposition of extracellular matrix proteins collagen IV and laminin that stabilize tubule formation were imaged, quantified, and found to be dependent upon PEG-ephrinA1 concentration. To confirm the importance of the EphA2-ephrinA1 interaction in tubule formation, soluble EphA2 was used to disrupt the EphA2-ephrinA1 interaction between a co-culture of HUVEC and human brain vascular pericyte cells. HUVECs seeded onto PEGDA hydrogels displayed a dose- dependent reduction in tubule formation in response to the soluble EphA2. Finally, hydrogels with releasable platelet derived growth factor (PDGF), immobilized RGDS, and covalently immobilized PEG-ephrinA1 were implanted into the mouse cornea micropocket. These hydrogels induced a more robust vascular response with an increase in vessel density as compared to hydrogels with releasable PDGF alone. As such, PEG-ephrinA1 may represent a promising molecule to regulate cell adhesion and migration for formation of a microvasculature in tissue engineered constructs.

RNA Sequencing Analysis Detection of a Novel Pathway of Endothelial Dysfunction in Pulmonary Arterial Hypertension

“…The latter have important functional consequences on angiogenesis and on the PA response to injury that relate directly to the development of PAH. We focused on COL4 and EFNA1 in particular, because both genes were downregulated in the RNAseq analysis, and both have important, intertwining roles in endothelial biology; COL4 is a major component of vascular basement membranes and EFNA1 is an EC guidance molecule (29) that induces the release of COL4 from these cells (21). Loss of BMPR2 signaling also impaired the ability of PAECs to produce COL4 and EFNA1 by a mechanism that was reproduced by reducing b-catenin, a previously reported downstream effector of BMPR2 gene regulation (4).…”

“…That is, with BMPR2 siRNA versus control siRNA, we recapitulated changes in three of four genes that were significantly up-regulated and three of six that were down-regulated in IPAH versus control PAECs. EFNA1 can regulate the release of COL4 from ECs (21) and therefore was selected along with COL4 for further functional analysis.…”

Tissue Engineering

“…One strategy towards this goal is to develop prevascularized constructs for implantation [7], while other strategies focus on rapid vascular invasion and perfusion [8]. The most promising research has combined these techniques to develop of rapidly vascularized scaffolds that recreate vascular space-filling properties similar to those found in normal, functional tissues, to provide adequate perfusion of implanted constructs [9][10][11].…”

Evaluation of photochemistry reaction kinetics to pattern bioactive proteins on hydrogels for biological applications

“…Theses radicals are critical for the reaction initiation but can also negatively affect proteins or cells that are present [11]. Two common photoinitiators used in bio-related PEG crosslinking include 2-dimethoxy-2-phenylacetophenone (DMPA) and lithium phenyl-2,4,6-trimethylbenzoylphosphinate (LAP) [12], [13]. Specifically, DMPA has been often used in combination with acrylate PEG chemistry [10], [12].…”

“…Two common photoinitiators used in bio-related PEG crosslinking include 2-dimethoxy-2-phenylacetophenone (DMPA) and lithium phenyl-2,4,6-trimethylbenzoylphosphinate (LAP) [12], [13]. Specifically, DMPA has been often used in combination with acrylate PEG chemistry [10], [12]. LAP has recently reported in studies utilizing thiol-ene reactions [13].…”