Impact of immobilizing of low molecular weight hyaluronic acid within gelatin-based hydrogel through enzymatic reaction on behavior of enclosed endothelial cells

Abstract

The hydrogels having the ability to promote migration and morphogenesis of endothelial cells (ECs) are useful for fabricating vascularized dense tissues in vitro. The present study explores the immobilization of low molecular weight hyaluronic acid (LMWHA) derivative within gelatin-based hydrogel to stimulate migration of ECs. The LMWHA derivative possessing phenolic hydroxyl moieties (LMWHA-Ph) was bound to gelatin-based derivative hydrogel through the horseradish peroxidase-catalyzed reaction. The motility of ECs was analyzed by scratch migration assay and microparticle-based cell migration assay. The incorporated LMWHA-Ph molecules within hydrogel was found to be preserved stably through covalent bonds during incubation. The free and immobilized LMWHA-Ph did not lose an inherent stimulatory effect on human umbilical vein endothelial cells (HUVECs). The immobilized LMWHA-Ph within gelatin-based hydrogel induced the high motility of HUVECs, accompanied by robust cytoskeleton extension, and cell subpopulation expressing CD44 cell receptor. In the presence of immobilized LMWHA-Ph, the migration distance and the number of existing HUVECs were demonstrated to be encouraged in dose-dependent and time-dependent manners. Based on the results obtained in this work, it was concluded that the enzymatic immobilization of LMWHA-Ph within gelatin-based hydrogel represents a promising approach to promote ECs' motility and further exploitation for vascular tissue engineering applications.

Keywords: Endothelial cells; Gelatin-based hydrogel; Horseradish peroxidase; Immobilization; Low molecular weight hyaluronic acid; Migration.