Human Umbilical Vein Endothelial Cells Form a Network on a Hyaluronic Acid/Gelatin Composite Hydrogel Moderately Crosslinked and Degraded by Hydrogen Peroxide

Abstract

The study of the capillary-like network formation of human umbilical vein endothelial cells (HUVECs) in vitro is important for understanding the factors that promote or inhibit angiogenesis. Here, we report the behavior of HUVECs on the composite hydrogels containing hyaluronic acid (HA) and gelatin with different degrees of degradation, inducing the different physicochemical properties of the hydrogels. The hydrogels were obtained through horseradish peroxidase (HRP)-catalyzed hydrogelation consuming hydrogen peroxide (H2O2, 16 ppm) supplied from the air, and the degradation degree was tuned by altering the exposure time to the air. The HUVECs on the composite hydrogel with intermediate stiffness (1.2 kPa) obtained through 120 min of the exposure were more elongated than those on the soft (0.4 kPa) and the stiff (2.4 kPa) composite hydrogels obtained through 15 min and 60 min of the exposure, respectively. In addition, HUVECs formed a capillary-like network only on the stiff composite hydrogel although those on the hydrogels with comparable stiffness but containing gelatin alone or alginate instead of HA did not form the network. These results show that the HA/gelatin composite hydrogels obtained through the H2O2-mediated crosslinking and degradation could be a tool for studies using HUVECs to understand the promotion and inhibition of angiogenesis.

Tuning the crosslinking and degradation of hyaluronic acid/gelatin hydrogels using hydrogen peroxide for muscle cell sheet fabrication

“…2a) can be attributed to alterations in the phenol moiety crosslinking and polymer chain degradation. These trends are consistent with our previous findings, 27,29 despite the different polymer contents of the hydrogels. The degradation of polymers by oxidative cleavage by H 2 O 2 is well known 36 and H 2 O 2 -induced degradation of Gelatin-Ph 28,35 and HA-Ph has been reported.…”

“…The chosen HRP concentration (1 U mL −1 ) facilitated the investigation of the correlations between HRP-catalysed crosslink formation and polymer chain degradation by H 2 O 2 . 27–29,35…”

“…The chosen HRP concentration (1 U mL À1 ) facilitated the investigation of the correlations between HRP-catalysed crosslink formation and polymer chain degradation by H 2 O 2 . [27][28][29]35 To modulate polymer crosslinking and degradation in the hydrogels with H 2 O 2 , we varied the exposure times of the HA-Ph, Gelatin-Ph, and HRP mixtures to air containing 16 ppm H 2 O 2 . The measurement of the mechanical properties has direct implications for the contradictory effects of H 2 O 2 .…”

“…We have reported that H 2 O 2 functions not only as an essential substrate for HRP-catalysed crosslinking of phenolic hydroxyl moieties but also degrades polymer chains in the hydrogels. 27–29 Our previous study demonstrated that C2C12 cell attachment and myogenic differentiation could be influenced by adjusting the degree of crosslinking and degradation of Gelatin-Ph molecules in hydrogels prepared from a solution containing Gelatin-Ph alone via H 2 O 2 -mediated reactions. 28…”

“…We have reported that H 2 O 2 functions not only as an essential substrate for HRP-catalysed crosslinking of phenolic hydroxyl moieties but also degrades polymer chains in the hydrogels. [27][28][29] Our previous study demonstrated that C2C12 cell attachment and myogenic differentiation could be influenced by adjusting the degree of crosslinking and degradation of Gelatin-Ph molecules in hydrogels prepared from a solution containing Gelatin-Ph alone via H 2 O 2 -mediated reactions. 28 The hypothesis for this study was that HA-Ph/Gelatin-Ph composite hydrogels with varying degrees of H 2 O 2 -mediated crosslinking and degradation would facilitate control over myogenic differentiation of myoblasts on the hydrogels and allow cell sheet harvesting using hyaluronidase (Fig.…”