Smart Design of Stable Extracellular Matrix Mimetic Hydrogel: Synthesis, Characterization, and In Vitro and In Vivo Evaluation for Tissue Engineering

Abstract

The simplicity and versatility of hydrazone crosslinking has made it a strategy of choice for the conjugation of bioactive molecules. However, the labile nature of hydrazone linkages and reversibility of this coupling reaction restricts its full potential. Based on the fundamental understanding of hydrazone stability, this problem is circumvented by resonance‐stabilization of a developing N2 positive charge in a hydrazone bond. A novel chemistry is presented to develop a resilient hydrazone bond that is stable and non‐ reversible under physiological conditions. A carbodihydrazide (CDH) type hydrazide derivative of the biomolecule forms intrinsically stabilized hydrazone‐linkages that are nearly 15‐fold more stable at pH 5 than conventional hydrazone. This chemoselective coupling reaction is catalyst‐free, instantaneous, and virtually non‐cleavable under physiological conditions, therefore can serve as a catalyst‐free alternative to click chemistry. This novel crosslinking reaction is used to tailor a hyaluronan hydrogel, which delivered exceptional hydrolytic stability, mechanical properties, low swelling, and controlled enzymatic degradation. These desired characteristics are achieved without increasing the chemical crosslinking. The in vivo evaluation of this hydrogel revealed neo‐bone with highly ordered collagen matrix mimicking natural bone regeneration. The proximity ligation assay or PLA is used to detect blood vessels, which highlighted the quality of engineered tissue.

Hyaluronic acid hydrogel scaffolds with a triple degradation behavior for bone tissue engineering

“…These results suggested that the degradation of the HA hydrogels was accelerated at acidic pH because of the pH-sensitive cleavage of hydrazone bonds. Similar phenomenon was observed in the hydrazide-aldehyde crosslinking system in the literature (Oommen et al, 2013). The degradation products of LEV-PEG-LEV are biocompatible PEG and LEV (Hami, Amini, Khansan, Rezayat, & Gilani, 2014).…”

“…Moreover, the characteristic peak of hydrazone bonds at 1638 cm −1 could be noticed in the FT-IR spectra of the HA hydrogels by comparing these spectra and that of DTPH-HA, although their typical sharp absorption peak was not observed. The reason for this is that the peak was overlaid by the two strong characteristic peaks of amide groups at 1663 and 1618 cm −1 (Oommen et al, 2013). The spectrum line between the two absorption peaks became more smooth as the degree of crosslinking of HA hydrogels increased.…”

“…Hyaluronic acid (HA) hydrogel scaffolds possess some advantages, including facilitating proliferation of seeded cells and wound healing, good anti-inflammatory ability and promoting intracellular signaling (Oommen et al, 2013;Collins & Birkinshaw, 2013a,b;Collins, 2014). HA hydrogels have been widely applied in the biomedical field because of their good biocompatibility and structural diversity (Collins & Birkinshaw, 2013a,b).…”

“…On the other side, polyethylene glycol (PEG) has been widely used to synthesize many types of hydrogels for tissue engineering due to its good biocompatibility, ability to promote tissue regeneration and longterm in vivo security (Byeon et al, 2014;Burdick & Anseth, 2002;Yu et al, 2014;Yu, Cao, Zeng, Zhang, & Chen, 2013;Rufaihah et al, 2013). A gentle hydrazone bond crosslinking method was recently reported to synthesize HA hydrogels by reacting HA aldehyde with HA adipic dihydrazide (Oommen et al, 2013). However, macromolecular aldehydes used for the preparation of hydrogels based on hydrazone bond crosslinking still show cytotoxicity and might hinder the migration of cells, although they display better biocompatibility than small molecular aldehydes (Mckinnon et al, 2014).…”

Imine Hydrogels with Tunable Degradability for Tissue Engineering

“…This is expected since it has been shown that changing the structure of the dihydrazide group used as a cross-linker can affect hydrazone bond stability by increasing resonance stabilization by shortening the carbon chain between hydrazide groups. 40 …”