Synthesis of Injectable, Thermally Responsive, Chondroitin Sulfate-Cross-Linked Poly(N-isopropylacrylamide) Hydrogels

Abstract

In this study, we describe the synthesis and characterization of a biosynthetic hydrogel system that consists of a thermally responsive macromer and biological cross-linkers. By combining a poly­(N-isopropylacrylamide)-based thermogelling macromer with epoxy pendant groups and chondroitin sulfate cross-linkers that are modified to contain either hydrazide or N-hydroxysuccinimide pendant groups, we successfully fabricated a system that undergoes gelation when the temperature is raised from room temperature to 37 °C and is further stabilized via covalent links between the macromers. The anionic charge on chondroitin sulfate contributed to a high degree of gel swelling, while the cross-linking reaction between the macromers prevented post-formation syneresis. The rate of degradation of CS-cross-linked hydrogels was dependent on the degree of substitution of hydrazide-modified chondroitin sulfate cross-linkers. A higher molar content of chondroitin sulfate led to a greater osmotic pressure within the hydrogel and thus a higher compressive modulus. On the other hand, excessive amounts of chondroitin sulfate caused time-dependent cytotoxicity, as confirmed by a leachables cytocompatibility study. Overall, the system described in this study provides a versatile platform to synthesize hydrogels with differing combinations of compressive moduli and rates of degradation, which is achievable by varying the degree of substitution of hydrazide groups on CS-based cross-linkers.

Evaluating the physicochemical effects of conjugating peptides into thermogelling hydrogels for regenerative biomaterials applications

“…Following hydrogel formation, the swelling and degradation behavior were assessed as previously described [ 1 , 4 , 9 , 11 , 14 , 27 , 29 , 30 ]. After crosslinking for 24 h at 37°C to form the hydrogels, the hydrogels were transferred and weighed in a pre-weighed 20 ml scintillation vial to obtain the formation weight ( W f ).…”

“…This concentration of collagenase has been previously established as being representative of the wound-healing environment [ 31 ]. Swelling and degradation were evaluated on terminal timepoints (Days 1, 3, 7, 14 and 21) as previously described with media changes performed every 3 days [ 1 , 9 , 11 , 14 , 30 ]. At each terminal time point, the swollen weights were measured, and the sacrificial hydrogels were frozen and lyophilized.…”

“…The LCST and crosslinking times of the hydrogel mixtures were evaluated using differential scanning calorimetry (DSC) 250 (TA Instruments) as previously described [ 9 , 11 , 13 ]. P(NiPAAm-co-GMA) and the respective crosslinker were kept on ice before mixing immediately before each run.…”

“…P(NiPAAm) is a polymer, which has been utilized extensively for a variety of thermogelling applications as it possesses a lower critical solution temperature (LCST) above which the network begins to gel [ 4 , 9 , 10 ]. This LCST can be modulated through the addition of varied co-monomers with more hydrophilic co-monomers increasing the LCST, and hydrophobic co-monomers lowering the LCST [ 4 , 11 ]. This has been demonstrated through the co-polymerization of NiPAAm with glycidyl methacrylate to form P(NiPAAm-co-GMA), which reduces the peak LCST from 32 to 30°C, and even a four-part macromer with acrylic acid, glycidyl methacrylate and dimethyl-γ-butyrolactone acrylate (DBA).…”

“…While efficacious, most thermogelling materials also employ chemical crosslinking. This creates a dual-gelling network in which thermal gelation space-fills a defect when injected into an environment above the LCST while chemical crosslinking secures and strengthens the network and prevents syneresis [ 1 , 11 , 13 ]. This dual-gelling strategy for injectable materials has proven to be useful in creating many robust tissue engineering materials capable of delivering cells, microparticles and bioactive factors.…”