Skip to main content
2024 BMES Annual Meeting Main banner
Icon Legend
This session is not in your schedule.
This session is in your schedule. Click again to remove it.
Back

Biomaterials

Biomaterials - Poster Session E

Poster O9 - Modifying thiolated gelatin hydrogels for improved material integration

Saturday, October 26, 2024
10:00 AM – 11:00 AM EST
Location: Exhibit Hall E, F & G

Introduction:  The enthesis transduces strain between the tendon-bone junction. Most common enthesis injuries happen in the rotator cuff and are treated by mechanically attaching the tendon and bone. In the US, approximately 250,000 rotator cuff surgeries are performed annually, resulting in up to 94% of retears, suggesting a need for new treatment approach. Previously, we have developed a triphasic biomaterial to treat enthesis injuries, featuring a thiolated polyethylene glycol (PEG-SH) hydrogel interface connecting mineralized (bone) and non-mineralized collagen (tendon) porous scaffolds to dissipate localized strain at the interface, however, PEG-SH is biologically inert. Gelatin has cell attachment and remodeling zones that can support cell metabolic processes but lack mechanical stability. We explored gelatin backbone modifications to modulate thiolated gelatin (Gel-SH) hydrogel crosslink kinetics and mechanical properties, aiming for a bioactive and tough material that can be integrated between collagen suspensions. Functional groups in gelatin are used as a crosslink moiety, as a target for functional group substitution or both, as presented here. We first substituted carboxylic groups for primary amines in the gelatin backbone to yield higher thiolation degrees. Gel-SH hydrogels (primary crosslinked) were formed by creating disulfide bonds on oxidized Gel-SH, catalyzed by horseradish peroxidase (HRP). A secondary crosslinked method was also applied to the Gel-SH system using carbodiimide bond strategies. We studied Gel-SH system rheological properties and will explore the benefits of incorporating it in the triphasic model.

Materials and

Methods:  Aminated gelatin was synthesized using ethylenediamine at different mass ratio (0, 0.35, 1.05 and 2.8 g of ethylenediamine) to g of gelatin and EDAC (1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride) with a 5.6:1 mass ratio (ethylenediamine: EDAC) for 24 hrs, dialyzed and lyophilized. Aminated gelatin was then thiolated to form Gel-SH by substituting primary amines on the gelatin backbone for thiols using Traut’s reagent, dialyzed and lyophilized. Gel-SH hydrogels were fabricated by creating disulfide bonds using HRP as a catalyst reagent to create phenolic radicals on tyramine, which oxidizes sulfhydryl groups on Gel-SH (Figure 1a). Hydrogels were then subjected to a secondary crosslink process 90 minutes in NHS/EDAC solution. Hydrogels were washed in PBS to remove any crosslinker debris. Time sweeps at 0.8 rad/s and 2% strain were performed to characterize kinetics parameters such as gelation time. Amplitude sweeps were performed to validate linear viscoelastic region and a frequency sweep was used to determine final elastic properties.

Results, Conclusions, and Discussions: Results and

Discussion: Gelatin based materials offer the advantage of high availability, fast degradation and high modularity. Here we proposed gelatin backbone modification to achieve materials with greater stability, which could be used to substitute synthetic polymers in a strain reducer triphasic design for rotator cuff injury. Higher conversions of thiols by substituting carboxylic acids for primary free amines were achieved (Figure 1b). Faster crosslink kinetics trends were observed on aminated Gel-SH versions; however, this was not followed by the most aminated version, 2.8g (Figure 1c-d). Lower degree of functionalization showed higher equilibrium elasticities on primary crosslinked hydrogels (Figure 1e). We believe these unexpected results could be due to high alterations in the gelatin backbone's ability to form physical bonding, or self-chain crosslink due to high concentrations of thiol groups. Carbodiimide crosslink had significant improvement on a non-aminated Gel-SH version, 0g; and no significant improvement was noticed on the other versions, this is due to a reduced number of carboxylic acid and amines ratio in the backbone that can make a potential crosslink (Figure 1e).

Conclusions: Naturally derived polymers often lack mechanical stability, but their biological compatibility and extracellular matrix-like properties are attractive for regenerative strategies. We exploited gelatin’s modularity and implemented a dual crosslink system to control their crosslink kinetics and the final elasticity. While lower elasticities were achieved on aminated Gel-SH versions, their potential to form faster crosslinks could improve the collagen suspension integration on the triphasic design, leading to a more robust and biologically active material. Current work is investigating Gel-SH's ability to reduce strain between collagen compartments through uniaxial tensile testing. This work expands methods to transition from synthetic materials to naturally derived and bioactive polymers like gelatin.

Acknowledgements (Optional):