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Biomaterials

Regenerative Medicine and Biomechanics

Engineering lipid mobility to enhance stress relaxation in hyaluronic acid hydrogels

Friday, October 13, 2023
4:15 PM – 4:30 PM PDT
Location: 333 (Center, Level 3)
Introduction:: This work presents a novel strategy to generate stress relaxation in hyaluronic acid (HA) hydrogels by leveraging lipid mobility. HA hydrogel matrices are naturally derived biomaterials used in disease modeling, 3D bioprinting, and tissue engineering 1–3. Matrix stress relaxation influences cell processes including cell proliferation, stem cell differentiation, and migration. Soft tissues have stress relaxation half-lives ranging from tens to hundreds of seconds. However, most synthetic HA hydrogels relax at significantly slower rates4. Conventional static covalent chemistries do not enable stress relaxation due to the irreversible nature of the crosslinking bonds. Recently, dynamic covalent chemistries have enabled the formation of hydrogels with enhanced stress relaxation. However, it remains difficult to achieve relaxation rates approaching those of soft tissues. New strategies are needed to augment stress relaxation independently of bond type.

Conventional hydrogels are formed by crosslinking polymer chains together to generate a network. The polymer chains themselves provide negligible stress relaxation, relying on reversible crosslinking chemistries to enable chain motion. In contrast, crosslinking directly through phospholipids in a liposome uses the mobility of individual lipids to enhance stress relaxation rates. We show that liposome nanoparticle crosslinked (LINC) hydrogels enable stress relaxation despite static covalent crosslinking chemistries. Lipid mobility and dynamic covalent chemistries synergize in HA-LINC gels to generate stress relaxation half-lives reaching values representative of soft tissues. We explore how liposome composition influences lipid mobility and thus hydrogel dynamics. Additionally, we demonstrate control of storage moduli while maintaining shear thinning, injectable, and self-healing properties.


Materials and Methods:: Liposomes were prepared using a thin film hydration and sonication method. The liposomes were surface functionalized with crosslinking groups (norbornene or hydrazine) to enable gelation when mixed with complementarily functionalized HA (tetrazine or benzaldehyde). Liposomes were composed of mixtures of DOPC, DOPE-Maleimide, and DOPE-PEG-Maleimide. Hydrogel mechanical properties were measured using small-angle oscillatory rheology.

Results, Conclusions, and Discussions:: Statically bonded HA hydrogels showed negligible stress relaxation, while statically bonded HA-LINC hydrogels have a relaxation half-life of 27 hours. This demonstrates that crosslinking directly through individual lipids enables stress relaxation independent of bond reversibility. HA hydrogels crosslinked through dynamic covalent hydrazone bonds (hydrazine + benzaldehyde) have a stress relaxation half-life of 14.7 hours. When this same crosslinking chemistry is used in HA-LINC hydrogels, lipid mobility and dynamic hydrazone bonding synergize to generate a stress relaxation half-life of 1.2 hours. This is significantly faster than either the relaxation due solely to lipid mobility or bond dynamics (27 and 14.7 hours, respectively).

We sought to engineer faster relaxation into the HA-LINC system by modifying lipid selection. Specifically, we hypothesized that adding a PEG linker between the lipids and the crosslinking group would add flexibility to the system and speed the stress relaxation rate. Adding a 1 kDa PEG linker between the crosslinking lipid and norbornene functionalization in the statically crosslinked HA-LINC hydrogel sped the stress relaxation half-life from 27 hours to 13.9 hours. Likewise, adding a PEG spacer before the hydrazine functionalization sped the stress relaxation half-life from 1.2 hours to 0.15 hours (540 seconds). Additionally, LINC hydrogels have controllable storage moduli (G’) ranging from 80-2000 Pa, while being shear-thinning, self-healing, and injectable.

Leveraging the molecular mobility of lipids gives us a new method of controlling stress relaxation independent of bond dynamics. Overall, LINC hydrogels illustrate a novel approach to engineering stress relaxation in hydrogel systems that can be used synergistically with existing hydrogel chemistries.


Acknowledgements (Optional): : This work was supported by the National Science Foundation Graduate Research Fellowship Program (NJB), the PhRMA Foundation Predoctoral Drug Delivery Fellowship (NJB), the National Institutes of Health, and the Stanford Cardiovascular Institute.


References (Optional): : (1) Abdalla, S.; Makhoul, G.; Duong, M.; Chiu, R. C. J.; Cecere, R. Hyaluronic Acid-Based Hydrogel Induces Neovascularization and Improves Cardiac Function in a Rat Model of Myocardial Infarction. Interact. Cardiovasc. Thorac. Surg. 2013, 17 (5), 767–772. https://doi.org/10.1093/icvts/ivt277.

(2) Vega, S. L.; Kwon, M. Y.; Song, K. H.; Wang, C.; Mauck, R. L.; Han, L.; Burdick, J. A. Combinatorial Hydrogels with Biochemical Gradients for Screening 3D Cellular Microenvironments. Nat. Commun. 2018, 9 (1), 614. https://doi.org/10.1038/s41467-018-03021-5.

(3) Wan, T.; Fan, P.; Zhang, M.; Shi, K.; Chen, X.; Yang, H.; Liu, X.; Xu, W.; Zhou, Y. Multiple Crosslinking Hyaluronic Acid Hydrogels with Improved Strength and 3D Printability. ACS Appl. Bio Mater. 2022, 5 (1), 334–343. https://doi.org/10.1021/acsabm.1c01141.

(4) Saraswathibhatla, A.; Indana, D.; Chaudhuri, O. Cell–Extracellular Matrix Mechanotransduction in 3D. Nat. Rev. Mol. Cell Biol. 2023, 1–22. https://doi.org/10.1038/s41580-023-00583-1.