Biology subjects

Gentry, J. L.

Publications and source records attributed to Gentry, J. L..

3 recordsLinked to original sources

bioRxiv2025

Norbornene homopolymerization limits cell spreading in thiol-ene photoclick hydrogels

Thiol-ene click chemistry is a powerful tool for designing hydrogels mimicking the mechanical and biochemical properties of 3D cellular microenvironments. The high selectivity of thiol-norbornene step-growth polymerization enables precise control of crosslinking mechanism, circumventing the alkene homopolymerization present in other systems that can prevent encapsulated cell spreading. Limited stress relaxation of a dynamically-crosslinked norbornene-modified hyaluronic acid (NorHA) hydrogel employing a thiol-norbornene photoclick reaction led us to investigate the prevalence of norbornene homopolymerization in this supposed click reaction. Norbornene conversion was quantified in multiple thiol plus norbornene-modified polymer system permutations, revealing higher norbornene conversion than expected for 1:1 thiol-ene addition. We showed that decreasing the number of norbornenes per NorHA chain (f) mitigated network formation via norbornene homopolymerization. Dynamic hydrogels fabricated with NorHA of f = 8 (Nor8HA) exhibited 93.0 {+/-} 1.6% relaxation, while those fabricated with NorHA of f = 40 (Nor40HA) achieved only 42.3 {+/-} 0.1% relaxation. As early as day 3 of culture, Nor8HA hydrogels facilitated spreading of encapsulated human mesenchymal stromal cells (hMSCs) into a spindle-like morphology (aspect ratio: 2.95 {+/-} 0.38), while Nor40HA hydrogels appeared to constrain cells into a spherical or compact star morphology (aspect ratio: 1.22 {+/-} 0.01). Inference of a single-cell morphological space derived from a shape-matching distance metric validated the two distinct hMSC morphological phenotypes primarily associated with polymer f. Despite its widespread use as a click reaction, radical-mediated thiol-norbornene crosslinking was found to not be stoichiometric in dilute aqueous conditions used to fabricate hydrogels. Altering network topology through polymer f enabled the rescue of hydrogel dynamic behavior and encapsulated hMSC spreading, despite the presence of norbornene homopolymerization, highlighting the need to consider network-level properties when designing engineered cellular microenvironments.

bioengineering↗

bioRxiv2023

Modular multiwell viscoelastic hydrogel platform for two- and three-dimensional cell culture applications

Hydrogels have gained significant popularity as model platforms to study the reciprocal interactions between cells and their microenvironment. While hydrogel tools to probe many characteristics of the extracellular space have been developed, fabrication approaches remain challenging and time-consuming, limiting multiplexing or widespread adoption. Thus, we have developed a modular fabrication approach to generate distinct hydrogel microenvironments within 96-well plates for increased throughput of fabrication as well as integration with existing high-throughput assay technologies. This approach enables in situ hydrogel mechanical characterization and was used to generate both elastic and viscoelastic hydrogels across a range of stiffnesses. Additionally, this fabrication method enabled a 3-fold reduction in polymer and up to an 8-fold reduction in fabrication time required per hydrogel replicate. The feasibility of this platform for cell culture applications was demonstrated by measuring both population-level and single cell-level metrics via microplate reader and high-content imaging. Finally, the 96-well hydrogel array was utilized for 3D cell culture, demonstrating the ability to support high cell viability. Together, this work demonstrates a versatile and easily adoptable fabrication approach that can support the ever-expanding tool kit of hydrogel technologies for cell culture applications.

bioengineering↗

bioRxiv2023

Supramolecular fibrous hydrogel augmentation of uterosacral ligament suspension for treatment of pelvic organ prolapse

Uterosacral ligament suspension (USLS) is a common surgical treatment for pelvic organ prolapse (POP). However, the relatively high failure rate of up to 40% underscores a strong clinical need for complementary treatment strategies, such as biomaterial augmentation. Herein, we describe the first hydrogel biomaterial augmentation of USLS in a recently established rat model using an injectable fibrous hydrogel composite. Supramolecularly-assembled hyaluronic acid (HA) hydrogel nanofibers encapsulated in a matrix metalloproteinase (MMP)-degradable HA hydrogel create an injectable scaffold showing excellent biocompatibility and hemocompatibility. The hydrogel can be successfully delivered and localized to the suture sites of the USLS procedure, where it gradually degrades over 6 weeks. In situ mechanical testing 24 weeks post-operative in the multiparous USLS rat model shows the ultimate load (load at failure) to be 1.70 {+/-} 0.36 N for the intact uterosacral ligament (USL), 0.89 {+/-} 0.28 N for the USLS repair, and 1.37 {+/-} 0.31 N for the USLS + hydrogel (USLS+H) repair (n = 8). These results indicate that the hydrogel composite significantly improves load required for tissue failure compared to the standard USLS, even after the hydrogel degrades, and that this hydrogel-based approach could potentially reduce the high failure rate associated with USLS procedures.

bioengineering↗