Interplay of Hydrogel Composition and Geometry on Human Mesenchymal Stem Cell Osteogenesis

Abstract

Microgels are emerging as an outstanding platform for tissue regeneration because they overcome issues associated with conventional bulk/macroscopic hydrogels such as limited cell−cell contact and cell communication and low diffusion rates. Owing to the enhanced mass transfer and injectability via a minimally invasive procedure, these microgels are becoming a promising approach for bone regeneration applications. Nevertheless, there still remains a huge gap between the understanding of how the hydrogel matrix composition can influence cell response and overall tissue formation when switching from bulk formats to microgel format, which is often neglected or rarely studied. Here, we fabricated polyethylene glycol-based microgels and bulk hydrogels incorporating gelatin and hyaluronic acid (HA), either individually or together, and assessed the impact of both hydrogel composition and format upon the osteogenic differentiation of encapsulated human bone marrow-derived mesenchymal stem cells (hBMSCs). Osteogenesis was significantly greater in microgels than bulk hydrogels for both gelatin alone (Gel) and gelatin HA composite (Gel:HA) hydrogels, as determined by the expression of Runt-related transcription factor (Runx2) and alkaline phosphatase (ALP) genes and mineral deposition. Interestingly, Gel and Gel:HA hydrogels behaved differently between bulk and microgel format. In bulk format, overall osteogenic outcomes were better in Gel:HA hydrogels, but in microgel format, while the level of osteogenic gene expression was equivalent between both compositions, the degree of mineralization was reduced in Gel:HA microgels. Investigation into the affinity of hydroxyapatite for the different matrix compositions indicated that the decreased mineralization of Gel:HA microgels was likely due to a low affinity of hydroxyapatite to bind to HA and support mineral deposition, which has a greater impact on microgels than bulk hydrogels. Together, these findings suggest that both hydrogel composition and format can determine the success of tissue formation and that there is a complex interplay of these two factors on both cell behavior and matrix deposition. This has important implications for tissue engineering, showing that hydrogel composition and geometry must be evaluated together when optimizing conditions for cell differentiation and tissue formation.

Cell Microencapsulation within Gelatin-PEG Microgels Using a Simple Pipet Tip-Based Device

“…These studies suggest that some cells are more vulnerable when exposed to fluid shear than others 32,33 suggesting that the effects should be verified for each individual cell type tested. However, in our previous work, the flow rate ratio of 4:1 mL −1 (oil versus hydrogel phase) was applied to several cell types (mesenchymal stromal cells, 5,26 human fetal chondroprogenitor cells, 29 human articular chondrocytes, 29 CAR-T cells, 27 coculture of hematopoietic stem cells, and OP9-DL cells (bone marrow stromal cells transduced to express the Notch ligand, Delta-like 1 or 4)) 22 without any negative effects on cell viability. A further reduction in microgel size was achieved by using minimal ID tubing (1.44 mm), which allowed the 33G needle to be used (Figure 4D).…”

“…We have primarily used hydrogels formed from norbornene-functionalized gelatin cross-linked with thiol-functionalized poly­(ethylene glycol), PEG. , These polymers can be cross-linked under blue (400–500 nm) or UV light (365 nm) in the presence of a biocompatible photoinitiator, lithium phenyl-2,4,6-trimethylbenzoylphosphinate (LAP), affording robust hydrogels. The gelatin provides cell recognition function for cell adhesion and remodeling, while the PEG improves the mechanical properties, enabling a highly biocompatible system that has been proven to support cell viability and function. , However, other polymer formulations, such as the inclusion of functionalized hyaluronic acid are possible, providing the hydrogel precursor can flow through the device and be cross-linked by light postmicrogel fabrication. The procedure can also be adapted for varying cell types. , …”

“…The gelatin provides cell recognition function for cell adhesion and remodeling, while the PEG improves the mechanical properties, enabling a highly biocompatible system that has been proven to support cell viability and function. 5,25 However, other polymer formulations, such as the inclusion of functionalized hyaluronic acid are possible, 26 providing the hydrogel precursor can flow through the device and be crosslinked by light postmicrogel fabrication. The procedure can also be adapted for varying cell types.…”

Micro-hydrogel injectables that deliver effective CAR-T immunotherapy against 3D solid tumor spheroids

“…Finally, the solution was lyophilized to obtain norbornene-conjugated gelatin. Thiol modified linear PEG (PEG(SH) 2 ) was synthesized via a three-step route based on our previously reported protocol [ 26 , 29 ].…”

Histone H3K9 demethylase JMJD2B/KDM4B promotes osteogenic differentiation of bone marrow-derived mesenchymal stem cells by regulating H3K9me2 on RUNX2

“…Through anodization and subsequent dip coating treatment, two highly ordered nano-pits with two different sizes were successfully constructed on the surface of 316LSS and then stimulated osteogenesis and angiogenesis ( Ni et al, 2020 ). Shrestha et al (2020) also found that both the composition and form of the hydrogel can determine the success of tissue formation, and these two factors have a complex interaction with cell behavior and matrix deposition. This has important implications for tissue engineering.…”