Combinatorial hydrogels with biochemical gradients for screening 3D cellular microenvironments

Abstract

3D microenvironmental parameters control cell behavior, but can be challenging to investigate over a wide range of conditions. Here, a combinatorial hydrogel platform is developed that uses light-mediated thiol-norbornene chemistry to encapsulate cells within hydrogels with biochemical gradients made by spatially varied light exposure. Specifically, mesenchymal stem cells are photoencapsulated in norbornene-modified hyaluronic acid hydrogels functionalized with gradients (0–5 mM) of peptides that mimic cell-cell or cell-matrix interactions, either as single or orthogonal gradients. Chondrogenesis varied spatially in these hydrogels based on the local biochemical formulation, as indicated by Sox9 and aggrecan expression levels. From 100 combinations investigated, discrete hydrogels are formulated and early gene expression and long-term cartilage-specific matrix production are assayed and found to be consistent with screening predictions. This platform is a scalable, high-throughput technique that enables the screening of the effects of multiple biochemical signals on 3D cell behavior.

Fine‐Tunable and Injectable 3D Hydrogel for On‐Demand Stem Cell Niche

“…We had chosen two different tracks for inducing MSC chondrogenesis, i.e., stimulation of receptor mediated signaling (TGF-β1) and cell-to-cell interactions (N-cadherin). [18,29] Though the HAV sequence alone is enough to induce MSC chondrogenesis, a previous study revealed that an elongated HAV sequence such as CLRAHAVDIN was more effective than the short HAV motif. [27] Out of all the peptide sequences in TGF-β1, the binding site exhibiting the maximum reactivity to the TGF-β1 receptor was selected for this study (CESPLKRQ).…”

“…Depending on the use of specific differentiation inducible factors, including chemicals, proteins, and peptide, the fate of stem cells and ultimate phenotype may vary (e.g., stem cells may differentiate into osteocytes, chondrocytes, myocytes, adipocytes, etc.). [18] Conversely, from the perspective of fine-tuning hydrogels with bioactive molecules, the modification of hydrogel for the stem cell differentiation leads to unpredictable molecular substitutions [19] or physical mixing of bioactive factors. [17] Although recently reported fine-tuned hydrogels were prepared to regulate stem cell differentiation, it is difficult to use them in vivo system owing to the requirement for use of transdermal UV irradiation, which is carcinogenic.…”

Recent Strategies in Fabrication of Gradient Hydrogels for Tissue Engineering Applications

“…To confirm the accuracy of the screening platform, they created a discrete hydrogel with a specific concentration of HAV and RGD, and analyzed differentiation of MSCs. As a result, they found that the outcome was consistent with that of the screening prediction 50. Similarly, Aizawa et al fabricated agarose hydrogels containing spatial gradient of vascular endothelial GF 165 (VEGF165) 51.…”

“…By using sliding optical mask or by controlling the intensity and exposure time of light, the total amount of energy transferred to hydrogel precursor could be controlled to form biochemical cue gradient. For example, Vega et al developed norbornene‐modified hyaluronic acid (NorHA) hydrogels with dual peptide gradient to investigate the behavior of MSCs over a wide range of conditions 50. Briefly, NorHA precursors containing dithiol (DTT) crosslinker and 0.5 wt% of Irgacure 2959 (Photoinitiator) were initially polymerized under UV‐light irradiation.…”

Injectable Supramolecular Hydrogel/Microgel Composites for Therapeutic Delivery

“…Norbornene‐modified HA (NorHA) was synthesized as previously described . Briefly, HA‐TBA was coupled with 5‐norbornene‐2‐methylamine (TCI Chemicals) in the presence of BOP in anhydrous DMSO under nitrogen for 2 h at room temperature.…”