Mechanical stretching of 3D hydrogels for neural stem cell differentiation

Abstract

While it is known that mechanical dynamics are influential in neural differentiation for critical processes like neurogenesis or neurodegeneration, studies on neural stem cell therapies usually focus on biochemical interactions rather than mechanical aspects, frequently resulting in low efficacy and unfulfilled potential. Therefore, current studies are attempting to elucidate the effect of mechanical stimulus on neural performance using conventional two-dimensional (2D) planar substrates. Yet, these 2D substrates fail to capture the defining three-dimensional (3D) characteristics of the in vivo neural stem cell environment.To complete this research gap, we synthesized a series of soft and elastic 3D hydrogels to mimic the neural tissue mechanical environment for 3D cell culture, using long-chain polyethylene glycol diacrylate (PEGDA) and gelatin-methacryloyl (GelMA). By varying the concentration of the polymer, we obtained biomimicking hydrogels with a tensile modulus as low as 10 kPa and a compressive modulus as low as 0.8 kPa. The in vitro results demonstrated that GelMA-PEGDA hydrogels have the high biocompatibility required to support neural cell growth, proliferation, and differentiation, as well as neurite outgrowth. We then studied the effect of mechanical stretching on the behaviors of neural cells and observed that mechanical stretching could significantly enhance neurite extension and axon elongation. In addition, the neurites were more directionally oriented to the stretching direction. Immunocytochemistry and relative gene expression data also suggested that mechanical tension could upregulate the expression of neural differentiation protein and genes, including GFAP and βIII-Tubulin. Overall, this study shows that in addition to the specific mechanical properties of GelMA-PEGDA that improve neural differentiation towards specific lineages, hydrogel stretching is also a potentially attractive strategy to improve the therapeutic outcomes of neural stem cell therapies.

Swelling Behavior of Novel Hydrogels Produced from Glucose-Based Ionic Monomers with Varying Cross-Linkers

“…Besides acrylamides, the second type of cross-linker studied were ethylene glycol diacrylates with different chain lengths. PEGDA and EGDMA (Figure ) are well-known in the literature as cross-linking agents for a number of hydrogels of different origins and show high biocompatibility in the 3D cross-linked state. − This is of major importance for subsequent applications in the fields of medicine or biotechnology. − …”

Advancing Peripheral Nerve Regeneration: 3D Bioprinting of GelMA-Based Cell-Laden Electroactive Bioinks for Nerve Conduits

“…Nevertheless, after a 5-day culture period, it was noted that the PEGDA-based compositions exhibited a slight reduction in cell proliferation. This outcome may be linked to the elevated ratio of PEGDA, which lacks crucial cell-attaching ligands (RGD peptides) necessary for facilitating effective cell adhesion and proliferation . Additionally, the morphology, cell-to-cell attachment, and cytoskeletal rearrangement of N2a cells over 5 days were investigated using fluorescence microscopy, as shown in Figure .…”

“…This outcome may be linked to the elevated ratio of PEGDA, which lacks crucial cell-attaching ligands (RGD peptides) necessary for facilitating effective cell adhesion and proliferation. 128 Additionally, the morphology, cell-to-cell attachment, and cytoskeletal rearrangement of N2a cells over 5 days were investigated using fluorescence microscopy, as shown in Figure 10. These observations were utilized to validate cell proliferation via quantitative assessment of cell density per unit area using the Fiji software package (NIH) and are shown in Figure S7.…”

Biofabrication of Composite Tendon Constructs with the Fibrous Arrangement, High Cell Density, and Enhanced Cell Alignment

“…Meanwhile, CTCs in groups 3 and 4 exhibited significantly enhanced protein expression after 3 days of cyclic stretching culture (Figure ). In many studies, mechanical stimulation has been shown to induce specific differentiation of primary cells. , For example, Czaplewski et al conducted the periodic tensile test on PLA-/PCL-woven scaffolds, which demonstrated that periodic stretching significantly upregulated the expression of tendon markers and inhibited osteoblastic differentiation . Chen et al performed cyclic stress experiments on woven silk/collagen scaffolds and found that MSCs derived from human embryonic stem cells expressed tendon-related proteins and genes under mechanical stimulation .…”