How Much Physical Guidance is Needed to Orient Growing Axons in 3D Hydrogels?

Abstract

Directing cells is essential to organize multi-cellular organisms that are built up from subunits executing specific tasks. This guidance requires a precisely controlled symphony of biochemical, mechanical, and structural signals. While many guiding mechanisms focus on 2D structural patterns or 3D biochemical gradients, injectable material platforms that elucidate how cellular processes are triggered by defined 3D physical guiding cues are still lacking but crucial for the repair of soft tissues. Herein, a recently developed anisotropic injectable hybrid hydrogel (Anisogel) contains rod-shaped microgels that orient in situ by a magnetic field and has propelled studying 3D cell guidance. Here, the Anisogel is used to investigate the dependence of axonal guidance on microgel dimensions, aspect ratio, and distance. While large microgels result in high material anisotropy, they significantly reduce neurite outgrowth and thus the guidance efficiency. Narrow and long microgels enable strong axonal guidance with maximal outgrowth including cell sensing over distances of tens of micrometers in 3D. Moreover, nerve cells decide to orient inside the Anisogel within the first three days, followed by strengthening of the alignment, which goes along with oriented fibronectin deposition. These findings demonstrate the potential of the Anisogel to tune structural and mechanical parameters for specific applications.

Bicyclic RGD peptides enhance nerve growth in synthetic PEG-based Anisogels

“…We define that nerves are highly aligned when the FWHM < 90°. 56 These observations are consistent with our previous report on fully PEG-based Anisogels tethered with fibronectin fragment. 20 …”

Controlling Structure with Injectable Biomaterials to Better Mimic Tissue Heterogeneity and Anisotropy

“…Owing to their high aspect ratio, the concentration of SPIONs required to align the microelements is very small, resulting in an overall SPION concentration of merely 1.8 ng µL −1 or 0.0046 vol% in the entire Anisogel when microgels occupy 0.45 vol%. [79] Neurons and fibroblasts cultured in Anisogels oriented in the direction of the aligned rod-shaped microgels or short fibers. [15] While the Strategies to obtain in vivo alignment.…”

“…Reproduced with permission. [79] Copyright 2020, John Wiley and Sons. microgels were fabricated from bioinert PEG using an in mold polymerization method, the short fibers were prepared from poly (lactide-co-glycolide) via an electrospinning/microcutting technique.…”

“…These 2.5 × 2.5 × 25 µm microgels only took up 0.45 vol% of the entire Anisogel (Figure 3H). [79] Besides the microgels, short fibers were produced and aligned inside the Anisogel resulting in spontaneous electrical activity of the aligned axons with calcium signals propagating along the anisotropy axis of the material. Short fibers have the advantage that their surface nanotopography can be adjusted between smooth, porous, and grooved, leading to higher neurite alignment in the case of nanogrooves [102] on the fibers and enhanced morphological aspect ratios and nuclear YAP shuttling in the case of fibroblasts.…”

“…Copyright 2018, 2014 Elsevier. Reproduced with permission [79]. Copyright 2020, John Wiley and Sons.…”