Highly Elastic Biodegradable Single-Network Hydrogel for Cell Printing

Abstract

Cell printing is becoming a common technique to fabricate cellularized printed scaffold for biomedical application. There are still significant challenges in soft tissue bioprinting using hydrogels, which requires live cells inside the hydrogels. Moreover, the resilient mechanical properties from hydrogels are also required to mechanically mimic the native soft tissues. Herein, we developed a visible-light cross-linked, single-network, biodegradable hydrogel with high elasticity and flexibility for cell printing, which is different from previous highly elastic hydrogel with double-network and two components. The single-network hydrogel using only one stimulus (visible light) to trigger gelation can greatly simplify the cell printing process. The obtained hydrogels possessed high elasticity, and their mechanical properties can be tuned to match various native soft tissues. The hydrogels had good cell compatibility to support fibroblast growth in vitro. Various human cells were bioprinted with the hydrogels to form cell-gel constructs, in which the cells exhibited high viability after 7 days of culture. Complex patterns were printed by the hydrogels, suggesting the hydrogel feasibility for cell printing. We believe that this highly elastic, single-network hydrogel can be simply printed with different cell types, and it may provide a new material platform and a new way of thinking for hydrogel-based bioprinting research.

Keywords: biodegradable hydrogel; cell printing; elasticity; single network; tissue regeneration.

Figure 1

Preparation and characterization of highly elastic, visible-light cross-linked, single-network, biodegradable hydrogel for cell printing. An acrylated PCL–PEG–PCL triblock polymer was synthesized and then cross-linked using visible light to form a highly elastic single-network biodegradable hydrogel. The hydrogel has attractive mechanical properties, and it is stretchable, compressible, and twistable. The hydrogel can also be bioprinted with various human cells and form complex patterns upon visible-light exposure.

Figure 2

¹H NMR spectra of (A) PEG-DA and (B) PEG–PCL(24k)-DA.

Figure 3

Photographs demonstrating the attractive mechanical properties of the PEG–PCL(24k)-DA hydrogel under stretching, compression, and twisting. (A) PEG-DA hydrogel was broken under stretching. (B) PEG–PCL(24k)-DA hydrogel could be stretched and recoiled back to the original length. (C) PEG-DA hydrogel was broken into pieces under compression at 80% strain. (D) PEG–PCL(24k)-DA hydrogel deformed and recovered under compression. (E) PEG-DA hydrogel was broken after twisting for four cycles. (F) PEG–PCL(24k)-DA hydrogel could be twisted for four cycles and recovered after releasing.

Figure 4

Mechanical properties of the PEG–PCL(24k)-DA hydrogel. (A) Compressive stress–strain curves of PEG–PCL-DA hydrogels. (B) Tensile stress–strain curves of PEG–PCL-DA hydrogels. (C) Cyclic stretching of PEG–PCL-DA hydrogels at 100% deformation for 10 cycles.

Figure 5

In vitro cytocompatibility of elastic hydrogels. (A) Live and dead stained 3T3 fibroblasts encapsulated in PEG-DA and PEG–PCL(24k)-DA hydrogels after 1 day and 3 days of culture. (B) Cell viability calculated as percentage of live cells (green) from the live and dead staining images. (C) Metabolic index of 3T3 fibroblasts encapsulated in PEG-DA and PEG–PCL(24k)-DA hydrogels.

Figure 6

Cytotoxicity and printability of elastic PEG–PCL(24k)-DA hydrogel using an extrusion bioprinter. (A) Viscosity curve of elastic PEG–PCL(24k)-DA precursor solution. (B, C) Cell viability of different cell types in printed 10% elastic PEG–PCL(24k)-DA hydrogel. Live/dead assay was performed immediately after gel polymerization and after 7 days in culture (scale bars represent 500 μm). (D) Effect of different needle sizes and precursor solution concentrations on viability of neonatal human lung fibroblasts. (E) Effect of shear stress on cell viability evaluated immediately after printing. (F, G) Sample shapes printed using a printer with different needle sizes (scale bars in (F) represent 2 mm and scale bars in (G) represent 5 mm).