3D bioprinted complex constructs reinforced by hybrid multilayers of electrospun nanofiber sheets

Abstract

Despite the usefulness of hydrogels for cell-based bioprinting, the fragility of their resulting constructs has hindered their practical applications in tissue engineering research. Here, we suggest a hybrid integration method based on cell-hydrogel bioprinting that includes alternate layering of flexible nanofiber (NF) sheets. Because the bioprinting was implemented on a nanofibrous surface, the hydrogel-based materials could be printed with enhanced shape resolution compared to printing on a bare hydrogel. Furthermore, the insertion of NF sheets was effective for alleviating the shrinkage distortion of the hydrogel construct, which is inherently generated during the crosslinking process, thereby enhancing shape fidelity throughout the three-dimensional (3D) architecture. In addition to the structural precision, the NF-embedded constructs improved the mechanical properties in terms of compressive strength, modulus, and resilience limit (up to four-fold enhancement). With structural and mechanical supports, we could 3D fabricate complex constructs, including fully opened internal channels, which provided a favorable perfusion condition for cell growth. We confirmed the enhanced bioactivity of the NF-embedded bioprinted construct via cell culture experiments with 80% enhanced proliferation compared to the monolithic one. The synergistic combination of the two flexible materials, NFs and hydrogels, is expected to have extensive applicability in soft tissue engineering.

Hydrogels for Bioprinting: A Systematic Review of Hydrogels Synthesis, Bioprinting Parameters, and Bioprinted Structures Behavior

“…Chemical cross-linking is mostly done to alginate (52 papers) with CaCl 2 solution (46 papers), and detailed concentrations can be seen in Figure 6 . Other (non-Ca 2+ ) solutions are used in two papers (Hafeez et al, 2018 ; Aydogdu et al, 2019 ) which use hydrazine and NaOH, respectively, and other two studies do not provide information (Aljohani et al, 2018 ; Yoon et al, 2019 ). Other specific cross-linking agents are genipin (Akkineni et al, 2016 ; Kim et al, 2016 ), mTgase (Tijore et al, 2018b ) or 1-ethyl-3-(3- dimethylaminopropyl)-carbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS) (Choi et al, 2018 ), and MAL-PEG-MAL (Yan et al, 2018 ) for gelatin, or different solute for other materials like thrombin (Zhang K. et al, 2016 ), mushroom tyrosinase (Das et al, 2015 ), and oxidative reactions (Shin et al, 2018 ).…”

“…Additionally, NIH3T3 is used in 15 papers where some analyze percentage of cell viability (7) (Bertassoni et al, 2014 ; Ersumo et al, 2016 ; Liu et al, 2018 ; Shin et al, 2018 ; Wang et al, 2018 ; Zhang X. et al, 2018 ; Krishnamoorthy et al, 2019 ), others report a qualitative scale of cell viability (5) (Ahn et al, 2017 ; Cochis et al, 2018 ; Raddatz et al, 2018 ; Xu C. et al, 2018 ; Yoon et al, 2019 ), and some do not perform any kind of test (3). These cell types are a fibroblast cell line used mostly because of its ease of growth.…”

“…This parameter can be considered an important factor for hydrogel printability. As it is known, viscosity indicates fluidity Blaeser et al, 2013;Bakarich et al, 2014;Narayanan et al, 2016;Freeman and Kelly, 2017;Gao et al, 2017;Li et al, 2017;Schütz et al, 2017;Yang et al, 2017;Ahlfeld et al, 2018;Habib et al, 2018;Hafeez et al, 2018;He et al, 2018;López-Marcial et al, 2018;Maiullari et al, 2018;Naghieh et al, 2018;Gonzalez-Fernandez et al, 2019;Kosik-Kozioł et al, 2019• Jia et al, 2014Markstedt et al, 2015;Nguyen et al, 2017;Aljohani et al, 2018;Datta et al, 2018;Ooi et al, 2018;Yu et al, 2018;Ji et al, 2019;Yoon et al, 2019• Zhang K. et al, 2016Aied et al, 2018;Choi et al, 2018;Li et al, 2018a,c;Shin et al, 2018;Tijore et al, 2018a and for this reason it is very important for hydrogel extrusion. So, the more the viscous, the more the inner pressure of hydrogel during the extrusion process and increased cell damage.…”

Recent progress in extrusion 3D bioprinting of hydrogel biomaterials for tissue regeneration: a comprehensive review with focus on advanced fabrication techniques

“…To overcome these issues, and also to mimic ECM matrix, EBB technique has been consolidated with electrospinning to develop scaffolds possessing advantages of different kinds of materials only in one construction [64][65][66][67]. In other words, joining 3D printing and electrospinning can make their particular advantages complementary and improve the capability of developing functional biomimetic scaffolds [68][69][70].…”

Multi-layer Scaffolds of Poly(caprolactone), Poly(glycerol sebacate) and Bioactive Glasses Manufactured by Combined 3D Printing and Electrospinning

“…It was instead necessary to partially melt one surface of the 3D scaffold and press it into the electrospun mat, with consequent changes in the morphology and porosity of this latter layer. In a recent work, PCL electrospun sheets have been treated with oxygen plasma, before being inserted in between cross-linked 3D-printed alginate layers [27]. The plasma treatment made the PCL fibres hydrophilic and prevented layer delamination.…”