Gel Microrods for 3D Tissue Printing

Abstract

the orientation of elongated gel fragments. The patterning of hundreds to tens of thousands of elongated microgel shapes in programmed orientations remains challenging. The various existing 3D-printing techniques, including microextrusion printing, [11] weaving, [12] inkjet printing, [13] laser-assisted printing, [14] and water-in-oil droplet printing, [15] cannot order elongated microgel shapes as repeating units.In this work, we present an innovative approach that uses a modified dropletbased microfluidics system to generate and print gelatin methacrylate (GelMa) [16,17] and Matrigel [18] microrods. Continuous-flow lithography can generate rigid elongated gel shapes but the setup is costly and challenging to integrate with 3D printing. [19] Droplet-based microfluidics has been used to fabricate rigid gel microrods, [20] but the published approaches were subject to channel clogging and unable to handle soft gel shapes, such as Matrigel microrods. Moreover, mm-length hydrogel microrods of high aspect ratio containing living cells have not been produced previously.In the present work, microrods were produced with a droplet-based microfluidics tubing (DMT) system that allowed high-throughput formation, transport and gelling of droplet plugs in polytetrafluoroethylene (PTFE) tubing. GelMa and Matrigel were gelled at 8 and 37 °C, respectively. Increased thermostability of the printed GelMa networks was achieved by photo-crosslinking. Cells in the microrods showed ≈90% viability after printing, and assumed expected morphologies. For example, human myofibroblasts cultured in GelMa rods underwent proliferation and elongation. Stromal cells in the Matrigel rods migrated to the periphery of the rods, forming an annular distribution. We patterned GelMa microrods in predefined geometries by using a micromanipulator or a rotating recipient container. Structures were stabilized by rapid fusion of the incompletely gelled rod-rod interfaces immediately after printing. GelMa and Matrigel microrods containing two or more segregated materials, Janus and ternary microrods, [21] were also produced. Finally, by using orientation and patterning, we also printed a microrod tube with luminal dimensions (≈2 cm in diameter) comparable to the human small intestine. Results Generation of GelMa MicrorodsOur DMT printing system allows the fabrication of gel microrods in microfluidics tubing and the synchronous 3D patterning Large scale 3D ordering of anisotropic gel objects, such as gel microrods, both rigid and soft, is in demand for the engineering of replica tissues but has not yet been achieved. Here, monodisperse gel microrods of gelatin methacrylate (GelMa) or Matrigel are generated by a droplet-based microfluidics tubing system. The microrods are 50-300 µm wide and 1-3 mm long; the GelMa versions are produced at up to 50 s −1 while the more fragile Matrigel versions are produced at up to 10 s −1 followed by 1 h of gelation. Upon ejection from the tubing, the rods can be printed into robust 3D structures of centimeter dimensions in w...

Microfluidics Fabrication of Soft Microtissues and Bottom‐Up Assembly

“…This renders a novel system of 3D bioprinting that is capable of patterning solid gel shapes of high aspect ratios. The new printing system is of particular interest to comprise tissues with elongated repeating units, such as the small intestine and muscle tissues . The fused interface upon contact of nonfully gelled microrods supports the structural integrity after bottom‐up patterning ( Figure 2 ).…”

“…When the microfluidics production, which includes the droplet formation, conduction, incubation, and gelation, is achieved on a single piece of plastic tubing, the droplet/rod takes the entire cross‐section of the interior of the tubing, thus avoiding oil bypassing the droplets and collision with each other, even at surfactant‐free conditions. The particle resolution can go down to 50 µm, which is suitable for producing identical microtissues, each as a culturing unit . As the droplets/rods are sequenced since production and evenly spaced, by integrating with a 3D bioprinting system, precision patterning of the structures can be achieved, individual rods being radially or parallelly aligned.…”

Cell Encapsulation in Soft, Anisometric Poly(ethylene) Glycol Microgels Using a Novel Radical‐Free Microfluidic System

“…These biocompatibility issues have been overcome by producing anisometric microgels from naturally occurring precursors, such as Matrigel and gelatin, which can be thermally crosslinked . Different cell types have been encapsulated, and shown to remain viable and proliferate for up to 10 days.…”

Compartmentalized Jet Polymerization as a High‐Resolution Process to Continuously Produce Anisometric Microgel Rods with Adjustable Size and Stiffness

“…In the case of drug delivery, particle size and shape have a substantial impact on cellular uptake, circulation in the blood, and distribution in biological systems . For tissue engineering, bottom‐up approaches use particle self‐assembly and self‐organization to fabricate and organize microgels into 3D macroscopic scaffolds . Recently, our research group introduced the Anisogel that induces mechanical and structural anisometry to guide cells and regenerating nerves in a unidirectional manner .…”

“…Here, (pre‐)polymer containing elongated plugs are generated by operating T‐junctions, cross‐junctions, or flow‐focusing devices in the dripping or plug flow regime The precursor solution inside these microfluidic plugs can be polymerized rapidly on chip via photoinitiation, resulting in covalently crosslinked disk or rod‐shaped gels and particles . Alternatively, noncovalent crosslinking can be triggered by calcium or temperature in the case of alginates or gelatin, respectively, or two‐component covalent Michael‐type addition can be employed to produce cell‐loaded soft microgel rods . Furthermore, microfluidic plug flow was utilized to obtain plug microreactors, in which solid anisometric nanocrystals were synthesized .…”