Multi-responsive hydrogel structures from patterned droplet networks

Downs, Florence G.; Lunn, David J.; Booth, Michael J.; Sauer, Joshua B.; Ramsay, William J.; Klemperer, R. George; Hawker, Craig J.; Bayley, Hagan

doi:10.1038/s41557-020-0444-1

Cited by 177 publications

(154 citation statements)

References 33 publications

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“…Recent advances in functionalized, tissue-like compartments could facilitate the development of such stimuli responsive technologies. [22][23][24] However, significant challenges remain in the development of such therapies, including the transcription and translation of active enzymes and localization to the central nervous system. Furthermore, problems with the instability of the enzymes must be overcome.…”

Section: (4 Of 5)mentioning

confidence: 99%

Cell‐Free Synthesis of Dopamine and Serotonin in Two Steps with Purified Enzymes

et al. 2020

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For example, the intermediates of the synthesis of dopamine and serotonin are L-DOPA (L-dihydroxy-phenylalanine) and 5-HTP (5-hydroxytryptophan), respectively, and have found use as administered precursors that are capable of crossing the blood-brain barrier. Although several studies have focused on the characterization of the separate enzymes involved in the biosynthesis of serotonin and dopamine, [3-5] surprisingly little effort has been put into developing enzymatic methods for the production of the final neurotransmitters starting from their proteinogenic amino acids precursors, i.e., L-tryptophan (Trp) and L-tyrosine (Tyr). If developed, such a system may open up new opportunities to build nanofactories and artificial cells for the treatment of neurological disorders. [6-8] To produce the monoamine neurotransmitters serotonin and dopamine in vitro, hydroxylases specific to the amino acid substrate were exploited. The catalytic domain of human tryptophan hydroxylase isoform 2 (TPH) was chosen for the synthesis of 5-HTP from Trp, [9] because this enzyme was previously shown to not be inhibited by high concentrations of the substrate. [10] Two versions of the catalytic domain of rat tyrosine hydroxylase were tested for the production of L-DOPA, including a recombinant, wild type version (rTH) and a truncated construct (ΔTH) that was not inhibited by substrate. [11] Additionally, a hydroxylase from the bacterium Chlamydia pneumoniae, Cpn1046, was tested. Cpn1046 has broader substrate specificity compared to TH and is homologous to eukaryotic aromatic amino acid hydroxylases. [12] Since the aromatic amino acid decarboxylase from Drosophila melanogaster (AADC) is active on both 5-HTP and L-DOPA, [13] a single enzyme was used for the decarboxylation step for the synthesis of both serotonin and dopamine. We found that TPH and ΔTH completely hydroxylated Trp and Tyr to produce the intermediates 5-HTP and L-DOPA, respectively. When TPH and ΔTH were coupled with AADC, serotonin and dopamine were efficiently produced in vitro. Each enzyme (TPH, rTH, ΔΤΗ, AADC, and Cpn1046) expressed well when fused to maltose binding protein (MBP), and each protein was purified in a single step with amylose resin (Figure S1, Supporting Information). However, multiple bands were observed on a SDS-PAGE of Cpn1046. Typically, 50 mg of purified protein was obtained per liter of bacterial culture expressing each construct. Conversely, the use of The synthesis of serotonin and dopamine with purified enzymes is described. Both pathways start from an amino acid substrate and synthesize the monoamine neurotransmitter in two enzymatic steps. The enzymes human tryptophan hydroxylase isoform 2, Rattus norvegicus tyrosine hydroxylase, Chlamydia pneumoniae Cpn1046, and aromatic amino acid decarboxylase from Drosophila melanogaster are recombinantly expressed, purified, and shown to be functional in vitro. The hydroxylases efficiently convert L-DOPA (L-dihydroxy-phenylalanine) and 5-HTP (5-hydroxytryptophan) from L-tyrosine and L-tryptophan, respectiv...

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Section: (4 Of 5)mentioning

confidence: 99%

Cell‐Free Synthesis of Dopamine and Serotonin in Two Steps with Purified Enzymes

et al. 2020

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“…[32] However, this technique is limited to the fabrication of thin sheets. More complex 3D structures can be achieved through patterned droplet networks [33] or jammed microgels, for example, using additive manufacturing techniques. [34][35][36][37] However, monodisperse spherical microgels have a small contact area such that the resulting superstructures are weak.…”

Section: Introductionmentioning

confidence: 99%

3D Printing of Strong and Tough Double Network Granular Hydrogels

Hirsch

Charlet

Amstad

2020

Adv Funct Materials

103

127

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Many soft natural tissues display a fascinating set of mechanical properties that remains unmatched by manmade counterparts. These unprecedented mechanical properties are achieved through an intricate interplay between the structure and locally varying the composition of these natural tissues. This level of control cannot be achieved in soft synthetic materials. To address this shortcoming, a novel 3D printing approach to fabricate strong and tough soft materials is introduced, namely double network granular hydrogels (DNGHs) made from compartmentalized reagents. This is achieved with an ink composed of microgels that are swollen in a monomer‐containing solution; after the ink is additive manufactured, these monomers are converted into a percolating network, resulting in a DNGH. These DNGHs are sufficiently stiff to repetitively support tensile loads up to 1.3 MPa. Moreover, they are more than an order of magnitude tougher than each of the pure polymeric networks they are made from. It is demonstrated that this ink enables printing macroscopic, strong, and tough objects, which can optionally be rendered responsive, with high shape fidelity. The modular and robust fabrication of DNGHs opens up new possibilities to design adaptive, strong, and tough hydrogels that have the potential to advance, for example, soft robotic applications.

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“…[ 21,22 ] Different droplet contents have been explored to produce droplet network derived synthetic and living tissues, with overall dimensions of hundreds of micrometers to millimeters. [ 22–26 ] Here, we report a modified droplet‐based method that uses molds to support large and hollow droplet networks for the production of collagen tubes with centimeter dimensions. Droplets, containing cells and collagen solution, generated in lipid‐containing oil, spontaneously acquired a monolayer of lipids ( Figure a).…”

Section: Resultsmentioning

confidence: 99%

Bioengineered Gastrointestinal Tissues with Fibroblast‐Induced Shapes

Zhou

Ruiz‐Puig

Jacobs

et al. 2020

Adv Funct Materials

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Bioengineered gastrointestinal (GI) tracts have potential applications in regenerative medicine and disease modeling. Methods for engineering tubular GI tracts containing natural extracellular matrix (ECM) are currently limited. Here, the fabrication of collagen tubes with designed shapes by using lipid bilayer supported droplet networks is reported. Droplets containing cells and collagen are arrayed in lipid‐containing oil to form droplet networks, which undergo thermal gelation to provide continuous collagen tubes. A variety of tubular GI tissues are fabricated. For example, human intestinal organoids embedded in the collagen tubes migrate to the luminal surfaces and fuse to form a continuous epithelial layer, mimicking aspects of intestinal tissue structure. Fibroblasts embedded in the collagen induce a cell density dependent contraction of the tubes. Complex tubular structures are produced by patterning droplets containing different densities of fibroblasts. The fibroblast‐containing collagen tubes are seeded with various epithelial cells at their luminal surfaces to form gastric and colonic tissues, which comprise monolayers or multilayers of epithelial cells and fibroblast‐containing subepithelial layers. The engineered gastric tissues are susceptible to infection with Helicobacter pylori. The versatile technique allows the construction of tubular GI tracts containing ECM and layered structures, with broad potential applications in disease research and regenerative medicine.

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Multi-responsive hydrogel structures from patterned droplet networks

Cited by 177 publications

References 33 publications

Cell‐Free Synthesis of Dopamine and Serotonin in Two Steps with Purified Enzymes

Cell‐Free Synthesis of Dopamine and Serotonin in Two Steps with Purified Enzymes

3D Printing of Strong and Tough Double Network Granular Hydrogels

Bioengineered Gastrointestinal Tissues with Fibroblast‐Induced Shapes

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