34 research outputs found
A photoreversible protein-patterning approach for guiding stem cell fate in three-dimensional gels
Although biochemically patterned hydrogels are capable of recapitulating many critical aspects of the heterogeneous cellular niche, exercising spatial and temporal control of the presentation and removal of biomolecular signalling cues in such systems has proved difficult. Here, we demonstrate a synthetic strategy that exploits two bioorthogonal photochemistries to achieve reversible immobilization of bioactive full-length proteins with good spatial and temporal control within synthetic, cell-laden biomimetic scaffolds. A photodeprotectionāoxime-ligation sequence permits user-defined quantities of proteins to be anchored within distinct subvolumes of a three-dimensional matrix, and an ortho-nitrobenzyl ester photoscission reaction facilitates subsequent protein removal. By using this approach to pattern the presentation of the extracellular matrix protein āvitronectin, we accomplished reversible differentiation of human mesenchymal stem cells to osteoblasts in a spatially defined manner. Our protein-patterning approach should provide further avenues to probe and direct changes in cell physiology in response to dynamic biochemical signalling
Dynamic biomacromolecular patterning of photoresponsive hydrogels
Polymer-based hydrogels have emerged as a unique class of biomaterials that enable stem cells to be cultured in three-dimensions within near-physiol., synthetic microenvironments. Recent strategies have been developed that permit bioepitopes (e.g., peptides, full-length proteins) to be introduced at any point in time and space to affect cell function spatiotemporally within user-defined subvolumes of the bulk material. While these techniques have been successfully utilized to direct a variety of basic cellular functions, advanced platforms that permit biol. cues to be both introduced and subsequently removed would be beneficial in recapitulating the dynamic abundance of signaling biomols. in the native, temporally-variable niche and in modulating complex cellular behavior. In this work, we demonstrate that the combination of two bioorthogonal light-based chemistries provides for the reversible immobilization of protein cues spatially within a hydrogel. The highlighted approach enables precise control over 4D biochem. functionalization of a synthetic polymer network in response to user-defined photonic stimuli. Results further highlight the versatility of such dynamic biomacromol. signal presentation in better understanding basic cell physiol
Peptide-Functionalized Click Hydrogels with Independently Tunable Mechanics and Chemical Functionality for 3D Cell Culture
Click chemistry offers highly selective and orthogonal reactions that proceed rapidly and under a variety of mild conditions with the opportunity to create highly defined and multifunctional materials. This work illustrates a strategy where step-growth networks are formed rapidly via a copper-free, azideāalkyne click chemistry between tetrafunctional poly(ethylene glycol) molecules and difunctionalized synthetic polypeptides. The molecular weight of the polymer precursors (10, 15, or 20 kDa PEG) and the stoichiometry of reactive end group functionalities (1.5:1 to 1:1.5) provide control over the material cross-linking density, enabling elastic materials with tunable moduli (<i>G</i>ā² = 1000ā6000 Pa). A sequential photochemically activated thiol-ene chemistry allows subsequent functionalization of the network through reaction with pendant alkene moieties on the peptide. Because the thiol-ene reaction is light-driven, the degree of modification is directly related to the dosage of light delivered to the system (0ā6 J cm<sup>ā2</sup>). We exploit this feature to create complex biochemical gradients of multiple peptides with well-defined magnitude and slope throughout the three-dimensional (3D) network. Since both reactions can occur in the presence of cells, this material ultimately enables independent and <i>in situ</i> tuning of biochemical and biomechanical properties of biomaterial networks, suggesting an avenue to direct cell function throughout specific regions within a 3D material
Streamlined Synthesis and Assembly of a Hybrid Sensing Architecture with Solid Binding Proteins and Click Chemistry
Combining bioorthogonal chemistry
with the use of proteins engineered
with adhesive and morphogenetic solid-binding peptides is a promising
route for synthesizing hybrid materials with the economy and efficiency
of living systems. Using optical sensing of chloramphenicol as a proof
of concept, we show here that a GFP variant engineered with zinc sulfide
and silica-binding peptides on opposite sides of its Ī²-barrel
supports the fabrication of protein-capped ZnS:Mn nanocrystals that
exhibit the combined emission signatures of organic and inorganic
fluorophores. Conjugation of a chloramphenicol-specific DNA aptamer
to the protein shell through strain-promoted azideāalkyne cycloaddition
and spontaneous concentration of the resulting nanostructures onto
SiO<sub>2</sub> particles mediated by the silica-binding sequence
enables visual detection of environmentally and clinically relevant
concentrations of chloramphenicol through analyte-mediated inner filtering
of sub-330 nm excitation light
Review: Synthetic scaffolds to control the biochemical, mechanical, and geometrical environment of stem cell-derived brain organoids
Stem cell-derived brain organoids provide a powerful platform for systematic studies of tissue functional architecture and the development of personalized therapies. Here, we review key advances at the interface of soft matter and stem cell biology on synthetic alternatives to extracellular matrices. We emphasize recent biomaterial-based strategies that have been proven advantageous towards optimizing organoid growth and controlling the geometrical, biomechanical, and biochemical properties of the organoid's three-dimensional environment. We highlight systems that have the potential to increase the translational value of region-specific brain organoid models suitable for different types of manipulations and high-throughput applications