Fully synthetic hydrogels promote robust crypt formation in intestinal organoids.

Initial landmark studies in the design of synthetic hydrogels for intestinal organoid culture identified precise matrix requirements for differentiation, namely decompression of matrix-imposed forces and supplementation of laminin. But beyond stating the necessity of laminin, organoid-laminin interactions have gone largely unstudied, as this ubiquitous requirement of exogenous laminin hinders investigation. In this work, we exploit a fast stress relaxing, boronate ester based synthetic hydrogel for the culture of intestinal organoids, and fortuitously discover that unlike all other synthetic hydrogels to date, laminin does not need to be supplemented for crypt formation.

Fibrillar adhesion dynamics govern the timescales of nuclear mechano-response via the vimentin cytoskeleton.

The cell nucleus is continuously exposed to external signals, of both chemical and mechanical nature. To ensure proper cellular response, cells need to regulate not only the transmission of these signals, but also their timing and duration. Such timescale regulation is well described for fluctuating chemical signals, but if and how it applies to mechanical signals reaching the nucleus is still unknown.

Supramolecular Linear-Dendritic Nanoreactors: Synthesis and Catalytic Activity in "Green" Suzuki-Miyaura Reactions.

This study describes the synthesis of novel amphiphilic linear-dendritic block copolymers and their self-assembly in water to form supramolecular nanoreactors capable of catalyzing Suzuki-Miyaura coupling reactions under "green" conditions. The block copolymers were formed through copper(I)-catalyzed alkyne-azide cycloaddition between azide functionalized poly(benzyl ether) dendrons as the perfectly branched blocks, as well as bis-alkyne modified poly(ethylene glycol), PEG, as the linear block. A first-generation poly(benzyl ether) dendron (G1) was coupled to a bis-alkyne modified PEG with molecular mass of 5 kDa, forming an ABA copolymer (G1)-PEG5k-(G1) (yield 62%), while a second-generation dendron (G2) was coupled to a 11 kDa bis-alkyne modified PEG to produce (G2)-PEG11k-(G2) (yield 49%).

Quantifying stiffness and forces of tumor colonies and embryos using a magnetic microrobot.

Stiffness and forces are two fundamental quantities essential to living cells and tissues. However, it has been a challenge to quantify both 3D traction forces and stiffness (or modulus) using the same probe in vivo. Here, we describe an approach that overcomes this challenge by creating a magnetic microrobot probe with controllable functionality.

In situ modulation of intestinal organoid epithelial curvature through photoinduced viscoelasticity directs crypt morphogenesis.

Spatiotemporally coordinated transformations in epithelial curvature are necessary to generate crypt-villus structures during intestinal development. However, the temporal regulation of mechanotransduction pathways that drive crypt morphogenesis remains understudied. Intestinal organoids have proven useful to study crypt morphogenesis in vitro, yet the reliance on static culture scaffolds limits the ability to assess the temporal effects of changing curvature.

Programming hydrogels to probe spatiotemporal cell biology.

The recapitulation of complex microenvironments that regulate cell behavior during development, disease, and wound healing is key to understanding fundamental biological processes. In vitro, multicellular morphogenesis, organoid maturation, and disease modeling have traditionally been studied using either non-physiological 2D substrates or 3D biological matrices, neither of which replicate the spatiotemporal biochemical and biophysical complexity of biology. Here, we provide a guided overview of the recent advances in the programming of synthetic hydrogels that offer precise control over the spatiotemporal properties within cellular microenvironments, such as advances in the control of cell-driven remodeling, bioprinting, or user-defined manipulation of properties (e.

4D Materials with Photoadaptable Properties Instruct and Enhance Intestinal Organoid Development.

Intestinal organoids are self-organized tissue constructs, grown , that resemble the structure and function of the intestine and are often considered promising as a prospective platform for drug testing and disease modeling. Organoid development is typically instructed by exogenous cues delivered from the media, but cellular responses also depend on properties of the surrounding microenvironmental niche, such as mechanical stiffness and extracellular matrix (ECM) ligands. In recent years, synthetic hydrogel platforms have been engineered to resemble the niche, with the goal of generating physiologically relevant environments that can promote mature and reproducible organoid development.

In Situ Super-Resolution Imaging of Organoids and Extracellular Matrix Interactions via Phototransfer by Allyl Sulfide Exchange-Expansion Microscopy (PhASE-ExM).

3D organoid models have recently seen a boom in popularity, as they can better recapitulate the complexity of multicellular organs compared to other in vitro culture systems. However, organoids are difficult to image because of the limited penetration depth of high-resolution microscopes and depth-dependent light attenuation, which can limit the understanding of signal transduction pathways and characterization of intimate cell-extracellular matrix (ECM) interactions. To overcome these challenges, phototransfer by allyl sulfide exchange-expansion microscopy (PhASE-ExM) is developed, enabling optical clearance and super-resolution imaging of organoids and their ECM in 3D.

Tissue geometry drives deterministic organoid patterning.

Epithelial organoids are stem cell–derived tissues that approximate aspects of real organs, and thus they have potential as powerful tools in basic and translational research. By definition, they self-organize, but the structures formed are often heterogeneous and irreproducible, which limits their use in the lab and clinic. We describe methodologies for spatially and temporally controlling organoid formation, thereby rendering a stochastic process more deterministic.

3D printing of sacrificial thioester elastomers using digital light processing for templating 3D organoid structures in soft biomatrices.

Biofabrication allows for the templating of structural features in materials on cellularly-relevant size scales, enabling the generation of tissue-like structures with controlled form and function. This is particularly relevant for growing organoids, where the application of biochemical and biomechanical stimuli can be used to guide the assembly and differentiation of stem cells and form architectures similar to the parent tissue or organ. Recently, ablative laser-scanning techniques was used to create 3D overhang features in collagen hydrogels at size scales of 10-100m and supported the crypt-villus architecture in intestinal organoids.

Secreted Factors From Proinflammatory Macrophages Promote an Osteoblast-Like Phenotype in Valvular Interstitial Cells.

Objective: Resident valvular interstitial cells (VICs) activate to myofibroblasts during aortic valve stenosis progression, which further promotes fibrosis or even differentiate into osteoblast-like cells that can lead to calcification of valve tissue. Inflammation is a hallmark of aortic valve stenosis, so we aimed to determine proinflammatory cytokines secreted from M1 macrophages that give rise to a transient VIC phenotype that leads to calcification of valve tissue. Approach and Results: We designed hydrogel biomaterials as valve extracellular matrix mimics enabling the culture of VICs in either their quiescent fibroblast or activated myofibroblast phenotype in response to the local matrix stiffness.

The Effect of Thiol Structure on Allyl Sulfide Photodegradable Hydrogels and their Application as a Degradable Scaffold for Organoid Passaging.

Intestinal organoids are useful in vitro models for basic and translational studies aimed at understanding and treating disease. However, their routine culture relies on animal-derived matrices that limit translation to clinical applications. In fact, there are few fully defined, synthetic hydrogel systems that allow for the expansion of intestinal organoids.

Relaxation of Extracellular Matrix Forces Directs Crypt Formation and Architecture in Intestinal Organoids.

Intestinal organoid protocols rely on the use of extracellular scaffolds, typically Matrigel, and upon switching from growth to differentiation promoting media, a symmetry breaking event takes place. During this stage, the first bud like structures analogous to crypts protrude from the central body and differentiation ensues. While organoids provide unparalleled architectural and functional complexity, this sophistication is also responsible for the high variability and lack of reproducibility of uniform crypt-villus structures.

Secondary Photocrosslinking of Click Hydrogels To Probe Myoblast Mechanotransduction in Three Dimensions.

Muscle cells sense the mechanical properties of their microenvironment, and these properties can change in response to injury or disease. Hydrogels with dynamic material properties can be used to study the effect of such varying mechanical signals. Here, we report the ability of azadibenzocyclooctyne to undergo a cytocompatible, photoinitiated crosslinking reaction.