Correction: Actuated 3D microgels for single cell mechanobiology.

Correction for 'Actuated 3D microgels for single cell mechanobiology' by Berna Özkale , , 2022, , 1962-1970, https://doi.org/10.1039/D2LC00203E.

Actuated 3D microgels for single cell mechanobiology.

We present a new cell culture technology for large-scale mechanobiology studies capable of generating and applying optically controlled uniform compression on single cells in 3D. Mesenchymal stem cells (MSCs) are individually encapsulated inside an optically triggered nanoactuator-alginate hybrid biomaterial using microfluidics, and the encapsulating network isotropically compresses the cell upon activation by light. The favorable biomolecular properties of alginate allow cell culture up to a week.

RNA-responsive elements for eukaryotic translational control.

The ability to control translation of endogenous or exogenous RNAs in eukaryotic cells would facilitate a variety of biotechnological applications. Current strategies are limited by low fold changes in transgene output and the size of trigger RNAs (trRNAs). Here we introduce eukaryotic toehold switches (eToeholds) as modular riboregulators.

Minimally instrumented SHERLOCK (miSHERLOCK) for CRISPR-based point-of-care diagnosis of SARS-CoV-2 and emerging variants.

The COVID-19 pandemic highlights the need for diagnostics that can be rapidly adapted and deployed in a variety of settings. Several SARS-CoV-2 variants have shown worrisome effects on vaccine and treatment efficacy, but no current point-of-care (POC) testing modality allows their specific identification. We have developed miSHERLOCK, a low-cost, CRISPR-based POC diagnostic platform that takes unprocessed patient saliva; extracts, purifies, and concentrates viral RNA; performs amplification and detection reactions; and provides fluorescent visual output with only three user actions and 1 hour from sample input to answer out.

Generation of the Compression-induced Dedifferentiated Adipocytes (CiDAs) Using Hypertonic Medium.

Current methods to obtain mesenchymal stem cells (MSCs) involve sampling, culturing, and expanding of primary MSCs from adipose, bone marrow, and umbilical cord tissues. However, the drawbacks are the limited numbers of total cells in MSC pools, and their decaying stemness during expansion. As an alternative resource, recent ceiling culture methods allow the generation of dedifferentiated fat cells (DFATs) from mature adipocytes.

Creating CRISPR-responsive smart materials for diagnostics and programmable cargo release.

Materials that sense and respond to biological signals in their environment have a broad range of potential applications in drug delivery, medical devices and diagnostics. Nucleic acids are important biological cues that encode information about organismal identity and clinically relevant phenotypes such as drug resistance. We recently developed a strategy to design nucleic acid-responsive materials using the CRISPR-associated nuclease Cas12a as a user-programmable sensor and material actuator.

Compression-induced dedifferentiation of adipocytes promotes tumor progression.

Dysregulated physical stresses are generated during tumorigenesis that affect the surrounding compliant tissues including adipocytes. However, the effect of physical stressors on the behavior of adipocytes and their cross-talk with tumor cells remain elusive. Here, we demonstrate that compression of cells, resulting from various types of physical stresses, can induce dedifferentiation of adipocytes via mechanically activating Wnt/β-catenin signaling.

A biomaterial-based vaccine eliciting durable tumour-specific responses against acute myeloid leukaemia.

Acute myeloid leukaemia (AML) is a malignancy of haematopoietic origin that has limited therapeutic options. The standard-of-care cytoreductive chemotherapy depletes AML cells to induce remission, but is infrequently curative. An immunosuppressive AML microenvironment in the bone marrow and the paucity of suitable immunotherapy targets limit the induction of effective immune responses.

Programmable CRISPR-responsive smart materials.

Stimuli-responsive materials activated by biological signals play an increasingly important role in biotechnology applications. We exploit the programmability of CRISPR-associated nucleases to actuate hydrogels containing DNA as a structural element or as an anchor for pendant groups. After activation by guide RNA-defined inputs, Cas12a cleaves DNA in the gels, thereby converting biological information into changes in material properties.

Programmable microencapsulation for enhanced mesenchymal stem cell persistence and immunomodulation.

Mesenchymal stem cell (MSC) therapies demonstrate particular promise in ameliorating diseases of immune dysregulation but are hampered by short in vivo cell persistence and inconsistencies in phenotype. Here, we demonstrate that biomaterial encapsulation into alginate using a microfluidic device could substantially increase in vivo MSC persistence after intravenous (i.v.

An injectable bone marrow-like scaffold enhances T cell immunity after hematopoietic stem cell transplantation.

Allogeneic hematopoietic stem cell transplantation (HSCT) is a curative treatment for multiple disorders, but deficiency and dysregulation of T cells limit its utility. Here we report a biomaterial-based scaffold that mimics features of T cell lymphopoiesis in the bone marrow. The bone marrow cryogel (BMC) releases bone morphogenetic protein-2 to recruit stromal cells and presents the Notch ligand Delta-like ligand-4 to facilitate T cell lineage specification of mouse and human hematopoietic progenitor cells.

Microfluidic Templated Multicompartment Microgels for 3D Encapsulation and Pairing of Single Cells.

Controlled encapsulation and pairing of single cells within a confined 3D matrix can enable the replication of the highly ordered cellular structure of human tissues. Microgels with independently controlled compartments that can encapsulate cells within separately confined hydrogel matrices would provide precise control over the route of pairing single cells. Here, a one-step microfluidic method is presented to generate monodisperse multicompartment microgels that can be used as a 3D matrix to pair single cells in a highly biocompatible manner.

Controlled self-assembly of alginate microgels by rapidly binding molecule pairs.

Controlled self-assembly of cell-encapsulating microscale polymeric hydrogels (microgels) could be advantageous in a variety of tissue engineering and regenerative medicine applications. Here, a method of assembly by chemical modification of alginate polymer with binding pair molecules (BPM) was explored. Alginate was modified with several types of BPM, specifically biotin and streptavidin and click chemistry compounds, and fabricated into 25-30 μm microgels using a microfluidic platform.

Single cell-laden protease-sensitive microniches for long-term culture in 3D.

Single cell-laden three-dimensional (3D) microgels that can serve to mimic stem cell niches in vitro, and are therefore termed microniches, can be efficiently fabricated by droplet-based microfluidics. In this technique an aqueous polymer solution along with a highly diluted cell solution is injected into a microfluidic device to create monodisperse pre-microgel droplets that are then solidified by a polymer crosslinking reaction to obtain monodisperse single cell-laden microniches. However, problems limiting this approach studying the fate of single cells include Poisson encapsulation statistics that result in mostly empty microniches, and cells egressing from the microniches during subsequent cell culture.

Deterministic encapsulation of single cells in thin tunable microgels for niche modelling and therapeutic delivery.

Existing techniques to encapsulate cells into microscale hydrogels generally yield high polymer-to-cell ratios and lack control over the hydrogel's mechanical properties. Here, we report a microfluidic-based method for encapsulating single cells in an approximately six-micrometre layer of alginate that increases the proportion of cell-containing microgels by a factor of ten, with encapsulation efficiencies over 90%. We show that in vitro cell viability was maintained over a three-day period, that the microgels are mechanically tractable, and that, for microscale cell assemblages of encapsulated marrow stromal cells cultured in microwells, osteogenic differentiation of encapsulated cells depends on gel stiffness and cell density.

Effects of substrate stiffness and cell-cell contact on mesenchymal stem cell differentiation.

The mechanical properties of the microenvironment and direct contact-mediated cell-cell interactions are two variables known to be important in the determination of stem cell differentiation fate, but little is known about the interplay of these cues. Here, we use a micropatterning approach on polyacrylamide gels of tunable stiffnesses to study how homotypic cell-cell contacts and mechanical stiffness affect different stages of osteogenesis of mesenchymal stem cells (MSCs). Nuclear localization of transcription factors associated with osteogenesis depended on substrate stiffness and was independent of the degree of cell-cell contact.

Regenerative medicine: Current therapies and future directions.

Organ and tissue loss through disease and injury motivate the development of therapies that can regenerate tissues and decrease reliance on transplantations. Regenerative medicine, an interdisciplinary field that applies engineering and life science principles to promote regeneration, can potentially restore diseased and injured tissues and whole organs. Since the inception of the field several decades ago, a number of regenerative medicine therapies, including those designed for wound healing and orthopedics applications, have received Food and Drug Administration (FDA) approval and are now commercially available.

Microfluidic Generation of Monodisperse, Structurally Homogeneous Alginate Microgels for Cell Encapsulation and 3D Cell Culture.

Monodisperse alginate microgels (10-50 μm) are created via droplet-based microfluidics by a novel crosslinking procedure. Ionic crosslinking of alginate is induced by release of chelated calcium ions. The process separates droplet formation and gelation reaction enabling excellent control over size and homogeneity under mild reaction conditions.

Linear patterning of mesenchymal condensations is modulated by geometric constraints.

The development of the vertebral column starts with the formation of a linear array of mesenchymal condensations, forming the blueprint for the eventual alternating pattern of bone and cartilage. Despite growing insight into the molecular mechanisms of morphogenesis, the impact of the physical aspects of the environment is not well understood. We hypothesized that geometric boundary conditions may play a pivotal role in the linear patterning of condensations, as neighbouring tissues provide physical constraints to the cell population.

Harnessing traction-mediated manipulation of the cell/matrix interface to control stem-cell fate.

Stem cells sense and respond to the mechanical properties of the extracellular matrix. However, both the extent to which extracellular-matrix mechanics affect stem-cell fate in three-dimensional microenvironments and the underlying biophysical mechanisms are unclear. We demonstrate that the commitment of mesenchymal stem-cell populations changes in response to the rigidity of three-dimensional microenvironments, with osteogenesis occurring predominantly at 11-30 kPa.