Mesenchymal stem cell-secretome laden photopolymerizable hydrogels for wound healing.

Mesenchymal stem cell-derived secretome represents an emerging acellular therapeutic which possess significant opportunity for clinical applications due to its anti-inflammatory, immunomodulatory, and wound healing properties. However, maintaining therapeutic efficacy and ensuring stability of cell-based products is challenging, requiring a robust delivery method. Therefore, we designed a hydrogel-based scaffold loaded with CK Cell Technologies' proprietary Mesenchymal stem cell-secretome for controlled release treatment of acute and chronic wounds. We incorporated both conditioned media (CM) and extracellular vesicles (EVs) into gelatin methacryloyl (GelMA) hydrogels and demonstrated how we can tune the diffusive release of the EVs from them. To demonstrate viability of the approach, we developed a wound healing scratch assay where we see in situ release of CM and EVs promote enhanced migration of human dermal fibroblasts (hDFs). We see the colocalization of these EVs in the fibroblasts using fluorescent microscopy. Finally, as a surrogate for in vivo neovascularization, we conducted an in vitro tube formation assay for the MSC-secretome using matrigel-embedded human microvascular endothelial cells. By adding CM and EVs, we observe an increase in tubulogenesis. Collectively, our data demonstrates by tuning the GelMA properties, we can influence the controlled release of the MSC-secretome for a wound dressing and bandage application for chronic and acute wounds.

ROS-mediated anticancer effects of EGFR-targeted nanoceria.

The therapeutic effectiveness of anticancer drugs, including nanomedicines, can be enhanced with active receptor-targeting strategies. Epidermal growth factor receptor (EGFR) is an important cancer biomarker, constitutively expressed in sarcoma patients of different histological types. The present work reports materials and in vitro biomedical analyses of silanized (passive delivery) and/or EGF-functionalized (active delivery) ceria nanorods exhibiting highly defective catalytically active surfaces. The EGFR-targeting efficiency of nanoceria was confirmed by receptor-binding studies. Increased cytotoxicity and reactive oxygen species (ROS) production were observed for EGF-functionalized nanoceria owing to enhanced cellular uptake by HT-1080 fibrosarcoma cells. The uptake was confirmed by TEM and confocal microscopy. Silanized nanoceria demonstrated negligible/minimal cytotoxicity toward healthy MRC-5 cells at 24 and 48 h, whereas this was significant at 72 h owing to a nanoceria accumulation effect. In contrast, considerable cytotoxicity toward the cancer cells was exhibited at all three times points. The ROS generation and associated cytotoxicity were moderated by the equilibrium between catalysis by ceria, generation of cell debris, and blockage of active sites. EGFR-targeting is shown to enhance the uptake levels of nanoceria by cancer cells, subsequently enhancing the overall anticancer activity and therapeutic performance of ceria.

Hierarchical assembly of tryptophan zipper peptides into stress-relaxing bioactive hydrogels.

Soft materials in nature are formed through reversible supramolecular assembly of biological polymers into dynamic hierarchical networks. Rational design has led to self-assembling peptides with structural similarities to natural materials. However, recreating the dynamic functional properties inherent to natural systems remains challenging. Here we report the discovery of a short peptide based on the tryptophan zipper (trpzip) motif, that shows multiscale hierarchical ordering that leads to emergent dynamic properties. Trpzip hydrogels are antimicrobial and self-healing, with tunable viscoelasticity and unique yield-stress properties that allow immediate harvest of embedded cells through a flick of the wrist. This characteristic makes Trpzip hydrogels amenable to syringe extrusion, which we demonstrate with examples of cell delivery and bioprinting. Trpzip hydrogels display innate bioactivity, allowing propagation of human intestinal organoids with apical-basal polarization. Considering these extensive attributes, we anticipate the Trpzip motif will prove a versatile building block for supramolecular assembly of soft materials for biotechnology and medicine.

Magnetic Nanofibrous Hydrogels for Dynamic Control of Stem Cell Differentiation.

The extracellular matrix in tissue consists of complex heterogeneous soft materials with hierarchical structure and dynamic mechanical properties dictating cell and tissue level function. In many natural matrices, there are nanofibrous structures that serve to guide cell activity and dictate the form and function of tissue. Synthetic hydrogels with integrated nanofibers can mimic the structural properties of native tissue; however, model systems with dynamic mechanical properties remain elusive. Here we demonstrate modular nanofibrous hydrogels that can be reversibly stiffened in response to applied magnetic fields. Iron oxide nanoparticles were incorporated into gelatin nanofibers through electrospinning, followed by chemical stabilization and fragmentation. These magnetoactive nanofibers can be mixed with virtually any hydrogel material and reversibly stiffen the matrix at a low fiber content (≤3%). In contrast to previous work, where a large quantity of magnetic material disallowed cell encapsulation, the low nanofiber content allows matrix stiffening with cells in 3D. Using adipose derived stem cells, we show how nanofibrous matrices are beneficial for both osteogenesis and adipogenesis, where stiffening the hydrogel with applied magnetic fields enhances osteogenesis while discouraging adipogenesis. Skeletal myoblast progenitors were used as a model of tissue morphogenesis with matrix stiffening augmenting myogenesis and multinucleated myotube formation. The ability to reversibly stiffen fibrous hydrogels through magnetic stimulation provides a useful tool for studying nanotopography and dynamic mechanics in cell culture, with a scope for stimuli responsive materials for tissue engineering.

Evaluation of the Immune Response to Chitosan-graft-poly(caprolactone) Biopolymer Scaffolds.

Biomimetic scaffolds recreating key elements of the architecture and biological activity of the extracellular matrix have enormous potential for soft tissue engineering applications. Combining appropriate mechanical properties with select biological cues presents a challenge for bioengineering, as natural materials are most bioactive but can lack mechanical integrity, while synthetic polymers have strength but are often biologically inert. Blends of synthetic and natural materials, aiming to combine the benefits of each, have shown promise but inherently require a compromise, diluting down favorable properties in each polymer to accommodate the other. Here, we electrospun a material comprising chitosan, a natural polysaccharide, and polycaprolactone (PCL), one of the most widely studied synthetic polymers used in materials engineering. In contrast to a classical blend, here PCL was chemically grafted onto the chitosan backbone to create chitosan-graft-polycaprolactone (CS-g-PCL) and then combined further with unmodified PCL to generate scaffolds with discreet chitosan functionalization. These small amounts of chitosan led to significant changes in scaffold architecture and surface chemistry, reducing the fiber diameter, pore size, and hydrophobicity. Interestingly, all CS-g-PCL-containing blends were stronger than control PCL, though with reduced elongation. In in vitro assessments, increasing the CS-g-PCL content led to significant improvements in in vitro blood compatibility compared to PCL alone while increasing fibroblast attachment and proliferation. In a mouse subcutaneous implantation model, a higher CS-g-PCL content improved the immune response to the implants. Macrophages in tissues surrounding CS-g-PCL scaffolds decreased proportionately to the chitosan content by up to 65%, with a corresponding decrease in pro-inflammatory cytokines. These results suggest that CS-g-PCL is a promising hybrid material comprising natural and synthetic polymers with tailorable mechanical and biological properties, justifying further development and in vivo evaluation.

Gas-modulating microcapsules for spatiotemporal control of hypoxia.

Oxygen is a vital molecule involved in regulating development, homeostasis, and disease. The oxygen levels in tissue vary from 1 to 14% with deviations from homeostasis impacting regulation of various physiological processes. In this work, we developed an approach to encapsulate enzymes at high loading capacity, which precisely controls the oxygen content in cell culture. Here, a single microcapsule is able to locally perturb the oxygen balance, and varying the concentration and distribution of matrix-embedded microcapsules provides spatiotemporal control. We demonstrate attenuation of hypoxia signaling in populations of stem cells, cancer cells, endothelial cells, cancer spheroids, and intestinal organoids. Varying capsule placement, media formulation, and timing of replenishment yields tunable oxygen gradients, with concurrent spatial growth and morphogenesis in a single well. Capsule containing hydrogel films applied to chick chorioallantoic membranes encourages neovascularization, providing scope for topical treatments or hydrogel wound dressings. This platform can be used in a variety of formats, including deposition in hydrogels, as granular solids for 3D bioprinting, and as injectable biomaterials. Overall, this platform's simplicity and flexibility will prove useful for fundamental studies of oxygen-mediated processes in virtually any in vitro or in vivo format, with scope for inclusion in biomedical materials for treating injury or disease.

Electrostatic Assembly of Multiarm PEG-Based Hydrogels as Extracellular Matrix Mimics: Cell Response in the Presence and Absence of RGD Cell Adhesive Ligands.

Synthetic hydrogels have been used widely as extracellular matrix (ECM) mimics due to the ability to control and mimic physical and biochemical cues observed in natural ECM proteins such as collagen, laminin, and fibronectin. Most synthetic hydrogels are formed via covalent bonding resulting in slow gelation which is incompatible with drop-on-demand 3D bioprinting of cells and injectable hydrogels for therapeutic delivery. Herein, we developed an electrostatically crosslinked PEG-based hydrogel system for creating high-throughput 3D in vitro models using synthetic hydrogels to mimic the ECM cancer environment. A 3-arm PEG-based polymer backbone was first modified with either permanent cationic charged moieties (2-(methacryloyloxy)ethyl trimethylammonium) or permanent anionic charged moieties (3-sulfopropyl methacrylate potassium salt). The resulting charged polymers can be conjugated further with various amounts of cell adhesive RGD motifs (0, 25, 75, and 98%) to study the influences of RGD motifs on breast cancer (MCF-7) spheroid formation. Formation, stability, and mechanical properties of hydrogels were tested with, and without, RGD to evaluate the cellular response to material parameters in a 3D environment. The hydrogels can be degraded in the presence of salts at room temperature by breaking the interaction of oppositely charged polymer chains. MCF-7 cells could be released with high viability through brief exposure to NaCl solution. Flow cytometry characterization demonstrated that embedded MCF-7 cells proliferate better in a softer (60 Pa) 3D hydrogel environment compared to those that are stiffer (1160 Pa). As the stiffness increases, the RGD motif plays a role in promoting cell proliferation in the stiffer hydrogel. Flow cytometry characterization demonstrated that embedded MCF-7 cells proliferate better in a softer (60 Pa) 3D hydrogel environment compared to those that are stiffer (1160 Pa). As the stiffness increases, the RGD motif plays a role in promoting cell proliferation in the stiffer hydrogel. Additionally, cell viability was not impacted by the tested hydrogel stiffness range between 60 to 1160 Pa. Taken together, this PEG-based tuneable hydrogel system shows great promise as a 3D ECM mimic of cancer extracellular environments with controllable biophysical and biochemical properties. The ease of gelation and dissolution through salt concentration provides a way to quickly harvest cells for further analysis at any given time of interest without compromising cell viability.

Biomechanical, biophysical and biochemical modulators of cytoskeletal remodelling and emergent stem cell lineage commitment.

Across complex, multi-time and -length scale biological systems, redundancy confers robustness and resilience, enabling adaptation and increasing survival under dynamic environmental conditions; this review addresses ubiquitous effects of cytoskeletal remodelling, triggered by biomechanical, biophysical and biochemical cues, on stem cell mechanoadaptation and emergent lineage commitment. The cytoskeleton provides an adaptive structural scaffold to the cell, regulating the emergence of stem cell structure-function relationships during tissue neogenesis, both in prenatal development as well as postnatal healing. Identification and mapping of the mechanical cues conducive to cytoskeletal remodelling and cell adaptation may help to establish environmental contexts that can be used prospectively as translational design specifications to target tissue neogenesis for regenerative medicine. In this review, we summarize findings on cytoskeletal remodelling in the context of tissue neogenesis during early development and postnatal healing, and its relevance in guiding lineage commitment for targeted tissue regeneration. We highlight how cytoskeleton-targeting chemical agents modulate stem cell differentiation and govern responses to mechanical cues in stem cells' emerging form and function. We further review methods for spatiotemporal visualization and measurement of cytoskeletal remodelling, as well as its effects on the mechanical properties of cells, as a function of adaptation. Research in these areas may facilitate translation of stem cells' own healing potential and improve the design of materials, therapies, and devices for regenerative medicine.

Hydrogel Microtumor Arrays to Evaluate Nanotherapeutics.

Nanoparticle drug formulations have many advantages for cancer therapy due to benefits in targeting selectivity, lack of systemic toxicity, and increased drug concentration in the tumor microenvironment after delivery. However, the promise of nanomedicine is limited by preclinical models that fail to accurately assess new drugs before entering human trials. In this work a new approach to testing nanomedicine using a microtumor array formed through hydrogel micropatterning is demonstrated. This technique allows partitioning of heterogeneous cell states within a geometric pattern-where boundary regions of curvature prime the stem cell-like fraction-allowing to simultaneously probe drug uptake and efficacy in different cancer cell fractions with high reproducibility. Using melanoma cells of different metastatic potential, a relationship between stem fraction and nanoparticle uptake is discovered. Deformation cytometry reveals that the stem cell-like population exhibits a more mechanically deformable cell membrane. Since the stem fraction in a tumor is implicated in drug resistance, recurrence, and metastasis, the findings suggest that nanoparticle drug formulations are well suited for targeting this dangerous cell population in cancer therapy.

In situ formation of osteochondral interfaces through "bone-ink" printing in tailored microgel suspensions.

Osteochondral tissue has a complex hierarchical structure spanning subchondral bone to articular cartilage. Biomaterials approaches to mimic and repair these interfaces have had limited success, largely due to challenges in fabricating composite hard-soft interfaces with living cells. Biofabrication approaches have emerged as attractive methods to form osteochondral analogues through additive assembly of hard and soft components. We have developed a unique printing platform that is able to integrate soft and hard materials concurrently through freeform printing of mineralized constructs within tunable microgel suspensions containing living cells. A library of microgels based on gelatin were prepared, where the stiffness of the microgels and a liquid "filler" phase can be tuned for bioprinting while simultaneously directing differentiation. Tuning microgel stiffness and filler content differentially directs chondrogenesis and osteogenesis within the same construct, demonstrating how this technique can be used to fabricate osteochondral interfaces in a single step. Printing of a rapidly setting calcium phosphate cement, so called "bone-ink" within a cell laden suspension bath further guides differentiation, where the cells adjacent to the nucleated hydroxyapatite phase undergo osteogenesis with cells in the surrounding medium undergoing chondrogenesis. In this way, bone analogues with hierarchical structure can be formed within cell-laden gradient soft matrices to yield multiphasic osteochondral constructs. This technique provides a versatile one-pot biofabrication approach without harsh post-processing which will aid efforts in bone disease modelling and tissue engineering. STATEMENT OF SIGNIFICANCE: This paper demonstrates the first example of a biofabrication approach to rapidly form osteochondral constructs in a single step under physiological conditions. Key to this advance is a tunable suspension of extracellular matrix microgels that are packed together with stem cells, providing a unique and modular scaffolding for guiding the simultaneous formation of bone and cartilage tissue. The physical properties of the suspension allow direct writing of a ceramic "bone-ink", resulting in an ordered structure of microscale hydrogels, living cells, and bone mimics in a single step. This platform reveals a simple approach to making complex skeletal tissue for disease modelling, with the possibility of repairing and replacing bone-cartilage interfaces in the clinic using a patient's own cells.

Ceramic Omnidirectional Bioprinting in Cell-laden Suspensions for the Generation of Bone Analogs.

Structurally, bone tissue is an inorganic-organic composite containing metabolically active cells embedded within a hierarchical, highly mineralized matrix. This organization is challenging to replicate due to the heterogeneous environment of bone. Ceramic omnidirectional bioprinting in cell-suspensions (COBICS) is a microgel-based bioprinting technique that uniquely replicates the mineral and cellular structure of bone. COBICS prints complex, biologically relevant constructs without the need for sacrificial support materials or harsh postprocessing steps (e.g., radiation and high-temperature sintering), which are two of the biggest challenges in the additive manufacturing of bone mimetic constructs. This technique is enabled via the freeform extrusion of a novel calcium phosphate-based ink within a gelatin-based microgel suspension. The yield-stress properties of the suspension allow deposition and support the printed bone structure. UV crosslinking and nanoprecipitation then "lock" it in place. The ability to print nanostructured bone-mimetic ceramics within cell-laden biomaterials provides spatiotemporal control over macro- and micro-architecture and facilitates the real-time fabrication of complex bone constructs in clinical settings.

Biomaterials directed activation of a cryostable therapeutic secretome in induced pluripotent stem cell derived mesenchymal stromal cells.

Mesenchymal stem cell therapy has suffered from wide variability in clinical efficacy, largely due to heterogeneous starting cell populations and large-scale cell death during and after implantation. Optimizing the manufacturing process has led to reproducible cell populations that can be cryopreserved for clinical applications. Nevertheless, ensuring a reproducible cell state that persists after cryopreservation remains a significant challenge, and is necessary to ensure reproducible clinical outcomes. Here we demonstrate how matrix-conjugated hydrogel cell culture materials can normalize a population of induced pluripotent stem cell derived mesenchymal stem cells (iPSC-MSCs) to display a defined secretory profile that promotes enhanced neovascularization in vitro and in vivo. Using a protein-conjugated biomaterials screen we identified two conditions-1 kPa collagen and 10 kPa fibronectin coated polyacrylamide gels-that promote reproducible secretion of pro-angiogenic and immunomodulatory cytokines from iPSC-MSCs that enhance tubulogenesis of endothelial cells in Geltrex and neovascularization in chick chorioallantoic membranes. Using defined culture substrates alone, we demonstrate maintenance of secretory activity after cryopreservation for the first time. This advance provides a simple and scalable approach for cell engineering and subsequent manufacturing, toward normalizing and priming a desired cell activity for clinical regenerative medicine.

Cell-Laden Gradient Microgel Suspensions for Spatial Control of Differentiation During Biofabrication.

During tissue development, stem and progenitor cells form functional tissue with high cellular diversity and intricate micro- and macro-architecture. Current approaches have attempted to replicate this process with materials cues or through spontaneous cell self-organization. However, cell-directed and materials-directed organization are required simultaneously to achieve biomimetic structure and function. Here, it is shown how integrating live adipose derived stem cells with gradient microgel suspensions steers divergent differentiation outcomes. Microgel matrices composed of small particles are found to promote adipogenic differentiation, while larger particles fostered increased cell spreading and osteogenic differentiation. Tuning the matrix formulation demonstrates that early cell adhesion and spreading dictate differentiation outcome. Combining small and large microgels into gradients spatially directs proliferation and differentiation over time. After 21 days of culture, osteogenic conditions foster significant mineralization within the individual microgels, thereby providing cell-directed changes in composition and mechanics within the gradient porous scaffold. Freeform printing of high-density cell suspensions is performed across these gradients to demonstrate the potential for hierarchical tissue biofabrication. Interstitial porosity influences cell expansion from the print and microgel size guides spatial differentiation, thereby providing scope to fabricate tissue gradients at multiple scales through integrated and printed cell populations.

A Tunable Tumor Microenvironment through Recombinant Bacterial Collagen-Hyaluronic Acid Hydrogels.

Laboratory models of the tumor microenvironment require control of mechanical and biochemical properties to ensure accurate mimicry of patient disease. In contrast to pure natural or synthetic materials, hybrid approaches that pair recombinant protein fragments with synthetic scaffolding show many advantages. Here we demonstrate production of a recombinant bacterial collagen-like protein (CLP) for thiol-ene pairing to norbornene functionalized hyaluronic acid (NorHA). The resultant hydrogel material shows an adjustable modulus with evidence for strain-stiffening behavior that resembles natural tumor matrices. Cysteine terminated peptide binding motifs are incorporated to adjust the cell-adhesion points. The modular hybrid gel shows good biocompatibility and was demonstrated to control cell adhesion, proliferation, and the invasive properties of MCF7 and MD-MBA-231 breast adenocarcinoma cells. The ease in which multiple structural and bioactive components can be integrated provides a robust framework to form models of the tumor microenvironment for fundamental studies and drug development.

Gallium Nanodroplets are Anti-Inflammatory without Interfering with Iron Homeostasis.

Gallium (Ga) compounds, as the source of Ga ions (Ga3+), have been historically used as anti-inflammatories. Currently, the widely accepted mechanisms of the anti-inflammatory effects for Ga3+ are rationalized on the basis of their similarities to ferric ions (Fe3+), which permits Ga3+ to bind with Fe-binding proteins and subsequently disturbs the Fe homeostasis in the immune cells. Here in contrast to the classic views, our study presents the mechanisms of Ga as anti-inflammatory by delivering Ga nanodroplets (GNDs) into lipopolysaccharide-induced macrophages and exploring the processes. The GNDs show a selective inhibition of nitric oxide (NO) production without affecting the accumulation of pro-inflammatory mediators. This is explained by GNDs disrupting the synthesis of inducible NO synthase in the activated macrophages by upregulating the levels of eIF2α phosphorylation, without interfering with the Fe homeostasis. The Fe3+ transferrin receptor-independent endocytosis of GNDs by the cells prompts a fundamentally different mechanism as anti-inflammatories in comparison to that imparted by Ga3+. This study reveals the fundamental molecular basis of GND-macrophage interactions, which may provide additional avenues for the use of Ga for anti-inflammatory and future biomedical and pharmaceutical applications.

Structural aspects controlling the mechanical and biological properties of tough, double network hydrogels.

Anticipating an increasing demand for hybrid double network (DN) hydrogels in biomedicine and biotechnology, this study evaluated the effects of each network on the mechanical and biological properties. Polyethylene glycol (PEG) (meth)acrylate hydrogels with varied monomer molecular weights and architectures (linear vs. 4-arm) were produced with and without an added ionically bonded alginate network and their mechanical properties were characterized using compression testing. The results showed that while some mechanical properties of PEG single network (SN) hydrogels decreased or changed negligibly with increasing molecular weight, the compressive modulus, strength, strain to failure, and toughness of DN hydrogels all significantly increased with increased PEG monomer molecular weight. At a fixed molecular weight (10 kDa), 4-arm PEG SN hydrogels exhibited better overall mechanical performance; however, this benefit was diminished for the corresponding DN hydrogels with comparable strength and toughness and lower strain to failure for the 4-arm case. Regardless of the PEG monomer structure, the alginate network made a relatively larger contribution to the overall DN mechanical properties when the covalent PEG network was looser with a larger mesh size (e.g., for larger monomer molecular weight and/or linear architecture) which presumably enabled more ionic crosslinking. Considering the biological performance, adipose derived stem cell cultures demonstrated monotonically increasing cell area and Yes-associated protein related mechanosensing with increasing amounts of alginate from 0 to 2 wt.%, demonstrating the possibility for using DN hydrogels in guiding musculoskeletal differentiation. These findings will be useful to design suitable hydrogels with controllable mechanical and biological properties for mechanically demanding applications. STATEMENT OF SIGNIFICANCE: Hydrogels are widely used in commercial applications, and recently developed hybrid double network hydrogels have enhanced strength and toughness that will enable further expansion into more mechanically demanding applications (e.g., medical implants, etc.). The significance of this work is that it uncovers some key principles regarding monomer molecular weight, architecture, and concentration for developing strong and tough hybrid double network hydrogels that would not be predicted from their single network counterparts or a linear combination of the two networks. Additionally, novel insight is given into the biological performance of hybrid double network hydrogels in the presence of adipose derived stem cell cultures which suggests new scope for using double network hydrogels in guiding musculoskeletal differentiation.

Pluripotent stem cell-derived mesenchymal stromal cells improve cardiac function and vascularity after myocardial infarction.

BACKGROUND AIMS: Mesenchymal stromal cells (MSCs) have been shown to improve cardiac function after injury and are the subject of ongoing clinical trials. In this study, the authors tested the cardiac regenerative potential of an induced pluripotent stem cell-derived MSC (iPSC-MSC) population (Cymerus MSCs) in a rat model of myocardial ischemia-reperfusion (I/R). Furthermore, the authors compared this efficacy with bone marrow-derived MSCs (BM-MSCs), which are the predominant cell type in clinical trials. METHODS: Four days after myocardial I/R injury, rats were randomly assigned to (i) a Cymerus MSC group (n = 15), (ii) a BM-MSC group (n = 15) or (iii) a vehicle control group (n = 14). For cell-treated animals, a total of 5 × 106 cells were injected at three sites within the infarcted left ventricular (LV) wall. RESULTS: One month after cell transplantation, Cymerus MSCs improved LV function (assessed by echocardiography) compared with vehicle and BM-MSCs. Interestingly, Cymerus MSCs enhanced angiogenesis without sustained engraftment or significant impact on infarct scar size. Suggesting safety, Cymerus MSCs had no effect on inducible tachycardia or the ventricular scar heterogeneity that provides a substrate for cardiac re-entrant circuits. CONCLUSIONS: The authors here demonstrate that intra-myocardial administration of iPSC-MSCs (Cymerus MSCs) provide better therapeutic effects compared with conventional BM-MSCs in a rodent model of myocardial I/R. Because of its manufacturing scalability, iPSC-MSC therapy offers an exciting opportunity for an "off-the-shelf" stem cell therapy for cardiac repair.

Heterotypic tumor models through freeform printing into photostabilized granular microgels.

The tissue microenvironment contains a complex assortment of multiple cell types, matrices, and vessel structures, which is difficult to reconstruct in vitro. Here, we demonstrate model tumor microenvironments formed through direct writing of vasculature channels and tumor cell aggregates, within a cell-laden microgel matrix. Photocrosslinkable microgels provide control over local and global mechanics, while enabling the integration of virtually any cell type. Direct writing of a Pluronic sacrificial ink into a stromal cell-microgel suspension is used to form vessel structures for endothelialization, followed by printing of melanoma aggregates. Tumor cells migrate into the prototype vessels as a function of spatial location, thereby providing a measure of invasive potential. The integration of perfusable channels with multiple spatially defined cell types provides new avenues for modelling development and disease, with scope for both fundamental research and drug development efforts.