Artificial niche microarrays for probing single stem cell fate in high throughput

Gobaa, Samy; Hoehnel, Sylke; Roccio, Marta; Negro, Andrea; Köbel, Stefan; Lütolf, Matthias P.

doi:10.1038/nmeth.1732

Cited by 376 publications

(328 citation statements)

References 29 publications

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“…The developed system, although requiring further investigations to specify the mode of establishment (e.g., a direct vs. indirect niche effect of the human cells in triggering homing of the mouse HSC), offers the opportunity to address a variety of critical questions at the interface between HSC biology and regenerative medicine. The model can be exploited to dissect cues that regulate HSC engraftment, by engineering marrow environments with defined and genetically controlled properties (e.g., by knockin/out of specific factors, possibly defined by high-throughput screening systems) (13,26). In fact, the capacity of the ectopic marrow to attract and retain rare circulating HSCs advocates its function as a "stem cell trap," ultimately providing a tool to investigate the physiological factors and mechanisms underlying the still debated concept of the HSC niche.…”

Section: Discussionmentioning

confidence: 99%

Engineering of a functional bone organ through endochondral ossification

Scotti

Piccinini

Takizawa

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

279

295

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Embryonic development, lengthening, and repair of most bones proceed by endochondral ossification, namely through formation of a cartilage intermediate. It was previously demonstrated that adult human bone marrow-derived mesenchymal stem/stromal cells (hMSCs) can execute an endochondral program and ectopically generate mature bone. Here we hypothesized that hMSCs pushed through endochondral ossification can engineer a scaled-up ossicle with features of a "bone organ," including physiologically remodeled bone, mature vasculature, and a fully functional hematopoietic compartment. Engineered hypertrophic cartilage required IL-1β to be efficiently remodeled into bone and bone marrow upon subcutaneous implantation. This model allowed distinguishing, by analogy with bone development and repair, an outer, cortical-like perichondral bone, generated mainly by host cells and laid over a premineralized area, and an inner, trabecular-like, endochondral bone, generated mainly by the human cells and formed over the cartilaginous template. Hypertrophic cartilage remodeling was paralleled by ingrowth of blood vessels, displaying sinusoid-like structures and stabilized by pericytic cells. Marrow cavities of the ossicles contained phenotypically defined hematopoietic stem cells and progenitor cells at similar frequencies as native bones, and marrow from ossicles reconstituted multilineage long-term hematopoiesis in lethally irradiated mice. This study, by invoking a "developmental engineering" paradigm, reports the generation by appropriately instructed hMSC of an ectopic "bone organ" with a size, structure, and functionality comparable to native bones. The work thus provides a model useful for fundamental and translational studies of bone morphogenesis and regeneration, as well as for the controlled manipulation of hematopoietic stem cell niches in physiology and pathology.

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Section: Discussionmentioning

confidence: 99%

Engineering of a functional bone organ through endochondral ossification

Scotti

Piccinini

Takizawa

et al. 2013

Proc. Natl. Acad. Sci. U.S.A.

279

295

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“…Such advances allow the introduction of spatially specific cues in hydrogels, making multicellular constructs, either through co-cultures or multilineage differentiation, a possibility [16][17][18] . Spatial patterning of hydrogels also provides an additional tool for the development of high-throughput screening technology for the rapid investigation of cell-material interactions [19][20][21][22] . High-throughput techniques may be particularly important as cell fate is simultaneously influenced by cell-cell interactions, soluble signals, cell-ECM interactions, cell shape and mechanical forces, and natural ECMs often present cells with many distinct signals simultaneously 23 .…”

Section: Static Hydrogels That Mimic Biophysical Cuesmentioning

confidence: 99%

Moving from static to dynamic complexity in hydrogel design

2012

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Hydrogels are water-swollen polymer networks that have found a range of applications from biological scaffolds to contact lenses. Historically, their design has consisted primarily of static systems and those that exhibit simple degradation. However, advances in polymer synthesis and processing have led to a new generation of dynamic systems that are capable of responding to artificial triggers and biological signals with spatial precision. These systems will open up new possibilities for the use of hydrogels as model biological structures and in tissue regeneration.H ydrogels are water-swollen polymer networks that have been used for many decades, with applications as varied as contact lenses and super-absorbant materials. As the field of biomedical engineering has developed, hydrogels have become a prime candidate for application as molecule delivery vehicles and as carriers for cells in tissue engineering, owing to their ability to mimic many aspects of the native cellular environment (for example, high water content, mechanical properties that match soft tissues). Traditional hydrogels, formed through the covalent and non-covalent crosslinking of polymer chains, were regarded as relatively inert materials, providing a simple biomimetic three-dimensional (3D) environment, either for tissue production by local resident cells or for positioning of cells delivered in vivo. However, the simplicity of these materials may have in fact hindered their application, restricting cellular interactions with the environment and preventing uniform extracellular matrix (ECM) production and proper tissue development. In addition, these materials were limited to modelling static environments and lacked the spatiotemporal dynamic properties relevant for complex tissue processes. Fortunately, during the last decade the concepts of hydrogel design and cellular interaction have evolved, shedding light on how they may control cell behaviour, particularly for tissue engineering applications.Hydrogels with a range of mechanical properties, and capable of incorporating a wide range of biologically relevant molecules, from individual functional groups to multidomain proteins, are currently in development 1 . In addition, hydrogels are being designed with spatial heterogeneity, to either replicate properties in native tissue structures or to produce constructs with distinct regionally specific cell behavior 2 . As a result, studies to date have clearly demonstrated the possibility of creating well-defined microenvironments with control over the 3D presentation of signals to cells 3 . However, recently there has been a focus on the concept of hydrogels that exhibit dynamic complexity. These materials should evolve with time and in response to

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“…Miniaturizing PEG hydrogels through microfabrication or microfluidics has emerged as a promising and versatile platform for biosensing 23 , tissue engineering 24 , drug delivery 25,26 , and high throughput screening 27 . The generation of structured hydrogels through bioprinting 28 , photolithography 29 , and liquid bridging 30 have been demonstrated to effectively generate functional platforms for cell studies.…”

Section: Introductionmentioning

confidence: 99%

A microfluidic-based cell encapsulation platform to achieve high long-term cell viability in photopolymerized PEGNB hydrogel microspheres

et al. 2017

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Cell encapsulation within photopolymerized polyethylene glycol (PEG)-based hydrogel scaffolds has been demonstrated as a robust strategy for cell delivery, tissue engineering, regenerative medicine, and developing in vitro platforms to study cellular behavior and fate. Strategies to achieve spatial and temporal control over PEG hydrogel mechanical properties, chemical functionalization, and cytocompatibility have advanced considerably in recent years. Recent microfluidic technologies have enabled the miniaturization of PEG hydrogels, thus enabling the fabrication of miniaturized cell-laden vehicles. However, rapid oxygen diffusive transport times on the microscale dramatically inhibit chain growth photopolymerization of polyethylene glycol diacrylate (PEGDA), thus decreasing the viability of cells encapsulated within these microstructures. Another promising PEG-based scaffold material, PEG norbornene (PEGNB), is formed by a step-growth photopolymerization and is not inhibited by oxygen. PEGNB has also been shown to be more cytocompatible than PEGDA and allows for orthogonal addition reactions. The step-growth kinetics, however, are slow and therefore challenging to fully polymerize within droplets flowing through microfluidic devices. Here, we describe a microfluidic-based droplet fabrication platform that generates consistently monodisperse cell-laden water-in-oil emulsions. Microfluidically generated PEGNB droplets are collected and photopolymerized under UV exposure in bulk emulsions. In this work, we compare this microfluidic-based cell encapsulation platform with a vortex-based method on the basis of microgel size, uniformity, post-encapsulation cell viability and long-term cell viability. Several factors that influence post-encapsulation cell viability were identified. Finally, long-term cell viability achieved by this platform was compared to a similar cell encapsulation platform using PEGDA. We show that this PEGNB microencapsulation platform is capable of generating cell-laden hydrogel microspheres at high rates with well-controlled size distributions and high long-term cell viability.

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Artificial niche microarrays for probing single stem cell fate in high throughput

Cited by 376 publications

References 29 publications

Engineering of a functional bone organ through endochondral ossification

Engineering of a functional bone organ through endochondral ossification

Moving from static to dynamic complexity in hydrogel design

A microfluidic-based cell encapsulation platform to achieve high long-term cell viability in photopolymerized PEGNB hydrogel microspheres

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