High-Throughput Toxicity and Phenotypic Screening of 3D Human Neural Progenitor Cell Cultures on a Microarray Chip Platform

Nierode, Gregory J.; Perea, Brian; McFarland, Sean; Pascoal, Jorge F.; Clark, Douglas S.; Schaffer, David V.; Dordick, Jonathan S.

doi:10.1016/j.stemcr.2016.10.001

Cited by 58 publications

(51 citation statements)

References 51 publications

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“…On-chip 3D cultures were prepared using an enclosed MicroSys 5100–4SQ (DigiLabs) non-contact robotic microarray liquids dispensing system with a cooling head modification, as described previously (Nierode et al, 2016). In short, 850 nL of complete growth media was printed into the microwell chips, covered with gas permeable sealing membranes (Diversified Biotech) and stored in an incubator (37 o C and 5% CO2).…”

Section: Methodsmentioning

confidence: 99%

“…The chips were then rinsed with sterile deionized water and air dried in a sterile laminar flow hood. Before use, the microwell chips were exposed to UV (302 nm; 9 cm from source) for 4 hr using a 96 W transilluminator (Syngene GVM-30) to reduce surface hydrophobicity, as outlined by Kohen, Little, & Healy (2009). On-chip 3D cultures were prepared using an enclosed MicroSys 5100-4SQ (DigiLabs) noncontact robotic microarray liquids dispensing system with a cooling head modification, as described previously (Nierode et al, 2016). In short, 850 nl of complete growth media was printed into the microwell chips, covered with gas permeable sealing membranes (Diversified Biotech) and stored in an incubator (37°C and 5% CO 2 ).…”

Section: Three-dimensional Microarray Culture Preparationmentioning

confidence: 99%

“…For example, Yang et al found that changing the encapsulating matrix altered directed differentiation outcomes of 3D encapsulated embryonic stem cells (Yang et al, 2010). There is, therefore, an important need to screen and optimize differentiation protocols, particularly for emerging 3D cell cultures, yet high-throughput screening of 3D cell cultures, including studies of stem cell differentiation, has only recently begun to emerge (Nierode et al, 2016; Ranga et al, 2014). …”

Section: Introductionmentioning

confidence: 99%

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High‐throughput identification of factors promoting neuronal differentiation of human neural progenitor cells in microscale 3D cell culture

Nierode

Gopal

Kwon

et al. 2018

Biotech & Bioengineering

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Identification of conditions for guided and specific differentiation of human stem cell and progenitor cells is important for continued development and engineering of in vitro cell culture systems for use in regenerative medicine, drug discovery, and human toxicology. Three-dimensional (3D) and organotypic cell culture models have been used increasingly for in vitro cell culture because they may better model endogenous tissue environments. However, detailed studies of stem cell differentiation within 3D cultures remain limited, particularly with respect to high-throughput screening. Herein, we demonstrate the use of a microarray chip-based platform to screen, in high-throughput, individual and paired effects of 12 soluble factors on the neuronal differentiation of a human neural progenitor cell line (ReNcell VM) encapsulated in microscale 3D Matrigel cultures. Dose-response analysis of selected combinations from the initial combinatorial screen revealed that the combined treatment of all-trans retinoic acid (RA) with the glycogen synthase kinase 3 inhibitor CHIR-99021 (CHIR) enhances neurogenesis while simultaneously decreases astrocyte differentiation, whereas the combined treatment of brain-derived neurotrophic factor and the small azide neuropathiazol enhances the differentiation into neurons and astrocytes. Subtype specification analysis of RA- and CHIR-differentiated cultures revealed that enhanced neurogenesis was not biased toward a specific neuronal subtype. Together, these results demonstrate a high-throughput screening platform for rapid evaluation of differentiation conditions in a 3D environment, which will aid the development and application of 3D stem cell culture models.

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Section: Methodsmentioning

confidence: 99%

Section: Three-dimensional Microarray Culture Preparationmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

High‐throughput identification of factors promoting neuronal differentiation of human neural progenitor cells in microscale 3D cell culture

Nierode

Gopal

Kwon

et al. 2018

Biotech & Bioengineering

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show abstract

“…That such individual floating cells can persist and fuel metastatic growth and/or relapse underscores the need to uncover their properties at clonal resolution. In this respect, the scalability of our method affords particular advantages, being uniquely suitable for high throughput plating, using microwells or micropillars 45,46 that allow to seed one cell at the time, thus optimizing the sample yield per experiment and thereby streamlining patient-specific drug screenings. scOCOs could thus be automatically seeded in well-dense plates and observed after controlled stimulation by drugs or molecular interference, isolating the best and worst responder for further multi-omics investigation.…”

Section: Discussionmentioning

confidence: 99%

Single cell derived organoids capture the self-renewing subpopulations of metastatic ovarian cancer

Velletri

Villa

Lupia

et al. 2018

Preprint

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High Grade Serous Ovarian cancer (HGSOC) is a major unmet need in oncology, due to its precocious dissemination and the lack of meaningful human models for the investigation of disease pathogenesis in a patient-specific manner. To overcome this roadblock, we present a new method to isolate and grow single cells directly from patients' ascites, establishing the conditions for propagating them as single-cell derived ovarian cancer organoids (scOCOs). By single cell RNA sequencing (scRNAseq) we define the cellular composition of metastatic ascites and trace its propagation in 2D and 3D culture paradigms, finding that scOCOs retain and amplify key subpopulations from the original patients' samples and recapitulate features of the original metastasis that do not emerge from classical 2D culture, including retention of individual patients' specificities. By enabling the enrichment of uniquely informative cell subpopulations from HGSOC metastasis and the clonal interrogation of their diversity at the functional and molecular level, this method transforms the prospects of precision oncology for ovarian cancer.

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“…Attempts have been made to generate thin‐layer Matrigel cultures using microfluidic technologies, microarray chips, polydimethylsiloxane molds, and wax‐printed chromatography paper . While these platforms offer high‐throughput production capabilities, specialized equipment and materials are required, which can limit accessibility of these techniques to the wider scientific community.…”

Section: Introductionmentioning

confidence: 99%

Fabrication of thin‐layer matrigel‐based constructs for three‐dimensional cell culture

Tsai

Frampton

2018

Biotechnology Progress

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Extracellular matrix‐based hydrogels such as Matrigel are easy‐to‐use, commercially available, and offer environments for three‐dimensional (3‐D) cell culture that mimic native tissue. However, manipulating small volumes of these materials to produce thin‐layer 3‐D culture systems suitable for analysis is difficult because of air–liquid‐substrate interfacial tension effects and evaporation. Here, we demonstrate two simple techniques that use standard liquid‐handling tools and nontreated 96‐well plates to produce uniform, thin‐layer constructs for 3‐D culture of cells in Matrigel. The first technique, the floating 3‐D cell culture method, uses phase‐separating polymers to form a barrier between the dispensed Matrigel, air, and cultureware surface to generate consistently thin hydrogels from volumes as low as 5 μL. These unanchored gels provide a useful assay for investigating airway smooth muscle cell contraction and may have future applications in studying asthma pathophysiology. The second technique, the fixed 3‐D cell culture method, provides an anchored gel system for culturing noncontractile cells (e.g., neurons) where 20 μL of Matrigel is dispensed into the bottom of a well filled with culture medium to form a thin gel containing embedded cells. This technique has potential widespread applications as an accessible 3‐D culture platform for high‐throughput production of disease models for evaluation of novel drug therapies. © 2018 American Institute of Chemical Engineers Biotechnol. Prog., 35: e2733, 2019

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High-Throughput Toxicity and Phenotypic Screening of 3D Human Neural Progenitor Cell Cultures on a Microarray Chip Platform

Cited by 58 publications

References 51 publications

High‐throughput identification of factors promoting neuronal differentiation of human neural progenitor cells in microscale 3D cell culture

High‐throughput identification of factors promoting neuronal differentiation of human neural progenitor cells in microscale 3D cell culture

Single cell derived organoids capture the self-renewing subpopulations of metastatic ovarian cancer

Fabrication of thin‐layer matrigel‐based constructs for three‐dimensional cell culture

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