Development-on-chip: in vitro neural tube patterning with a microfluidic device

Embryogenesis is a highly regulated process in which the precise spatial and temporal release of soluble cues directs differentiation of multipotent stem cells into discrete populations of specialized adult cell types. In the spinal cord, neural progenitor cells are directed to differentiate into adult neurons through the action of mediators released from nearby organizing centers, such as the floor plate and paraxial mesoderm. These signals combine to create spatiotemporal diffusional landscapes that precisely regulate the development of the central nervous system (CNS). Currently, in vivo and ex vivo studies of these signaling factors present some inherent ambiguity. In vitro methods are preferred for their enhanced experimental clarity but often lack the technical sophistication required for biological realism. In this article, we present a versatile microfluidic platform capable of mimicking the spatial and temporal chemical environments found in vivo during neural tube development. Simultaneous opposing and/or orthogonal gradients of developmental morphogens can be maintained, resulting in neural tube patterning analogous to that observed in vivo. Summary: A microfluidic device mimics the spatial and temporal environment of neural tube development in vivo and enables the correct spatial organization of neural tube formation from stem cells in vitro.

[1]  A. Martinez Arias,et al.  Generation of Aggregates of Mouse Embryonic Stem Cells that Show Symmetry Breaking, Polarization and Emergent Collective Behaviour In Vitro , 2015, Journal of visualized experiments : JoVE.

[2]  L. Rubin,et al.  Motoneurons Derived from Embryonic Stem Cells Express Transcription Factors and Develop Phenotypes Characteristic of Medial Motor Column Neurons , 2006, The Journal of Neuroscience.

[3]  K. Eggan,et al.  How to make spinal motor neurons , 2014, Development.

[4]  T. Jessell,et al.  Regulation of the neural patterning activity of sonic hedgehog by secreted BMP inhibitors expressed by notochord and somites. , 2000, Development.

[5]  T. Jessell,et al.  Graded sonic hedgehog signaling and the specification of cell fate in the ventral neural tube. , 1997, Cold Spring Harbor symposia on quantitative biology.

[6]  C. Henderson,et al.  Motor neuron diversity in development and disease. , 2010, Annual review of neuroscience.

[7]  S. Price,et al.  The generation and diversification of spinal motor neurons: signals and responses , 2004, Mechanisms of Development.

[8]  Silvia Arber,et al.  Requirement for the Homeobox Gene Hb9 in the Consolidation of Motor Neuron Identity , 1999, Neuron.

[9]  Mohammad A. Qasaimeh,et al.  Integration of Shallow Gradients of Shh and Netrin-1 Guides Commissural Axons , 2015, PLoS biology.

[10]  H. Wichterle,et al.  A Requirement for Retinoic Acid-Mediated Transcriptional Activation in Ventral Neural Patterning and Motor Neuron Specification , 2003, Neuron.

[11]  Emily Gale,et al.  Opposing FGF and Retinoid Pathways Control Ventral Neural Pattern, Neuronal Differentiation, and Segmentation during Body Axis Extension , 2003, Neuron.

[12]  J. Briscoe,et al.  Canonical BMP7 activity is required for the generation of discrete neuronal populations in the dorsal spinal cord , 2012, Development.

[13]  D. Beebe,et al.  Biological implications of polydimethylsiloxane-based microfluidic cell culture. , 2009, Lab on a chip.

[14]  R. Kamm,et al.  Extracellular Matrix Heterogeneity Regulates Three‐Dimensional Morphologies of Breast Adenocarcinoma Cell Invasion , 2012, Advanced healthcare materials.

[15]  Martin Dufva,et al.  Poly(Dimethylsiloxane) (PDMS) Affects Gene Expression in PC12 Cells Differentiating into Neuronal-Like Cells , 2013, PloS one.

[16]  James Briscoe,et al.  Pattern formation in the vertebrate neural tube: a sonic hedgehog morphogen-regulated transcriptional network , 2008, Development.

[17]  R. Broome,et al.  Wnt/β-catenin and FGF signalling direct the specification and maintenance of a neuromesodermal axial progenitor in ensembles of mouse embryonic stem cells , 2014, Development.

[18]  T. Jessell,et al.  Floor plate and motor neuron induction by different concentrations of the amino-terminal cleavage product of sonic hedgehog autoproteolysis , 1995, Cell.

[19]  Raymond Turner,et al.  Specification , 2011, Minds and Machines.

[20]  C. Stern.  Neural induction: 10 years on since the 'default model'. , 2006, Current opinion in cell biology.

[21]  Olivier Pourquié,et al.  Faculty Opinions recommendation of Opposing FGF and retinoid pathways control ventral neural pattern, neuronal differentiation, and segmentation during body axis extension. , 2004 .

[22]  L. Wilson,et al.  The mechanisms of dorsoventral patterning in the vertebrate neural tube. , 2005, Developmental biology.

[23]  G. Schoenwolf,et al.  Neurulation: coming to closure , 1997, Trends in Neurosciences.

[24]  J. Briscoe,et al.  Temporal control of BMP signalling determines neuronal subtype identity in the dorsal neural tube , 2013, Development.

[25]  S. D. Collins,et al.  Microfluidic device for the combinatorial application and maintenance of dynamically imposed diffusional gradients , 2010 .

[26]  Ya-min Wu,et al.  Directed differentiation of embryonic stem cells into motor neurons by gene , 2004 .

[27]  T. Jessell.  Neuronal specification in the spinal cord: inductive signals and transcriptional codes , 2000, Nature Reviews Genetics.

[28]  Sonja Nowotschin,et al.  Symmetry breaking, germ layer specification and axial organisation in aggregates of mouse embryonic stem cells , 2014, Development.

[29]  Jong-Hoon Kim,et al.  Differentiation of Neural Progenitor Cells in a Microfluidic Chip‐Generated Cytokine Gradient , 2009, Stem cells.

[30]  A. McMahon,et al.  Notochord-derived Shh concentrates in close association with the apically positioned basal body in neural target cells and forms a dynamic gradient during neural patterning , 2008, Development.

[31]  D. Gottlieb,et al.  Retinoic acid promotes neural and represses mesodermal gene expression in mouse embryonic stem cells in culture. , 1996, Biochemical and biophysical research communications.

[32]  James Briscoe,et al.  Establishing and interpreting graded Sonic Hedgehog signaling during vertebrate neural tube patterning: the role of negative feedback. , 2009, Cold Spring Harbor perspectives in biology.

[33]  Surajit Sinha,et al.  Purmorphamine activates the Hedgehog pathway by targeting Smoothened , 2006, Nature chemical biology.

[34]  K. Storey,et al.  Opposing FGF and retinoid pathways: a signalling switch that controls differentiation and patterning onset in the extending vertebrate body axis , 2004, BioEssays : news and reviews in molecular, cellular and developmental biology.

[35]  J. Kleinjung,et al.  In Vitro Generation of Neuromesodermal Progenitors Reveals Distinct Roles for Wnt Signalling in the Specification of Spinal Cord and Paraxial Mesoderm Identity , 2014, PLoS biology.

[36]  Richard T. Lee,et al.  A low resistance microfluidic system for the creation of stable concentration gradients in a defined 3D microenvironment , 2010, Biomedical microdevices.

[37]  James Briscoe,et al.  Dynamic Assignment and Maintenance of Positional Identity in the Ventral Neural Tube by the Morphogen Sonic Hedgehog , 2010, PLoS biology.

[38]  A. McMahon,et al.  The Morphogen Sonic Hedgehog Is an Axonal Chemoattractant that Collaborates with Netrin-1 in Midline Axon Guidance , 2003, Cell.

[39]  J. Briscoe,et al.  Morphogens and the Control of Cell Proliferation and Patterning in the Spinal Cord , 2007, Cell cycle.

[40]  J. Nickerson,et al.  Diffusion coefficients of retinoids , 2002, Current eye research.

[41]  T. Jessell,et al.  Assigning the Positional Identity of Spinal Motor Neurons Rostrocaudal Patterning of Hox-c Expression by FGFs, Gdf11, and Retinoids , 2001, Neuron.

[42]  T. Jessell,et al.  Retinoid Receptor Signaling in Postmitotic Motor Neurons Regulates Rostrocaudal Positional Identity and Axonal Projection Pattern , 2003, Neuron.

[43]  H. Lodish.  Molecular Cell Biology , 1986 .

[44]  Harvey F. Lodish,et al.  MOLECULAR.CELL.BIOLOGY 5TH.ED , 2003 .

[45]  J. Rossant,et al.  Distinct functions of BMP4 during different stages of mouse ES cell neural commitment. , 2010, Development.

[46]  H. Wichterle,et al.  Directed Differentiation of Embryonic Stem Cells into Motor Neurons , 2002, Cell.

[47]  Natascha Bushati,et al.  Regulation of Neuronal Subtype Identity in the Vertebrate Neural Tube (Neuronal Subtype Identity Regulation) , 2012 .

[48]  Hynek Wichterle,et al.  Functional diversity of ESC-derived motor neuron subtypes revealed through intraspinal transplantation. , 2010, Cell stem cell.