Patterned cell culture inside microfluidic devices

Rhee, Seog Woo; Taylor, Anne Marion; Tu, Christina H.; Cribbs, David H.; Cotman, Carl W.; Jeon, Noo Li

doi:10.1039/b403091e

Cited by 260 publications

(206 citation statements)

References 29 publications

(40 reference statements)

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“…The high fluidic resistance of the microgrooves produces a small but sustained flow between the compartments that counteracts diffusion 23,24 . To demonstrate fluidic integrity of the compartments, we incubated Texas red dextran (3,000 Da; 20 μM) in the axonal compartment for over 20 h. We detected no trace of fluorescence within the somal compartment by either UV spectroscopy (equivalent to background) or confocal microscopy (Fig.…”

Section: Fluidic Isolation Within the Axonal Compartmentmentioning

confidence: 99%

A microfluidic culture platform for CNS axonal injury, regeneration and transport

et al. 2005

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Investigation of axonal biology in the central nervous system (CNS) is hindered by a lack of an appropriate in vitro method to probe axons independently from cell bodies. Here we describe a microfluidic culture platform that polarizes the growth of CNS axons into a fluidically isolated environment without the use of targeting neurotrophins. In addition to its compatibility with live cell imaging, the platform can be used to (i) isolate CNS axons without somata or dendrites, facilitating biochemical analyses of pure axonal fractions and (ii) localize physical and chemical treatments to axons or somata. We report the first evidence that presynaptic (Syp) but not postsynaptic (Camk2a) mRNA is localized to developing rat cortical and hippocampal axons. The platform also serves as a straightforward, reproducible method to model CNS axonal injury and regeneration. The results presented here demonstrate several experimental paradigms using the microfluidic platform, which can greatly facilitate future studies in axonal biology.Neurons in the CNS extend axons over considerable distances and through varying extracellular microenvironments to form synapses, the basis of neuronal connectivity. Axonal damage is critical to the etiology of CNS injuries and neurodegenerative disease (for example, spinal cord injury and Alzheimer disease) 1-3 ; therefore, considerable effort focuses on the molecular and cellular mechanisms that influence axonal plasticity and response to injury. In vitro models facilitate the study of axonal biology in the peripheral nervous system (PNS), but no suitable method has been developed for the study of the CNS because of the challenges associated with culturing CNS neurons.In vitro studies using compartmentalized 'Campenot' chambers have greatly improved the understanding of axonal biology within the PNS 4-6 . Campenot chambers use a compartmented Teflon divider attached to a collagen-coated petri dish via a thinly applied silicone grease layer; typically nerve growth factor (NGF) promotes neuritic growth through the grease layer. Much of the work involving Campenot chambers focused on the influence and transport of NGF.

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Section: Fluidic Isolation Within the Axonal Compartmentmentioning

confidence: 99%

A microfluidic culture platform for CNS axonal injury, regeneration and transport

et al. 2005

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“…Microcontact printing of biomaterial patterns prior to enclosure within PDMS microchannels has previously been used. 25,26 However, progress with in chip biomaterial patterning is in its infancy, and is hampered by aggressive bonding conditions which damage biomaterials, the incompatibility of photoresists and processing conditions with biomolecules, and the requirement to align the biomaterial pattern with the microfluidic structures. This is especially problematic with elastomeric PDMS microstructures.…”

Section: Introductionmentioning

confidence: 99%

Microfluidic construction of minimalistic neuronal co-cultures

et al. 2013

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In this paper we present compartmentalized neuron arraying (CNA) microfluidic circuits for the preparation of neuronal networks using minimal cellular inputs (10-100-fold less than existing systems).The approach combines the benefits of microfluidics for precision single cell handling with biomaterial patterning for the long term maintenance of neuronal arrangements. A differential flow principle was used for cell metering and loading along linear arrays. An innovative water masking technique was developed for the inclusion of aligned biomaterial patterns within the microfluidic environment. For patterning primary neurons the technique involved the use of meniscus-pinning micropillars to align a water mask for plasma stencilling a poly-amine coating. The approach was extended for patterning the human SH-SY5Y neuroblastoma cell line using a poly(ethylene glycol) (PEG) back-fill and for dopaminergic LUHMES neuronal precursors by the further addition of a fibronectin coating. The patterning efficiency E patt was .75% during lengthy in chip culture, with y85% of the outgrowth channels occupied by neurites. Neurons were also cultured in next generation circuits which enable neurite guidance into all outgrowth channels for the formation of extensive inter-compartment networks. Fluidic isolation protocols were developed for the rapid and sustained treatment of the different cellular and sub-cellular compartments. In summary, this research demonstrates widely applicable microfluidic methods for the construction of compartmentalized brain models with single cell precision. These minimalistic ex vivo tissue constructs pave the way for high throughput experimentation to gain deeper insights into pathological processes such as Alzheimer and Parkinson Diseases, as well as neuronal development and function in health.

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“…26,27 Figure 1 shows a schematic view of the experimental setup including controlled axonal outgrowth and fluidic isolation property. Because of the difference (30-fold) in height between the multicompartmental channels (100 μm high) and the microgrooves (3 μm high), high fluidic resistance was achieved and one could apply drugs only at a local region of neurons, for example, proximal, distal axon, or growth cones.…”

Section: ■ Results and Discussionmentioning

confidence: 99%

Quantitative Analysis of Axonal Transport by Using Compartmentalized and Surface Micropatterned Culture of Neurons

Kim

Park

Byun

et al. 2012

ACS Chem. Neurosci.

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Mitochondria, synaptic vesicles, and other cytoplasmic constituents have to travel long distance along the axons from cell bodies to nerve terminals. Interruption of this axonal transport may contribute to many neurodegenerative diseases including Alzheimer's disease (AD). It has been recently shown that exposure of cultured neurons to β-amyloid (Aβ) resulted in severe impairment of mitochondrial transport. This Letter describes an integrated microfluidic platform that establishes surface patterned and compartmentalized culture of neurons for studying the effect of Aβ on mitochondria trafficking in full length of axons. We have successfully quantified the trafficking of fluorescently labeled mitochondria in distal and proximal axons using image processing. Selective treatment of Aβ in the somal or axonal compartments resulted in considerable decrease in mitochondria movement in a location dependent manner such that mitochondria trafficking slowed down more significantly proximal to the location of Aβ exposure. Furthermore, this result suggests a promising application of microfluidic technology for investigating the dysfunction of axonal transport related to neurodegenerative diseases.

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Patterned cell culture inside microfluidic devices

Cited by 260 publications

References 29 publications

A microfluidic culture platform for CNS axonal injury, regeneration and transport

A microfluidic culture platform for CNS axonal injury, regeneration and transport

Microfluidic construction of minimalistic neuronal co-cultures

Quantitative Analysis of Axonal Transport by Using Compartmentalized and Surface Micropatterned Culture of Neurons

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