Summary
Cerebral tracts connect separated regions within a brain and serve as fundamental structures that support integrative brain functions. However, understanding the mechanisms of cerebral tract development, macro-circuit formation, and related disorders has been hampered by the lack of an
in vitro
model. Here, we developed a human stem cell-derived model of cerebral tracts, which is composed of two spheroids of cortical neurons and a robust fascicle of axons linking these spheroids reciprocally. In a microdevice, two spheroids of cerebral neurons extended axons into a microchannel between the spheroids and spontaneously formed an axon fascicle, mimicking a cerebral tract. We found that the formation of axon fascicle was significantly promoted when two spheroids extended axons toward each other compared with axons extended from only one spheroid. The two spheroids were able to communicate electrically through the axon fascicle. This model tissue could facilitate studies of cerebral tract development and diseases.
Versatile
methods for patterning multiple types of cells with single-cell
resolution have become an increasingly important technology for cell
analysis, cell-based device construction, and tissue engineering.
Here, we present a photoactivatable material based on poly(ethylene
glycol) (PEG)-lipids for patterning a variety of cells, regardless
of their adhesion abilities. In this study, PEG-lipids bearing dual
fatty acid chains were first shown to perfectly suppress cell anchoring
on their coated substrate surfaces whereas those with single-chain
lipids stably anchored cells through lipid–cell membrane interactions.
From this finding, a PEG-lipid with one each of both normal and photocleavable
fatty acid chains was synthesized as a material that could convert
the chain number from two to one by exposure to light. On the photoconvertible
PEG-lipid surface, cell anchoring was activated by light exposure.
High-speed atomic force microscopy measurements revealed that this
photocaging of the lipid–cell membrane interaction occurs because
the hydrophobic dual chains self-assemble into nanoscale structures
and cooperatively inhibit the anchoring. Light-induced dissociation
of the lipid assembly achieved the light-guided fine patterning of
multiple cells through local photoactivation of the anchoring interactions.
Using this surface, human natural killer cells and leukemia cells
could be positioned to interact one-by-one. The cytotoxic capacity
of single immune cells was then monitored via microscopy, showing
the proof-of-principle for applications in the high-throughput analysis
of the heterogeneity in individual cell–cell communications.
Thus, the substrate coated with our photoactivatable material can
serve as a versatile platform for the accurate and rapid patterning
of multiple-element cells for intercellular communication-based diagnostics.
Molecular networks on the cytoplasmic faces of cellular plasma membranes are critical research topics in biological sciences and medicinal chemistry. However, the selective permeability of the cell membrane restricts the researchers from accessing to the intact intracellular factors on the membrane from the outside. Here, a microfluidic method to prepare cell membrane sheets was developed as a promising tool for direct examination of the cytoplasmic faces of cell membranes. Mammalian cells immobilized on a poly(ethylene glycol)-lipid coated substrate were rapidly and efficiently fractured, with the sheer stress of laminar flow in microchannels, resulting in isolation of the bottom cell membrane sheets with exposed intact cytoplasmic faces. On these faces of the cell membrane sheets, both ligand-induced phosphorylation of receptor tyrosine kinases and selective enzymatic modification of a G-protein coupling receptor were directly observed. Thus, the present cell membrane sheet should serve as a unique platform for studies providing new insights into juxta-membrane molecular networks and drug discovery.
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