Relative
to two-dimensional (2D) culture, three-dimensional (3D)
culture of primary neurons has yielded increasingly physiological
responses from cells. Electrospun nanofiber scaffolds are frequently
used as a 3D biomaterial support for primary neurons in neural tissue
engineering, while hydrophobic surfaces typically induce aggregation
of cells. Poly-l-lactic acid (PLLA) was electrospun as aligned
PLLA nanofiber scaffolds to generate a structure with both qualities.
Primary cortical neurons from E18 Sprague–Dawley rats cultured
on aligned PLLA nanofibers generated 3D clusters of cells that extended
highly aligned, fasciculated neurite bundles within 10 days. These
clusters were viable for 28 days and responsive to AMPA and GABA.
Relative to the 2D culture, the 3D cultures exhibited a more developed
profile; mass spectrometry demonstrated an upregulation of proteins
involved in cortical lamination, polarization, and axon fasciculation
and a downregulation of immature neuronal markers. The use of artificial
neural network inference suggests that the increased formation of
synapses may drive the increase in development that is observed for
the 3D cell clusters. This research suggests that aligned PLLA nanofibers
may be highly useful for generating advanced 3D cell cultures for
high-throughput systems.
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