Three-dimensional cell culture models have either relied on the self-organizing properties of mammalian cells1,2,3,4,5,6 or used bioengineered constructs to arrange cells in an organ-like configuration7,8. While self-organizing organoids excel at recapitulating early developmental events, bioengineered constructs reproducibly generate desired tissue architectures. Here, we combine these two approaches to reproducibly generate human forebrain tissue while maintaining its self-organizing capacity. We use poly(lactide-co-glycolide) copolymer (PLGA) fiber microfilaments as a floating scaffold to generate elongated embryoid bodies. Microfilament-engineered cerebral organoids (enCORs) display enhanced neuroectoderm formation and improved cortical development. Furthermore, reconstitution of the basement membrane leads to characteristic cortical tissue architecture, including formation of a polarized cortical plate and radial units. Thus, enCORs model the distinctive radial organization of the cerebral cortex and allow for the study of neuronal migration. Our data demonstrate that combining 3D cell culture with bioengineering can increase reproducibility and improve tissue architecture.
Gene Expression Omnibus
We thank members of the Lancaster laboratory for helpful discussion and technical support, especially S. Giandomenico and M. Sutcliffe, as well as A.G. Gianni for sea sponge sample. We also thank members of the Knoblich laboratory for insight and technical help, particularly A. Peer. T.O. was supported by the Cambridge Wellcome Trust PhD program in developmental biology, and F.J.L. is a Wellcome Trust Investigator. M.A.L. was funded by a Marie Curie Postdoctoral fellowship, and work in M.A.L.'s laboratory is supported by the Medical Research Council (MC_UP_1201/9). N.S.C. was funded by an EMBO long-term fellowship and a Deutsche Forschungsgemeinschaft research fellowship (DFG CO 1324/1-1). Work in J.A.K.'s laboratory is supported by the Austrian Academy of Sciences, the Austrian Science Fund (grants I_1281-B19 and Z_153_B09), and an advanced grant from the European Research Council. pCAGEN and pCAG-GFP were gifts from Connie Cepko, Harvard Medical School. pT2/HB was a gift from Perry Hackett, University of Minnesota. pENTR-EGFP2 was a gift from Nathan Lawson, University of Massachusetts Medical School. pCMV(CAT)T7-SB100 was a gift from Zsuzsanna Izsvak, Max Delbruck Center for Molecular Medicine.
Integrated supplementary information
Live imaging of GFP electroporated slice culture of H1 1009 enCOR electroporated with GFP on day 63, followed by vibratome sectioning four 1010 days later and live imaging 24-hours later
Live image of GFP electroporated H9 organoid displaying several labeled radial glial processes and their endfeet (asterisks) as well as several migrating neurons (arrows).
Higher magnification of the multipolar neuron marked by blue arrow in Supplementary Video 2.
Live imaging of farnesyl-GFP electroporated cells in slice culture taken 13 days after electroporation
Higher magnification of live imaging of farnesyl-GFP electroporated slice culture shown in Supplementary Video 4
Higher magnification of the neuron marked by the orange arrow in Supplementary Video 2
Live imaging of spontaneous calcium surges using the calcium dye Fluo-4 in a slice culture taken 13 days after electroporation