Fig. 1: DV patterned µSCLS.
From: A controllable human spinal cord model with full dorsoventral patterning
a Schematic of embryonic spinal cord being patterned along the DV axis. b Schematic of microfluidic device with µSCLS embedded in the microfluidic channel. c Protocol for generation of µSCLS. d Representative brightfield images showing development of µSCLS. e Bar plots showing tissue length (left) and projected area (right) of µSCLS from day 2 to 12. nµSCLS = 54. f Representative stitched confocal micrographs showing µSCLS on day 12 immunostained for indicated HOX genes. Similar observations were made in n = 3 independent experiments. g Representative stitched confocal micrograph showing DV patterned µSCLS on day 12 immunostained for indicated markers. Zoom-in views of boxed regions are shown on the right. Arrowheads mark a subset of cells co-expressing PAX3 and PAX6. Intensity maps show relative mean expression levels of indicated markers as a function of relative DV position in µSCLS on day 12. To generate expression heatmaps, nPAX3 = 33, nPAX6 = 33, nOLIG2 = 53 tissues from n = 3 experiments. h Representative confocal micrographs showing NCCs (box I), RP (box II), and FP (box III) in µSCLS on day 12. i Bar plot of patterning efficiencies of µSCLS from three different hPSC lines combined. Different symbols represent each line. n = 3 independent experiments for each cell line. j Representative brightfield image showing neurite-like processes in µSCLS on day 12. Arrowheads mark subset of such processes. Similar observations were made in n = 3 independent experiments. k Representative confocal micrographs showing neurites expressing βIII-TUBULIN. Arrowheads mark subset of neurites. Similar observations were made in n = 3 independent experiments. l Representative confocal micrographs of µSCLS on day 12 immunostained for indicated lineage markers. Top panels show neural crest region, while bottom panels show pMN domain. Green arrowheads mark subset of OLIG2−HB9−ISL1+ sensory neurons. Orange arrowheads mark subset of OLIG2−HB9+ISL1+ motor neuron. White arrowheads mark subset of OLIG2+HB9+ISL1+ early motor neurons. In (e and i), error bars represent mean ± s.d. Scale bars, 500 µm (d, g), 200 µm (j, k), 50 µm (h, l).
