Extended Data Fig. 10: Effect of retinoic acid exposure and inhibition on hTLS.
From: Modelling co-development between the somites and neural tube in human trunk-like structures
(a) Representative brightfield images of 120 h LaminB1::mTagRFP-T hTLS with replacement of RAL with all trans-retinoic acid (ATRA), without retinal (RAL), or with treatment with BMS493 (RA inhibitor) at 72 h (top) and 96 h (bottom). Scale bars, 50 µm. (n = 29-135 per condition) (b) Quantification of hTLS area following RA signalling modulation. Box plots outline the mean (horizontal line), lower and upper quartiles (box hinges), 1.5x interquartile range (whiskers) and outlying datapoints (full circles). * indicates P < 0.05 and **** indicates P < 0.0001 obtained by two-sided Wilcoxon test between control and ATRA (W = 1536, p = 0.1647), No Retinal (W = 2033, p = 0.3842) and BMS-72h (W = 722, p = 3.439e−7), or control and BMS-96h (W = 552, p = 0.03378) (n = 18-127 per condition). (c) Projected images of 120 h LaminB1::mTagRFP-T hTLS HCR-stained following RA signalling pathway modulation. (n = 4-8 per condition). Dashed lines outline hTLS. (d, e) Effect of RA modulation on neurogenesis as assessed by DLL1 + HCR staining in the neural tube of 120 h LaminB1::mTagRFP-T hTLS. (n = 2-7 per condition) (f) Quantification of SOX2 + DLL1+ cells by HCR. * indicates P < 0.05 and ** indicates P < 0.005 obtained by two-sided Wilcoxon test between the control and treatment groups at 72 h (W = 28, p-value = 0.01038) and 96 h (W = 62, p-value = 0.001436; n = 27). (g) Effect of ATRA addition on 120 h LaminB1::mTagRFP-T hTLS, including PAX6 expression. (n = 3-5 per condition). RA, Retinoic acid; ATRA, all-trans Retinoic Acid; RAL, Retinal. Scale bars, 100 µm, unless otherwise stated.
