Extended Data Fig. 1: Characterization of SARS-CoV-2-exposed cerebral organoids.
From: Brain exposure to SARS-CoV-2 virions perturbs synaptic homeostasis
(a) Three-dimensional imaging of a cerebral organoid differentiated for 45 days stained for neural progenitor (Pax6, green), and neurons (CTIP2, purple). (b–d) Cerebral organoids differentiated for 45 days were mock-infected or infected for 10 days (B) or at indicated time (C-D) with SARS-CoV-2 at 105 pfu/organoid in the presence or absence of 10 μM Remdesivir. RNA extraction and RT-qPCR were performed and the data corresponds to SARS-CoV-2 RNA levels using the N1 primers and normalized by RPL27 (C) from n of 2 organoids per condition in duplicates or to SARS-CoV-2 RNA levels using the sgRNA E primers and normalized by GAPDH (D) from n of 2 organoids per condition in duplicates. Data are presented as mean values +/− SD (C) or +/− SEM (D). (C) Two-tailed p value (Kruskal-Wallis test) was 0.0045 (**) or 0.3268 (ns). (D) Two-tailed p value from paired t test was< 0.0385 (*). (e–i) Cerebral organoids differentiated for 45 days were mock-infected or infected for 10 days with SARS-CoV-2 at 105 pfu/organoid. (E) The organoid supernatant or dissociated organoids were used for titration by plaque assay on Vero E6 cells. The labels pure to -6 correspond to log10 dilution factors. (f) LDH release was measured in the supernatant of the organoids. The data are from a n of two organoids per condition in duplicates and shows no toxicity in any condition, except for Ctrl, a positive control used to account for maximum cytotoxicity. Data are presented as mean values +/− SD. (g) Cerebral organoids were fixed, permeabilized and stained with the anti-cleaved Caspase 3 antibody (orange, a cell death marker, and Dapi (blue). The micrographs show marginal apoptotic staining in both conditions. (h) The organoid size was measured every 2 days for 10 days. Data are presented as mean values +/− SD and each dot corresponds to single organoids with n = 6 organoid per time point. No significant difference in size was found between mock and SARS-CoV-2 infected organoids over time. (i) Cerebral organoids differentiated for 45 days were and stain for Spike and the neuronal marker CTIP2. The micrograph and insets show CTIP2-positive infected cells. The pie chart made from 3 fields of view shows that ≈ 90% of the infected cells were positive for CTIP2.
