Fig. 1: scRNA-seq analysis unveils a GBM radiotherapy-induced cell state transition.
From: VC-resist glioblastoma cell state: vessel co-option as a key driver of chemoradiation resistance
A Cell state heterogeneity in MGG4 gliomaspheres: astrocyte (AC)-like, oligodendrocyte progenitor cells (OPC)-like, mesenchymal (MES)-like and neural progenitor cells (NPC)-like. Schematics created with BioRender.com. B UMAP dimensionality reduction plot of scRNA-seq for MGG4 gliospheres exposed to 5 Gy IR or not, at day3 or 5 of culture (n = 2 independent experiments). C Effect of IR on each cluster (p = 0,0005; Chi-square). D, E RNA velocity and trajectory analysis of irradiated or naïve MGG4 cells at day5. F Expression of CL3 geneset in the Neftel’ representation of cell states. GBM cell positions indicate relative scores for meta-modules, with colors reflecting CL3 geneset expression. Violin and box-and-whiskers plot (Tukey) from 28 GBM patients produced with https://singlecell.broadinstitute.org. G Feature plot for CL3 signature in the harmonized database with over 1 M GBM cells. H Feature plot of Nestin expression. I Enrichment of NesHI cell population upon IR (2 and 5 Gy) in NestinP-dTomato MGG4 cells analyzed by real-time microscopy. Data are means ± SEM (n = 3 independent experiments, technical duplicates per experiment; p < 0.0001; one-way ANOVA, Turkey’s multiple comparisons test). J Enrichment of NesHI cell population upon IR (2, 5, 8, 10, 12 Gy) analyzed at day5 by FACS in NestinP-dTomato MGG4 and MGG18 cells. Data are means ± SEM (MGG4 n = 4; MGG18 n = 3; ns, non-significant; *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001; one-way ANOVA, Turkey’s multiple comparisons test). K Spearman correlation analysis of basal Nestin expression and fold change (FC) of Nestin expression after irradiation (5 Gy) in 10 GBM cell lines. X-axis: 1/basal CT values for Nestin expression determined by RT-PCR; Y-axis: Nestin FC after IR determined by RT-PCR. Patient-derived (blue) mouse (green) cell lines (n = 3 per cell line, R = 0,88; p = 0,001, Spearman test). L Time-lapse micrographs of FACS-sorted MGG4 NesLO cells showing Nestin reporter activation (arrows). M, N Enrichment of MGG4 NesHI cell population overtime upon IR (5 Gy) or not (Naive) in FACS-sorted MGG4 NesLO cells analyzed by real-time microscopy and FACS. Data are means ± SEM (n = 3 biologically independent experiments, p < 0.0001, Pearson test). O Enrichment of NesHI cell population under IR (5 Gy) in FACS-sorted MGG18 and GL261 NesLO cells analyzed at day3 by FACS. Data are means ± SEM (MGG18 n = 4; GL261 n = 3; ***p < 0.001; ****p < 0.0001; unpaired two-sided t test).
