Journal: bioRxiv
Article Title: Dynamic changes in chromosome and nuclear architecture during maturation of normal and ALS C9orf72 motor neurons
doi: 10.1101/2025.09.22.677835
Figure Lengend Snippet: a) Brightfield images of three different cell-types (scale bar=100 μm). b) Schematic view of neural differentiation from IPSCs into motor neuron and motor neuron maturation. IPSCs are treated with listed small molecules for 14 days. At day 14, cells were dissociated and sorted. NCAM+ and EPCAM-cells (population highlighted in red) were collected and plated as immature post-mitotic neurons that were cultured 6 more weeks for maturation (SB-431542: TGF-beta/Smad inhibitor; LDN-193189: BMP pathway inhibitor; RA: retinoic acid; SAG: Smoothened agonist; DAPT: γ-secretase inhibitor; SU-5402: FGFR1 inhibitor; BDNF: brain-derived neurotrophic factor; GDNF: glial-derived neurotrophic factor; CNTF: ciliary neurotrophic factor. c) Schematic view of the study design: primary fibroblasts of two healthy controls and two ALS patients with C9orf72 mutation were reprogrammed into IPSCs. Two independent IPSC clones for each line were picked for neural differentiations. Hi-C and RNA-seq libraries were prepared for each clone. d) TPM counts of PF related genes were shown for PF (red), IPSC (salmon) and MN (blue) cell-types. e) TPM counts of IPSC related genes were shown for PF, IPSC and MN cell-types. f) TPM counts of MN and glial related genes were shown for PF, IPSC and MN cell-types.
Article Snippet: The sorted EpCAM-negative NCAM-positive cells were collected and plated onto Poly-L-lysine coated plates (Sigma, P5899) with motor neuron media (Neurobasal medium, 1x N-2 supplement, 1x B-27 supplement, 1x GlutaMax and 1x MEM NEAA solution) with 10 μM ROCK inhibitor and 10 ng per ml of the following neurotrophic factors: glial cell-derived neurotrophic factor (GDNF) (R&D systems, cat. no. 212-GD-010/CF), brain-derived neurotrophic factor (BDNF) (R&D systems, cat. no. 11166-BD-010) and ciliary neurotrophic factor (CNTF) (R&D systems, cat. no. 257-NT-010/CF).
Techniques: Cell Culture, Derivative Assay, Mutagenesis, Clone Assay, Hi-C, RNA Sequencing
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