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. 2020 Jan 31;11:630. doi: 10.1038/s41467-020-14466-y

Fig. 4. Fibrinogen αC domain requirement for NSPC differentiation into astrocytes.

Fig. 4

a Immunoblot for P-Smad1/5/8 and Smad1 in NSPCs pretreated with an increasing concentration of Noggin before fibrinogen stimulation. BMP-2 served as positive control. b Scheme illustrating fibrinogen fractions isolated from plasma. Human Des-αC-fibrinogen lacks most of the residues of the C-terminus of the Aα chain including the RGD sequence. c Immunoblot for P-Smad1/5/8 and total Smad1 protein expression in NSPCs 2 days after fibrinogen or Des-αC-fibrinogen treatment. Values are mean of n = 3 independent experiments. d GFAP + astrocytes in fibrinogen- and Des-αC-fibrinogen-treated NSPC cultures 2 days after initiation of differentiation. Scale bar, 125 μm. Quantification of GFAP + astrocytes derived from fibrinogen- or Des-αC-fibrinogen-treated NSPCs. Results are from four independent experiments (mean ± s.e.m, one-way ANOVA and Bonferroni’s multiple comparisons test, ****P < 0.0001). e Active β1-integrin (green) immunostaining in WT NSPCs treated for 15 min with AF594-fibrinogen (red). Scale bar, 2 μm. Representative images of two independent experiments are shown. f GFAP + astrocytes in untreated and fibrinogen-treated NSPC cultures pretreated with the integrin-blocking RGDS or the control PDEA peptides for 2 days. Scale bar, 60 μm. Quantification of GFAP + astrocytes derived from untreated and fibrinogen-treated NSPCs pretreated with the PDEA (control) and RGDS peptides. Results from three independent experiments performed in duplicates are shown (mean ± s.e.m, one-way ANOVA and Bonferroni’s multiple comparisons test, *P < 0.05, **P < 0.01, ns not significant).