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. 2009 Dec 1;20(23):4941–4950. doi: 10.1091/mbc.E09-03-0200

Figure 6.

Figure 6.

The percentage of Nestin+ NPCs is increased by overexpression of RhoG-V12 but is not affected by knockdown of RhoG. (A) Cells electroporated with control, RhoG-WT, or RhoG-V12 vector at E13 were dispersed at E14 and stained with anti-Nestin (red), anti-GFP (green), and Hoechst 33258 (blue) after 48-h culture. Arrows indicate the Nestin+ NPC clusters. Scale bar, 50 μm. (B) Twenty-four hours after control, RhoG-WT, or RhoG-V12 vector was electroporated at E13, the regions of cortices containing EYFP+ cells were dissociated and dispersed on PDL-coated coverslips. Cells were fixed 3, 24, or 48 h after plating and stained with anti-Nestin antibody. The numbers of Nestin+EYFP+ and EYFP+ cells were counted, and the percentages of Nestin+ cells in the total number of EYFP+ cells were calculated. Control, n = 9; RhoG-WT, n = 5; RhoG-V12, n = 6. Values indicate means ± SEM; *p < 0.05 (Student's t test). (C) Cells electroporated with control, RhoG-WT, or RhoG-V12 vector at E13 were dispersed at E14 and stained with anti-Tuj1 (red), anti-GFP (green), and Hoechst 33258 (blue) after 48-h culture. Arrowheads indicate the Tuj1− cell clusters. Scale bar, 50 μm. (D) Cells were cultured and fixed as described in B and then stained with anti-Tuj1 antibody. The numbers of Tuj1+EYFP+ and EYFP+ cells were counted, and the percentages of Tuj1+ cells in the total number of EYFP+ cells were calculated. Control, n = 3; RhoG-WT, n = 3; RhoG-V12, n = 6. Values indicate means ± SEM; *p < 0.05 (Student's t test). (E) shLuc, shRG-A, or -B vector was electroporated at E13, and the percentages of Nestin+ cells were quantified as described in B. shLuc, n = 6; shRG-A, n = 5; shRG-B, n = 5. Values indicate means ± SEM. (F) shLuc, shRG-A, or -B vector was electroporated at E13, and the percentages of Tuj1+ cells were quantified as described in D. shLuc, n = 6; shRG-A, n = 6; shRG-B, n = 3. Values indicate means ± SEM. (G) shLuc, shRG-A, or -B vector was electroporated at E14, and brains were labeled with BrdU at E15 and fixed at E16. Cell cycle exit index indicate the percentages of EYFP+BrdU+Ki67− cells in EYFP+BrdU+ cells. shLuc, n = 3; shRG-A, n = 3; shRG-B, n = 3. Values indicate means ± SEM.