Fig. 5

Ablation of ctgfa⁺ cells during SC regeneration. (A) Experimental timeline to establish a genetic ablation model for ctgfa⁺ cells after SCI. Transgenic ctgfa:mCherry-2A-NTR animals and control siblings were subjected to complete SC transection and metronidazole (MTZ) treatment at 4 and 7 dpi to ablate ctgfa⁺ cells. TUNEL staining, PCNA staining and ctgfa in situ hybridization were performed to evaluate the extent of cell death and cell proliferation at 9 dpi. (B,C) TUNEL staining in ctgfa:mCherry-2A-NTR animals (Tg⁺) 9 dpi. Tissue sections from vehicle- and MTZ-treated animals are shown. Representative micrographs shown in B are 150 µm rostral to the lesion. The proportion of mCherry⁺ TUNEL⁺ cells was normalized to total nuclei number and shown in C. (D,E) PCNA staining in ctgfa:mCherry-2A-NTR animals (Tg⁺) at 9 dpi. Tissue sections from vehicle- and MTZ-treated animals are shown. Representative micrographs shown in D are 450 µm rostral to the lesion. The proportion of mCherry⁺ PCNA⁺ cells was normalized to total nuclei number and shown in E. In B,D, arrowheads in high-magnification insets indicate mCherry⁺ TUNEL⁺ and mCherry⁺ PCNA⁺ cells in triple-, double- or single-channel views. (F,G) ctgfa in situ hybridization in ctgfa:mCherry-2A-NTR animals (Tg⁺) at 9 dpi. Tissue sections from vehicle- and MTZ-treated animals are shown. Representative micrographs shown in F are 450 µm rostral to the lesion. Dashed red lines delineate central canal edges. Quantification of ctgfa⁺ expression domains is shown in G. For all quantifications, dots represent individual animals and animal numbers are indicated in parentheses. Error bars represent s.e.m. **P<0.01; *P<0.05 (unpaired t-test with Welch's correction). Scale bars: 100 µm.