Figure 3

The spontaneous recovery of brain injury after ICH is associated with increased nestin expression. (A) Kaplan Meier analysis shows no difference in survival between WT, ICH− and ICH+ larvae at 21 dpf (93% ICH− and WT, 92% ICH+, N = 35 larvae per group at 5 dpf). (B) Representative images of annexinV expression in an ICH+ larvae at 2, 3, 4 and 5 dpf show apoptotic cells within the head during early development. A dense cluster of dying cells associated with ICH (denoted by white dotted line) at 3 dpf (24 h post-ICH) is resolved by 4 dpf. Scale bar = 100 µm. (C) Representative images of nestin positive larvae from both ICH− and ICH+ groups at 2–5 dpf. Global expression of nestin in the developing larval brain is lost over time with the greatest reduction in expression between 2 and 3 dpf. Overall, ICH+ larvae show a greater intensity of nestin expression. Yellow arrow denotes the absence of nestin expression between hemispheres and white arrows denote regions of intense expression of interest. Scale bar = 100 µm (n = 20, n = 12 at 5 dpf). (D) Quantification of the intensity fluorescence of nestin signal in the brain (n = 12–20). Data were analysed using a one-way ANOVA, **P = 0.0063, ***P = 0.0003, F(3,71) = 19.12. (E) Comparison of nestin expression in the brain at 3 and 4 dpf between ICH− and ICH+ larvae. Data were analysed using a one-way ANOVA ****P < 0.0001, F(3, 110) = 100.9. (F) Paired 3 and 4 dpf data show no difference in brain region of nestin expression. ICH−, intracerebral haemorrhage negative; ICH+, intracerebral haemorrhage positive; WT, wildtype; dpf, days post-fertilisation; ns, non-significant; ALLG, anterior lateral line ganglion; MHB, mid-hindbrain boundary.