176 Chapter 6 of the ROI in G. Each yellow dot represents one counted axon in the main fibre direction and a green dot in the perpendicular fibre direction. Data on the average axon counts are displayed for each donor in the graph, and group results are expressed as mean ± standard error of the mean (SEM). * corresponds to a q-value < 0.05 and ** to a q-value < 0.01 *** to a q-value < 0.001. The scale bars represent two mm for A, D and G, and 100 µm for B, C, E, F, H. Non-significantly lower axonal density colour intensity (in the Bielschowsky silver staining) was seen in narcolepsy type 1 compared to controls (Table 4). Post-hoc analyses per region showed no significantly lower colour intensity after correction for multiple comparisons (Appendix A, Supplementary Table 2). No differences in axonal morphology became apparent when qualitatively inspecting the axons in the different sections with the Bielschowsky silver staining. Table 4: Colour intensity and axonal injury area percentage stained Staining β Standard error 95% CI t-value p-value Effect size Axonal density (Bielschowsky silver staining colour intensity) -0.02 0.02 -0.06, 0.03 -0.83 0.426 0.20 Axonal injury (SMI312 area percentage stained) 0.60 0.59 -0.67, 1.86 1.02 0.331 0.20 Myelin integrity (PLP colour intensity) -0.001 0.02 -0.03, 0.03 -0.10 0.924 0.02 Myelin integrity (LFB colour intensity) 0.002 0.03 -0.07, 0.07 0.06 0.955 0.02 PLP = proteolipid protein; LFB = luxol fast blue. Axonal injury No significant differences were found in the SMI312 axonal injury staining in narcolepsy type 1 compared to controls in the overall analysis (Table 4), and in the post-hoc per ROI analyses (Appendix A, Supplementary Table 2). Myelin integrity The PLP and LFB stainings investigating myelin integrity showed no significant differences between groups. Post-hoc per ROI analyses only showed significantly lower PLP colour intensity (-9.6% colour intensity; p = 0.001, q = 0.009) in the secondary visual cortex (V2) in narcolepsy type 1 compared
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