Fig. 2: Blocking CV2/CRMP5-Abs with epitope peptides prevents the sensitization of sensory neurons and mechanical hypersensitivity in rats.

A Heatmap of the immunoreactivity of CV2/CRMP5-Abs sera hybridized on a CRMP5 peptide array mapping the entire sequence of the protein in 15-mer peptides with three amino acid increments. Four main epitopes on CRMP5 are targeted by the CV2/CRMP5-Abs. B Bar graph with scatter plot showing that peptides 53, 142, and 146 can block the binding of CV2/CRMP5-Abs from patients #8 and #4 to purified CRMP5. 0.03% DMSO is the vehicle, purified CRMP5 was used as a positive control to achieve maximal displacement of the CV2/CRMP5-Abs. n = 3 independent measures from an average of three repeats. *p < 0.05, one-way ANOVA. C Micrograph of a rat DRG immunolabelled with CV2/CRMP5-Abs positive serum (1/100) from patient #1 and then with blocking peptides 53, 142, 146. Blocking peptides abolished the immunoreactivity of CV2/CRMP5-Abs for their protein target CRMP5. The experiment was repeated across five rats with consistent results. D Representative recordings of evoked action potentials recorded from small-diameter DRG neurons treated with serum from patient #1 (1/100 dilution) in combination with 100 ng/ml of peptides 53, 142, and 146 as indicated, overnight in response to depolarizing current injection of 30, 60, and 90 pA. E Quantification of the number of evoked action potentials in response to 0–100 pA of injected current. *p < 0.05, multiple Mann–Whitney tests. DMSO n = 15 cells, Patient #1 n = 15 cells, DMSO + peptides n = 14 cells, Patient #1 + peptides n = 16 cells. F Representative traces of rheobase recordings from cells treated with the serum from patient #1 and blocking peptides as indicated. G Bar graph with scatter plot showing unchanged rheobase in cells treated with the serum from patient #1 and with blocking peptides as indicated. *p < 0.05, Kruskal–Wallis test. DMSO n = 15 cells, Patient #1 n = 14 cells, Patient #1 + peptides n = 16 cells from at least three rats. H Graph showing the paw withdrawal threshold of rats injected with 15 µl of the indicated CV2/CRMP5-Abs positive sera (1/10 dilution). I Graph showing the paw withdrawal threshold of rats injected with CV2/CRMP5-Abs positive sera with blocking peptides 53, 142, and 146 (300 ng/ml). *p < 0.05, two-way ANOVA. J Bar graph with scatter plot showing the area under the curve of the data in (H, I) and color coded per treatment groups (n = 3 rats each), *p < 0.05, Mann–Whitney. All data are shown with error bars that indicate mean ± SEM. Experimenters were blind to the treatment groups. See Supplementary Data 1 for additional statistical details. Source data are provided as a Source Data file.