Supplementary Figure 13: Extended data from Fig. 7. mAb 10.5.6 neutralizes SFRP1 activity.

Cultured cells release into the culture medium the sAPPα peptide derived from ADAM10-mediated proteolytic processing of APP ¹⁶. Sfrp1 binds to ADAM10 and prevents APP processing thus decreasing the amount of sAPPα in the media of cultured cells ¹⁴. A) The panels show Western blot analysis of media from wt and Sfrp1^-/- cultures immunoprecipitated with anti-Aβ (6E10) antibody and analyzed with the same antibody. Telencephalic neuroepithelial cells express high level of Sfrp1 mRNA ¹⁵ therefore cultured telencephalic cells from E13.5 wt embryos release undetectable levels sAPPα into the medium because the activity of ADAM10, a ubiquitously expressed protein, is inhibited. Sfrp1^-/- telencephalic cultures instead release detectable sAPPα levels (compare control lanes in the two cultures) because ADAM10 is free to process APP. The addition of the 10.5.6 IgG1 against Sfrp1, but not that of an unspecific IgG of the same isotype to the medium of wt cultures enhanced sAPPα release, indicating its ability of neutralizing Sfrp1 function. No changes in the amount of sAPPα were instead observed in the Sfrp1^-/- cultures. B) Quantification of the changes observed in three independent experiments performed in the same conditions as those illustrated in A. Data are means ± SEM analyzed with two-sided Student-t test; α-SFRP1 vs C P=0.00047; α-SFRP1 vs IgG1 P=0.0058., **p≤0.01; ***p≤0.001; n.s. not statistically significant.