Fig. 4.

The effect of rPrP and its fragments on the cytotoxicity of Aβ oligomers in rat primary hippocampal neurons. (A) The efficiency of the knockdown of PrPC in primary neurons after 72 h treatment with Prnp siRNA as evaluated by Western blot analysis using 3F10 antibody (for prion protein) and C4 antibody (for actin). Neurons treated with scrambled siRNA were used as controls. (B and C) Primary neurons (pretreated in the presence or absence of Prnp siRNA) were treated with Aβ oligomers alone and two different preparations of Aβ mixtures with rPrP or its fragments N1 and rPrP90-231. Data shown in panel B were obtained using Aβ mixtures with rPrP or its fragments prepared by adding these proteins to freshly disaggregated Aβ and incubating the samples for 3 h at 37 °C , as described in Preparation of Aβ Oligomers for Preparation A. Data shown in panel C represent experiments using Aβ mixtures with rPrP and its fragments prepared by adding rPrP, N1 or rPrP90-231 to Aβ oligomers that were preformed in the absence of these proteins (Preparation B described in Preparation of Aβ Oligomers). The final concentration of Aβ in each case was 500 nM, and the molar ratio of rPrP or its fragments to Aβ was 1:20. The cytotoxic effect was assessed by measuring LDH release. Mean values are based on at least five independent experiments; error bars represent SEM. ***, p<0.001; **, p<0.01; *, p<0.05.