Blocking the apolipoprotein E/amyloid-β interaction as a potential therapeutic approach for Alzheimer's disease

1. 

   View larger version:

   • In this window
   • In a new window

   • Download as PowerPoint Slide

   Fig. 1.

   Aβ12-28P binds to apoE and abolishes its effect on Aβ fibrillization. (a) Shown is a solid-phase binding assay of lipidated human apoE4 isoform to Aβ1-40, Aβ12-28P, and Aβ12-28 synthesized from l-amino acids. K _D values represent mean ± SEM from three independent experiments. (b) Shown is dose-dependent inhibition of the apoE4/Aβ binding by increasing concentrations of Aβ12-28. P values represent mean ± SEM from three independent experiments. (c) Thioflavin-T aggregation assay demonstrates the effect of Aβ12-28P on the apoE/Aβ interaction. Adding the human lipidated apoE4 complexes dramatically increases the amount of Aβ1-40 fibrils formed over time (P = 0.007, repeated measures ANOVA; P < 0.01 for specific post hoc comparison of Aβ1-40 + apoE4 versus Aβ1-40 alone). Preincubation of apoE with Aβ12-28P abolishes the apoE effect on Aβ1-40 aggregation (P < 0.01 and nonsignificant post hoc analysis for the specific effect of Aβ+apoE/Aβ12-28P versus Aβ + apoE and Aβ, respectively). Aβ12-28P alone has no effect on the aggregation of Aβ1-40 (nonsignificant). Aβ12-28P does not aggregate over time. (d) Shown is a lack of direct effect of Aβ12-28P on Aβ1-40 aggregation. Aβ1-40 (200 μmol/liter) was incubated in the presence of Aβ12-28P concentrations ranging from 0 to 200 μmol/liter (repeated measures ANOVA P = 0.573).

   
   
2. 

   View larger version:

   • In this window
   • In a new window

   • Download as PowerPoint Slide

   Fig. 2.

   Treatment with Aβ12-28P rescues APP_K670N/M671L mice from memory decline. Aβ12-28P-treated APP_K670N/M671L mice performed comparably to WT, age- and sex-matched littermates on radial arm maze testing. Both groups performed statistically better than APP_K670N/M671L mice treated with vehicle; ANOVA P < 0.0001, post hoc Aβ12-28P vs. WT nonsignificant, Aβ12-28P vs. vehicle P < 0.001, WT vs. vehicle P < 0.001 (n = 11 for vehicle and Aβ12-28P-treated Tg groups, n = 12 for WT).

   
   
3. 

   View larger version:

   • In this window
   • In a new window

   • Download as PowerPoint Slide

   Fig. 3.

   Treatment with Aβ12-28P reduces Aβ deposition in APP_K670N/M671L and APP_K670N/M671L/PS1_M146L mice. (a) Decrease in the total Aβ burden in the neocortex, the cingulated cortex, and the hippocampus as quantified by unbiased hierarchical sampling (n = 11; ∗, P < 0.01; ∗∗, P < 0.001). (b) Hemispheric sections from 7-month-old APP_K670N/M671L/PS1_M146L mice treated with vehicle (Left) and Aβ12-28P (Right) depict the difference in Aβ burden. Immunostaining was done with a mixture of 4G8 and 6E10 anti-Aβ mAbs. (c) Shown is a reduction in the fibrillar Aβ burden in Aβ12-28P-treated animals (n = 11; ∗, P < 0.05). (d) Shown is the cingulate cortex and the neocortex of 18-month-old vehicle-treated (Left) and Aβ12-28P-treated (Right) APP_K670N/M671L mice stained with Thioflavin-S. There is a clearly visible reduction in the burden of parenchymal Aβ deposits (yellow arrowhead) and CAA (white arrowhead). Both types of deposits were quantified separately. (e) Shown is a decrease in the CAA burden in Aβ12-28P-treated APP_K670N/M671L mice (∗, P < 0.05). (f) Shown is Perls staining of parenchymal vessels in Aβ12-28P-treated APP_K670N/M671L mice, revealing a lack of microhemorrhages.

   
   
4. 

   View larger version:

   • In this window
   • In a new window

   • Download as PowerPoint Slide

   Fig. 4.

   Treatment with Aβ12-28P reduces the amount of apoE present in Aβ deposits. (a) Shown is double immunofluorescent staining colocalizating apoE in Aβ deposits in the hippocampus of APP_K670N/M671L mice. Only a minority of plaques were apoE-negative in both groups (see white arrowheads). (b) Shown is a reduction in the burden of apoE-positive deposits in Aβ12-28P-treated animals (∗, P < 0.01; ∗∗, P < 0.001). Values are averaged for all three areas of interest. (c) Shown is the reduction in the mean optic density (O.D.) index of apoE deposits in Aβ12-28P-treated animals (∗, P < 0.05).

   
   
5. 

   View larger version:

   • In this window
   • In a new window

   • Download as PowerPoint Slide

   Fig. 5.

   Treatment with Aβ12-28P decreases the levels of total Aβ40 and Aβ42 but does not alter levels of the soluble Aβ fraction or Aβ oligomers. (a) Shown is a statistically significant decrease in the levels of FA-extracted (FA_extr) Aβ40 and Aβ42 (total Aβ) in APP_K670N/M671L mice (Left) and APP_K670N/M671L/PS1_M146L mice (Right). The level of soluble Aβ40 and Aβ42 fractions extracted with DEA (DEA_extr) did not differ between groups (n = 11; ∗, P < 0.05; ns, not significant). (b) A Western blot of brain homogenates stained with A11 oligomer-specific polyclonal antibody. The density and thickness of oligomer bands did not differ between vehicle-and Aβ12-28P-treated 18-month-old APP_K670N/M671L mice. No oligomers were detected in age-matched WT littermates. (c) The densitometric analysis of oligomer-specific bands. There is no significant difference between Aβ12-28P- and vehicle-treated groups in either Tg model.