Fig 3. Evaluation of AAV-αAβ IgG neuronal expression and neurotoxicity.

A) LCMSMS analysis showing detected peptides from huIgG heavy and light chain from hemibrain lysates of SCID mice injected with AAV-αAβ IgG compared to animals injected with PBS (Sham), or Sham brain homogenate spiked with equivalent levels of huIgG as in the AAV-αAβ IgG group. Graphs at right show quantification of functional huIgG compared to total huIgG expressed in SCID mice either centrally (2E10 total GC injected into hippocampus) or peripherally (1E12 total GC injected IV). Levels of functional Aβ antibody in brain extracts from AAV-αAβ IgG expressing SCID mice were quantified by antigen (Ag) ELISA and compared in parallel with a pan-huIgG ELISA (IgG). Levels of mean huIgG bound to antigen accounted for only 21% of mean total huIgG when expressed from the brain whereas this discrepancy is not detected in sera one month following peripheral expression of the vector. Data are presented as mean +/- SEM. **p<0.01, unpaired Students t-test. B) H&E staining of C57BL/6 mouse brain hippocampus following intra-hippocampal injection of the indicated vectors compared to PBS control. H&E staining revealed neuronal, eosinophilic to hyaline-like inclusions reminiscent of glycoprotein accumulation only in brains injected with the antibody expression vectors, while PBS and AAV-Empty vector injected animals did not have this pathology. Inset shows detail, arrows point to representative hyaline inclusions. Scale bar = 100μm. Results are summarized in the table at right as the number of animals scored with or without this pathology. C) Quantitative IHC for GFAP+ area. C57BL/6 mice were injected with either PBS or AAV-αAβ msIgG (2E10 GC into hippocampus), and 5μm sagittal brain sections were collected 16 weeks later. Each circle represents one mouse. Bars indicate group mean +/- SEM of GFAP+ area normalized to PBS. ***p<0.001, unpaired Students t-test, n = 8 mice per group.