Journal: The Journal of neuroscience : the official journal of the Society for Neuroscience

Article Title: Soluble A? seeds are potent inducers of cerebral ?-amyloid deposition

doi: 10.1523/JNEUROSCI.3088-11.2011

Figure Lengend Snippet: Brain extracts from an aged APP23 tg mouse (Tg extract), an aged non-tg mouse (Wt extract) spiked with synthetic Aβ fibrils (syn. Aβ fibrils in Wt extract; 10µM Aβ), and synthetic Aβ fibrils in PBS (syn. Aβ fibrils in PBS; 10µM) were treated with 50µg/ml proteinase K (PK) for 0min, 30min, 1h, or 2h at 37°C. (A) Silver staining of a 12% Bis-Tris NuPage® gel for total protein reveals the digestion of the majority of proteins in the brain extracts and of synthetic Aβ fibrils already after 30 min PK treatment. (B) Immunoblot analysis with an antibody specific to human Aβ (6E10) shows that Aβ in the Tg extract is largely PK-resistant for up to 2h. Aβ in the spiked Wt extract revealed some, but significantly less PK-resistance, whereas synthetic Aβ fibrils in PBS were almost completely digested by the PK treatment. (C) Densitometric quantification of Aβ-immunoblots after PK treatment of 5 Tg extracts [each from a different animal] and 3 independent synthetic Aβ fibril preparations are shown. Each preparation was tested at least 3 times and the mean was taken. Aβ concentration at time point 0 was designated as 100%. Indicated is the mean ± SEM. Repeated Measures 2-way ANOVA revealed a significant difference between the groups (F (2,34) = 112.2). Multiple Bonferroni post hoc tests showed significances after PK digestion for all group comparisons, *** p < 0.001. (D–F) In vivo seeding activity of brain extracts with or without PK treatment (50µg/ml PK for 30min) was tested by intrahippocampal injections into young, predepositing 4-month-old APP23 mice. Brains were immunohistochemically analyzed for Aβ deposition 5 months later. Shown is the dentate gyrus of the hippocampus, which revealed robust β-amyloid induction by the Tg extract (D) and the PK treated Tg extract (E), but not with Wt extract (F). Note that extracts were boiled before injection to deactivate residual PK activity. The small insert shows double staining for Aβ immunoreactivity and Congo-red binding of the induced β-amyloid in an adjacent section. Scale bars: 200µm a n d 2 0µm. (G) Stereological quantification of Aβ load in the hippocampus. Although PK treatment reduced the β-amyloid induction, the PK-resistant Tg material still showed potent seeding activity compared to the Wt extract (n=5–6 mice per group; mean ± SEM: ANOVA followed by Bonferroni post hoc tests revealed a significant difference between the extracts (F (2, 14) = 18.95, *** p < 0.001; * p < 0.05). Note that neither PK-treated nor untreated Wt extract induced any amyloid, and therefore these groups were combined.

Article Snippet: APP23 mice currently bred at the Hertie Institute for Clinical Brain Research first develop individual β-amyloid plaques in the neocortex at 7 months of age.

Techniques: Silver Staining, Western Blot, Concentration Assay, In Vivo, Activity Assay, Injection, Double Staining, Binding Assay

Journal: The Journal of neuroscience : the official journal of the Society for Neuroscience

Article Title: Soluble A? seeds are potent inducers of cerebral ?-amyloid deposition

doi: 10.1523/JNEUROSCI.3088-11.2011

Figure Lengend Snippet: (A) Brain extracts (10% [w/v]) from aged APP23 mice (Tg extract) were subjected to 100,000 × g ultracentrifugation (1h, 4°C). Left panel: Silver staining of a 12%Bis-Tris gel shows somewhat less protein in the pellet fraction (P) compared to the supernatant (SN) fraction (this was confirmed by total protein measurements revealing approximately 60% of the protein in the supernatant and 40% in the pellet, see results). Right panel: Immunoblotting for human Aβ (antibody 6E10) using 12% Bis-Tris gels reveals that most Aβ is retained in the pellet fraction. Aβ in the supernatant was below the level of detection by this immunoblot, but was estimated to be <0.05% of total Aβ concentration in the original Tg extract using ELISA (see Table 1). Loading control: synthetic Aβ, 15ng/lane. (B–D) In vivo seeding activity of the supernatant and pellet fractions was tested by intrahippocampal injections into young, predepositing 4-month-old APP23 mice. Brains were immunohistochemically analyzed for Aβ deposition 4 months post-injection. Shown is the dentate gyrus of the hippocampus. While the Tg extract (B) induced robust Aβ deposition, the SN fraction (C) induced largely diffuse Congo red-negative Aβ deposits. Aβ deposition induced by the pellet fraction (D) is similar to that induced by the Tg extract. The small insert shows double staining of Aβ and Congo red in seeded β-amyloid in an adjacent section. Note that the β-amyloid-induction by the SN was largely Congo red-negative after this 4-month incubation period, but was partly Congo red-positive after a 6-month incubation (see panel G below). (E) Stereological quantification of Aβ load in the hippocampus 4 months post-injection (n = 5 mice per group; mean ± SEM; ANOVA followed by Bonferroni post hoc test (F [2, 13] = 11.93; ** p < 0.01, n.s. [not significant]). Note that an additive effect of the β-amyloid induction by the SN and P fractions cannot be assumed (Meyer-Luehmann et al., 2006). (F) β-amyloid induction by the SN fraction 6 months post-injection. (G) The majority of the SN fraction-induced β-amyloid-deposits at 6 months post-injection were still Congo red negative, however a subset of the induced deposits were congophilic (arrow) and surrounded by activated, darkly-stained Iba-1-positive microglia, hypertrophic GFAP-positive astrocytes, and APP-positive dystrophic boutons and neurites (arrowheads). Shown are adjacent sections to (F). Scale bars: 200µm, 20µm (B-D, F) and 50µm (G).

Article Snippet: APP23 mice currently bred at the Hertie Institute for Clinical Brain Research first develop individual β-amyloid plaques in the neocortex at 7 months of age.

Techniques: Silver Staining, Western Blot, Concentration Assay, Enzyme-linked Immunosorbent Assay, Control, In Vivo, Activity Assay, Injection, Double Staining, Incubation, Staining

Journal: The Journal of neuroscience : the official journal of the Society for Neuroscience

Article Title: Soluble A? seeds are potent inducers of cerebral ?-amyloid deposition

doi: 10.1523/JNEUROSCI.3088-11.2011

Figure Lengend Snippet: (A) Brain extracts (10% [w/v]) from aged APP23 mice (Tg extract) and the 100,000 × g supernatant (SN) of the Tg extract (ultracentrifugation, 1h, 4°C) were treated with 50µg PK/ml for 30 minutes at 37°C. Immunoblotting for human Aβ (antibody 6E10) using 12% Bis-Tris gels reveals the expected PK-resistance of the Tg extract. In contrast, only a minor fraction of the SN was not cleaved by PK (see long exposure). Note that the Tg extract was diluted 10-fold, and that 10-fold more of the SN fraction was loaded compared to that in Figs. 1B and ​and2A,2A, respectively. Also note that there is an empty lane between the two Tg extract lanes and the two lanes with the SN. (B) Densitometric analysis of Aβ immunoblots after PK treatment (4 Tg extracts [each from a different animal] and the corresponding 100,000 × g supernatant were used; each preparation was tested at least 2 times and the mean was taken). Aβ concentration at time point 0 was defined as 100%. Indicated is the mean ± SEM. (C, D) In vivo seeding activity of brain extracts with or without PK treatment was tested by intrahippocampal injections into young, predepositing 3 month-old APP23 mice. Brains were immunohistochemically analyzed for Aβ-deposition 6 months later. Shown is the dentate gyrus of the hippocampus. SN of Tg brain extract was boiled for 5min at 95°C (C). PK-treated (30min) and boiled SN of Tg brain extract (D). The small insert shows double labeling of Aβ immunoreactivity and Congo red binding of the induced β-amyloid in an adjacent section. Scale bars: 200µm and 20µm. (E) Stereological quantification of Aβ load in the hippocampus. PK treatment of the 100,000 × g supernatant of Tg brain extract diminished the seeding activity. t-test revealed a significant difference between the two groups (n = 4–5 mice per group, mean ± SEM, t[7] = 4.713; ** p < 0.01).

Article Snippet: APP23 mice currently bred at the Hertie Institute for Clinical Brain Research first develop individual β-amyloid plaques in the neocortex at 7 months of age.

Techniques: Western Blot, Concentration Assay, In Vivo, Activity Assay, Labeling, Binding Assay

Journal: The Journal of neuroscience : the official journal of the Society for Neuroscience

Article Title: Soluble A? seeds are potent inducers of cerebral ?-amyloid deposition

doi: 10.1523/JNEUROSCI.3088-11.2011

Figure Lengend Snippet: Brain extracts (10% [w/v]) from aged APP23 mice (Tg extract) were subjected to additional sonication (3 × 20sec). (A) Immunoblot analysis with an antibody specific to human Aβ reveals no change in total Aβ between the original Tg extract (−) and the extract with extended sonication (+). However, more Aβ was found in the supernatant (SN) of the extra-sonicated extract after ultracentrifugation (100,000 × g; 30 min) (long exposure). (B–C) In vivo seeding activity of the original and extra-sonicated extracts was tested by intrahippocampal injections into young, pre-depositing APP23 mice. Brains were immunohistochemically analyzed for Aβ deposition 4 months post-injection. Shown is the dentate gyrus of the hippocampus. Injection of original Tg extract (B) induced robust congophilic Aβ deposition with a filamentous and dense pattern, while Aβ induction with the extra-sonicated Tg extract generated more punctate and small deposits (C). The insert shows double labeling of Aβ immunoreactivity and Congo red binding of the induced β-amyloid. Scale bars: 200µm and 20µm. (D) Stereological quantification of Aβ load in the hippocampus revealed a significant increase in β-amyloid induction by extended sonication. (n = 5–6 mice per group; mean ± SEM, t[9]=3.188; * p < 0.05).

Article Snippet: APP23 mice currently bred at the Hertie Institute for Clinical Brain Research first develop individual β-amyloid plaques in the neocortex at 7 months of age.

Techniques: Sonication, Western Blot, In Vivo, Activity Assay, Injection, Generated, Labeling, Binding Assay

Journal: Journal of Cerebral Blood Flow & Metabolism

Article Title: Neuropathological correlates of MRI-observed hypointense lesions in the APP23 mouse model of cerebral amyloid angiopathy

doi: 10.1177/0271678X261424057

Figure Lengend Snippet: Aged Tg APP23 mice have higher numbers of hypointense lesions on MGE 9.4 T MRI compared to WT littermates: (a) a representative example of an MGE image in a 24-month-old Tg APP23 male mouse, showing three cortical (white squares) and one deep (orange square) hypointense lesions, (b) the number of cortical hypointense lesions on MGE MRI was higher in the Tg mice compared to the WT littermates (β = 2.57 (95% CI: 1.45–3.86), z = 4.25, p < 0.001), and (c) the number of deep hypointense lesions on MGE MRI was also higher in the Tg mice compared to the WT littermates (β = 1.24 (95% CI: 0.50–2.02), z = 3.37, p < 0.001). These effects are corrected for age and sex using a negative binomial regression model ( n = 29 mice). Circles and triangles indicate female and male mice, respectively. MGE: multi-gradient echo; WT: wildtype; Tg: transgenic.

Article Snippet: The APP23 mouse line is available at Jackson Laboratories (reference number #030504).

Techniques: Transgenic Assay

Journal: Journal of Cerebral Blood Flow & Metabolism

Article Title: Neuropathological correlates of MRI-observed hypointense lesions in the APP23 mouse model of cerebral amyloid angiopathy

doi: 10.1177/0271678X261424057

Figure Lengend Snippet: Histopathological correlates of cortical and deep hypointense lesions: (a) MGE image of a cortical hypointense lesion (white arrow) in a 24-month-old Tg APP23 female mouse that corresponded to an iron-positive microbleed with associated hemosiderin deposits (black arrows on H&E) on histopathology (a’: adjacent PB and H&E stains shown), (b) MGE image of a deep hypointense lesion (white arrow) in a 24-month-old Tg APP23 male mouse that corresponded to an iron-positive microbleed with associated hemosiderin deposits (black arrows on H&E) on histopathology (b’: adjacent PB and H&E stains shown), and (c) MGE image of two deep hypointense lesions (white arrows) in a 24-month-old Tg APP23 female mouse (same as in (a)) that corresponded to calcifications on histopathology (c’: adjacent H&E and VK stains shown). MGE: multi-gradient echo; PB: Perls’ Prussian Blue; H&E: hematoxylin & eosin; VK: Von Kossa.

Article Snippet: The APP23 mouse line is available at Jackson Laboratories (reference number #030504).

Techniques: Histopathology

Journal: Journal of Cerebral Blood Flow & Metabolism

Article Title: Neuropathological correlates of MRI-observed hypointense lesions in the APP23 mouse model of cerebral amyloid angiopathy

doi: 10.1177/0271678X261424057

Figure Lengend Snippet: Microbleeds in Tg APP23 mice show signs of vascular remodeling and lower vascular amyloid-β burden at the rupture site: (a, b) examples of two separate microbleeds that demonstrated signs of vascular remodeling (arrows in (a, b)), and absence of amyloid-β at the presumed rupture site (arrows in (a′, b′)) and (c, d) examples of two separate microbleeds (arrows in (c, d) point at hemosiderin deposits) for which the culprit vessel or the rupture site could not be identified (c′, d′). H&E: hematoxylin & eosin; Aβ: amyloid-β.

Article Snippet: The APP23 mouse line is available at Jackson Laboratories (reference number #030504).

Techniques:

Journal: Journal of Cerebral Blood Flow & Metabolism

Article Title: Neuropathological correlates of MRI-observed hypointense lesions in the APP23 mouse model of cerebral amyloid angiopathy

doi: 10.1177/0271678X261424057

Figure Lengend Snippet: Remodeled vessels are observed in Tg APP23 mice: (a) an example of a leptomeningeal blood vessel at the level of the pial surface with evidence of vascular remodeling, (b) lower burden of amyloid-β compared to neighboring vessels, (c) deposition of fibrin(ogen) in the wall, suggestive of blood–brain barrier leakage, and (d) perivascular inflammation in the form of Iba-1-positive cells (arrows). H&E: hematoxylin & eosin; Aβ: amyloid-β; Iba-1: ionized calcium-binding adapter molecule 1.

Article Snippet: The APP23 mouse line is available at Jackson Laboratories (reference number #030504).

Techniques: Binding Assay

Journal: Journal of Cerebral Blood Flow & Metabolism

Article Title: Neuropathological correlates of MRI-observed hypointense lesions in the APP23 mouse model of cerebral amyloid angiopathy

doi: 10.1177/0271678X261424057

Figure Lengend Snippet: 3D visualization of cortical blood vessel in a microbleed region: (a) MGE image of microbleed in a 24-month-old Tg APP23 male mouse, (b) MGE image co-registered to 3D Lightsheet microscopy image in region of microbleed. Box surrounding vessel shown in (c/d). Vessel with discontinuity (arrow) in which no amyloid-β, SMA, or Glut1 staining was observed (c, d). Rotation of blood vessel in 3D is shown in Supplemental Movie 1 . MGE: multi-gradient echo; SMA: smooth muscle actin; Glut1: glucose transporter 1.

Article Snippet: The APP23 mouse line is available at Jackson Laboratories (reference number #030504).

Techniques: Microscopy, Staining

Journal: Journal of nuclear medicine : official publication, Society of Nuclear Medicine

Article Title: Neuroinflammation Appears Early on PET Imaging and Then Plateaus in a Mouse Model of Alzheimer Disease.

doi: 10.2967/jnumed.117.197608

Figure Lengend Snippet: Figure 1. 11C-PIB binding in the brain of tg and wt APP23 mice. In vivo PET images of 11C-PIB in 26-

Article Snippet: APP23 mice (n = 36; 30 female and 6 male) (Novartis Inc., Switzerland) express human APP751swe driven by the neuronspecific mouse Thy-1.2 gene fragment as a promoter.

Techniques: Binding Assay, In Vivo

Journal: Journal of nuclear medicine : official publication, Society of Nuclear Medicine

Article Title: Neuroinflammation Appears Early on PET Imaging and Then Plateaus in a Mouse Model of Alzheimer Disease.

doi: 10.2967/jnumed.117.197608

Figure Lengend Snippet: Figure 2. 18F-GE-180 binding in the brain of tg and wt APP23 and mice. In vivo PET images of 18F-

Article Snippet: APP23 mice (n = 36; 30 female and 6 male) (Novartis Inc., Switzerland) express human APP751swe driven by the neuronspecific mouse Thy-1.2 gene fragment as a promoter.

Techniques: Binding Assay, In Vivo

Journal: Journal of nuclear medicine : official publication, Society of Nuclear Medicine

Article Title: Neuroinflammation Appears Early on PET Imaging and Then Plateaus in a Mouse Model of Alzheimer Disease.

doi: 10.2967/jnumed.117.197608

Figure Lengend Snippet: Figure 4. Representative ex vivo autoradiography images of 11C-PIB binding in wt and tg APP23 mice

Article Snippet: APP23 mice (n = 36; 30 female and 6 male) (Novartis Inc., Switzerland) express human APP751swe driven by the neuronspecific mouse Thy-1.2 gene fragment as a promoter.

Techniques: Ex Vivo, Autoradiography, Binding Assay