- Email: [email protected]
VASCULOTIDE TREATMENT ACCELERATES BLOOD-BRAIN BARRIER RESTORATION AFTER FOCUSED ULTRASOUND IN A MOUSE MODEL OF ALZHEIMER'S DISEASE
P556
Poster Presentations: Sunday, July 24, 2016
blood donors, have been shown to reduce AD-related pathologies in mouse models. Despite these promising results, a Phase III clinical trial failed to meet critical endpoints, specifically in improving cognition in AD patients. These suboptimal results of IVIg treatments may be due to the restricted ability of therapeutics to cross the blood-brain barrier (BBB). We propose to address this problem by using focused ultrasound (FUS), to alter BBB permeability and deliver IVIg to specific brain regions. Our hypothesis is that combination of FUS with IVIg therapy will enhance neuronal plasticity and decrease amyloid pathology in an amyloidosis mouse model. Methods: We used a transgenic (Tg) mouse model of amyloidosis. Animal cohorts included IVIg or saline treatments given intravenously, with and without FUS, in Tg mice and their non-Tg littermates. Treatments targeted the bilateral hippocampi and were given once a week for two consecutive weeks. Neurogenesis, neuronal activation and amyloid pathology were evaluated using post-mortem immunohistochemistry. Results: Post-mortem immunohistochemistry analysis shows that compared to animals treated with saline alone, FUS+IVIg treated animals had reduced Abeta pathology (p<0.05), increased neurogenesis (p<0.05) and increased neuronal activation (p<0.05). Conclusions: These findings are favorable as they signify that IVIg drug delivery to the brain by FUS enhances the beneficial effects of IVIg therapy related to Alzheimer’s disease. Furthermore, these results point to the utility of FUS as a drug delivery tool for other neurodegenerative diseases. P1-334
REGIONAL DISTRIBUTION OF AMYLOID-BETA, SOLUBLE APP AND SYNAPTIC MARKERS IN HUMAN BRAINS
Mitsuru Shinohara, Dennis W. Dickson, Guojun Bu, Mayo Clinic, Jacksonville, FL, USA. Contact e-mail: [email protected] Background: We recently reported the strong regional association be-
tween postsynaptic markers and Abeta accumulation in human brains, suggesting that synapses play important roles in Abeta accumulation (Shinohara et al., Acta Neuropathologica 2013; 125(4):535-547). To further reveal potential mechanism underlying the involvement of synapses in Abeta accumulation, we determined the regional distribution of soluble APPalpha and APPbeta by specific ELISAs and addressed their association with the regional distribution of Abeta, synaptic markers and other related molecules. Methods: Twelve brain regions including neocortical, limbic and subcortical areas were dissected from brains of non-demented individuals with or without Abeta accumulation (n ¼ 27, age ¼ 89.7 6 9.4 years) and sporadic Alzheimer’s disease (AD) patients (n¼19, age ¼ 84.7 6 7.8 years) extracted according to increasing insolubility by a sequential three-step method (TBS, Triton-X and guanidine). The levels of Abeta40, Abeta42, sAPPalpha, sAPPbeta, APP, BACE1, PSD95, synaptophysin, LRP1 were determined by ELISAs. Results: In non-demented individuals, the regional distribution of sAPPalpha and sAPPbeta correlated with one another, while their correlation with full-length APP was weak. Importantly, the correlations between insoluble Abeta (especially Abeta42) and sAPPalpha or sAPPbeta were strong. Moreover, the postsynaptic marker PSD95 showed strong regional correlation with sAPPalpha and sAPPbeta. No significant association was observed between BACE1 and sAPPbeta. The distribution of sAPPalpha and sAPPbeta also showed strong regional association with Abeta accumulation in AD patients. Conclusions: These results suggest a potential involvement of synaptic function in the metabolism of soluble APP, associated with the accumulation of Abeta during the development of sporadic Alzheimer’s disease.
We are currently addressing how synapses regulate the metabolism of soluble APP by in vivo microdialysis or another experimental system. P1-335
VALUE OF NEUROSURGERYNEUROPATHOLOGY COLLABORATION IN IDENTIFICATION OF VASCULAR B-AMYLOID PROTEINOPATHIES AND IN NEURORADIOLOGY CORRELATION
Kathy Newell, Paul Camarata, Stephen Blatt, University of Kansas Medical Center, Kansas City, KS, USA. Contact e-mail: [email protected] Background: Cerebral b-amyloid angiopathy (CAA) may be clini-
cally silent for long periods. Typical clinical presentations include large lobar hemorrhages or multiple microhemorrhages. Definitive diagnosis of CAA is based on pathologic confirmation; however, noninvasive neuroimaging techniques offer the potential for valuable insights into the diagnosis and study of this progressive disease. Methods: We have identified and studied 8 neurosurgical cases of pathology-confirmed CAA with diverse clinical presentations. CAA-related lesions were not suspected or implicated preoperatively. Imaging studies included CT, MRI, and/or arteriography. The clinical and radiologic records were reviewed. The brain tissue specimens were processed for paraffin embedding. All histologic sections were stained with H&E and b-amyloid immunostains. Other antibodies utilized varied as to case, and included: neurofilament, CD45, CD3, CD20, phosphotau, alpha-synuclein, smooth muscle actin, and GFAP. Results: The patients ranged from 55-76 years of age. Clinical presentations included confusion, headache, ataxia, memory loss, apathy, abulia, visual change, somnolence, behavioral change, and increased seizure activity. For one subject, the preoperative clinical diagnosis was encephalopathy, possibly prion disease. The differential diagnoses for the cases were multiple: diffuse cerebral edema (1), atypical demyelination (1), leuokencephalopathy/posterior reversible encephalopathy syndrome (1), vasculitis (1), tumor/mass lesion (1), intracranial hematoma of unknown etiology (3), vascular malformation (2), and recurrent glioblastoma (1). All cases had pathologic findings of vascular amyloid on H&E stains, confirmed with b-amyloid immunostains. In three cases, there was histologic evidence of amyloid-b related angiitis. Two of these subjects plus a third subject, whose biopsy did not contain obvious inflammation, improved clinically with immunosuppressive therapy. The biopsy of the patient with the treated glioblastoma had both tumor recurrence and a focus of CAA with hemorrhage in the focus of enhancement. The subject with Down syndrome had severe CAA and cortical plaques. b-amyloid plaques were detected in 6 of the 8 cases. Conclusions: CAA may present with diverse clinical presentations that necessitate neurosurgical procedures and pathologic evaluation. Feedback from the neuropathology-neurosurgery collaboration is essential for completing the picture of the clinical-neuroradiological spectrum of CAA-related disease. P1-336
VASCULOTIDE TREATMENT ACCELERATES BLOOD-BRAIN BARRIER RESTORATION AFTER FOCUSED ULTRASOUND IN A MOUSE MODEL OF ALZHEIMER’S DISEASE
Madelaine Lynch1,2, Meaghan O’Reilly1,2, Kelly Coultes1, Paul Van Slyke3, Dan Dumont1,2, Kullervo Hynynen1,2, Isabelle Aubert1,2, 1 Sunnybrook Research Institute, Toronto, ON, Canada; 2University of Toronto, Toronto, ON, Canada; 3Vasomune Therapeutics, Toronto, ON, Canada. Contact e-mail: [email protected]
Poster Presentations: Sunday, July 24, 2016 Background: Focused ultrasound (FUS), in presence of microbub-
bles, transiently induces blood-brain barrier (BBB) permeability to allow for minimally invasive delivery of therapeutics, to targeted areas of the brain. Restoration of the BBB following FUS occurs rapidly in healthy models, however, the primary clinical use would be in neurological disorders, such as Alzheimer’s disease (AD), where BBB integrity may be compromised. The use of FUS to deliver AD therapeutics could have beneficial clinical implications, however, it is unknown whether the plasticity of the BBB and its capacity for repair is maintained in the presence of AD pathology. Furthermore, we sought to accelerate BBB restoration using Vasculotide (VT), a synthetic angiopoietin-1 mimetic that activates the Tie2 signaling cascade; a pathway known to promote vascular stability and cell survival in peripheral organs. Methods: Using a transgenic (Tg) mouse model of amyloidosis and their non-Tg littermates, VT was injected, every 48 hours for 2-3 months. We used FUS to induce transient increases in BBB permeability. The entry of gadolinium in the brain was monitored by MRI and quantified using MATLAB at 6, 12 and 20 hours post-FUS. Lastly, in a separate cohort of mice, Evans Blue dye was injected to evaluate BBB closure 24 hours post-FUS. Results: VT significantly
P557
accelerated BBB restoration, with no significant difference between Tg and non-Tg mice. Furthermore, VT reduced the initial enhancement and acoustic pressure required to induce BBB permeability, with no difference between Tg and non-Tg mice. Finally, Evans Blue dye injected 24 hours post-FUS confirmed BBB closure. Conclusions: Our research is the first to examine the effects of VT on BBB permeability and provides a better understanding of the effects of FUS. This research presents a novel method of facilitating BBB closure that could lead to a VT pre-treatment with FUS delivery of AD therapeutics.
P1-337
AFRICAN AMERICAN BRAIN AUTOPSIES FROM THE NATIONAL ALZHEIMER’S COORDINATING CENTER
Neill R. Graff-Radford1, Lilah M. Besser2, Julia E. Crook1, Walter A. Kukull2,3, Dennis W. Dickson1, 1Mayo Clinic, Jacksonville, FL, USA; 2University of Washington, Seattle, WA, USA; 3National Alzheimer’s Coordinating Center, Seattle, WA, USA. Contact e-mail: graffradford. [email protected]
Table 4 AD neuropathology among African American and White subjects with autopsy data and dementia at last visit before death.
AD neuropathology* NIA/Reagan neuropathological criteria High likelihood dementia due to AD Intermediate likelihood dementia due to AD Low likelihood dementia due to AD Criteria not met CERAD neuropathological criteria Definite AD Probable AD Possible AD Criteria not met Braak stage for neurofibrillary degeneration Stages 5-6 Stages 3-4 Stages 1-2 Stage 0 CERAD score for density of neuritic plaques Frequent Moderate Sparse None CERAD semi-quantitative score for diffuse plaques Frequent Moderate Sparse None
African Americans
Whites
n (%)
n (%)
63 (65.6%)
207 (54.1%)
18 (18.8%) 9 (9.4%) 6 (6.3%)
63 (16.5%) 44 (11.5%) 69 (18.0%)
66 (71.0%) 12 (12.9%) 7 (7.5%) 8 (8.6%)
206 (58.7%) 49 (14.0%) 23 (6.6%) 73 (20.8%)
81 (74.3%) 18 (16.5%) 6 (5.5%) 4 (3.7%)
243 (55.7%) 93 (21.3%) 72 (16.5%) 28 (6.4%)
77 (70.0%) 18 (16.4%) 7 (6.4%) 8 (7.3%)
243 (55.0%) 75 (17.0%) 35 (7.9%) 89 (20.1%)
64 (68.1%) 21 (22.3%) 4 (4.3%) 5 (5.3%)
236 (58.6%) 65 (16.1%) 42 (10.4%) 60 (14.9%)
Uadjustedy
Adjusted without APOEy,z
Adjusted with APOEy,x
0.01
0.03
0.15
0.02
0.02
0.15
<.001
0.001
0.04
0.002
0.002
0.05
0.03
0.02
0.11
NC, Not calculated (most subjects missing Thal phase) * Missing data (African American; white): Braak stage (n¼2; n¼8); Density of neuritic plaques (n¼1; n¼2); Thal phase (n¼98; n¼390); Diffuse neuritic plaques (n¼17; n¼41) y Ordinal logistic regression z Adjusted for age at death, education, and CDR sum of boxes (complete case analysis; number of subjects missing: some covariates and/or: NIA/Reagan criteria (n¼83,15.0%); CERAD criteria (n¼117, 21.1%) Braak stage (n¼19, 3.4%); neuritic plaques (n¼12, 2.2%); diffuse plaques (n¼67,12.1%) x Adjusted forage at death, education, CDR sum of boxes, and number of APOE e4 alleles (complete case analysis; number of subjects missing: some covariates and/or: NIA/Reagan criteria (n¼177, 31.9%); CERAD criteria (n¼202, 36.4%); Braak stage (n¼131, 23.6%); neuritic plaques (n¼125, 22.5%); diffuse plaques (n¼170, 30.6%)
Poster Presentations: Sunday, July 24, 2016
blood donors, have been shown to reduce AD-related pathologies in mouse models. Despite these promising results, a Phase III clinical trial failed to meet critical endpoints, specifically in improving cognition in AD patients. These suboptimal results of IVIg treatments may be due to the restricted ability of therapeutics to cross the blood-brain barrier (BBB). We propose to address this problem by using focused ultrasound (FUS), to alter BBB permeability and deliver IVIg to specific brain regions. Our hypothesis is that combination of FUS with IVIg therapy will enhance neuronal plasticity and decrease amyloid pathology in an amyloidosis mouse model. Methods: We used a transgenic (Tg) mouse model of amyloidosis. Animal cohorts included IVIg or saline treatments given intravenously, with and without FUS, in Tg mice and their non-Tg littermates. Treatments targeted the bilateral hippocampi and were given once a week for two consecutive weeks. Neurogenesis, neuronal activation and amyloid pathology were evaluated using post-mortem immunohistochemistry. Results: Post-mortem immunohistochemistry analysis shows that compared to animals treated with saline alone, FUS+IVIg treated animals had reduced Abeta pathology (p<0.05), increased neurogenesis (p<0.05) and increased neuronal activation (p<0.05). Conclusions: These findings are favorable as they signify that IVIg drug delivery to the brain by FUS enhances the beneficial effects of IVIg therapy related to Alzheimer’s disease. Furthermore, these results point to the utility of FUS as a drug delivery tool for other neurodegenerative diseases. P1-334
REGIONAL DISTRIBUTION OF AMYLOID-BETA, SOLUBLE APP AND SYNAPTIC MARKERS IN HUMAN BRAINS
Mitsuru Shinohara, Dennis W. Dickson, Guojun Bu, Mayo Clinic, Jacksonville, FL, USA. Contact e-mail: [email protected] Background: We recently reported the strong regional association be-
tween postsynaptic markers and Abeta accumulation in human brains, suggesting that synapses play important roles in Abeta accumulation (Shinohara et al., Acta Neuropathologica 2013; 125(4):535-547). To further reveal potential mechanism underlying the involvement of synapses in Abeta accumulation, we determined the regional distribution of soluble APPalpha and APPbeta by specific ELISAs and addressed their association with the regional distribution of Abeta, synaptic markers and other related molecules. Methods: Twelve brain regions including neocortical, limbic and subcortical areas were dissected from brains of non-demented individuals with or without Abeta accumulation (n ¼ 27, age ¼ 89.7 6 9.4 years) and sporadic Alzheimer’s disease (AD) patients (n¼19, age ¼ 84.7 6 7.8 years) extracted according to increasing insolubility by a sequential three-step method (TBS, Triton-X and guanidine). The levels of Abeta40, Abeta42, sAPPalpha, sAPPbeta, APP, BACE1, PSD95, synaptophysin, LRP1 were determined by ELISAs. Results: In non-demented individuals, the regional distribution of sAPPalpha and sAPPbeta correlated with one another, while their correlation with full-length APP was weak. Importantly, the correlations between insoluble Abeta (especially Abeta42) and sAPPalpha or sAPPbeta were strong. Moreover, the postsynaptic marker PSD95 showed strong regional correlation with sAPPalpha and sAPPbeta. No significant association was observed between BACE1 and sAPPbeta. The distribution of sAPPalpha and sAPPbeta also showed strong regional association with Abeta accumulation in AD patients. Conclusions: These results suggest a potential involvement of synaptic function in the metabolism of soluble APP, associated with the accumulation of Abeta during the development of sporadic Alzheimer’s disease.
We are currently addressing how synapses regulate the metabolism of soluble APP by in vivo microdialysis or another experimental system. P1-335
VALUE OF NEUROSURGERYNEUROPATHOLOGY COLLABORATION IN IDENTIFICATION OF VASCULAR B-AMYLOID PROTEINOPATHIES AND IN NEURORADIOLOGY CORRELATION
Kathy Newell, Paul Camarata, Stephen Blatt, University of Kansas Medical Center, Kansas City, KS, USA. Contact e-mail: [email protected] Background: Cerebral b-amyloid angiopathy (CAA) may be clini-
cally silent for long periods. Typical clinical presentations include large lobar hemorrhages or multiple microhemorrhages. Definitive diagnosis of CAA is based on pathologic confirmation; however, noninvasive neuroimaging techniques offer the potential for valuable insights into the diagnosis and study of this progressive disease. Methods: We have identified and studied 8 neurosurgical cases of pathology-confirmed CAA with diverse clinical presentations. CAA-related lesions were not suspected or implicated preoperatively. Imaging studies included CT, MRI, and/or arteriography. The clinical and radiologic records were reviewed. The brain tissue specimens were processed for paraffin embedding. All histologic sections were stained with H&E and b-amyloid immunostains. Other antibodies utilized varied as to case, and included: neurofilament, CD45, CD3, CD20, phosphotau, alpha-synuclein, smooth muscle actin, and GFAP. Results: The patients ranged from 55-76 years of age. Clinical presentations included confusion, headache, ataxia, memory loss, apathy, abulia, visual change, somnolence, behavioral change, and increased seizure activity. For one subject, the preoperative clinical diagnosis was encephalopathy, possibly prion disease. The differential diagnoses for the cases were multiple: diffuse cerebral edema (1), atypical demyelination (1), leuokencephalopathy/posterior reversible encephalopathy syndrome (1), vasculitis (1), tumor/mass lesion (1), intracranial hematoma of unknown etiology (3), vascular malformation (2), and recurrent glioblastoma (1). All cases had pathologic findings of vascular amyloid on H&E stains, confirmed with b-amyloid immunostains. In three cases, there was histologic evidence of amyloid-b related angiitis. Two of these subjects plus a third subject, whose biopsy did not contain obvious inflammation, improved clinically with immunosuppressive therapy. The biopsy of the patient with the treated glioblastoma had both tumor recurrence and a focus of CAA with hemorrhage in the focus of enhancement. The subject with Down syndrome had severe CAA and cortical plaques. b-amyloid plaques were detected in 6 of the 8 cases. Conclusions: CAA may present with diverse clinical presentations that necessitate neurosurgical procedures and pathologic evaluation. Feedback from the neuropathology-neurosurgery collaboration is essential for completing the picture of the clinical-neuroradiological spectrum of CAA-related disease. P1-336
VASCULOTIDE TREATMENT ACCELERATES BLOOD-BRAIN BARRIER RESTORATION AFTER FOCUSED ULTRASOUND IN A MOUSE MODEL OF ALZHEIMER’S DISEASE
Madelaine Lynch1,2, Meaghan O’Reilly1,2, Kelly Coultes1, Paul Van Slyke3, Dan Dumont1,2, Kullervo Hynynen1,2, Isabelle Aubert1,2, 1 Sunnybrook Research Institute, Toronto, ON, Canada; 2University of Toronto, Toronto, ON, Canada; 3Vasomune Therapeutics, Toronto, ON, Canada. Contact e-mail: [email protected]
Poster Presentations: Sunday, July 24, 2016 Background: Focused ultrasound (FUS), in presence of microbub-
bles, transiently induces blood-brain barrier (BBB) permeability to allow for minimally invasive delivery of therapeutics, to targeted areas of the brain. Restoration of the BBB following FUS occurs rapidly in healthy models, however, the primary clinical use would be in neurological disorders, such as Alzheimer’s disease (AD), where BBB integrity may be compromised. The use of FUS to deliver AD therapeutics could have beneficial clinical implications, however, it is unknown whether the plasticity of the BBB and its capacity for repair is maintained in the presence of AD pathology. Furthermore, we sought to accelerate BBB restoration using Vasculotide (VT), a synthetic angiopoietin-1 mimetic that activates the Tie2 signaling cascade; a pathway known to promote vascular stability and cell survival in peripheral organs. Methods: Using a transgenic (Tg) mouse model of amyloidosis and their non-Tg littermates, VT was injected, every 48 hours for 2-3 months. We used FUS to induce transient increases in BBB permeability. The entry of gadolinium in the brain was monitored by MRI and quantified using MATLAB at 6, 12 and 20 hours post-FUS. Lastly, in a separate cohort of mice, Evans Blue dye was injected to evaluate BBB closure 24 hours post-FUS. Results: VT significantly
P557
accelerated BBB restoration, with no significant difference between Tg and non-Tg mice. Furthermore, VT reduced the initial enhancement and acoustic pressure required to induce BBB permeability, with no difference between Tg and non-Tg mice. Finally, Evans Blue dye injected 24 hours post-FUS confirmed BBB closure. Conclusions: Our research is the first to examine the effects of VT on BBB permeability and provides a better understanding of the effects of FUS. This research presents a novel method of facilitating BBB closure that could lead to a VT pre-treatment with FUS delivery of AD therapeutics.
P1-337
AFRICAN AMERICAN BRAIN AUTOPSIES FROM THE NATIONAL ALZHEIMER’S COORDINATING CENTER
Neill R. Graff-Radford1, Lilah M. Besser2, Julia E. Crook1, Walter A. Kukull2,3, Dennis W. Dickson1, 1Mayo Clinic, Jacksonville, FL, USA; 2University of Washington, Seattle, WA, USA; 3National Alzheimer’s Coordinating Center, Seattle, WA, USA. Contact e-mail: graffradford. [email protected]
Table 4 AD neuropathology among African American and White subjects with autopsy data and dementia at last visit before death.
AD neuropathology* NIA/Reagan neuropathological criteria High likelihood dementia due to AD Intermediate likelihood dementia due to AD Low likelihood dementia due to AD Criteria not met CERAD neuropathological criteria Definite AD Probable AD Possible AD Criteria not met Braak stage for neurofibrillary degeneration Stages 5-6 Stages 3-4 Stages 1-2 Stage 0 CERAD score for density of neuritic plaques Frequent Moderate Sparse None CERAD semi-quantitative score for diffuse plaques Frequent Moderate Sparse None
African Americans
Whites
n (%)
n (%)
63 (65.6%)
207 (54.1%)
18 (18.8%) 9 (9.4%) 6 (6.3%)
63 (16.5%) 44 (11.5%) 69 (18.0%)
66 (71.0%) 12 (12.9%) 7 (7.5%) 8 (8.6%)
206 (58.7%) 49 (14.0%) 23 (6.6%) 73 (20.8%)
81 (74.3%) 18 (16.5%) 6 (5.5%) 4 (3.7%)
243 (55.7%) 93 (21.3%) 72 (16.5%) 28 (6.4%)
77 (70.0%) 18 (16.4%) 7 (6.4%) 8 (7.3%)
243 (55.0%) 75 (17.0%) 35 (7.9%) 89 (20.1%)
64 (68.1%) 21 (22.3%) 4 (4.3%) 5 (5.3%)
236 (58.6%) 65 (16.1%) 42 (10.4%) 60 (14.9%)
Uadjustedy
Adjusted without APOEy,z
Adjusted with APOEy,x
0.01
0.03
0.15
0.02
0.02
0.15
<.001
0.001
0.04
0.002
0.002
0.05
0.03
0.02
0.11
NC, Not calculated (most subjects missing Thal phase) * Missing data (African American; white): Braak stage (n¼2; n¼8); Density of neuritic plaques (n¼1; n¼2); Thal phase (n¼98; n¼390); Diffuse neuritic plaques (n¼17; n¼41) y Ordinal logistic regression z Adjusted for age at death, education, and CDR sum of boxes (complete case analysis; number of subjects missing: some covariates and/or: NIA/Reagan criteria (n¼83,15.0%); CERAD criteria (n¼117, 21.1%) Braak stage (n¼19, 3.4%); neuritic plaques (n¼12, 2.2%); diffuse plaques (n¼67,12.1%) x Adjusted forage at death, education, CDR sum of boxes, and number of APOE e4 alleles (complete case analysis; number of subjects missing: some covariates and/or: NIA/Reagan criteria (n¼177, 31.9%); CERAD criteria (n¼202, 36.4%); Braak stage (n¼131, 23.6%); neuritic plaques (n¼125, 22.5%); diffuse plaques (n¼170, 30.6%)