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  Glial and vascular contributions to neurodegenerative diseases
Effects of ACE1 knockout on tau pathology, amyloid deposition, and neurodegeneration in AD-relevant mouse models
Leah K Cuddy1, Alia Alia1, Miranda Salvo1, Jelena Popovic1, Dmitry Prokopenko3, Rudolph Tanzi4, Robert Vassar1,2
1The Ken and Ruth Davee Department of Neurology, Northwestern University Feinberg School of Medicine, Chicago, USA , 2Mesulam Center for Cognitive Neurology and Alzheimer's Disease, Northwestern University Feinberg School of Medicine, Chicago , USA , 3Genetics and Aging Unit and McCance Center for Brain Health, Department of Neurology, Massachusetts General Hospital, Boston, USA, 4Genetics and Aging Unit and McCance Center for Brain Health, Department of Neurology, Massachusetts General Hospital, Boston, USA
 Background: Recent GWAS have identified ACE as an Alzheimer's disease (AD) risk locus. ACE encodes angiotensin I converting enzyme (ACE1) known for regulating blood pressure through the renin-angiotensin system (RAS). Using whole-genome sequencing, we identified rare ACE coding variants and investigated one, ACE1 R1279Q, in knockin (KI) mice. Increased ACE1 in ACE1 KI brains caused neurodegeneration within the hippocampus with aging [1]. Amyloidosis in 5XFAD mice crossed to KI mice accelerated hippocampal atrophy, although Aβ deposition was unchanged [1]. ACE1 is increased in human AD brains, and antihypertensive medications that inhibit ACE1/RAS prevents abnormal tau pathology, lowers Aβ, and slows the conversion of MCI to AD. Although the mechanism of ACE1 elevation in AD is unknown, increased ACE1 appears to be pathological and may selectively accelerate hippocampal neurodegeneration.
Materials and Methods: To further dissect the role of ACE1 in AD pathogenesis, we generated mice with ACE1 conditionally knocked out (cKO) in excitatory forebrain neurons and crossed these mice to the 5XFAD mouse model of amyloid pathology and to tau P301S tauopathy mice.
Results: Although ACE1 was knocked out in hippocampal and cortical neurons, RAS signaling was selectively disrupted in the hippocampus. RAS pathway inhibition associated with hippocampal dependent memory deficits. ACE1 cKO slightly reduced Aβ and neuroinflammation in 5XFAD mice, yet markedly enhanced CA neuron loss in the hippocampus in tau P301S mice resulting in a phenotype comparable to ACE1 KI mice.
Discussion/Conclusion: Antihypertensive medications are generally protective toward AD. Unexpectedly, we found unexpectedly that complete KO of ACE1 in excitatory neurons enhanced tau-mediated neurodegeneration and impaired memory. Our results suggest that physiological levels of neuronal ACE1 are required for protecting against selective hippocampal neurodegeneration in AD.
1.Cuddy LK, Prokopenko D, Cunningham EP, Brimberry R, Song P, Kirchner R, et al. Aβ-accelerated neurodegeneration caused by Alzheimer's -associated. Sci Transl Med. 2020;12(563).
Background: BACE1 initiates production of β-amyloid peptides (Aβ), which is associated with cognitive dysfunction in Alzheimer’s disease (AD) due to abnormal oligomerization and aggregation. While BACE1 inhibitors show strong reduction in Aβ deposition, they fail to improve cognitive function in patients, largely due to its role in synaptic function. Materials and Methods: We used BACE1 inhibitors Verabucestat and Lanabecestat alone or in combination with mGluR1 PAM in C57BL/6 mice to study the effect on synaptic plasticity using electrophysiology, electron microscope and biochemistry techniques. Details methodology can be obtained at (1).
Results: We show BACE1 is required for optimal release of synaptic vesicles. BACE1 deficiency or inhibition decreases synaptic vesicle docking in the synaptic active zones. BACE1-null mice or mice treated with BACE1 inhibitors Verabucestat and Lanabecestat exhibit severe reduction in hippocampal LTP and learning behaviors. To counterbalance this synaptic deficit, we discovered that BACE1-null mice treated with positive allosteric modulators (PAM) of metabotropic glutamate receptor-1 (mGluR1), whose levels reduced in BACE1-null mice, significantly improved LTP and cognitive behaviors. Similarly, mice treated with mGluR1 PAM showed significantly mitigated synaptic deficits caused by BACE1 inhibitors.
Discussion: BACE1 is a prime target for AD therapy, it is elevated in brains of AD patients, accumulated in axons and its cleavage of APP is the rate-limiting step in Aβ production (2-4). However, clinical trials of BACE1 inhibitors in AD therapy have not been successful because of their failures to improve cognitive functions, despite reducing plaque load (5, 6). Therefore, it is critical to find solutions that will take advantage of this plaque reduction while overcoming the unwanted side effects like worsening cognitive functions/scores.
Conclusions: Data suggest that a therapy combining BACE1 inhibitors for reducing amyloid deposition and a mGluR1 PAM for counteracting BACE1-mediated synaptic deficits appears to be an effective approach for treating AD patients.
Background: The AD brain is characterized by amyloid plaques consisting of the β-amyloid peptide and neurofibrillary tangles containing hyperphosphorylated, aggregated tau. Amyloid plaques form first and likely give rise to tangles, but the mechanistic link between them is unclear. The peri-plaque environment is toxic to neurons, characterized by synaptic loss, activated microglia, and vesicle-filled dystrophic neurites, which accumulate aggregation-prone phosphorylated forms of tau. We hypothesize that axonal contact with plaque β-amyloid causes membrane damage, leading to calcium influx, kinase activation, microtubule disruption, trafficking impairment, tau hyperphosphorylation, and dystrophic neurites. Here we describe the effects of membrane repair protein annexin A6 on dystrophic neurites in AD.
Materials/Methods: Primary neurons from mice expressing genomically encoded annexin A6-GFP were subjected to laser injury to induce membrane damage. Localization of genomic A6-GFP and endogenous A6 in 5XFAD brains was determined using immunofluorescence. Overexpression of A6-GFP was induced by intracerebroventricular injection of P0 mouse pups which were harvested at 4 months old for analysis by immunoblot and immunofluorescence.
Results: After membrane injury, genomic and recombinant annexin A6 localized to the site of damage. In 5XFAD mouse model of amyloidosis, genomic A6-tGFP and endogenous A6 localized to plasma membranes of large neurons and of dystrophic neurites. In mice that overexpress annexin A6-GFP in neurons, LAMP1 and ptau- 181 positive dystrophic neurites per plaque are significantly reduced, while microglia and astrocytes around the plaques remain unchanged. Intracerbroventricularly injected recombinant A6 localizes to outer membrane of dystrophic neurites.
Discussion: Overexpression of annexin A6 reduced dystrophic neurites resulting in decreased p-tau 181, suggesting that targeting dystrophic neurites could decrease the toxicity of amyloid plaques by slowing seeding of tau tangles, which are associated with cognitive decline. Conclusions: Further work will explore the ability of annexin A6 to prevent tau spreading, and the use of recombinant A6 as an AD therapeutic.
Annexin A6 in membrane resealing in Alzheimer's disease
Katherine Sadleir1, Ammaarah Khatri, Regan Andringa-Seed,
Alexis Demonbreun, Elizabeth McNally, Robert Vassar
1Northwestern University, Chicago, USA
BACE1 controls synaptic function through modulating release of synaptic vesicles Brati Das1, Neeraj Singh1, Annie Y. Yao1, John Zhou,
Wanxia He, Xaingyou Hu, Riqiang Yan1
1UConn Health Center, USA
 54 • ISMND 2022

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