Aß Amyloid Scaffolds the Accumulation of Matrisome and Additional Proteins in Alzheimer's Disease.

We report a highly significant correlation in brain proteome changes between Alzheimers disease (AD) and CRND8 APP695NL/F transgenic mice. However, integrating protein changes observed in the CRND8 mice with co-expression networks derived from human AD, reveals both conserved and divergent module changes. For the most highly conserved module (M42, matrisome) we find many proteins accumulate in plaques, cerebrovascular amyloid (CAA), dystrophic processes, or a combination thereof. Overexpression of two M42 proteins, midkine (Mdk) and pleiotrophin (PTN), in CRND8 mice brains leads to increased accumulation of A ß ; in plaques and in CAA; further, recombinant MDK and PTN enhance A ß ; aggregation into amyloid. Multiple M42 proteins, annotated as heparan sulfate binding proteins, bind to fibrillar A ß 42 and a non-human amyloid fibril in vitro. Supporting this binding data, MDK and PTN co-accumulate with transthyretin (TTR) amyloid in the heart and islet amyloid polypeptide (IAPP) amyloid in the pancreas. Our findings establish several critical insights. Proteomic changes in modules observed in human AD brains define an A ß ; amyloid responsome that is well conserved from mouse model to human. Further, distinct amyloid structures may serve as scaffolds, facilitating the co-accumulation of proteins with signaling functions. We hypothesize that this co-accumulation may contribute to downstream pathological sequalae. Overall, this contextualized understanding of proteomic changes and their interplay with amyloid deposition provides valuable insights into the complexity of AD pathogenesis and potential biomarkers and therapeutic targets.

Modeling the Competition between Misfolded Aß Conformers That Produce Distinct Types of Amyloid Pathology in Alzheimer's Disease.

The amyloid pathology characteristic of Alzheimer's disease (AD) can be broadly classified as either fibrillary amyloid or diffuse amyloid. Fibrillary amyloid is found in cored-neuritic deposits, fibrillar deposits, and vascular deposits, and binds strongly to the amyloid revealing dyes Thioflavin-S or Congo Red. Diffuse amyloid can appear as wispy dispersed deposits or compact tufted deposits dispersed in neuropil, and binds amyloid dyes weakly if at all. In AD brains, both types of pathology are detected. Homogenates from AD brains, or the brains of transgenic mice modeling AD-amyloidosis, have been used to seed pathology in vulnerable host transgenic models. These studies suggest that pathologies may arise from distinct conformers or strains of misfolded Aß, similar to propagating prions. Using Aß strains sourced from four different AD-amyloidosis models, we injected pathological seeds into the brains of newborn mice from three different transgenic hosts with distinctive Aß pathologies. Two of the seeding sources were from mice that primarily develop cored-neuritic Aß deposits (cored strain) while the other two seeding sources were from mice that develop diffuse Aß deposits (diffuse strain). These seeds were injected into host APP mice in which the resident strain was either diffuse or cored-neuritic pathology. Seeding-homogenates were injected into the brains of newborn mice to initiate propagation as early as possible. Depending upon the level of transgene expression in the host, we show that the injected strains of misfolded Aß from the seeding homogenate were able to outcompete the resident strain of the APP host model. In serial passaging experiments, it appeared that the diffuse strain was more easily propagated than the cored strain. Collectively, our studies align with the idea that different types of Aß pathology in AD brains arise from different populations of Aß conformers that compete to populate the brain.

TAPPing into the potential of inducible tau/APP transgenic mice.

AIMS: Our understanding of the pathological interactions between amyloidosis and tauopathy in Alzheimer's disease is incomplete. We sought to determine if the relative timing of the amyloidosis and tauopathy is critical for amyloid-enhanced tauopathy. METHODS: We crossed an inducible tauopathy model with two ß-amyloid models utilising the doxycycline-repressible transgenic system to modulate timing and duration of human tau expression in the context of amyloidosis and then assessed tauopathy, amyloidosis and gliosis. RESULTS: We combined inducible rTg4510 tau with APPswe/PS1dE9 [Line 85 (L85)] mice to examine the interactions between Aß and tauopathy at different stages of amyloidosis. When we initially suppressed mutant human tau expression for 14-15 months and subsequently induced tau expression for 6 months, severe amyloidosis with robust tauopathy resulted in rTg4510/L85 but not rTg4510 mice. When we suppressed mutant tau for 7 months before inducing expression for a subsequent 6 months in another cohort of rTg4510/L85 and rTg4510 mice, only rTg4510/L85 mice displayed robust tauopathy. Lastly, we crossed rTg4510 mice to tet-regulated APPswe/ind [Line 107 (L107)] mice, using doxycycline to initially suppress both transgenes for 1 month before inducing expression for 5 months to model early amyloidosis. In contrast to rTg4510, rTg4510/L107 mice rapidly developed amyloidosis, accompanied by robust tauopathy. CONCLUSIONS: These data suggest that tau misfolding is exacerbated by both newly forming Aß deposits in younger brain and mature deposits in older brains. Refined use and repurposing of these models provide new tools to explore the intersection of ageing, amyloid and tauopathy and to test interventions to disrupt the amyloid cascade.

Il-10 signaling reduces survival in mouse models of synucleinopathy.

Parkinson's disease (PD) and related synucleinopathies are characterized by chronic neuroinflammation leading to the premise that anti-inflammatory therapies could ameliorate synucleinopathy and associated sequelae. To test this idea, we used recombinant adeno-associated viruses (AAV) to express the anti-inflammatory cytokine, Interleukin (Il)-10, in Line M83 transgenic mice that expresses the PD-associated A53T mutant human α-synuclein (αSyn). Contrary to our expectations, we observed that intraspinal Il-10 expression initiated at birth upregulated microgliosis and led to early death in homozygous M83+/+ mice. We further observed that Il-10 preconditioning led to reduced lifespan in the hemizygous M83+/- mice injected with preformed αSyn aggregates in hindlimb muscles. To determine the mechanistic basis for these adverse effects, we took advantage of the I87A variant Il-10 (vIl-10) that has predominantly immunosuppressive properties. Sustained intraspinal expression of vIl-10 in preformed αSyn-aggregate seeded M83+/- mice resulted in earlier death, accelerated αSyn pathology, pronounced microgliosis, and increased apoptosis compared to control mice. AAV-vIl-10 expression robustly induced p62 and neuronal LC3B accumulation in these mice, indicating that Il-10 signaling mediated preconditioning of the neuraxis can potentially exacerbate αSyn accumulation through autophagy dysfunction in the neurons. Together, our data demonstrate unexpected adverse effects of both Il-10 and its immunosuppressive variant, vIl-10, in a mouse model of PD, highlighting the pleiotropic functions of immune mediators and their complex role in non-cell autonomous signaling in neurodegenerative proteinopathies.

Reactive astrocytes as treatment targets in Alzheimer's disease-Systematic review of studies using the APPswePS1dE9 mouse model.

Astrocytes regulate synaptic communication and are essential for proper brain functioning. In Alzheimer's disease (AD) astrocytes become reactive, which is characterized by an increased expression of intermediate filament proteins and cellular hypertrophy. Reactive astrocytes are found in close association with amyloid-beta (Aß) deposits. Synaptic communication and neuronal network function could be directly modulated by reactive astrocytes, potentially contributing to cognitive decline in AD. In this review, we focus on reactive astrocytes as treatment targets in AD in the APPswePS1dE9 AD mouse model, a widely used model to study amyloidosis and gliosis. We first give an overview of the model; that is, how it was generated, which cells express the transgenes, and the effect of its genetic background on Aß pathology. Subsequently, to determine whether modifying reactive astrocytes in AD could influence pathogenesis and cognition, we review studies using this mouse model in which interventions were directly targeted at reactive astrocytes or had an indirect effect on reactive astrocytes. Overall, studies specifically targeting astrocytes to reduce astrogliosis showed beneficial effects on cognition, which indicates that targeting astrocytes should be included in developing novel therapies for AD.

Remodeling Alzheimer-amyloidosis models by seeding.

Alzheimer's disease (AD) is among the most prevalent neurodegenerative diseases, with brain pathology defined by extracellular amyloid beta deposits and intracellular tau aggregates. To aid in research efforts to improve understanding of this disease, transgenic murine models have been developed that replicate aspects of AD pathology. Familial AD is associated with mutations in the amyloid precursor protein and in the presenilins (associated with amyloidosis); transgenic amyloid models feature one or more of these mutant genes. Recent advances in seeding methods provide a means to alter the morphology of resultant amyloid deposits and the age that pathology develops. In this review, we discuss the variety of factors that influence the seeding of amyloid beta pathology, including the source of seed, the time interval after seeding, the nature of the transgenic host, and the preparation of the seeding inoculum.

IL-10 based immunomodulation initiated at birth extends lifespan in a familial mouse model of amyotrophic lateral sclerosis.

Inflammatory signaling is thought to modulate the neurodegenerative cascade in amyotrophic lateral sclerosis (ALS). We have previously shown that expression of Interleukin-10 (IL-10), a classical anti-inflammatory cytokine, extends lifespan in the SOD1-G93A mouse model of familial ALS. Here we test whether co-expression of the decoy chemokine receptor M3, that can scavenge inflammatory chemokines, augments the efficacy of IL-10. We found that recombinant adeno-associated virus (AAV)-mediated expression of IL-10, alone, or in combination with M3, resulted in modest extension of lifespan relative to control SOD1-G93A cohort. Interestingly neither AAV-M3 alone nor AAV-IL-10 + AAV-M3 extend survival beyond that of the AAV-IL-10 alone cohort. Focused transcriptomic analysis revealed induction of innate immunity and phagocytotic pathways in presymptomatic SOD1-G93A mice expressing IL-10 + M3 or IL-10 alone. Further, while IL-10 expression increased microglial burden, the IL-10 + M3 group showed lower microglial burden, suggesting that M3 can successfully lower microgliosis before disease onset. Our data demonstrates that over-expression of an anti-inflammatory cytokine and a decoy chemokine receptor can modulate inflammatory processes in SOD1-G93A mice, modestly delaying the age to paralysis. This suggests that multiple inflammatory pathways can be targeted simultaneously in neurodegenerative disease and supports consideration of adapting these approaches to treatment of ALS and related disorders.

Diversity in Aß deposit morphology and secondary proteome insolubility across models of Alzheimer-type amyloidosis.

A hallmark pathology of Alzheimer's disease (AD) is the formation of amyloid ß (Aß) deposits that exhibit diverse localization and morphologies, ranging from diffuse to cored-neuritic deposits in brain parenchyma, with cerebral vascular deposition in leptomeningeal and parenchymal compartments. Most AD brains exhibit the full spectrum of pathologic Aß morphologies. In the course of studies to model AD amyloidosis, we have generated multiple transgenic mouse models that vary in the nature of the transgene constructs that are expressed; including the species origin of Aß peptides, the levels and length of Aß that is deposited, and whether mutant presenilin 1 (PS1) is co-expressed. These models recapitulate features of human AD amyloidosis, but interestingly some models can produce pathology in which one type of Aß morphology dominates. In prior studies of mice that primarily develop cored-neuritic deposits, we determined that Aß deposition is associated with changes in cytosolic protein solubility in which a subset of proteins become detergent-insoluble, indicative of secondary proteome instability. Here, we survey changes in cytosolic protein solubility across seven different transgenic mouse models that exhibit a range of Aß deposit morphologies. We find a surprisingly diverse range of changes in proteome solubility across these models. Mice that deposit human Aß40 and Aß42 in cored-neuritic plaques had the most robust changes in proteome solubility. Insoluble cytosolic proteins were also detected in the brains of mice that develop diffuse Aß42 deposits but to a lesser extent. Notably, mice with cored deposits containing only Aß42 had relatively few proteins that became detergent-insoluble. Our data provide new insight into the diversity of biological effects that can be attributed to different types of Aß pathology and support the view that fibrillar cored-neuritic plaque pathology is the more disruptive Aß pathology in the Alzheimer's cascade.

Intracerebral Expression of AAV-APOE4 Is Not Sufficient to Alter Tau Burden in Two Distinct Models of Tauopathy.

Apolipoprotein E4 (APOE4) is the major genetic risk factor for sporadic Alzheimer's disease (AD), which is characterized by amyloid ß (Aß) plaques and tau tangles. Though the role of APOE4 in Aß pathogenesis has been mechanistically defined in rodent models, much less is known regarding the relationship of APOE4 to tau pathogenesis. Recent studies have indicated a possible correlation between APOE isoform-dependent alterations in tau pathology and neurodegeneration. To explore whether neuronal expression of APOE4 triggers tauopathy, here we delivered adeno-associated viruses (AAV) expressing human APOE4 in two different models of tauopathy-rTg4510 and PS19 lines. Intracerebroventricular delivery of AAV-APOE4 in neonatal rTg4510 and PS19 mice resulted in increased APOE4 protein in neurons but did not result in altered phosphorylated tau burden, pretangle tau pathology, or silver-positive tangle pathology. Biochemical analysis of synaptic proteins did not reveal substantial alterations. Our results indicate that over-expression of APOE4 in neurons, using an AAV-mediated approache, is not sufficient to accelerate or otherwise alter the inherent tau pathology that occurs in mice overexpressing mutant human tau.

Aberrant accrual of BIN1 near Alzheimer's disease amyloid deposits in transgenic models.

Bridging integrator 1 (BIN1) is the most significant late-onset Alzheimer's disease (AD) susceptibility locus identified via genome-wide association studies. BIN1 is an adaptor protein that regulates membrane dynamics in the context of endocytosis and membrane remodeling. An increase in BIN1 expression and changes in the relative levels of alternatively spliced BIN1 isoforms have been reported in the brains of patients with AD. BIN1 can bind to Tau, and an increase in BIN1 expression correlates with Tau pathology. In contrast, the loss of BIN1 expression in cultured cells elevates Aß production and Tau propagation by insfluencing endocytosis and recycling. Here, we show that BIN1 accumulates adjacent to amyloid deposits in vivo. We found an increase in insoluble BIN1 and a striking accrual of BIN1 within and near amyloid deposits in the brains of multiple transgenic models of AD. The peri-deposit aberrant BIN1 localization was conspicuously different from the accumulation of APP and BACE1 within dystrophic neurites. Although BIN1 is highly expressed in mature oligodendrocytes, BIN1 association with amyloid deposits occurred in the absence of the accretion of other oligodendrocyte or myelin proteins. Finally, super-resolution microscopy and immunogold electron microscopy analyses highlight the presence of BIN1 in proximity to amyloid fibrils at the edges of amyloid deposits. These results reveal the aberrant accumulation of BIN1 is a feature associated with AD amyloid pathology. Our findings suggest a potential role for BIN1 in extracellular Aß deposition in vivo that is distinct from its well-characterized function as an adaptor protein in endocytosis and membrane remodeling.

Changes in proteome solubility indicate widespread proteostatic disruption in mouse models of neurodegenerative disease.

The deposition of pathologic misfolded proteins in neurodegenerative disorders such as Alzheimer's disease, Parkinson's disease, frontotemporal dementia and amyotrophic lateral sclerosis is hypothesized to burden protein homeostatic (proteostatic) machinery, potentially leading to insufficient capacity to maintain the proteome. This hypothesis has been supported by previous work in our laboratory, as evidenced by the perturbation of cytosolic protein solubility in response to amyloid plaques in a mouse model of Alzheimer's amyloidosis. In the current study, we demonstrate changes in proteome solubility are a common pathology to mouse models of neurodegenerative disease. Pathological accumulations of misfolded tau, α-synuclein and mutant superoxide dismutase 1 in CNS tissues of transgenic mice were associated with changes in the solubility of hundreds of CNS proteins in each model. We observed that changes in proteome solubility were progressive and, using the rTg4510 model of inducible tau pathology, demonstrated that these changes were dependent upon sustained expression of the primary pathologic protein. In all of the models examined, changes in proteome solubility were robust, easily detected, and provided a sensitive indicator of proteostatic disruption. Interestingly, a subset of the proteins that display a shift towards insolubility were common between these different models, suggesting that a specific subset of the proteome is vulnerable to proteostatic disruption. Overall, our data suggest that neurodegenerative proteinopathies modeled in mice impose a burden on the proteostatic network that diminishes the ability of neural cells to prevent aberrant conformational changes that alter the solubility of hundreds of abundant cellular proteins.

Short Aß peptides attenuate Aß42 toxicity in vivo.

Processing of amyloid-ß (Aß) precursor protein (APP) by γ-secretase produces multiple species of Aß: Aß40, short Aß peptides (Aß37-39), and longer Aß peptides (Aß42-43). γ-Secretase modulators, a class of Alzheimer's disease therapeutics, reduce production of the pathogenic Aß42 but increase the relative abundance of short Aß peptides. To evaluate the pathological relevance of these peptides, we expressed Aß36-40 and Aß42-43 in Drosophila melanogaster to evaluate inherent toxicity and potential modulatory effects on Aß42 toxicity. In contrast to Aß42, the short Aß peptides were not toxic and, when coexpressed with Aß42, were protective in a dose-dependent fashion. In parallel, we explored the effects of recombinant adeno-associated virus-mediated expression of Aß38 and Aß40 in mice. When expressed in nontransgenic mice at levels sufficient to drive Aß42 deposition, Aß38 and Aß40 did not deposit or cause behavioral alterations. These studies indicate that treatments that lower Aß42 by raising the levels of short Aß peptides could attenuate the toxic effects of Aß42.

Quantitative Comparison of Dense-Core Amyloid Plaque Accumulation in Amyloid-ß Protein Precursor Transgenic Mice.

There exist several dozen lines of transgenic mice that express human amyloid-ß protein precursor (AßPP) with Alzheimer's disease (AD)-linked mutations. AßPP transgenic mouse lines differ in the types and amounts of Aß that they generate and in their spatiotemporal patterns of expression of Aß assemblies, providing a toolkit to study Aß amyloidosis and the influence of Aß aggregation on brain function. More complete quantitative descriptions of the types of Aß assemblies present in transgenic mice and in humans during disease progression should add to our understanding of how Aß toxicity in mice relates to the pathogenesis of AD. Here, we provide a direct quantitative comparison of amyloid plaque burdens and plaque sizes in four lines of AßPP transgenic mice. We measured the fraction of cortex and hippocampus occupied by dense-core plaques, visualized by staining with Thioflavin S, in mice from young adulthood through advanced age. We found that the plaque burdens among the transgenic lines varied by an order of magnitude: at 15 months of age, the oldest age studied, the median cortical plaque burden in 5XFAD mice was already ∼4.5 times that of 21-month-old Tg2576 mice and ∼15 times that of 21-24-month-old rTg9191 mice. Plaque-size distributions changed across the lifespan in a line- and region-dependent manner. We also compared the dense-core plaque burdens in the mice to those measured in a set of pathologically-confirmed AD cases from the Nun Study. Cortical plaque burdens in Tg2576, APPSwePS1ΔE9, and 5XFAD mice eventually far exceeded those measured in the human cohort.

Generation of a new transgenic mouse model for assessment of tau gene silencing therapies.

BACKGROUND: Targeting the expression of genes has emerged as a potentially viable therapeutic approach to human disease. In Alzheimer's disease, therapies that silence the expression of tau could be a viable strategy to slow disease progression. METHODS: We produced a novel strain of transgenic mice that could be used to assess the efficacy of gene knockdown therapies for human tau, in live mice. We designed a tetracycline-regulated transgene construct in which the cDNA for human tau was fused to ubiquitin and to luciferase to create a single fusion polyprotein, termed TUL. RESULTS: When expressed in brain, the TUL polyprotein was cleaved by ubiquitin-processing enzymes to release the luciferase as an independent protein, separating the half-life of luciferase from the long-lived tau protein. Treatment of bigenic tTA/TUL mice with doxycycline produced rapid declines in luciferase levels visualized by in vivo imaging and ex vivo enzyme measurement. CONCLUSIONS: This new mouse model can be used as a discovery tool in optimizing gene targeting therapeutics directed to reduce human tau mRNA levels.

Murine Aß over-production produces diffuse and compact Alzheimer-type amyloid deposits.

INTRODUCTION: Transgenic overexpression of amyloid precursor protein (APP) genes that are either entirely human in sequence or have humanized Aß sequences can produce Alzheimer-type amyloidosis in mice, provided the transgenes also encode mutations linked to familial Alzheimer's Disease (FAD). Although transgenic mice have been produced that overexpress wild-type mouse APP, no mice have been generated that express mouse APP with FAD mutations. Here we describe two different versions of such mice that produce amyloid deposits consisting of entirely of mouse Aß peptides. One line of mice co-expresses mouse APP-Swedish (moAPPswe) with a human presenilin exon-9 deleted variant (PS1dE9) and another line expresses mouse APP-Swedish/Indiana (APPsi) using tetracycline-regulated vectors (tet.moAPPsi). RESULTS: Both lines of mice that produce mouse Aß develop amyloid deposits, with the moAPPswe/PS1dE9 mice developing extracellular compact, cored, neuritic deposits that primarily localize to white matter tracts and meningial layers, whereas the tet.moAPPsi mice developed extracellular diffuse cortical/hippocampal deposits distributed throughout the parenchyma. CONCLUSIONS: These findings demonstrate that murine Aß peptides have the capacity to produce amyloid deposits that are morphologically similar to deposits found in human AD provided the murine APP gene harbors mutations linked to human FAD.

RAN Translation in Huntington Disease.

Huntington disease (HD) is caused by a CAG ⋅ CTG expansion in the huntingtin (HTT) gene. While most research has focused on the HTT polyGln-expansion protein, we demonstrate that four additional, novel, homopolymeric expansion proteins (polyAla, polySer, polyLeu, and polyCys) accumulate in HD human brains. These sense and antisense repeat-associated non-ATG (RAN) translation proteins accumulate most abundantly in brain regions with neuronal loss, microglial activation and apoptosis, including caudate/putamen, white matter, and, in juvenile-onset cases, also the cerebellum. RAN protein accumulation and aggregation are length dependent, and individual RAN proteins are toxic to neural cells independent of RNA effects. These data suggest RAN proteins contribute to HD and that therapeutic strategies targeting both sense and antisense genes may be required for efficacy in HD patients. This is the first demonstration that RAN proteins are expressed across an expansion located in an open reading frame and suggests RAN translation may also contribute to other polyglutamine diseases.

Behavioral abnormalities in APPSwe/PS1dE9 mouse model of AD-like pathology: comparative analysis across multiple behavioral domains.

Alzheimer's disease (AD) is characterized by dysfunction in cognitive and noncognitive domains with clinical diagnosis based on multiple neuropsychological tests. Here, we evaluated cognitive and noncognitive behaviors in 2 age cohorts (8 and 14 months at the start of the study) of APPSwe/PS1dE9 transgenic mice that model AD-like amyloidosis. We used a battery of tests that included fear-conditioned context and tone memories, swimming activity, and orientation to a proximal cue in a visible platform water maze test and burrowing and nest building activity. To compare the performance of mice across all tests, we used z-score normalization of data. The analyses revealed that the behavior of the transgenic mice was significantly compromised in cognitive as well as in noncognitive domains. Combining scores across multiple behavioral tests produced an integrated index characterizing the overall phenotypic abnormality in this model of AD-like amyloidosis. Assessing multiple behavioral domains provides a broader view of the breadth of impairments in multiple behavioral systems. Greater implementation of such approaches could enable reliable and clinically predictive evaluation of therapeutics in mouse models of amyloidosis.

Widespread and efficient transduction of spinal cord and brain following neonatal AAV injection and potential disease modifying effect in ALS mice.

The architecture of the spinal cord makes efficient delivery of recombinant adeno-associated virus (rAAV) vectors throughout the neuraxis challenging. We describe a paradigm in which small amounts of virus delivered intraspinally to newborn mice result in robust rAAV-mediated transgene expression in the spinal cord. We compared the efficacy of rAAV2/1, 2/5, 2/8, and 2/9 encoding EGFP delivered to the hindlimb muscle (IM), cisterna magna (ICM), or lumbar spinal cord (IS) of neonatal pups. IS injection of all four capsids resulted in robust transduction of the spinal cord with rAAV2/5, 2/8, and 2/9 vectors appearing to be transported to brain. ICM injection resulted in widespread expression of EGFP in the brain, and upper spinal cord. IM injection resulted in robust muscle expression, with only rAAV2/8 and 2/9 transducing spinal motor and sensory neurons. As proof of concept, we use the IS paradigm to express murine Interleukin (IL)-10 in the spinal cord of the SOD1-G93A transgenic mouse model of amyotrophic lateral sclerosis. We show that expression of IL-10 in the spinal axis of SOD1-G93A mice altered the immune milieu and significantly prolonged survival. These data establish an efficient paradigm for somatic transgene delivery of therapeutic biologics to the spinal cord of mice.

Capsid serotype and timing of injection determines AAV transduction in the neonatal mice brain.

Adeno-associated virus (AAV) mediated gene expression is a powerful tool for gene therapy and preclinical studies. A comprehensive analysis of CNS cell type tropism, expression levels and biodistribution of different capsid serotypes has not yet been undertaken in neonatal rodents. Our previous studies show that intracerebroventricular injection with AAV2/1 on neonatal day P0 results in widespread CNS expression but the biodistribution is limited if injected beyond neonatal day P1. To extend these observations we explored the effect of timing of injection on tropism and biodistribution of six commonly used pseudotyped AAVs delivered in the cerebral ventricles of neonatal mice. We demonstrate that AAV2/8 and 2/9 resulted in the most widespread biodistribution in the brain. Most serotypes showed varying biodistribution depending on the day of injection. Injection on neonatal day P0 resulted in mostly neuronal transduction, whereas administration in later periods of development (24-84 hours postnatal) resulted in more non-neuronal transduction. AAV2/5 showed widespread transduction of astrocytes irrespective of the time of injection. None of the serotypes tested showed any microglial transduction. This study demonstrates that both capsid serotype and timing of injection influence the regional and cell-type distribution of AAV in neonatal rodents, and emphasizes the utility of pseudotyped AAV vectors for translational gene therapy paradigms.

Comment on "ApoE-directed therapeutics rapidly clear ß-amyloid and reverse deficits in AD mouse models".

Cramer et al. (Reports, 23 March 2012, p. 1503; published online 9 February 2012) demonstrates short-term bexarotene treatment clearing preexisting ß-amyloid deposits from the brains of APP/PS1ΔE9 mice with low amyloid burden, providing a rationale for repurposing this anticancer agent as an Alzheimer's disease (AD) therapeutic. Using a nearly identical treatment regimen, we were unable to detect any evidence of drug efficacy despite demonstration of target engagement.