Disease Transgenic Mice Research Articles

FBXL16: a new regulator of neuroinflammation and cognition in Alzheimer’s disease through the ubiquitination-dependent degradation of amyloid precursor protein

BackgroundActivating the ubiquitin-proteasome system to dismantle disease- related proteins such as tau, β-amyloid, APP, and α-synuclein is an important focus in the research of neurodegenerative proteinopathy. By analyzing the serum RNA extracted from wild-type and Alzheimer’s disease (AD) transgenic mice at different ages (4, 8, and 12 months), this study revealed a new protective role of FBXL16 in AD, primarily through facilitating the degradation of disease-related proteins via the ubiquitin proteasome system.MethodsProteomic analysis were conducted using protein lysates from HEK293 cells overexpressing FBXL16 to identify potential interacting proteins that interact with FBXL16. Subsequent experiments demonstrated that FBXL16 promotes the proteasomal degradation of the APP protein, as evidenced by co-immunoprecipitation with MG132 and cycloheximide (CHX), immunohistochemistry (IHC) and immunocytochemistry (ICC). Memory and cognitive improvements were observed in 3×Tg AD mice through the use of a lentivirus-mediated approach to generate a brain-specific AD mouse model overexpressing FBXL16 via stereotaxic injection. Furthermore, a brain-specific conditional knockout (cko) FBXL16 mouse model was generated and employed to further confirm the functional role of FBXL 16 in AD via various behavioral tests including Morris water maze and Y-maze.ResultsThe level of FBXL16 in the brains of transgenic APP/PSEN mice with AD decreased with age. Accelerated degradation of APP was observed when FBXL16 was overexpressed in the hippocampi of these AD mice via a lentivirus. This process led to notable improvements in cognitive impairments and reductions in neuroinflammation. Further studies using proteomics and bioinformatics techniques identified transcription factors and binding proteins associated with FBXL16, providing deeper insights into the potential role of FBXL16 in the regulation of AD. Finally, the in vivo effects of FBXL16 deficiency were further substantiated in cko mice, which overexpress Aβ but specifically lack FBXL16 in the brain region.ConclusionsThese findings suggest that FBXL16 could be a new regulator of AD. These findings provide a foundation for further research into drug development and potential therapeutic strategies to combat other related neurodegenerative proteinopathies.

A pro-drug derivative of carnosic acid activates the Nrf2 transcriptional pathway and shows efficacy in Alzheimer’s disease transgenic mice

A pro-drug derivative of carnosic acid activates the Nrf2 transcriptional pathway and shows efficacy in Alzheimer’s disease transgenic mice

High-Density Lipoprotein Mimetic Peptide 4F Reduces Toxic Amyloid-Beta Exposure to the Blood-Brain Barrier Endothelium in Alzheimer's Disease Transgenic Mice.

Aβ accumulation in the blood-brain barrier (BBB) endothelium, which lines the cerebrovascular lumen, is a significant contributor to cerebrovascular dysfunction in Alzheimer's disease (AD). Reduced high-density lipoprotein (HDL) levels are associated with increased AD risk, and the HDL mimetic peptide 4F has been developed as a promising therapeutic agent to improve cerebrovascular health in AD. In this study, we evaluated the impact of 4F on 125I-Aβ42 blood-to-brain distribution using dynamic SPECT/CT imaging in both wild-type and APP/PS1 transgenic mice. Graphical analysis of the imaging data demonstrated that 4F significantly reduced the blood-to-brain influx rate in wild-type mice and the distribution of 125I-Aβ42 in the BBB endothelium in APP/PS1 mice. To elucidate the molecular mechanisms underlying the effect of 4F, we evaluated its impact on the p38 pathway and its role in mediating Aβ42 trafficking in human BBB endothelial cell monolayers. Treatment with 4F significantly decreased Aβ42 induced p38 activation in BBB endothelial cells. Furthermore, inhibition of p38 kinase significantly reduced endothelial accumulation of fluorescence-labeled Aβ42 and luminal-to-abluminal permeability across the cell monolayer. While our previous publication has hinted at the potential of 4F to reduce Aβ accumulation in the brain parenchyma, the current findings demonstrated the protective effect of 4F in reducing Aβ42 accumulation in the BBB endothelium of AD transgenic mice. These findings revealed the impact of a clinically tested agent, the HDL mimetic peptide 4F, on Aβ exposure to the BBB endothelium and offer novel mechanistic insights into potential therapeutic strategies to treat cerebrovascular dysfunction in AD.

Delivering DNA Aptamers Across the Blood-Brain Barrier Reveals Heterogeneous Decreased ATP in Different Brain Regions of Alzheimer's Disease Mouse Models.

DNA aptamers have been developed as sensors to detect metabolites with high sensitivity, selectivity, and biocompatibility. While they are effective in sensing important targets in the brain, the lack of methods for their efficient delivery across the blood-brain barrier (BBB) has significantly hindered their applications in brain research. To address this issue, we herein report the development of brain cell-derived exosomes as endogenous BBB delivery vehicles to deliver an ATP-responsive aptamer across the BBB of live mice for noninvasive live brain imaging. We found that the system uses endosome recycling to transfer the sensors between the delivered exosomes and native recycling endosomes, resulting in high delivery efficiencies. Using this system, we observed unique signal distributions for ATP across different brain regions, with significant accumulation in the subiculum and cortex in healthy mice. In an Alzheimer's disease transgenic mouse model, ATP levels decreased in the subiculum and cortex, demonstrating this method's capability to determine metabolite location and relative abundance with high spatial resolution in vivo. Since DNA aptamers have been obtained for many other targets, the method developed in this work can be applied to deliver sensors across the BBB to image a wide range of other brain-related metabolites.

Integrated Genome-Wide Analysis of DNA Methylation and Gene Expression in the Hippocampi of 5xFAD Alzheimer's Disease Mouse Model.

DNA methylation forms 5-methylcytosine and its regulation in the hippocampus is critical for learning and memory. Indeed, dysregulation of DNA methylation is associated with neurological diseases. Alzheimer's disease (AD) is the predominant of dementia and a neurodegenerative disorder. We examined the learning and memory function in 3- and 9-month-old wild-type and 5xfamiliar Alzheimer's disease (5xFAD) transgenic mice by performing the object recognition memory and Y-maze tests, and identified the hippocampal amyloid beta burden. To investigate the epigenetically regulated genes involved in the development or neuropathology of AD, we performed genome-wide DNA methylation sequencing and RNA sequencing analyses in the hippocampus of 9-month-old wild-type and 5xFAD tg mice. To validate the genes inversely regulated by epigenetics, we confirmed their methylation status and mRNA levels. At 9 months of age, 5xFAD tg mice showed significant cognitive impairment and amyloid-beta plaques in the hippocampus. DNA methylation sequencing identified a total of 13,777 differentially methylated regions, including 4484 of hyper- and 9293 of hypomethylated regions, that are associated with several gene ontology (GO) terms including 'nervous system development' and 'axon guidance'. In RNA sequencing analysis, we confirmed a total of 101 differentially expressed genes, including 52 up- and 49 downregulated genes, associated with GO functions such as 'positive regulation of synaptic transmission, glutamatergic' and 'actin filament organization'. Through further integrated analysis of DNA methylation and RNA sequencing, three epigenetically regulated genes were selected: thymus cell antigen 1, theta (Thy1), myosin VI (Myo6), and filamin A-interacting protein 1-like (Filip1l). The methylation level of Thy1 decreased and its mRNA levels increased, whereas that of Myo6 and Filip1l increased and their mRNA levels decreased. The common functions of these three genes may be associated with the neural cytoskeleton and synaptic plasticity. We suggest that the candidate genes epigenetically play a role in AD-associated neuropathology (i.e., amyloid-beta plaques) and memory deficit by influencing neural structure and synaptic plasticity. Furthermore, counteracting dysregulated epigenetic changes may delay or ameliorate AD onset or symptoms.

Feasibility of measuring blood-brain barrier permeability using ultra-short echo time radial magnetic resonance imaging.

The purpose of this study is to evaluate the feasibility of using 3-dimensional (3D) ultra-short echo time (UTE) radial imaging method for measurement of the permeability of the blood-brain barrier (BBB) to gadolinium-based contrast agent. In this study, we propose to use the golden-angle radial sparse parallel (GRASP) method with 3D center-out trajectories for UTE, hence named as 3D UTE-GRASP. We first examined the feasibility of using 3D UTE-GRASP dynamic contrast-enhanced (DCE)-magnetic resonance imaging (MRI) for differentiating subtle BBB disruptions induced by focused ultrasound (FUS). Then, we examined the BBB permeability changes in Alzheimer's disease (AD) pathology using Alzheimer's disease transgenic mice (5xFAD) at different ages. For FUS experiments, we used four Sprague Dawley rats at similar ages where we compared BBB permeability of each rat receiving the FUS sonication with different acoustic power (0.4-1.0MPa). For AD transgenic mice experiments, we included three 5xFAD mice (6, 12, and 16 months old) and three wild-type mice (4, 8, and 12 months old). The result from FUS experiments showed a progressive increase in BBB permeability with increase of acoustic power (p <.05), demonstrating the sensitivity of DCE-MRI method for detecting subtle changes in BBB disruption. Our AD transgenic mice experiments suggest an early BBB disruption in 5xFAD mice, which is further impaired with aging. The results in this study substantiate the feasibility of using the proposed 3D UTE-GRASP method for detecting subtle BBB permeability changes expected in neurodegenerative diseases, such as AD.

Neuroprotective Effects and Cognitive Enhancement of Allomargaritarine in 5xFAD Alzheimer's Disease Mice Model

Here, we report the results of an investigation of the neuroprotective effects of securinine with tryptamine conjugate-allomargaritarine (2b), previously selected as the leading compound among a wide range of natural derivatives. 2b was synthesized from securinine using various Lewis acids as catalysts. In addition to the antioxidant and cytoprotective properties previously shown for 2b, in this work,&lt;em&gt; in vitro&lt;/em&gt; analysis of the biological activity of the compound demonstrated that this conjugate is also able to influence the primary pathogenetic mechanism of Alzheimer's disease - proteinopathy, modulating the homeostasis of β-amyloid peptide. In particular, it was found that 2b is an effective inhibitor of β-secretase 1 - an enzyme responsible for initiating the generation of pathological forms of β-amyloid peptide, as well as directly preventing the pathological aggregation of Aβ&lt;sub&gt;1-42&lt;/sub&gt;. As a compound with a promising biological activity profile found &lt;em&gt;in vitro&lt;/em&gt;, 2b has also demonstrated excellent neuroprotective effects on the &lt;em&gt;in vivo&lt;/em&gt; 5xFAD Alzheimer's disease transgenic mice model. Thus, 2b effectively restored cognitive dysfunction: short-term and long-term episodic and spatial memory, which in the &lt;em&gt;post-mortem&lt;/em&gt; studies was also accompanied by a decrease in the number of amyloid deposits and the intensity of oxidative stress in brain samples. These results provide an opportunity to draw a line under years of research on the neuroprotective potential of 2b as a viable therapy for Alzheimer's disease.

Striatal dopamine neurotransmission is altered in age- and region-specific manner in a Parkinson’s disease transgenic mouse

Dopamine (DA) plays a critical role in striatal motor control. The drop in DA level within the dorsal striatum is directly associated with the appearance of motor symptoms in Parkinson’s disease (PD). The progression of the disease and inherent disruption of the DA neurotransmission has been closely related to accumulation of the synaptic protein α-synuclein. However, it is still unclear how α-synuclein affects dopaminergic terminals in different areas of dorsal striatum. Here we demonstrate that the overexpression of human α-synuclein (h-α-syn) interferes with the striatal DA neurotransmission in an age‐dependent manner, preferentially in the dorsolateral striatum (DLS) of PDGF-h-α-syn mice. While 3-month-old mice showed an increase at the onset of h-α-syn accumulation in the DLS, 12-month-old mice revealed a decrease in electrically-evoked DA release. The enhanced DA release in 3-month-old mice coincided with better performance in a behavioural task. Notably, DA amplitude alterations were also accompanied by a delay in the DA clearance independently from the animal age. Structurally, dopamine transporter (DAT) was found to be redistributed in larger DAT-positive clumps only in the DLS of 3- and 12-month-old mice. Together, our data provide new insight into the vulnerability of DLS and suggest DAT-related dysfunctionalities from the very early stages of h-α-syn accumulation.

A novel BODIPY-based theranostic agent for in vivo fluorescence imaging of cerebral Aβ and ameliorating Aβ-associated disorders in Alzheimer's disease transgenic mice.

β-Amyloid (Aβ) aggregation is increasingly recognized as both a biomarker and an inducer of the progression of Alzheimer's disease (AD). Here, we describe a novel fluorescent probe P14, developed based on the BODIPY structure, capable of simultaneous visualization and inhibition of Aβ aggregation in vivo. P14 shows high binding affinity to Aβ aggregates and selectively labels Aβ plaques in the brain slices of APP/PS1 mice. Moreover, P14 is able to visualize overloaded Aβ in both APP/PS1 and 5 × FAD transgenic mice in vivo. From the aspect of potential therapeutic effects, P14 administration inhibits Aβ aggregation and alleviates Aβ-induced neuronal damage in vitro, as well as reduces central Aβ deposition and ameliorates cognitive impairment in APP/PS1 transgenic mice in vivo. Finally, P14 is applied to monitor the progression of Aβ aggregation in the brain of 5 × FAD transgenic mice and the intervention effect itself by fluorescence imaging. In summary, the discovery of this fluorescent agent might provide important clues for the future development of theranostic drug candidates targeting Aβ aggregation in AD.

Amyloid-β mediates intestinal dysfunction and enteric neurons loss in Alzheimer's disease transgenic mouse.

Alzheimer's disease (AD) is traditionally considered as a brain disorder featured by amyloid-β (Aβ) deposition. The current study on whether pathological changes of AD extend to the enteric nervous system (ENS) is still in its infancy. In this study, we found enteric Aβ deposition, intestinal dysfunction, and colonic inflammation in the young APP/PS1 mice. Moreover, these mice exhibited cholinergic and nitrergic signaling pathways damages and enteric neuronal loss. Our data show that Aβ42 treatment remarkably affected the gene expression of cultured myenteric neurons and the spontaneous contraction of intestinal smooth muscles. The intra-colon administration of Aβ42 induced ENS dysfunction, brain gliosis, and β-amyloidosis-like changes in the wild-type mice. Our results suggest that ENS mirrors the neuropathology observed in AD brains, and intestinal pathological changes may represent the prodromal events, which contribute to brain pathology in AD. In summary, our findings provide new opportunities for AD early diagnosis and prevention.

Reduced slow-wave activity and autonomic dysfunction during sleep precede cognitive deficits in Alzheimer’s disease transgenic mice

Occurrence of amyloid-β (Aβ) aggregation in brain begins before the clinical onset of Alzheimer’s disease (AD), as preclinical AD. Studies have reported that sleep problems and autonomic dysfunction associate closely with AD. However, whether they, especially the interaction between sleep and autonomic function, play critical roles in preclinical AD are unclear. Therefore, we investigated how sleep patterns and autonomic regulation at different sleep–wake stages changed and whether they were related to cognitive performance in pathogenesis of AD mice. Polysomnographic recordings in freely-moving APP/PS1 and wild-type (WT) littermates were collected to study sleep patterns and autonomic function at 4 (early disease stage) and 8 months of age (advanced disease stage), cognitive tasks including novel object recognition and Morris water maze were performed, and Aβ levels in brain were measured. APP/PS1 mice at early stage of AD pathology with Aβ aggregation but without significant differences in cognitive performance had frequent sleep–wake transitions, lower sleep-related delta power percentage, lower overall autonomic activity, and lower parasympathetic activity mainly during sleep compared with WT mice. The same phenomenon was observed in advanced-stage APP/PS1 mice with significant cognitive deficits. In mice at both disease stages, sleep-related delta power percentage correlated positively with memory performance. At early stage, memory performance correlated positively with sympathetic activity during wakefulness; at advanced stage, memory performance correlated positively with parasympathetic activity during both wakefulness and sleep. In conclusion, sleep quality and distinction between wake- and sleep-related autonomic function may be biomarkers for early AD detection.

Maternal separation differentially modulates early pathology by sex in 5xFAD Alzheimer’s disease-transgenic mice

Alzheimer’s disease (AD) is the most common neurodegenerative disease. Most cases of AD are considered idiopathic and likely due to a combination of genetic, environmental, and lifestyle-related risk factors. Despite occurring decades before the typical age of an AD diagnosis, early-life stress (ELS) has been suggested to have long-lasting effects that may contribute to AD risk and pathogenesis. Still, the mechanisms that underlie the role of ELS on AD risk remain largely unknown. Here, we used 5xFAD transgenic mice to study relatively short-term alterations related to ELS in an AD-like susceptible mouse model at 6 weeks of age. To model ELS, we separated pups from their dams for 3 h per day from postnatal day 2–14. Around 6 weeks of age, we found that maternally separated (MS) 5xFAD mice, particularly female mice, displayed increased amyloid-β-immunoreactivity in the anterior cingulate cortex (ACC) and basolateral amygdala (BLA). In anterior cingulate cortex, we also noted significantly increased intraneuronal amyloid-β-immunoreactivity associated with MS but only in female mice. Moreover, IBA1-positive DAPI density was significantly increased in relation to MS in ACC and BLA, and microglia in BLA of MS mice had significantly different morphology compared to microglia in non-MS 5xFAD mice. Cytokine analysis showed that male MS mice, specifically, had increased levels of neuroinflammatory markers CXCL1 and IL-10 in hippocampal extracts compared to non-MS counterparts. Additionally, hippocampal extracts from both male and female MS 5xFAD mice had decreased levels of synapse- and activity-related markers Bdnf, 5htr6, Cox2, and Syp in hippocampus. Lastly, we performed behavioral tests to evaluate anxiety- and depressive-like behavior and working memory but could not detect any significant differences between groups. Overall, we detected several sex-specific molecular and cellular alterations in 6-week-old adolescent 5xFAD mice associated with MS that may help explain the connection between ELS and AD risk.

Second passage experiments of chronic wasting disease in transgenic mice overexpressing human prion protein

Chronic wasting disease (CWD) is a prion disease of cervids including deer, elk, reindeer, and moose. Human consumption of cervids is common, therefore assessing the risk potential of CWD transmission to humans is critical. In a previous study, we tested CWD transmission via intracerebral inoculation into transgenic mice (tg66 and tgRM) that over-expressed human prion protein. Mice screened by traditional prion detection assays were negative. However, in a group of 88 mice screened by the ultrasensitive RT-QuIC assay, we identified 4 tg66 mice that produced inconsistent positive RT-QuIC reactions. These data could be false positive reactions, residual input inoculum or indicative of subclinical infections suggestive of cross species transmission of CWD to humans. Additional experiments were required to understand the nature of the prion seeding activity in this model. In this manuscript, second passage experiments using brains from mice with weak prion seeding activity showed they were not infectious to additional recipient tg66 mice. Clearance experiments showed that input CWD prion seeding activity was eliminated by 180 days in tg66 mice and PrPKO mice, which are unable to replicate prion protein, indicating that the weak positive levels of seeding activity detected at later time points was not likely residual inoculum. The failure of CWD prions to cause disease in tg66 after two sequential passages suggested that a strong species barrier prevented CWD infection of mice expressing human prion protein.

Bona fide atypical scrapie faithfully reproduced for the first time in a rodent model

Atypical Scrapie, which is not linked to epidemics, is assumed to be an idiopathic spontaneous prion disease in small ruminants. Therefore, its occurrence is unlikely to be controlled through selective breeding or other strategies as it is done for classical scrapie outbreaks. Its spontaneous nature and its sporadic incidence worldwide is reminiscent of the incidence of idiopathic spontaneous prion diseases in humans, which account for more than 85% of the cases in humans. Hence, developing animal models that consistently reproduce this phenomenon of spontaneous PrP misfolding, is of importance to study the pathobiology of idiopathic spontaneous prion disorders. Transgenic mice overexpressing sheep PrPC with I112 polymorphism (TgShI112, 1–2 × PrP levels compared to sheep brain) manifest clinical signs of a spongiform encephalopathy spontaneously as early as 380 days of age. The brains of these animals show the neuropathological hallmarks of prion disease and biochemical analyses of the misfolded prion protein show a ladder-like PrPres pattern with a predominant 7–10 kDa band. Brain homogenates from spontaneously diseased transgenic mice were inoculated in several models to assess their transmissibility and characterize the prion strain generated: TgShI112 (ovine I112 ARQ PrPC), Tg338 (ovine VRQ PrPC), Tg501 (ovine ARQ PrPC), Tg340 (human M129 PrPC), Tg361 (human V129 PrPC), TgVole (bank vole I109 PrPC), bank vole (I109I PrPC), and sheep (AHQ/ARR and AHQ/AHQ churra-tensina breeds). Our analysis of the results of these bioassays concludes that the strain generated in this model is indistinguishable to that causing atypical scrapie (Nor98). Thus, we present the first faithful model for a bona fide, transmissible, ovine, atypical scrapie prion disease.

Longitudinal spatial mapping of lipid metabolites reveals pre-symptomatic changes in the hippocampi of Huntington's disease transgenic mice

In Huntington's disease (HD), a key pathological feature includes the development of inclusion-bodies of fragments of the mutant huntingtin protein in the neurons of the striatum and hippocampus. To examine the molecular changes associated with inclusion-body formation, we applied MALDI-mass spectrometry imaging and deuterium pulse labelling to determine lipid levels and synthesis rates in the hippocampus of a transgenic mouse model of HD (R6/1 line). The R6/1 HD mice lacked inclusions in the hippocampus at 6 weeks of age (pre-symptomatic), whereas inclusions were pervasive by 16 weeks of age (symptomatic). Hippocampal subfields (CA1, CA3 and DG), which formed the highest density of inclusion formation in the mouse brain showed a reduction in the relative abundance of neuron-enriched lipids that have roles in neurotransmission, synaptic plasticity, neurogenesis, and ER-stress protection. Lipids involved in the adaptive response to ER stress (phosphatidylinositol, phosphatidic acid, and ganglioside classes) displayed increased rates of synthesis in HD mice relative to WT mice at all the ages examined, including prior to the formation of the inclusion bodies. Our findings, therefore, support a role for ER stress occurring pre-symptomatically and potentially contributing to pathological mechanisms underlying HD.

Dysregulation of sphingosine-1-phosphate (S1P) and S1P receptor 1 signaling in the 5xFAD mouse model of Alzheimer’s disease

Sphingolipid-1-phosphate (S1P) signaling through the activation S1P receptors (S1PRs) plays critical roles in cellular events in the brain. Aberrant S1P metabolism has been identified in the brains of Alzheimer’s disease (AD) patients. Our recent studies have shown that treatment with fingolimod, an analog of sphingosine, provides neuroprotective effects in five familiar Alzheimer disease (5xFAD) transgenic mice, resulting in the reduction of amyloid-β (Aβ) neurotoxicity, inhibition of activation of microglia and astrocytes, increased hippocampal neurogenesis, and improved learning and memory. However, the pathways by which dysfunctional S1P and S1PR signaling may associate with the development of AD-like pathology remain unknown.In this study, we investigated the alteration of signaling of S1P/S1P receptor 1 (S1PR1), the most abundant S1PR subtype in the brain, in the cortex of 5xFAD transgenic mice at 3, 8, and 14 months of age. Compared to non-transgenic wildtype (WT) littermates, we found significant decreased levels of sphingosine kinases (SphKs), increased S1P lyase (S1PL), and increased S1PR1 in 8- and 14-month-old, but not in 3-month-old 5xFAD mice. Furthermore, we detected increased activation of the S1PR1 downstream pathway of Akt/mTor/Tau signaling in aging 5xFAD mice. Treatment with fingolimod from 1 to 8 months of age reversed the levels of SphKs, S1PL, and furthermore, those of S1PR1 and its downstream pathway of Akt/mTor/Tau signaling. Together the data reveal that dysregulation of S1P and S1PR signaling may associate with the development of AD-like pathology through Akt/mTor/Tau signaling.

Transcriptomic Profiling Identifies CD8+ T Cells in the Brain of Aged and Alzheimer's Disease Transgenic Mice as Tissue-Resident Memory T Cells.

Peripheral immune cell infiltration into the brain is a prominent feature in aging and various neurodegenerative diseases such as Alzheimer's disease (AD). As AD progresses, CD8+ T cells infiltrate into the brain parenchyma, where they tightly associate with neurons and microglia. The functional properties of CD8+ T cells in the brain are largely unknown. To gain further insights into the putative functions of CD8+ T cells in the brain, we explored and compared the transcriptomic profile of CD8+ T cells isolated from the brain and blood of transgenic AD (APPswe/PSEN1dE9, line 85 [APP-PS1]) and age-matched wild-type (WT) mice. Brain CD8+ T cells of APP-PS1 and WT animals had similar transcriptomic profiles and substantially differed from blood circulating CD8+ T cells. The gene signature of brain CD8+ T cells identified them as tissue-resident memory (Trm) T cells. Gene Ontology enrichment and Kyoto Encyclopedia of Genes and Genomes pathway analysis on the significantly upregulated genes revealed overrepresentation of biological processes involved in IFN-β signaling and the response to viral infections. Furthermore, brain CD8+ T cells of APP-PS1 and aged WT mice showed similar differentially regulated genes as brain Trm CD8+ T cells in mouse models with acute virus infection, chronic parasite infection, and tumor growth. In conclusion, our profiling of brain CD8+ T cells suggests that in AD, these cells exhibit similar adaptive immune responses as in other inflammatory diseases of the CNS, potentially opening the door for immunotherapy in AD.

Tolfenamic acid inhibits ROS-generating oxidase Nox1-regulated p53 activity in intrastriatal injection of malonic acid rats

It has been reported that wild-type p53-induced gene 1 (Wig1), which is downstream of p53, regulates the expression of mutant huntingtin protein (mHtt) in Huntington’s disease (HD) patients and transgenic mouse brains. Intrastriatal injection of malonic acid in rats is often used as a model to study the pathological changes of Huntington’s disease, and this model has the advantages of a fast preparation and low cost. Therefore, in this study, we used intrastriatal injections of 6 μM malonic acid in rats to evaluate the effect of tolfenamic acid on motor and cognitive deficits and the effect of 6 mg/kg and 32 mg/kg tolfenamic acid on p53 and its downstream targets, such as Wig1. The results showed that 32 mg/kg tolfenamic acid attenuated motor and spatial memory dysfunction, prevented Nox1-mediated reactive oxygen species (ROS) production, and downregulated the activity of p53 by increasing the phosphorylation level at the Ser378 site and decreasing the acetylation level at the Lys382 site. Tolfenamic acid reduced mouse double minute 2 (Mdm2), phosphatase and tensin homologue (Pten), P53-upregulated modulator of apoptosis (Puma) and Bcl2-associated X (Bax) at the mRNA level to inhibit apoptosis and downregulated sestrin 2 (Sesn2) and hypoxia inducible factor 1, alpha subunit (Hif-1α) mRNA levels to exert antioxidative stress effects. In addition, 32 mg/kg tolfenamic acid played a role in neuroprotection by decreasing the terminal deoxynucleotidyl transferase-mediated dUTP nick end labelling (TUNEL)-positive cell numbers. However, there was no difference in the Wig mRNA level among all groups, and tolfenamic acid could not decrease the protein level of Wig1. In conclusion, tolfenamic acid inhibited the ROS-generating oxidase Nox1-regulated p53 activity and attenuated motor and spatial memory deficits in malonic acid-injected rats.

Bryostatin Placebo-Controlled Trials Indicate Cognitive Restoration Above Baseline for Advanced Alzheimer's Disease in the Absence of Memantine1.

Background:In pre-clinical studies of Alzheimer’s disease (AD) transgenic mice, bryostatin restored synaptic connections, prevented neuronal death, reduced amyloid plaques, and reduced neurofibrillary tangles.Objective:Within pre-specified cohorts of advanced AD patients in two double-blind placebo-controlled bryostatin Phase II trials, to conduct exploratory statistical analyses of patients with identical conditions of enrollment and treatment.Methods:Severe Impairment Battery (SIB) scores above baseline at 5, 9, and 13 weeks were analyzed initially in the complete cases, with multiple imputation methods based on an iterative Markov chain Monte Carlo algorithm used for missing SIB scores. To mitigate confounding by a chance imbalance of 4.9 SIB baseline scores (Study #203), each patient was used as their own control with differences in 13-week SIB from baseline in single trial and pooled analyses to measure benefit at 13 weeks using general estimating equations (GEE) modeling.Results:Patients treated with bryostatin pre-specified at Mini-Mental State Examination scores 10–14, without memantine, showed baseline balance, complete safety, and SIB improvements at 13 weeks with multiple imputation analysis: Study #203 = 4.1 SIB points above baseline (p = 0.005), and Study #202 = 4.2 SIB points above baseline (p = 0.016). An increased power (N = 95) “pooled analysis” showed an increased SIB over time and a higher mean SIB at 13 weeks in the bryostatin treatment group (p < 0.001) but not significant (NS) for the placebo patients.Conclusion:Pre-specified exploratory analyses for the individual trials and the pooled trials confirmed significant bryostatin-induced improvement over baseline (treatment p < 0.001, placebo NS).

An integrative analysis of miRNA and mRNA expression in the brains of Alzheimer's disease transgenic mice after real-world PM2.5 exposure

Fine particulate matter (PM2.5) is associated with increased risks of Alzheimer's disease (AD), yet the toxicological mechanisms of PM2.5 promoting AD remain unclear. In this study, wild-type and APP/PS1 transgenic mice (AD mice) were exposed to either filtered air (FA) or PM2.5 for eight weeks with a real-world exposure system in Taiyuan, China (mean PM2.5 concentration in the cage was 61 µg/m3). We found that PM2.5 exposure could remarkably aggravate AD mice's ethological and brain ultrastructural damage, along with the elevation of the pro-inflammatory cytokines (IL-6 and TNF-α), Aβ-42 and AChE levels and the decline of ChAT levels in the brains. Based on high-throughput sequencing results, some differentially expressed (DE) mRNAs and DE miRNAs in the brains of AD mice after PM2.5 exposure were screened. Using RT-qPCR, seven DE miRNAs (mmu-miR-193b-5p, 122b-5p, 466h-3p, 10b-5p, 1895, 384–5p, and 6412) and six genes (Pcdhgb8, Unc13b, Robo3, Prph, Pter, and Tbata) were evidenced the and verified. Two miRNA-target gene pairs (miR-125b-Pcdhgb8 pair and miR-466h-3p-IL-17Rα/TGF-βR2/Aβ-42/AChE pairs) were demonstrated that they were more related to PM2.5-induced brain injury. Results of Gene Ontology (GO) pathways and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways predicted that synaptic and postsynaptic regulation, axon guidance, Wnt, MAPK, and mTOR pathways might be the possible regulatory mechanisms associated with pathological response. These revealed that PM2.5-elevated pro-inflammatory cytokine levels and PM2.5-altered neurotransmitter levels in AD mice could be the important causes of brain damage and proposed the promising miRNA and mRNA biomarkers and potential miRNA-mRNA interaction networks of PM2.5-promoted AD.