Loss of TREM2 rescues hyperactivation of microglia, but not lysosomal deficits and neurotoxicity in models of progranulin deficiency.

Haploinsufficiency of the progranulin (PGRN)-encoding gene (GRN) causes frontotemporal lobar degeneration (GRN-FTLD) and results in microglial hyperactivation, TREM2 activation, lysosomal dysfunction, and TDP-43 deposition. To understand the contribution of microglial hyperactivation to pathology, we used genetic and pharmacological approaches to suppress TREM2-dependent transition of microglia from a homeostatic to a disease-associated state. Trem2 deficiency in Grn KO mice reduced microglia hyperactivation. To explore antibody-mediated pharmacological modulation of TREM2-dependent microglial states, we identified antagonistic TREM2 antibodies. Treatment of macrophages from GRN-FTLD patients with these antibodies led to reduced TREM2 signaling due to its enhanced shedding. Furthermore, TREM2 antibody-treated PGRN-deficient microglia derived from human-induced pluripotent stem cells showed reduced microglial hyperactivation, TREM2 signaling, and phagocytic activity, but lysosomal dysfunction was not rescued. Similarly, lysosomal dysfunction, lipid dysregulation, and glucose hypometabolism of Grn KO mice were not rescued by TREM2 ablation. Synaptic loss and neurofilament light-chain (NfL) levels, a biomarker for neurodegeneration, were further elevated in the Grn/Trem2 KO cerebrospinal fluid (CSF). These findings suggest that TREM2-dependent microglia hyperactivation in models of GRN deficiency does not promote neurotoxicity, but rather neuroprotection.

A custom-built step exergame training programme to prevent falls in people with multiple sclerosis: A multicentre randomised controlled trial.

BACKGROUND: Cognitive-motor step training can improve stepping, balance and mobility in people with multiple sclerosis (MS), but effectiveness in preventing falls has not been demonstrated. OBJECTIVES: This multisite randomised controlled trial aimed to determine whether 6 months of home-based step exergame training could reduce falls and improve associated risk factors compared with usual care in people with MS. METHODS: In total, 461 people with MS aged 22-81 years were randomly allocated to usual care (control) or unsupervised home-based step exergame training (120 minutes/week) for 6 months. The primary outcome was rate of falls over 6 months from randomisation. Secondary outcomes included physical, cognitive and psychosocial function at 6 months and falls over 12 months. RESULTS: Mean (standard deviation (SD)) weekly training duration was 70 (51) minutes over 6 months. Fall rates did not differ between intervention and control groups (incidence rates (95% confidence interval (CI)): 2.13 (1.57-2.69) versus 2.24 (1.35-3.13), respectively, incidence rate ratio: 0.96 (95% CI: 0.69-1.34, p = 0.816)). Intervention participants performed faster in tests of choice-stepping reaction time at 6 months. No serious training-related adverse events were reported. CONCLUSION: The step exergame training programme did not reduce falls among people with MS. However, it significantly improved choice-stepping reaction time which is critical to ambulate safely in daily life environment.

Patient-initiated follow-up for high-risk cutaneous squamous cell carcinoma. How we do it and two years of outcome data.

Why do we follow-up patients with high-risk cutaneous squamous-cell carcinomas (cSCC)? Current guidance suggests we offer post-treatment follow-up appointments at regular intervals for 24 months. Is this to improve prognosis, provide psychological support or find the next cancer? Recent data confirms recurrence and metastasis to be rarer events. We question whether these follow-up practices are truly 'needed' by introducing an option of patient-initiated follow-up (PIFU). We enrolled 476 cSCC patients (January 2020 - January 2023) who fulfilled the definition of high-risk cSCC based on guidelines in use at the time. 59 patients were excluded. 53% (n=250) did not re-contact us during the 2-year period, with no clinical record of complications or recurrences. 35% (n=167) utilised the PIFU pathway, of which 119 patients only required 1 face-to-face appointment. 7 patients (1.5%) developed metastatic disease, 11 patients developed recurrence, and 68 patients developed cSCC at another site. All lesions were identified by the patient via PIFU. We saved 1250 follow-up appointments from those that did not contact us (n=250); financially equating to £181,462.50. Our data implies that PIFU can be considered safe alternative practice for patients with cSCC. Patients independently identified the need for review without scheduled follow-up, making these appointments available to other patient cohorts. Although follow-up appointments may provide mental health support, they can be inconvenient and not the ideal use of our health resources. Our findings support a call for revision of existing skin cancer health policies, to cope with and subsequently improve our practices, to better patient care.

The relationship between micronutrient status, frailty, systemic inflammation, and clinical outcomes in patients admitted to hospital with COVID-19.

BACKGROUND: Micronutrients have been associated with disease severity and poorer clinical outcomes in patients with COVID-19. However, there is a paucity of studies examining if the relationship with micronutrient status and clinical outcomes is independent of recognised prognostic factors, specifically frailty and the systemic inflammatory response (SIR). The aim of the present study was to examine the relationship between micronutrient status, frailty, systemic inflammation, and clinical outcomes in patients admitted with COVID-19. METHODS: Retrospective analysis of prospectively collected data was performed on patients with confirmed COVID-19, admitted to hospital between the 1st April 2020-6th July 2020. Clinicopathological characteristics, frailty assessment, biochemical and micronutrient laboratory results were recorded. Frailty status was determined using the Clinical Frailty scale. SIR was determined using serum CRP. Clinical outcomes of interest were oxygen requirement, ITU admission and 30-day mortality. Categorical variables were analysed using chi-square test and binary logistics regression analysis. Continuous variables were analysed using the Mann-Whitney U or Kruskal Wallis tests. RESULTS: 281 patients were included. 55% (n = 155) were aged ≥ 70 years and 39% (n = 109) were male. 49% (n = 138) of patients were frail (CFS > 3). 86% (n = 242) of patients had a serum CRP > 10 mg/L. On univariate analysis, frailty was significantly associated with thirty-day mortality (p < 0.001). On univariate analysis, serum CRP was found to be significantly associated with an oxygen requirement on admission in non-frail patients (p = 0.004). Over a third (36%) of non-frail patients had a low vitamin B1, despite having normal reference range values of red cell B2, B6 and selenium. Furthermore, serum CRP was found to be significantly associated with a lower median red cell vitamin B1 (p = 0.029). CONCLUSION: Vitamin B1 stores may be depleted in COVID-19 patients experiencing a significant SIR and providing rationale for thiamine supplementation. Further longitudinal studies are warranted to delineate the trend in thiamine status following COVID-19.

Catalysis-dependent and redundant roles of Dma1 and Dma2 in maintenance of genome stability in Saccharomyces cerevisiae.

DNA double-strand breaks (DSBs) are among the deleterious lesions that are both endogenous and exogenous in origin and are repaired by nonhomologous end joining or homologous recombination. However, the molecular mechanisms responsible for maintaining genome stability remain incompletely understood. Here, we investigate the role of two E3 ligases, Dma1 and Dma2 (homologs of human RNF8), in the maintenance of genome stability in budding yeast. Using yeast spotting assays, chromatin immunoprecipitation and plasmid and chromosomal repair assays, we establish that Dma1 and Dma2 act in a redundant and a catalysis-dependent manner in the maintenance of genome stability, as well as localize to transcribed regions of the genome and increase in abundance upon phleomycin treatment. In addition, Dma1 and Dma2 are required for the normal kinetics of histone H4 acetylation under DNA damage conditions, genetically interact with RAD9 and SAE2, and are in a complex with Rad53 and histones. Taken together, our results demonstrate the requirement of Dma1 and Dma2 in regulating DNA repair pathway choice, preferentially affecting homologous recombination over nonhomologous end joining, and open up the possibility of using these candidates in manipulating the repair pathways toward precision genome editing.

Variation in Model-Based Economic Evaluations of Low-Dose Computed Tomography Screening for Lung Cancer: A Methodological Review.

OBJECTIVES: There is significant heterogeneity in the results of published model-based economic evaluations of low-dose computed tomography (LDCT) screening for lung cancer. We sought to understand and demonstrate how these models differ. METHODS: An expansion and update of a previous systematic review (N = 19). Databases (including MEDLINE and Embase) were searched. Studies were included if strategies involving (single or multiple) LDCT screening were compared with no screening or other imaging modalities, in a population at risk of lung cancer. More detailed data extraction of studies from the previous review was conducted. Studies were critically appraised using the Consensus Health Economic Criteria list. RESULTS: A total of 16 new studies met the inclusion criteria, giving a total of 35 studies. There are geographic and temporal differences and differences in screening intervals and eligible populations. Studies varied in the types of models used, for example, decision tree, Markov, and microsimulation models. Most conducted a cost-effectiveness analysis (using life-years gained) or cost-utility analysis. The potential for overdiagnosis was considered in many models, unlike with other potential consequences of screening. Some studies report considering lead-time bias, but fewer mention length bias. Generally, the more recent studies, involving more complex modeling, tended to meet more of the critical appraisal criteria, with notable exceptions. CONCLUSIONS: There are many differences across the economic evaluations contributing to variation in estimates of the cost-effectiveness of LDCT screening for lung cancer. Several methodological factors and evidence needs have been highlighted that will require consideration in future economic evaluations to achieve better agreement.

Systematic review of the impact of cannabinoids on neurobehavioral outcomes in preclinical models of traumatic and nontraumatic spinal cord injury.

STUDY DESIGN: Systematic review. OBJECTIVES: To evaluate the impact of cannabinoids on neurobehavioral outcomes in preclinical models of nontraumatic and traumatic spinal cord injury (SCI), with the aim of determining suitability for clinical trials involving SCI patients. METHODS: A systematic search was performed in MEDLINE and Embase databases, following registration with PROPSERO (CRD42019149671). Studies evaluating the impact of cannabinoids (agonists or antagonists) on neurobehavioral outcomes in preclinical models of nontraumatic and traumatic SCI were included. Data extracted from relevant studies, included sample characteristics, injury model, neurobehavioural outcomes assessed and study results. PRISMA guidelines were followed and the SYRCLE checklist was used to assess risk of bias. RESULTS: The search returned 8714 studies, 19 of which met our inclusion criteria. Sample sizes ranged from 23 to 390 animals. WIN 55,212-2 (n = 6) and AM 630 (n = 8) were the most used cannabinoid receptor agonist and antagonist respectively. Acute SCI models included traumatic injury (n = 16), ischaemia/reperfusion injury (n = 2), spinal cord cryoinjury (n = 1) and spinal cord ischaemia (n = 1). Assessment tools used assessed locomotor function, pain and anxiety. Cannabinoid receptor agonists resulted in statistically significant improvement in locomotor function in 9 out of 10 studies and pain outcomes in 6 out of 6 studies. CONCLUSION: Modulation of the endo-cannabinoid system has demonstrated significant improvement in both pain and locomotor function in pre-clinical SCI models; however, the risk of bias is unclear in all studies. These results may help to contextualise future translational clinical trials investigating whether cannabinoids can improve pain and locomotor function in SCI patients.

Telemetric Intracranial Pressure: A Snapshot Does not Give the Full Story.

Telemetric intracranial pressure (ICP) monitors are useful tools in the management of complex hydrocephalus and idiopathic intracranial hypertension (IIH). Clinicians may use them as a "snapshot" screening tool to assess shunt function or ICP. We compared "snapshot" telemetric ICP recordings with extended, in-patient periods of monitoring to determine whether this practice is safe and useful for clinical decision making.

Development, validation and effectiveness of diagnostic prediction tools for colorectal cancer in primary care: a systematic review.

BACKGROUND: Tools based on diagnostic prediction models are available to help general practitioners (GP) diagnose colorectal cancer. It is unclear how well they perform and whether they lead to increased or quicker diagnoses and ultimately impact on patient quality of life and/or survival. The aim of this systematic review is to evaluate the development, validation, effectiveness, and cost-effectiveness, of cancer diagnostic tools for colorectal cancer in primary care. METHODS: Electronic databases including Medline and Web of Science were searched in May 2017 (updated October 2019). Two reviewers independently screened titles, abstracts and full-texts. Studies were included if they reported the development, validation or accuracy of a prediction model, or assessed the effectiveness or cost-effectiveness of diagnostic tools based on prediction models to aid GP decision-making for symptomatic patients presenting with features potentially indicative of colorectal cancer. Data extraction and risk of bias were completed by one reviewer and checked by a second. A narrative synthesis was conducted. RESULTS: Eleven thousand one hundred thirteen records were screened and 23 studies met the inclusion criteria. Twenty-studies reported on the development, validation and/or accuracy of 13 prediction models: eight for colorectal cancer, five for cancer areas/types that include colorectal cancer. The Qcancer models were generally the best performing. Three impact studies met the inclusion criteria. Two (an RCT and a pre-post study) assessed tools based on the RAT prediction model. The third study looked at the impact of GP practices having access to RAT or Qcancer. Although the pre-post study reported a positive impact of the tools on outcomes, the results of the RCT and cross-sectional survey found no evidence that use of, or access to, the tools was associated with better outcomes. No study evaluated cost effectiveness. CONCLUSIONS: Many prediction models have been developed but none have been fully validated. Evidence demonstrating improved patient outcome of introducing the tools is the main deficiency and is essential given the imperfect classification achieved by all tools. This need is emphasised by the equivocal results of the small number of impact studies done so far.

Perisynaptic Schwann cells phagocytose nerve terminal debris in a mouse model of Guillain-Barré syndrome.

In mouse models of acute motor axonal neuropathy, anti-ganglioside antibodies (AGAbs) bind to motor axons, notably the distal nerve, and activate the complement cascade. While complement activation is well studied in this model, the role of inflammatory cells is unknown. Herein we aimed to investigate the contribution of phagocytic cells including macrophages, neutrophils and perisynaptic Schwann cells (pSCs) to distal nerve pathology. To observe this, we first created a subacute injury model of sufficient duration to allow inflammatory cell recruitment. Mice were injected intraperitoneally with an anti-GD1b monoclonal antibody that binds strongly to mouse motor nerve axons. Subsequently, mice received normal human serum as a source of complement. Dosing was titrated to allow humane survival of mice over a period of 3 days, yet still induce the characteristic neurological impairment. Behaviour and pathology were assessed in vivo using whole-body plethysmography and post-sacrifice by immunofluorescence and flow cytometry. ex vivo nerve-muscle preparations were used to investigate the acute phagocytic role of pSCs following distal nerve injury. Following complement activation at distal intramuscular nerve sites in the diaphragm macrophage localisation or numbers are not altered, nor do they shift to a pro- or anti-inflammatory phenotype. Similarly, neutrophils are not significantly recruited. Instead, ex vivo nerve-muscle preparations exposed to AGAb plus complement reveal that pSCs rapidly become phagocytic and engulf axonal debris. These data suggest that pSCs, rather than inflammatory cells, are the major cellular vehicle for axonal debris clearance following distal nerve injury, in contrast to larger nerve bundles where macrophage-mediated clearance predominates.

DIS3 isoforms vary in their endoribonuclease activity and are differentially expressed within haematological cancers.

DIS3 (defective in sister chromatid joining) is the catalytic subunit of the exosome, a protein complex involved in the 3'-5' degradation of RNAs. DIS3 is a highly conserved exoribonuclease, also known as Rrp44. Global sequencing studies have identified DIS3 as being mutated in a range of cancers, with a considerable incidence in multiple myeloma. In this work, we have identified two protein-coding isoforms of DIS3. Both isoforms are functionally relevant and result from alternative splicing. They differ from each other in the size of their N-terminal PIN (PilT N-terminal) domain, which has been shown to have endoribonuclease activity and tether DIS3 to the exosome. Isoform 1 encodes a full-length PIN domain, whereas the PIN domain of isoform 2 is shorter and is missing a segment with conserved amino acids. We have carried out biochemical activity assays on both isoforms of full-length DIS3 and the isolated PIN domains. We find that isoform 2, despite missing part of the PIN domain, has greater endonuclease activity compared with isoform 1. Examination of the available structural information allows us to provide a hypothesis to explain this altered behaviour. Our results also show that multiple myeloma patient cells and all cancer cell lines tested have higher levels of isoform 1 compared with isoform 2, whereas acute myeloid leukaemia and chronic myelomonocytic leukaemia patient cells and samples from healthy donors have similar levels of isoforms 1 and 2. Taken together, our data indicate that significant changes in the ratios of the two isoforms could be symptomatic of haematological cancers.

Conditional Deletion of Prnp Rescues Behavioral and Synaptic Deficits after Disease Onset in Transgenic Alzheimer's Disease.

Biochemical and genetic evidence implicate soluble oligomeric amyloid-ß (Aßo) in triggering Alzheimer's disease (AD) pathophysiology. Moreover, constitutive deletion of the Aßo-binding cellular prion protein (PrPC) prevents development of memory deficits in APPswe/PS1ΔE9 mice, a model of familial AD. Here, we define the role of PrPC to rescue or halt established AD endophenotypes in a therapeutic disease-modifying time window after symptom onset. Deletion of Prnp at either 12 or 16 months of age fully reverses hippocampal synapse loss and completely rescues preexisting behavioral deficits by 17 months. In contrast, but consistent with a neuronal function for Aßo/PrPC signaling, plaque density, microgliosis, and astrocytosis are not altered. Degeneration of catecholaminergic neurons remains unchanged by PrPC reduction after disease onset. These results define the potential of targeting PrPC as a disease-modifying therapy for certain AD-related phenotypes after disease onset.SIGNIFICANCE STATEMENT The study presented here further elucidates our understanding of the soluble oligomeric amyloid-ß-Aßo-binding cellular prion protein (PrPC) signaling pathway in a familial form of Alzheimer's disease (AD) by implicating PrPC as a potential therapeutic target for AD. In particular, genetic deletion of Prnp rescued several familial AD (FAD)-associated phenotypes after disease onset in a mouse model of FAD. This study underscores the therapeutic potential of PrPC deletion given that patients already present symptoms at the time of diagnosis.

Virus Infection of Plants Alters Pollinator Preference: A Payback for Susceptible Hosts?

Plant volatiles play important roles in attraction of certain pollinators and in host location by herbivorous insects. Virus infection induces changes in plant volatile emission profiles, and this can make plants more attractive to insect herbivores, such as aphids, that act as viral vectors. However, it is unknown if virus-induced alterations in volatile production affect plant-pollinator interactions. We found that volatiles emitted by cucumber mosaic virus (CMV)-infected tomato (Solanum lycopersicum) and Arabidopsis thaliana plants altered the foraging behaviour of bumblebees (Bombus terrestris). Virus-induced quantitative and qualitative changes in blends of volatile organic compounds emitted by tomato plants were identified by gas chromatography-coupled mass spectrometry. Experiments with a CMV mutant unable to express the 2b RNA silencing suppressor protein and with Arabidopsis silencing mutants implicate microRNAs in regulating emission of pollinator-perceivable volatiles. In tomato, CMV infection made plants emit volatiles attractive to bumblebees. Bumblebees pollinate tomato by 'buzzing' (sonicating) the flowers, which releases pollen and enhances self-fertilization and seed production as well as pollen export. Without buzz-pollination, CMV infection decreased seed yield, but when flowers of mock-inoculated and CMV-infected plants were buzz-pollinated, the increased seed yield for CMV-infected plants was similar to that for mock-inoculated plants. Increased pollinator preference can potentially increase plant reproductive success in two ways: i) as female parents, by increasing the probability that ovules are fertilized; ii) as male parents, by increasing pollen export. Mathematical modeling suggested that over a wide range of conditions in the wild, these increases to the number of offspring of infected susceptible plants resulting from increased pollinator preference could outweigh underlying strong selection pressures favoring pathogen resistance, allowing genes for disease susceptibility to persist in plant populations. We speculate that enhanced pollinator service for infected individuals in wild plant populations might provide mutual benefits to the virus and its susceptible hosts.

RNA-seq reveals post-transcriptional regulation of Drosophila insulin-like peptide dilp8 and the neuropeptide-like precursor Nplp2 by the exoribonuclease Pacman/XRN1.

Ribonucleases are critically important in many cellular and developmental processes and defects in their expression are associated with human disease. Pacman/XRN1 is a highly conserved cytoplasmic exoribonuclease which degrades RNAs in a 5'-3' direction. In Drosophila, null mutations in pacman result in small imaginal discs, a delay in onset of pupariation and lethality during the early pupal stage. In this paper, we have used RNA-seq in a genome-wide search for mRNAs misregulated in pacman null wing imaginal discs. Only 4.2% of genes are misregulated ±>2-fold in pacman null mutants compared to controls, in line with previous work showing that Pacman has specificity for particular mRNAs. Further analysis of the most upregulated mRNAs showed that Pacman post-transcriptionally regulates the expression of the secreted insulin-like peptide Dilp8. Dilp8 is related to human IGF-1, and has been shown to coordinate tissue growth with developmental timing in Drosophila. The increased expression of Dilp8 is consistent with the developmental delay seen in pacman null mutants. Our analysis, together with our previous results, show that the normal role of this exoribonuclease in imaginal discs is to suppress the expression of transcripts that are crucial in apoptosis and growth control during normal development.

Refinement and revalidation of the demoralization scale: The DS-II-external validity.

BACKGROUND: The recently refined Demoralization Scale-II (DS-II) is a 16-item, self-report measure of demoralization. Its 2 factors-Meaning and Purpose and Distress and Coping Ability-demonstrate sound internal validity, including item fit, unidimensionality, internal consistency, and test-retest reliability. The convergent and discriminant validity of the DS-II with various measures is reported here. METHODS: Patients who had cancer or other progressive diseases and were receiving palliative care (n = 211) completed a battery of questionnaires, including the DS-II and measures of symptom burden, quality of life, depression, and attitudes toward the end of life. Spearman ρ correlations were determined to assess convergent validity. Mann-Whitney U tests with calculated effect sizes were used to examine discriminant validity and establish the minimal clinically important difference (MCID). Cross-tabulation frequencies with chi-square analyses were used to examine discriminant validity with major depression. RESULTS: The DS-II demonstrated convergent validity with measures of psychological distress, quality of life, and attitudes toward the end of life. It also demonstrated discriminant validity, as the DS-II differentiated patients who had different functional performance levels and high/low symptoms, with a difference of 2 points between groups on the DS-II considered clinically meaningful. Furthermore, discriminant validity was demonstrated, as comorbidity with depression was not observed at moderate levels of demoralization. CONCLUSIONS: The DS-II has sound psychometric properties and is an appropriate measure of demoralization. Given its structural simplicity and brevity, it is likely to be a useful tool in meaning-centered therapies. Cancer 2016;122:2260-7. © 2016 American Cancer Society.

Refinement and revalidation of the demoralization scale: The DS-II-internal validity.

BACKGROUND: The Demoralization Scale (DS) was initially validated in 2004 to enable the measurement of demoralization in patients with advanced cancer. Subsequent shortcomings indicated the need for psychometric strengthening. Here, the authors report on the refinement and revalidation of the DS to form the DS-II, specifically reporting the scale's internal validity. METHODS: Patients with cancer or other progressive diseases who were receiving palliative care (n = 211) completed a revised version of the 24-item DS and a measure of symptom burden (the Memorial Symptom Assessment Scale). Exploratory factor analysis and Rasch modeling were used to evaluate, modify, and revalidate the scale, providing information about dimensionality, suitability of response format, item fit, item bias, and item difficulty. Test-retest reliability was examined for 58 symptomatically stable patients at a 5-day follow-up. RESULTS: Exploratory factor analysis supported a 22-item, 2-component model. Separate Rasch modeling of each component resulted in collapsing the response option categories and removing 3 items from each component. Both final 8-item subscales met Rasch model expectations and were appropriate to sum as a 16-item total score. The DS-II demonstrated internal consistency and test-retest reliability (Meaning and Purpose subscale: α = .84; intraclass correlation [ICC] = 0.68; Distress and Coping Ability subscale: α = .82; ICC = 0.82; total DS: α = .89; ICC = 0.80). CONCLUSIONS: The DS-II is a 3-point response, self-report scale comprising 16 items and 2 subscales. Given its revalidation, psychometric strengthening, and simplification, the DS-II is an improved and more practical measure of demoralization for research and clinical use. External validation of the DS-II will be reported subsequently. Cancer 2016;122:2251-9. © 2016 American Cancer Society.

Fyn inhibition rescues established memory and synapse loss in Alzheimer mice.

OBJECTIVE: Currently no effective disease-modifying agents exist for the treatment of Alzheimer disease (AD). The Fyn tyrosine kinase is implicated in AD pathology triggered by amyloid-ß oligomers (Aßo) and propagated by Tau. Thus, Fyn inhibition may prevent or delay disease progression. Here, we sought to repurpose the Src family kinase inhibitor oncology compound, AZD0530, for AD. METHODS: The pharmacokinetics and distribution of AZD0530 were evaluated in mice. Inhibition of Aßo signaling to Fyn, Pyk2, and Glu receptors by AZD0530 was tested by brain slice assays. After AZD0530 or vehicle treatment of wild-type and AD transgenic mice, memory was assessed by Morris water maze and novel object recognition. For these cohorts, amyloid precursor protein (APP) metabolism, synaptic markers (SV2 and PSD-95), and targets of Fyn (Pyk2 and Tau) were studied by immunohistochemistry and by immunoblotting. RESULTS: AZD0530 potently inhibits Fyn and prevents both Aßo-induced Fyn signaling and downstream phosphorylation of the AD risk gene product Pyk2, and of NR2B Glu receptors in brain slices. After 4 weeks of treatment, AZD0530 dosing of APP/PS1 transgenic mice fully rescues spatial memory deficits and synaptic depletion, without altering APP or Aß metabolism. AZD0530 treatment also reduces microglial activation in APP/PS1 mice, and rescues Tau phosphorylation and deposition abnormalities in APP/PS1/Tau transgenic mice. There is no evidence of AZD0530 chronic toxicity. INTERPRETATION: Targeting Fyn can reverse memory deficits found in AD mouse models, and rescue synapse density loss characteristic of the disease. Thus, AZD0530 is a promising candidate to test as a potential therapy for AD.

5' isomiR variation is of functional and evolutionary importance.

We have sequenced miRNA libraries from human embryonic, neural and foetal mesenchymal stem cells. We report that the majority of miRNA genes encode mature isomers that vary in size by one or more bases at the 3' and/or 5' end of the miRNA. Northern blotting for individual miRNAs showed that the proportions of isomiRs expressed by a single miRNA gene often differ between cell and tissue types. IsomiRs were readily co-immunoprecipitated with Argonaute proteins in vivo and were active in luciferase assays, indicating that they are functional. Bioinformatics analysis predicts substantial differences in targeting between miRNAs with minor 5' differences and in support of this we report that a 5' isomiR-9-1 gained the ability to inhibit the expression of DNMT3B and NCAM2 but lost the ability to inhibit CDH1 in vitro. This result was confirmed by the use of isomiR-specific sponges. Our analysis of the miRGator database indicates that a small percentage of human miRNA genes express isomiRs as the dominant transcript in certain cell types and analysis of miRBase shows that 5' isomiRs have replaced canonical miRNAs many times during evolution. This strongly indicates that isomiRs are of functional importance and have contributed to the evolution of miRNA genes.

Proteomics of mouse brain endothelium uncovers dysregulation of vesicular transport pathways during aging.

Age-related decline in brain endothelial cell (BEC) function contributes critically to neurological disease. Comprehensive atlases of the BEC transcriptome have become available, but results from proteomic profiling are lacking. To gain insights into endothelial pathways affected by aging, we developed a magnetic-activated cell sorting-based mouse BEC enrichment protocol compatible with proteomics and resolved the profiles of protein abundance changes during aging. Unsupervised cluster analysis revealed a segregation of age-related protein dynamics with biological functions, including a downregulation of vesicle-mediated transport. We found a dysregulation of key regulators of endocytosis and receptor recycling (most prominently Arf6), macropinocytosis and lysosomal degradation. In gene deletion and overexpression experiments, Arf6 affected endocytosis pathways in endothelial cells. Our approach uncovered changes not picked up by transcriptomic studies, such as accumulation of vesicle cargo and receptor ligands, including Apoe. Proteomic analysis of BECs from Apoe-deficient mice revealed a signature of accelerated aging. Our findings provide a resource for analysing BEC function during aging.

Peripheral expression of brain-penetrant progranulin rescues pathologies in mouse models of frontotemporal lobar degeneration.

Progranulin (PGRN) haploinsufficiency is a major risk factor for frontotemporal lobar degeneration with TAR DNA-binding protein 43 (TDP-43) pathology (FTLD-GRN). Multiple therapeutic strategies are in clinical development to restore PGRN in the CNS, including gene therapy. However, a limitation of current gene therapy approaches aimed to alleviate FTLD-associated pathologies may be their inefficient brain exposure and biodistribution. We therefore developed an adeno-associated virus (AAV) targeting the liver (L) to achieve sustained peripheral expression of a transferrin receptor (TfR) binding, brain-penetrant (b) PGRN variant [AAV(L):bPGRN] in two mouse models of FTLD-GRN, namely, Grn knockout and GrnxTmem106b double knockout mice. This therapeutic strategy avoids potential safety and biodistribution issues of CNS-administered AAVs and maintains sustained concentrations of PGRN in the brain after a single dose. AAV(L):bPGRN treatment reduced several FTLD-GRN-associated pathologies including severe motor function deficits, aberrant TDP-43 phosphorylation, dysfunctional protein degradation, lipid metabolism, gliosis, and neurodegeneration in the brain. The potential translatability of our findings was tested in an in vitro model using cocultured human induced pluripotent stem cell (hiPSC)-derived microglia lacking PGRN and TMEM106B and wild-type hiPSC-derived neurons. As in mice, aberrant TDP-43, lysosomal dysfunction, and neuronal loss were ameliorated after treatment with exogenous TfR-binding protein transport vehicle fused to PGRN (PTV:PGRN). Together, our studies suggest that peripherally administered brain-penetrant PGRN replacement strategies ameliorate FTLD-GRN relevant phenotypes including TDP-43 pathology, neurodegeneration, and behavioral deficits. Our data provide preclinical proof of concept for the use of this AAV platform for treatment of FTLD-GRN and potentially other CNS disorders.