Microglial P2Y6 calcium signaling promotes phagocytosis and shapes neuroimmune responses in epileptogenesis.

Microglial calcium signaling is rare in a baseline state but strongly engaged during early epilepsy development. The mechanism(s) governing microglial calcium signaling are not known. By developing an in vivo uridine diphosphate (UDP) fluorescent sensor, GRABUDP1.0, we discovered that UDP release is a conserved response to seizures and excitotoxicity across brain regions. UDP can signal through the microglial-enriched P2Y6 receptor to increase calcium activity during epileptogenesis. P2Y6 calcium activity is associated with lysosome biogenesis and enhanced production of NF-κB-related cytokines. In the hippocampus, knockout of the P2Y6 receptor prevents microglia from fully engulfing neurons. Attenuating microglial calcium signaling through calcium extruder ("CalEx") expression recapitulates multiple features of P2Y6 knockout, including reduced lysosome biogenesis and phagocytic interactions. Ultimately, P2Y6 knockout mice retain more CA3 neurons and better cognitive task performance during epileptogenesis. Our results demonstrate that P2Y6 signaling impacts multiple aspects of myeloid cell immune function during epileptogenesis.

Hyperfunction of post-synaptic density protein 95 promotes seizure response in early-stage aß pathology.

Accumulation of amyloid-beta (Aß) can lead to the formation of aggregates that contribute to neurodegeneration in Alzheimer's disease (AD). Despite globally reduced neural activity during AD onset, recent studies have suggested that Aß induces hyperexcitability and seizure-like activity during the early stages of the disease that ultimately exacerbate cognitive decline. However, the underlying mechanism is unknown. Here, we reveal an Aß-induced elevation of postsynaptic density protein 95 (PSD-95) in cultured neurons in vitro and in an in vivo AD model using APP/PS1 mice at 8 weeks of age. Elevation of PSD-95 occurs as a result of reduced ubiquitination caused by Akt-dependent phosphorylation of E3 ubiquitin ligase murine-double-minute 2 (Mdm2). The elevation of PSD-95 is consistent with the facilitation of excitatory synapses and the surface expression of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors induced by Aß. Inhibition of PSD-95 corrects these Aß-induced synaptic defects and reduces seizure activity in APP/PS1 mice. Our results demonstrate a mechanism underlying elevated seizure activity during early-stage Aß pathology and suggest that PSD-95 could be an early biomarker and novel therapeutic target for AD.

Oligodendrocyte calcium signaling promotes actin-dependent myelin sheath extension.

Myelin is essential for rapid nerve signaling and is increasingly found to play important roles in learning and in diverse diseases of the CNS. Morphological parameters of myelin such as sheath length are thought to precisely tune conduction velocity, but the mechanisms controlling sheath morphology are poorly understood. Local calcium signaling has been observed in nascent myelin sheaths and can be modulated by neuronal activity. However, the role of calcium signaling in sheath formation remains incompletely understood. Here, we use genetic tools to attenuate oligodendrocyte calcium signaling during myelination in the developing mouse CNS. Surprisingly, genetic calcium attenuation does not grossly affect the number of myelinated axons or myelin thickness. Instead, calcium attenuation causes myelination defects resulting in shorter, dysmorphic sheaths. Mechanistically, calcium attenuation reduces actin filaments in oligodendrocytes, and an intact actin cytoskeleton is necessary and sufficient to achieve accurate myelin morphology. Together, our work reveals a cellular mechanism required for accurate CNS myelin formation and may provide mechanistic insight into how oligodendrocytes respond to neuronal activity to sculpt and refine myelin sheaths.

Microglial P2Y6 calcium signaling promotes phagocytosis and shapes neuroimmune responses in epileptogenesis.

Microglial calcium signaling is rare in a baseline state but shows strong engagement during early epilepsy development. The mechanism and purpose behind microglial calcium signaling is not known. By developing an in vivo UDP fluorescent sensor, GRABUDP1.0, we discovered that UDP release is a conserved response to seizures and excitotoxicity across brain regions. UDP signals to the microglial P2Y6 receptor for broad increases in calcium signaling during epileptogenesis. UDP-P2Y6 signaling is necessary for lysosome upregulation across limbic brain regions and enhances production of pro-inflammatory cytokines-TNFα and IL-1ß. Failures in lysosome upregulation, observed in P2Y6 KO mice, can also be phenocopied by attenuating microglial calcium signaling in Calcium Extruder ("CalEx") mice. In the hippocampus, only microglia with P2Y6 expression can perform full neuronal engulfment, which substantially reduces CA3 neuron survival and impairs cognition. Our results demonstrate that calcium activity, driven by UDP-P2Y6 signaling, is a signature of phagocytic and pro-inflammatory function in microglia during epileptogenesis.

Oligodendrocyte calcium signaling sculpts myelin sheath morphology.

Myelin is essential for rapid nerve signaling and is increasingly found to play important roles in learning and in diverse diseases of the CNS. Morphological parameters of myelin such as sheath length and thickness are regulated by neuronal activity and can precisely tune conduction velocity, but the mechanisms controlling sheath morphology are poorly understood. Local calcium signaling has been observed in nascent myelin sheaths and can be modulated by neuronal activity. However, the role of calcium signaling in sheath formation and remodeling is unknown. Here, we used genetic tools to attenuate oligodendrocyte calcium signaling during active myelination in the developing mouse CNS. Surprisingly, we found that genetic calcium attenuation did not grossly affect the number of myelinated axons or myelin thickness. Instead, calcium attenuation caused striking myelination defects resulting in shorter, dysmorphic sheaths. Mechanistically, calcium attenuation reduced actin filaments in oligodendrocytes, and an intact actin cytoskeleton was necessary and sufficient to achieve accurate myelin morphology. Together, our work reveals a novel cellular mechanism required for accurate CNS myelin formation and provides mechanistic insight into how oligodendrocytes may respond to neuronal activity to sculpt myelin sheaths throughout the nervous system.

Exploring the causal role of the immune response to varicella-zoster virus on multiple traits: a phenome-wide Mendelian randomization study.

BACKGROUND: The immune response to infections could be largely driven by the individual's genes, especially in the major histocompatibility complex (MHC) region. Varicella-zoster virus (VZV) is a highly communicable pathogen. In addition to infection, the reactivations of VZV can be a potential causal factor for multiple traits. Identification of VZV immune response-related health conditions can therefore help elucidate the aetiology of certain diseases. METHODS: A phenome-wide Mendelian randomization (MR) study of anti-VZV immunoglobulin G (IgG) levels with 1370 traits was conducted to explore the potential causal role of VZV-specific immune response on multiple traits using the UK Biobank cohort. For the robustness of the results, we performed MR analyses using five different methods. To investigate the impact of the MHC region on MR results, the analyses were conducted using instrumental variables (IVs) inside (IVmhc) and outside (IVno.mhc) the MHC region or all together (IVfull). RESULTS: Forty-nine single nucleotide polymorphisms (IVfull) were associated with anti-VZV IgG levels, of which five (IVmhc) were located in the MHC region and 44 (IVno.mhc) were not. Statistical evidence (false discovery rate < 0.05 in at least three of the five MR methods) for a causal effect of anti-VZV IgG levels was found on 22 traits using IVmhc, while no evidence was found when using IVno.mhc or IVfull. The reactivations of VZV increased the risk of Dupuytren disease, mononeuropathies of the upper limb, sarcoidosis, coeliac disease, teeth problems and earlier onset of allergic rhinitis, which evidence was concordant with the literature. Suggestive causal evidence (P < 0.05 in at least three of five MR methods) using IVfull, IVmhc and IVno.mhc was detected in 92, 194 and 56 traits, respectively. MR results from IVfull correlated with those from IVmhc or IVno.mhc. However, the results between IVmhc and IVno.mhc were noticeably different, as evidenced by causal associations in opposite directions between anti-VZV IgG and ten traits. CONCLUSIONS: In this exploratory study, anti-VZV IgG was causally associated with multiple traits. IVs in the MHC region might have a substantial impact on MR, and therefore, could be potentially considered in future MR studies.

Astrocytes amplify neurovascular coupling to sustained activation of neocortex in awake mice.

Functional hyperemia occurs when enhanced neuronal activity signals to increase local cerebral blood flow (CBF) to satisfy regional energy demand. Ca2+ elevation in astrocytes can drive arteriole dilation to increase CBF, yet affirmative evidence for the necessity of astrocytes in functional hyperemia in vivo is lacking. In awake mice, we discovered that functional hyperemia is bimodal with a distinct early and late component whereby arteriole dilation progresses as sensory stimulation is sustained. Clamping astrocyte Ca2+ signaling in vivo by expressing a plasma membrane Ca2+ ATPase (CalEx) reduces sustained but not brief sensory-evoked arteriole dilation. Elevating astrocyte free Ca2+ using chemogenetics selectively augments sustained hyperemia. Antagonizing NMDA-receptors or epoxyeicosatrienoic acid production reduces only the late component of functional hyperemia, leaving brief increases in CBF to sensory stimulation intact. We propose that a fundamental role of astrocyte Ca2+ is to amplify functional hyperemia when neuronal activation is prolonged.

Molecular basis of astrocyte diversity and morphology across the CNS in health and disease.

Astrocytes, a type of glia, are abundant and morphologically complex cells. Here, we report astrocyte molecular profiles, diversity, and morphology across the mouse central nervous system (CNS). We identified shared and region-specific astrocytic genes and functions and explored the cellular origins of their regional diversity. We identified gene networks correlated with astrocyte morphology, several of which unexpectedly contained Alzheimer's disease (AD) risk genes. CRISPR/Cas9-mediated reduction of candidate genes reduced astrocyte morphological complexity and resulted in cognitive deficits. The same genes were down-regulated in human AD, in an AD mouse model that displayed reduced astrocyte morphology, and in other human brain disorders. We thus provide comprehensive molecular data on astrocyte diversity and mechanisms across the CNS and on the molecular basis of astrocyte morphology in health and disease.

Enhancing GAT-3 in thalamic astrocytes promotes resilience to brain injury in rodents.

Inflammatory processes induced by brain injury are important for recovery; however, when uncontrolled, inflammation can be deleterious, likely explaining why most anti-inflammatory treatments have failed to improve neurological outcomes after brain injury in clinical trials. In the thalamus, chronic activation of glial cells, a proxy of inflammation, has been suggested as an indicator of increased seizure risk and cognitive deficits that develop after cortical injury. Furthermore, lesions in the thalamus, more than other brain regions, have been reported in patients with viral infections associated with neurological deficits, such as SARS-CoV-2. However, the extent to which thalamic inflammation is a driver or by-product of neurological deficits remains unknown. Here, we found that thalamic inflammation in mice was sufficient to phenocopy the cellular and circuit hyperexcitability, enhanced seizure risk, and disruptions in cortical rhythms that develop after cortical injury. In our model, down-regulation of the GABA transporter GAT-3 in thalamic astrocytes mediated this neurological dysfunction. In addition, GAT-3 was decreased in regions of thalamic reactive astrocytes in mouse models of cortical injury. Enhancing GAT-3 in thalamic astrocytes prevented seizure risk, restored cortical states, and was protective against severe chemoconvulsant-induced seizures and mortality in a mouse model of traumatic brain injury, emphasizing the potential of therapeutically targeting this pathway. Together, our results identified a potential therapeutic target for reducing negative outcomes after brain injury.

A suggested shared aetiology of dementia - a colocalization study.

Identification of shared causal genes between dementia and its related clinical outcomes can help understand shared aetiology and multimorbidity surrounding dementia. We performed the HyPrColoc colocalization analysis to detect possible shared causal genes between dementia or Alzheimer's disease (AD) and 5 selected traits: stroke, diabetes, atherosclerosis, cholesterol level, and alcohol consumption within 601 dementia or AD associated genetic regions using summary results of the UK Biobank genome-wide association studies. Functional analysis was performed on the candidate causal genes to explore potential biological pathways. Rs150562240 in the LPIN3 gene was identified as a candidate shared causal variant across dementia, AD and atherosclerosis. Evidence for pairwise colocalization between dementia and stroke, dementia (or AD) and atherosclerosis, and dementia (or AD) and diabetes was found in 2, 6 and 2 genetic regions respectively. Colocalization signals between diabetes and the other 3 non-dementia/AD traits were detected in 5 regions. The colocalization evidence shown in our study suggested shared aetiology between dementia and related diseases such as stroke, atherosclerosis, and diabetes.

Hippocampal astrocytes represent navigation space.

Hippocampal place cells, which display location-specific activity, are known to encode spatial information. A recent study in PLOS Biology by Curreli and colleagues shows that hippocampal astrocytes are implicated in encoding complementary spatial information, suggesting the existence of glial place cells.

TCMNER and PubMed: A Novel Chinese Character-Level-Based Model and a Dataset for TCM Named Entity Recognition.

Intelligent traditional Chinese medicine (TCM) has become a popular research field by means of prospering of deep learning technology. Important achievements have been made in such representative tasks as automatic diagnosis of TCM syndromes and diseases and generation of TCM herbal prescriptions. However, one unavoidable issue that still hinders its progress is the lack of labeled samples, i.e., the TCM medical records. As an efficient tool, the named entity recognition (NER) models trained on various TCM resources can effectively alleviate this problem and continuously increase the labeled TCM samples. In this work, on the basis of in-depth analysis, we argue that the performance of the TCM named entity recognition model can be better by using the character-level representation and tagging and propose a novel word-character integrated self-attention module. With the help of TCM doctors and experts, we define 5 classes of TCM named entities and construct a comprehensive NER dataset containing the standard content of the publications and the clinical medical records. The experimental results on this dataset demonstrate the effectiveness of the proposed module.

Specific and behaviorally consequential astrocyte Gq GPCR signaling attenuation in vivo with ißARK.

Astrocytes respond to neurotransmitters and neuromodulators using G-protein-coupled receptors (GPCRs) to mediate physiological responses. Despite their importance, there has been no method to genetically, specifically, and effectively attenuate astrocyte Gq GPCR pathways to explore consequences of this prevalent signaling mechanism in vivo. We report a 122-residue inhibitory peptide from ß-adrenergic receptor kinase 1 (ißARK; and inactive D110A control) to attenuate astrocyte Gq GPCR signaling. ißARK significantly attenuated Gq GPCR Ca2+ signaling in brain slices and, in vivo, altered behavioral responses, spared other GPCR responses, and did not alter astrocyte spontaneous Ca2+ signals, morphology, electrophysiological properties, or gene expression in the striatum. Furthermore, brain-wide attenuation of astrocyte Gq GPCR signaling with ißARK using PHP.eB adeno-associated viruses (AAVs), when combined with c-Fos mapping, suggested nuclei-specific contributions to behavioral adaptation and spatial memory. ißARK extends the toolkit needed to explore functions of astrocyte Gq GPCR signaling within neural circuits in vivo.

Local and CNS-Wide Astrocyte Intracellular Calcium Signaling Attenuation In Vivo with CalExflox Mice.

Astrocytes exist throughout the CNS and affect neural circuits and behavior through intracellular Ca2+ signaling. Studying the function(s) of astrocyte Ca2+ signaling has proven difficult because of the paucity of tools to achieve selective attenuation. Based on recent studies, we generated and used male and female knock-in mice for Cre-dependent expression of mCherry-tagged hPMCA2w/b to attenuate astrocyte Ca2+ signaling in genetically defined cells in vivo (CalExflox mice for Calcium Extrusion). We characterized CalExflox mice following local AAV-Cre microinjections into the striatum and found reduced astrocyte Ca2+ signaling (∼90%) accompanied with repetitive self-grooming behavior. We also crossed CalExflox mice to astrocyte-specific Aldh1l1-Cre/ERT2 mice to achieve inducible global CNS-wide Ca2+ signaling attenuation. Within 6 d of induction in the bigenic mice, we observed significantly altered ambulation in the open field, disrupted motor coordination and gait, and premature lethality. Furthermore, with histologic, imaging, and transcriptomic analyses, we identified cellular and molecular alterations in the cerebellum following mCherry-tagged hPMCA2w/b expression. Our data show that expression of mCherry-tagged hPMCA2w/b with CalExflox mice throughout the CNS resulted in substantial attenuation of astrocyte Ca2+ signaling and significant behavioral alterations in adult mice. We interpreted these findings candidly in relation to the ability of CalEx to attenuate astrocyte Ca2+ signaling, with regards to additional mechanistic interpretations of the data, and their relation to past studies that reduced astrocyte Ca2+ signaling throughout the CNS. The data and resources provide complementary ways to interrogate the function(s) of astrocytes in multiple experimental scenarios.SIGNIFICANCE STATEMENT Astrocytes represent a significant fraction of all brain cells and tile the entire central nervous system. Unlike neurons, astrocytes lack propagated electrical signals. Instead, astrocytes are proposed to use diverse and dynamic intracellular Ca2+ signals to communicate with other cells. An open question concerns if and how astrocyte Ca2+ signaling regulates behavior in adult mice. We approached this problem by generating a new transgenic mouse line to achieve inducible astrocyte Ca2+ signaling attenuation in vivo We report our data with this mouse line and we interpret the findings candidly in relation to past studies and within the framework of different mechanistic interpretations.

Long-term results of uncemented allograft prosthesis composite reconstruction for the tumor in proximal femur: a minimum follow-up of sixty-five months.

BACKGROUND: Uncemented allograft prosthesis composite (APC) has been applied for tumorous bone defect reconstruction in the proximal femur. However, the long-term results are rarely reported. This study aimed to evaluate long-term outcomes of uncemented APC. METHODS: Eighteen patients who received uncemented APC reconstruction in the proximal femur after tumor resections were retrospectively reviewed. RESULTS: The average resection length was 110 mm (80-154) and the average follow-up was 106.7 months (65-141). Bone union achieved in all patients with an average duration of 7.6 months (5-10). The average HHS, MSTS score and gluteus medius strength at one-year follow-up were 88.0 (80-94), 25.2 (22-28) and 4 (3-5), respectively. While at the last follow-up, the HHS, MSTS score and gluteus medius strength were 83.0 (48-100), 24.0 (10-30) and 4 (2-5), respectively. Five intraoperative fractures were fixed with cerclage wires. Two postoperative periprosthetic and prosthetic fractures received a revision. Three local recurrent patients received a secondary surgery. One of these three lung metastatic patients underwent lung metastatic tumor resection. Another two patients were diagnosed with both bone and lung metastases, only one of them underwent amputation. Two greater trochanteric fractures received no treatment. There were10 severe, 3 moderate and 5 mild allograft resorptions without treatment. CONCLUSION: Uncemented APC is a reliable reconstruction for neoplastic bone defect of the proximal femur, especially for the young patient who expected long-life expectancy and good function. Though allograft resorption and trochanteric fracture are the common complications, they seem no effect on the function.

Behaviorally consequential astrocytic regulation of neural circuits.

Astrocytes are a large and diverse population of morphologically complex cells that exist throughout nervous systems of multiple species. Progress over the last two decades has shown that astrocytes mediate developmental, physiological, and pathological processes. However, a long-standing open question is how astrocytes regulate neural circuits in ways that are behaviorally consequential. In this regard, we summarize recent studies using Caenorhabditis elegans, Drosophila melanogaster, Danio rerio, and Mus musculus. The data reveal diverse astrocyte mechanisms operating in seconds or much longer timescales within neural circuits and shaping multiple behavioral outputs. We also refer to human diseases that have a known primary astrocytic basis. We suggest that including astrocytes in mechanistic, theoretical, and computational studies of neural circuits provides new perspectives to understand behavior, its regulation, and its disease-related manifestations.

Design Comorbidity Portfolios to Improve Treatment Cost Prediction of Asthma Using Machine Learning.

Comorbidity is an important factor to consider when trying to predict the cost of treating asthma patients. When an asthmatic patient suffered from comorbidity, the cost of treating such a patient becomes dependent on the nature of the comorbidity. Therefore, lack of recognition of comorbidity on asthmatic patient poses a challenge in predicting the cost of treatment. In this study, we proposed a comorbidity portfolio design that improves the prediction cost of treating asthmatic patients by regrouping frequently occurred comorbidities in different cost groups. In the experiment, predictive models, including logistic regression, random forest, support vector machine, classification regression tree, and backpropagation neural network were trained with real-world data of asthmatic patients from 2012 to 2014 in a large city of China. The 10-fold cross validation and random search algorithm were employed to optimize the hyper-parameters. We recorded significant improvements using our model, which are attributed to comorbidity portfolios in area under curve (AUC) and sensitivity increase of 46.89% (standard deviation: 4.45%) and 101.07% (standard deviation: 44.94%), respectively. In risk analysis of comorbidity on cost, respiratory diseases with a cumulative proportion in the adjusted odds ratio of 36.38% (95%CI: 27.61%, 47.86%) and circulatory diseases with a cumulative proportion in the adjusted odds ratio of 23.83% (95%CI: 15.95%, 35.22%) are the dominant risks of asthmatic patients that affects the treatment cost. It is found that the comorbidity portfolio is robust, and provides a better prediction of the high-cost of treating asthmatic patients. The preliminary characterization of the joint risk of multiple comorbidities posed on cost are also reported. This study will be of great help in improving cost prediction and comorbidity management.

Context-Specific Striatal Astrocyte Molecular Responses Are Phenotypically Exploitable.

Astrocytes tile the central nervous system and are widely implicated in brain diseases, but the molecular mechanisms by which astrocytes contribute to brain disorders remain incompletely explored. By performing astrocyte gene expression analyses following 14 experimental perturbations of relevance to the striatum, we discovered that striatal astrocytes mount context-specific molecular responses at the level of genes, pathways, and upstream regulators. Through data mining, we also identified astrocyte pathways in Huntington's disease (HD) that were reciprocally altered with respect to the activation of striatal astrocyte G protein-coupled receptor (GPCR) signaling. Furthermore, selective striatal astrocyte stimulation of the Gi-GPCR pathway in vivo corrected several HD-associated astrocytic, synaptic, and behavioral phenotypes, with accompanying improvement of HD-associated astrocyte signaling pathways, including those related to synaptogenesis and neuroimmune functions. Overall, our data show that astrocytes are malleable, using context-specific responses that can be dissected molecularly and used for phenotypic benefit in brain disorders.