An Experimental Performance Assessment of Temporal Convolutional Networks for Microphone Virtualization in a Car Cabin

In this paper, the experimental results on microphone virtualization in realistic automotive scenarios are presented. A Temporal Convolutional Network (TCN) was designed in order to estimate the acoustic signal at the driver’s ear positions based on the knowledge of monitoring microphone signals at different positions—a technique known as virtual microphone. An experimental setup was implemented on a popular B-segment car to acquire the acoustic field within the cabin while running on smooth asphalt at variable speeds. In order to test the potentiality of the TCN, microphone signals were recorded in two different scenarios, either with or without the front passenger. Our experimental results show that, when training is performed in both scenarios, the adopted TCN is able to robustly adapt to different conditions and guarantee a good average performance. Furthermore, an investigation on the parameters of the Neural Network (NN) that guarantee the sufficient accuracy of the estimation of the virtual microphone signals while maintaining a low computational complexity is presented

Myeloid C/EBPβ deficiency reshapes microglial gene expression and is protective in experimental autoimmune encephalomyelitis

Background CCAAT/enhancer binding protein β (C/EBPβ) is a transcription factor that regulates the expression of important pro-inflammatory genes in microglia. Mice deficient for C/EBPβ show protection against excitotoxic and ischemic CNS damage, but the involvement in this neuroprotective effect of the various C/EBPβ-expressing cell types is not solved. Since C/EBPβ-deficient microglia show attenuated neurotoxicity in culture, we hypothesized that specific C/EBPβ deficiency in microglia could be neuroprotective in vivo. In this study, we have tested this hypothesis by generating mice with myeloid C/EBPβ deficiency. Methods Mice with myeloid C/EBPβ deficiency were generated by crossing LysMCre and C/EBPβfl/fl mice. Primary microglial cultures from C/EBPβfl/fl and LysMCre-C/EBPβfl/fl mice were treated with lipopolysaccharide ± interferon γ (IFNγ) for 6 h, and gene expression was analyzed by RNA sequencing. Gene expression and C/EBPβ deletion were analyzed in vivo in microglia isolated from the brains of C/EBPβfl/fl and LysMCre-C/EBPβfl/fl mice treated systemically with lipolysaccharide or vehicle. Mice of LysMCre-C/EBPβfl/fl or control genotypes were subjected to experimental autoimmune encephalitis and analyzed for clinical signs for 52 days. One- or two-way ANOVA or Kruskal-Wallis with their appropriate post hoc tests were used. Results LysMCre-C/EBPβfl/fl mice showed an efficiency of C/EBPβ deletion in microglia of 100 and 90% in vitro and in vivo, respectively. These mice were devoid of female infertility, perinatal mortality and reduced lifespan that are associated to full C/EBPβ deficiency. Transcriptomic analysis of C/EBPβ-deficient primary microglia revealed C/EBPβ-dependent expression of 1068 genes, significantly enriched in inflammatory and innate immune responses GO terms. In vivo, microglial expression of the pro-inflammatory genes Cybb, Ptges, Il23a, Tnf and Csf3 induced by systemic lipopolysaccharide injection was also blunted by C/EBPβ deletion. CNS expression of C/EBPβ was upregulated in experimental autoimmune encephalitis and in multiple sclerosis samples. Finally, LysMCre-C/EBPβfl/fl mice showed robust attenuation of clinical signs in experimental autoimmune encephalitis. Conclusion This study provides new data that support a central role for C/EBPβ in the biology of activated microglia, and it offers proof of concept for the therapeutic potential of microglial C/EBPβ inhibition in multiple sclerosis

Fast and efficient neural conversion of human hematopoietic cells

Neurons obtained directly from human somatic cells hold great promise for disease modeling and drug screening. Available protocols rely on overexpression of transcription factors using integrative vectors and are often slow, complex, and inefficient. We report a fast and efficient approach for generating induced neural cells (iNCs) directly from human hematopoietic cells using Sendai virus. Upon SOX2 and c-MYC expression, CD133-positive cord blood cells rapidly adopt a neuroepithelial morphology and exhibit high expansion capacity. Under defined neurogenic culture conditions, they express mature neuronal markers and fire spontaneous action potentials that can be modulated with neurotransmitters. SOX2 and c-MYC are also sufficient to convert peripheral blood mononuclear cells into iNCs. However, the conversion process is less efficient and resulting iNCs have limited expansion capacity and electrophysiological activity upon differentiation. Our study demonstrates rapid and efficient generation of iNCs from hematopoietic cells while underscoring the impact of target cells on conversion efficiency

Kv7 channels are upregulated during striatal neuron development and promote maturation of human iPSC-derived neurons

Kv7 channels determine the resting membrane potential of neurons and regulate their excitability. Even though dysfunction of Kv7 channels has been linked to several debilitating childhood neuronal disorders, the ontogeny of the constituent genes, which encode Kv7 channels (KNCQ), and expression of their subunits have been largely unexplored. Here, we show that developmentally regulated expression of specific KCNQ mRNA and Kv7 channel subunits in mouse and human striatum is crucial to the functional maturation of mouse striatal neurons and human-induced pluripotent stem cell-derived neurons. This demonstrates their pivotal role in normal development and maturation, the knowledge of which can now be harnessed to synchronise and accelerate neuronal differentiation of stem cell-derived neurons, enhancing their utility for disease modelling and drug discovery

Pro-inflammatory gene expression and neurotoxic effects of activated microglia are attenuated by absence of CCAAT/enhancer binding protein β

Background. Microglia and astrocytes respond to homeostatic disturbances with profound changes of gene expression. This response, known as glial activation or neuroinflammation, can be detrimental to the surrounding tissue. The transcription factor CCAAT/enhancer binding protein ß (C/EBPß) is an important regulator of gene expression in inflammation but little is known about its involvement in glial activation. To explore the functional role of C/EBPß in glial activation we have analyzed pro-inflammatory gene expression and neurotoxicity in murine wild type and C/EBPß-null glial cultures. Methods. Due to fertility and mortality problems associated with the C/EBPß-null genotype we developed a protocol to prepare mixed glial cultures from cerebral cortex of a single mouse embryo with high yield. Wild-type and C/EBPß-null glial cultures were compared in terms of total cell density by Hoechst-33258 staining; microglial content by CD11b immunocytochemistry; astroglial content by GFAP western blot; gene expression by quantitative real-time PCR, western blot, immunocytochemistry and Griess reaction; and microglial neurotoxicity by estimating MAP2 content in neuronal/microglial cocultures. C/EBPß DNA binding activity was evaluated by electrophoretic mobility shift assay and quantitative chromatin immunoprecipitation. Results. C/EBPß mRNA and protein levels, as well as DNA binding, were increased in glial cultures by treatment with lipopolysaccharide (LPS) or LPS + interferon ¿ (IFN¿). Quantitative chromatin immunoprecipitation showed binding of C/EBPß to pro-inflammatory gene promoters in glial activation in a stimulus- and gene-dependent manner. In agreement with these results, LPS and LPS+IFN¿ induced different transcriptional patterns between pro-inflammatory cytokines and NO synthase-2 genes. Furthermore, the expressions of IL-1ß and NO synthase-2, and consequent NO production, were reduced in the absence of C/EBPß. In addition, neurotoxicity elicited by LPS+IFN¿-treated microglia co-cultured with neurons was completely abolished by the absence of C/EBPß in microglia

Human-BasedNewApproachMethodologiesin DevelopmentalToxicityTesting:AStepAheadfromtheState oftheArtwithaFeto–PlacentalOrgan-on-ChipPlatform

Download PDFsettingsOrder Article Reprints Open AccessReview Human-Based New Approach Methodologies in Developmental Toxicity Testing: A Step Ahead from the State of the Art with a Feto–Placental Organ-on-Chip Platform by Michaela Luconi 1,2,†ORCID,Miguel A. Sogorb 3,†ORCID,Udo R. Markert 4ORCID,Emilio Benfenati 5,Tobias May 6,Susanne Wolbank 7ORCID,Alessandra Roncaglioni 5,Astrid Schmidt 4,Marco Straccia 8ORCID andSabrina Tait 9,*ORCID 1 Department of Experimental and Clinical Biomedical Sciences “Mario Serio”, University of Florence, Viale Pieraccini 6, 50139 Florence, Italy 2 I.N.B.B. (Istituto Nazionale Biostrutture e Biosistemi), Viale Medaglie d’Oro 305, 00136 Rome, Italy 3 Instituto de Bioingeniería, Universidad Miguel Hernández de Elche, Avenida de la Universidad s/n, 03202 Elche, Spain 4 Placenta Lab, Department of Obstetrics, University Hospital Jena, Am Klinikum 1, 07747 Jena, Germany 5 Department of Environmental Health Sciences, Istituto di Ricerche Farmacologiche Mario Negri IRCCS, Via Mario Negri 2, 20156 Milan, Italy 6 InSCREENeX GmbH, Inhoffenstr. 7, 38124 Braunschweig, Germany 7 Ludwig Boltzmann Institut for Traumatology, The Research Center in Cooperation with AUVA, Austrian Cluster for Tissue Regeneration, Donaueschingenstrasse 13, 1200 Vienna, Austria 8 FRESCI by Science&Strategy SL, C/Roure Monjo 33, Vacarisses, 08233 Barcelona, Spain 9 Centre for Gender-Specific Medicine, Istituto Superiore di Sanità, 00161 Rome, Italy * Author to whom correspondence should be addressed. † These authors contributed equally to this work. Int. J. Environ. Res. Public Health 2022, 19(23), 15828; https://doi.org/10.3390/ijerph192315828 Submission received: 24 October 2022 / Revised: 20 November 2022 / Accepted: 25 November 2022 / Published: 28 November 2022 (This article belongs to the Section Toxicology and Public Health) Downloadkeyboard_arrow_down Browse Figures Review Reports Versions Notes Abstract Developmental toxicity testing urgently requires the implementation of human-relevant new approach methodologies (NAMs) that better recapitulate the peculiar nature of human physiology during pregnancy, especially the placenta and the maternal/fetal interface, which represent a key stage for human lifelong health. Fit-for-purpose NAMs for the placental–fetal interface are desirable to improve the biological knowledge of environmental exposure at the molecular level and to reduce the high cost, time and ethical impact of animal studies. This article reviews the state of the art on the available in vitro (placental, fetal and amniotic cell-based systems) and in silico NAMs of human relevance for developmental toxicity testing purposes; in addition, we considered available Adverse Outcome Pathways related to developmental toxicity. The OECD TG 414 for the identification and assessment of deleterious effects of prenatal exposure to chemicals on developing organisms will be discussed to delineate the regulatory context and to better debate what is missing and needed in the context of the Developmental Origins of Health and Disease hypothesis to significantly improve this sector. Starting from this analysis, the development of a novel human feto–placental organ-on-chip platform will be introduced as an innovative future alternative tool for developmental toxicity testing, considering possible implementation and validation strategies to overcome the limitation of the current animal studies and NAMs available in regulatory toxicology and in the biomedical field

CD200 is up-regulated in R6/1 transgenic mouse model of Huntington's disease

In Huntington's disease (HD), striatal medium spiny neurons (MSNs) are particularly sensitive to the presence of a CAG repeat in the huntingtin (HTT) gene. However, there are many evidences that cells from the peripheral immune system and central nervous system (CNS) immune cells, namely microglia, play an important role in the etiology and the progression of HD. However, it remains unclear whether MSNs neurodegeneration is mediated by a non-cell autonomous mechanism. The homeostasis in the healthy CNS is maintained by several mechanisms of interaction between all brain cells. Neurons can control microglia activation through several inhibitory mechanisms, such as the CD200-CD200R1 interaction. Due to the complete lack of knowledge about the CD200-CD200R1 system in HD, we determined the temporal patterns of CD200 and CD200R1 expression in the neocortex, hippocampus and striatum in the HD mouse models R6/1 and HdhQ111/7 from pre-symptomatic to manifest stages. In order to explore any alteration in the peripheral immune system, we also studied the levels of expression of CD200 and CD200R1 in whole blood. Although CD200R1 expression was not altered, we observed and increase in CD200 gene expression and protein levels in the brain parenchyma of all the regions we examined, along with HD pathogenesis in R6/1 mice. Interestingly, the expression of CD200 mRNA was also up-regulated in blood following a similar temporal pattern. These results suggest that canonical neuronal-microglial communication through CD200-CD200R1 interaction is not compromised, and CD200 up-regulation in R6/1 brain parenchyma could represent a neurotrophic signal to sustain or extend neuronal function in the latest stages of HD as pro-survival mechanism

Advanced Non-animal Models in Biomedical Research

It is widely recognised that immuno-oncology has revolutionised cancer treatment, nevertheless there are major hurdles in addressing several key medical questions. Among these, there is a lack of preclinical animal models capable of mimicking patient conditions and predicting responses to such new therapies. Therefore, there is an ongoing need to develop more comprehensive, functional non-animal models. The JRC’s EU Reference Laboratory for alternatives to animal testing (EURL ECVAM) conducted a study to review the state-of-the-art of advanced non-animal models in use for immuno-oncology research. In this study around 130,000 peer-reviewed publications on immuno-oncology were initially retrieved and screened for representative papers describing innovative and promising advanced non-animal models.JRC.F.3 - Systems Toxicolog

Developmental alterations in Huntington's disease neural cells and pharmacological rescue in cells and mice

Neural cultures derived from Huntington's disease (HD) patient-derived induced pluripotent stem cells were used for 'omics' analyses to identify mechanisms underlying neurodegeneration. RNA-seq analysis identified genes in glutamate and GABA signaling, axonal guidance and calcium influx whose expression was decreased in HD cultures. One-third of gene changes were in pathways regulating neuronal development and maturation. When mapped to stages of mouse striatal development, the profiles aligned with earlier embryonic stages of neuronal differentiation. We observed a strong correlation between HD-related histone marks, gene expression and unique peak profiles associated with dysregulated genes, suggesting a coordinated epigenetic program. Treatment with isoxazole-9, which targets key dysregulated pathways, led to amelioration of expanded polyglutamine repeat-associated phenotypes in neural cells and of cognitive impairment and synaptic pathology in HD model R6/2 mice. These data suggest that mutant huntingtin impairs neurodevelopmental pathways that could disrupt synaptic homeostasis and increase vulnerability to the pathologic consequence of expanded polyglutamine repeats over time