Using transcriptomics, anatomical studies, imaging and ELISA, Morin-Brureau et al. examine microglia in patients with temporal lobe epilepsies. In highly sclerotic regions such as CA1, the ...anti-inflammatory cytokine IL-10 regulates microglial phenotype. Seizures induce a transient microglial phenotype associated with secretion of inflammatory cytokines including human CXCL8.
Abstract
Microglia, the immune cells of the brain, are highly plastic and possess multiple functional phenotypes. Differences in phenotype in different regions and different states of epileptic human brain have been little studied. Here we use transcriptomics, anatomy, imaging of living cells and ELISA measurements of cytokine release to examine microglia from patients with temporal lobe epilepsies. Two distinct microglial phenotypes were explored. First we asked how microglial phenotype differs between regions of high and low neuronal loss in the same brain. Second, we asked how microglial phenotype is changed by a recent seizure. In sclerotic areas with few neurons, microglia have an amoeboid rather than ramified shape, express activation markers and respond faster to purinergic stimuli. The repairing interleukin, IL-10, regulates the basal phenotype of microglia in the CA1 and CA3 regions with neuronal loss and gliosis. To understand changes in phenotype induced by a seizure, we estimated the delay from the last seizure until tissue collection from changes in reads for immediate early gene transcripts. Pseudotime ordering of these data was validated by comparison with results from kainate-treated mice. It revealed a local and transient phenotype in which microglia secrete the human interleukin CXCL8, IL-1B and other cytokines. This secretory response is mediated in part via the NRLP3 inflammasome.
Similarities were noted between images presented in this article 1 and two additional articles published by the same research group 2, 3: * The schematics of postanal day-3 and adult cochlear sensory ...epithelia presented in Fig 1 1 are identical to the schematics of postanal day-3 and adult cochlear sensory epithelia presented in Fig 1A 2 and the schematics presented in Gene Expression Patterns 3. * The immunohistochemistry images of undifferentiated CGR8 mouse embryonic stem cells in Fig 4D–4F 1 are identical to the immunohistochemistry images of undifferentiated CGR8 mouse embryonic stem cells in Fig 1B 2. “Culture of mouse embryonic stem cells (mESCs) The undifferentiated mESCs (CGR8 line kindly provided by Bernard Binetruy, Aix-Marseille University, France) were expanded in the absence of feeder cells in DMEM culture medium (Gibco by Life Technologies) supplemented with LIF (leukemia inhibitory factor) on gelatin-coated plates. The correct sentence now reads: “We thank Drs T. Van de Water (University of Miami) for his critical reading, B. Binetruy (University Aix Marseille) for the ES CGR8 cells, M. Narita (Cambridge Institute) for the HMGA2 antibody and A. Dos Santos (UMR 7260, University Aix Marseille) for cell culture.” * The authors have indicated that in addition to data within the paper and its Supporting Information files additional data are accessible at the gene expression Omnibus (GEO) repository https://www.ncbi.nlm.nih.gov/geo through the provisional accession series number GSE32963.
Similarities were noted between images presented in this article 1 and two additional articles published by the same research group 2, 3: * The schematics of postanal day-3 and adult cochlear sensory ...epithelia presented in Fig 1A 1 are identical to the schematics of postanal day-3 and adult cochlear sensory epithelia presented in Fig 1 2 and the schematics presented in Gene Expression Patterns 3. * The immunohistochemistry images of undifferentiated CGR8 mouse embryonic stem cells in Fig 1B 1 are identical to the immunohistochemistry images of undifferentiated CGR8 mouse embryonic stem cells in Fig 4D–4F 2. The following information has been added to the Materials and Methods section: “Culture of mouse embryonic stem cells (mESCs) The undifferentiated mESCs (CGR8 line kindly provided by Bernard Binetruy, Aix-Marseille University, France) were expanded in the absence of feeder cells in DMEM culture medium (Gibco by Life Technologies) supplemented with LIF (leukemia inhibitory factor) on gelatin-coated plates. The undifferentiated and untreated cells used for immunohistochemistry were harvested from passage 2 cell cultures and fixed in paraformaldehyde 4% in Phosphate Buffer Solution for 20 min at room temperature.
Hmga2 protein belongs to the non-histone chromosomal high-mobility group (HMG) protein family. HMG proteins have been shown to function as architectural transcription regulators, facilitating ...enhanceosome formation on a variety of mammalian promoters. Hmga2 are expressed at high levels in embryonic and transformed cells. Terminally differentiated cells, however, have been reported to express only minimal, if any, Hmga2. Our previous affymetrix array data showed that Hmga2 is expressed in the developing and adult mammalian cochleas. However, the spatio-temporal expression pattern of Hmga2 in the murine cochlea remained unknown. In this study, we report the expression of Hmga2 in developing and adult cochleas using immunohistochemistry and quantitative real time PCR analysis. Immunolabeling of Hmga2 in the embryonic, postnatal, and mature cochleas showed broad Hmga2 expression in embryonic cochlea (E14.5) at the level of the developing organ of Corti in differentiating hair cells, supporting cells, in addition to immature cells in the GER and LER areas. By postnatal stage (P0-P3), Hmga2 is predominantly expressed in the hair and supporting cells, in addition to cells in the LER area. By P12, Hmga2 immunolabeling is confined to the hair cells and supporting cells. In the adult ear, Hmga2 expression is maintained in the hair and supporting cell subtypes (i.e. Deiters' cells, Hensen cells, pillar cells, inner phalangeal and border cells) in the cochlear epithelium. Using quantitative real time PCR, we found a decrease in transcript level for Hmga2 comparable to other known inner ear developmental genes (Sox2, Atoh1, Jagged1 and Hes5) in the cochlear epithelium of the adult relative to postnatal ears. These data provide for the first time the tissue-specific expression and transcription level of Hmga2 during inner ear development and suggest its potential dual role in early differentiation and maintenance of both hair and supporting cell phenotypes.
The adult mammalian cochlea lacks regenerative ability and the irreversible degeneration of cochlear sensory hair cells leads to permanent hearing loss. Previous data show that early postnatal ...cochlea harbors stem/progenitor-like cells and shows a limited regenerative/repair capacity. These properties are progressively lost later during the postnatal development. Little is known about the genes and pathways that are potentially involved in this difference of the regenerative/repair potentialities between early postnatal and adult mammalian cochlear sensory epithelia (CSE). The goal of our study is to investigate the transcriptomic profiles of these two stages. We used Mouse Genome 430 2.0 microarray to perform an extensive analysis of the genes expressed in mouse postnatal day-3 (P3) and adult CSE. Statistical analysis of microarray data was performed using SAM (Significance Analysis of Microarrays) software. We identified 5644 statistically significant differentially expressed transcripts with a fold change (FC) >2 and a False Discovery Rate (FDR) ≤0.05. The P3 CSE signature included 3,102 transcripts, among which were known genes in the cochlea, but also new transcripts such as, Hmga2 (high mobility group AT-hook 2) and Nrarp (Notch-regulated ankyrin repeat protein). The adult CSE overexpressed 2,542 transcripts including new transcripts, such as Prl (Prolactin) and Ar (Androgen receptor), that previously were not known to be expressed in the adult cochlea. Our comparative study revealed important genes and pathways differentially expressed between the developing and adult CSE. The identification of new candidate genes would be useful as potential markers of the maintenance or the loss of stem cells and regenerative/repair ability during mammalian cochlear development.
The presubiculum, located between hippocampus and entorhinal cortex, plays a fundamental role in representing spatial information, notably head direction. Little is known about GABAergic interneurons ...of this region. Here, we used three transgenic mouse lines, Pvalb-Cre, Sst-Cre, and X98, to examine distinct interneurons labeled with tdTomato or green fluorescent protein. The distribution of interneurons in presubicular lamina for each animal line was compared to that in the GAD67-GFP knock-in animal line. Labeling was specific in the Pvalb-Cre line with 87% of labeled interneurons immunopositive for parvalbumin (PV). Immunostaining for somatostatin (SOM) revealed good specificity in the X98 line with 89% of fluorescent cells, but a lesser specificity in Sst-Cre animals where only 71% of labeled cells were immunopositive. A minority of ∼6% of interneurons co-expressed PV and SOM in the presubiculum of Sst-Cre animals. The electrophysiological and morphological properties of fluorescent interneurons from Pvalb-Cre, Sst-Cre, and X98 mice differed. Distinct physiological groups of presubicular interneurons were resolved by unsupervised cluster analysis of parameters describing passive properties, firing patterns and AP shapes. One group consisted of SOM-positive, Martinotti type neurons with a low firing threshold (cluster 1). Fast spiking basket cells, mainly from the Pvalb-Cre line, formed a distinct group (cluster 3). Another group (cluster 2) contained interneurons of intermediate electrical properties and basket-cell like morphologies. These labeled neurons were recorded from both Sst-Cre and Pvalb-Cre animals. Thus, our results reveal a wide variation in anatomical and physiological properties for these interneurons, a real overlap of interneurons immuno-positive for both PV and SOM as well as an off-target recombination in the Sst-Cre line, possibly linked to maternal cre inheritance.
Over the years, microwave radiation has emerged as an efficient source of energy for material processing. This technology provides a rapid and a volumetric heating of material. However, the main ...issues that prevent microwave technology from being widespread in material processing are temperature control regulation and heating distribution within the sample. Most of the experimental works are usually manually monitored, and their reproducibility is rarely evaluated and discussed. In this work, an originally designed 915 MHz microwave single-mode applicator for high-temperature processing is presented. The overall microwave system is described in terms of an equivalent electrical circuit. This circuit has allowed to point out the different parameters which need to be adjusted to get a fully controlled heating process. The basic principle of regulation is then depicted in terms of a block function diagram. From it, the process has been developed and tested to sinter zirconia- and spinel-based ceramics. It is clearly shown that the process can be successfully used to program multistep temperature cycles up to ∼1550°C, improving significantly the reproducibility and the ease of use of this emerging high-temperature process technology.
Loss of hair cells in the mammalian cochlea leads to permanent sensori-neural hearing loss. Hair cells degenerate and their places are taken by phalangeal scars formed by non-sensory supporting ...cells. Current data indicate that early postnatal post-mitotic supporting cells can proliferate and differentiate into hair cell-like cells in culture.
In this study, we used GFAP and nestin promoter-GFP transgenic mice in combination with other stem cell markers to characterize supporting cell subtypes in the postnatal day-3 (P3) and adult organs of Corti with potential stem/progenitor cell phenotype.
In P3 organ of Corti, we show GFAP-GFP signal in all the supporting cell subtypes while the nestin-GFP was restricted to the supporting cells in the inner hair cell area. At this stage, GFAP and selected stem/progenitor markers displayed overlapping expression pattern in the supporting cell population.
In the adult, GFAP expression is down-regulated from the supporting cells in the outer hair cell area and nestin expression is down-regulated in the supporting cells of the inner hair cell area. Sox2 and Jagged1 expression is maintained in the mature supporting cells, while Abcg2 was down-regulated in these cells. In contrast, GFAP and Abcg2 expression was up-regulated in the inner sulcus limbal cells outside the mature organ of Corti’s area. Using quantitative reverse transcription-PCR, we found a decrease in transcripts for Jagged1 and Sox2 in adult cochleae. Our findings suggest that the loss of regenerative capacity of the adult organ of Corti is related to down-regulation of stem/progenitor key-markers from the mature supporting cells.
Zn
1−
x
Ni
x
O dense ceramics were prepared from Zn
1−
x
Ni
x
O nanoparticles with
x varying from 0 to 0.06. These nanoparticles were synthesized by liquid route. In the sintered samples, the ...solubility limit of Ni in the Zn
1−
x
Ni
x
O wurtzite structure was found to be 0.03. The increase of
x until 0.03 led to a significant raise in both electrical conductivity (
σ) and absolute value of Seebeck coefficient (|
S|). Ni-richer samples (
x
>
0.03) contained in addition a small amount of Ni rich secondary phase (Zn
y
Ni
z
O) with a cubic structure similar to NiO. The thermoelectric properties of all samples were investigated from room temperature to 1000
K. All doped samples showed a n-type semiconducting conductivity. For Ni contents higher than
x
=
0.03, the increase of the secondary phase content induced a decrease in
σ and |
S|. The highest power factor (0.6
mW
m
−1
K
−2) and
ZT (0.09) were found for Zn
0.97Ni
0.03O at 1000
K.
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