IL-1β and IL-1ra mRNA in first phase and remission of cr-EAE.

<p>Film autoradiographs of affected brain regions in cr-EAE rats after <i>in situ</i> hybridization for IL-1β (left pannels) or IL-1ra (middle pannels) or with sense probes (right panels). Upper and middle pannels are taken from animals in first disease phase, lower panels from animals in remission. <b>(A, B)</b> clusters of IL-1β and IL-1ra mRNA in triangular septal nucleus (TS) and ventral hippocampal commissure (vhc); <b>(D, E)</b> clusters of IL-1β and IL-1ra mRNA in stria medullaris of the thalamus (sm) and habenular nuclei (Hb), and <b>(G, H)</b> IL-1β and IL-1ra mRNA in optic tract (OT). <b>(C, F)</b> sections hybridized with a sense probe for IL-1β, (<b>I</b>) section hybridized with a sense probe for IL-1ra. Scale bar (<b>A–I)</b> = 2 mm.</p

IL-1β expression, CD68 immunoreactivity and Oil-Red O staining.

<p>Expression of mRNA for IL-1β, presence of activated macrophages/microglial cells, and lipid fragmentation. All pictures were taken from animals at the first phase of cr-EAE. <b>(A)</b> IL-1β mRNA in cerebellar lobule. <b>(B)</b> Oil-Red O in cerebellar lobule. Arrows indicate lipid laden cells. <b>(C)</b> CD68 in cerebellar lobule. <b>(D)</b> IL-1β mRNA in the habenula. <b>(E)</b> Absence of IL-1β mRNA in the habenula of a control animal <b>(F)</b> Oil-Red O in the habenula. Sections are counterstained with Cresyl Violet <b>(A, D)</b> or with Mayer's hematoxylin <b>(B, E, F)</b> D3V: dorsal third ventricle, Hb habenula nuclei, LPMR: lateral posterior thalamic nucleus, sm: stria medullaris. Scale bars = 20 µm.</p

CD68 positive cells in the brain stem and the spinal cord during cr-EAE.

<p>Left panels: first phase of disease, middle panels: remission, and right panels: relapse. <b>(A–C)</b> CD68 positive cells in the brain stem, and <b>(D–F)</b> in the spinal cord. Note the difference in the morphology of CD68 positive cells in the brain stem and spinal cord during the relapse phase. Scale bar = 20 µm. Arrows in <b>C</b> indicate ramified CD68 positive cells; arrowheads in <b>F</b> indicated cells with an amoeboid morphology.</p

IL-1β and IL-1ra mRNA in CD68 positive cells.

<p>Expression of IL-1β and IL-1ra mRNA in CD68 positive activated macrophages/microglial cells during the first disease phase of EAE. <b>(A, B)</b> IL-1β mRNA (black grains) combined with CD68 immunoreactivity (brown) in the vestibular nucleus of the brain stem. <b>(C, D</b>) IL-1ra mRNA (black grains) combined with CD68 immunoreactivity in the spinal cord. Scale bars = 20 µm.</p

BRCA1-Dependent Translational Regulation in Breast Cancer Cells

<div><p>BRCA1 (Breast Cancer 1) has been implicated in a number of cellular processes, including transcription regulation, DNA damage repair and protein ubiquitination. We previously demonstrated that BRCA1 interacts with PABP1 (Poly(A)-Binding Protein 1) and that BRCA1 modulates protein synthesis through this interaction. To identify the mRNAs that are translationally regulated by BRCA1, we used a microarray analysis of polysome-bound mRNAs in BRCA1-depleted and non-depleted MCF7 cells. Our findings show that BRCA1 modifies the translational efficiency of approximately 7% of the mRNAs expressed in these cells. Further analysis revealed that several processes contributing to cell surveillance such as cell cycle arrest, cell death, cellular growth and proliferation, DNA repair and gene expression, are largely enriched for the mRNAs whose translation is impacted by BRCA1. The BRCA1-dependent translation of these species of mRNAs therefore uncovers a novel mechanism through which BRCA1 exerts its onco-suppressive role. In addition, the BRCA1-dependent translation of mRNAs participating in unexpected functions such as cellular movement, nucleic acid metabolism or protein trafficking is indicative of novel functions for BRCA1. Finally, this study contributes to the identification of several markers associated with BRCA1 deficiency and to the discovery of new potential anti-neoplastic therapeutic targets.</p></div

Distribution of IL-1β and IL-1ra mRNA in affected brain regions of cr-EAE rats.

<p>Grey areas represent localization of IL-1β and IL-1ra mRNA in the <b>(1)</b> triangular septal nucleus, <b>(2)</b> septofimbrial nucleus, <b>(3)</b> ventral hippocampal commissure, <b>(4)</b> subfornical organ, <b>(5)</b> paraventricular thalamic nucleus, <b>(6)</b> stria medullaris of the thalamus, <b>(7)</b> 3^rd ventricle, <b>(8)</b> medial preoptic area, <b>(9)</b> nucleus of diagonal band, <b>(10)</b> suprachiasmatic nucleus, <b>(11)</b> supra optic hypothalamic nucleus, <b>(12)</b> optic chiasm, <b>(13)</b> medial habenular nucleus, <b>(14)</b> lateral habenular nucleus, <b>(15)</b> intermediodorsal thalamic nucleus, <b>(16)</b> lateral thalamic nuclei, <b>(17)</b> optic tract, <b>(18)</b> habenular commissure, <b>(19)</b> cerebellar lobules, <b>(20)</b> cerebellar peduncles, <b>(21)</b> parabrachial nucleus, <b>(22)</b> central grey pons, <b>(23)</b> trigeminal nucleus, <b>(24)</b> cochlear nucleus, <b>(25)</b> predorsal bundle, <b>(26)</b> pontine reticular nucleus, <b>(27)</b> vestibular nucleus, and <b>(28)</b> spinal trigeminal tract.</p

Histopathological features of CNS lesions during the first disease phase of cr-EAE.

<p>Left panels: PLP immunoreactivity. Middle panels: CD68 immunoreactivity. Right panels: CD3 immunoreactivity during the first disease phase of cr-EAE in (<b>A–C</b>) the trigeminal tract (WM), and (<b>D–F</b>) the trigeminal nucleus (GM). Scale bar = 60 µm.</p

SNAIL proteins confer a survival advantage to MCF10A cells under low-adherence conditions.

<p>(A) MCF10A cells infected with SNAIL-protein-encoding constructs, as indicated at the top, were cultured in ultra-low attachment dishes for different periods of time as indicated on the right. Cells were then stained with annexin V-FITC and propidium iodide and analyzed by flow cytometry. The results shown are representative of three independent experiments. (B) Histogram showing percentages of apoptotic cells (including annexin V⁺/PI⁻ and annexin V⁺/PI⁺ cells) means with SD of triplicate experiments. (<b>C</b>) Analysis of cleaved caspase-3 fragment by western blotting.</p

SNAIL proteins promote MCF10A commitment to EMT.

<p>MCF10A cells were infected with constructs encoding SNAIL proteins as indicated at the top and characterized. (A) Representative photomicrographs of cells obtained by phase contrast microscopy. (B) Analysis of epithelial (E-caherin, β-catenin) and mesenchymal (fibronectin, vimentin) markers by immunofluorescence. (C) Analysis by flow cytometry of CD44 and CD24. (D) Invasion assay. Percentages of invasive cells are indicated. (E) Upper panels: analysis of ectopic <i>SNAI</i> expression by qRT-PCR. Transcript levels are expressed with respect to transcripts of the <i>HPRT1</i> housekeeping gene. Lower panels: western blot analysis of SNAIL proteins. Proteins of interest are indicated with stars. (F) Analysis of epithelial and mesenchymal markers by western blotting.</p

SNAIL proteins confer a survival advantage to HMEC-hTERT cells under low-adherence conditions.

<p>(A) Upper panels: HMEC-derived cell lines were cultured in ultra-low attachment dishes for different periods of time as indicated on the right. The cells were then stained with annexin V-FITC and propidium iodide and analyzed by flow cytometry. (B) Percentages of apoptotic cells (including Annexin V⁺/PI⁻ and Annexin V⁺/PI⁺ cells) are indicated as means ±SD of triplicate experiments. (C) Analysis of the cleaved caspase-3 fragment by western blotting. (D) Examination by western blotting of the status of the ERK and AKT pathways. P-ERK and P-AKT stand for phospho-T202, Y204 ERK1/2 and phospho-S473 AKT respectively. (E) Expression analysis of <i>ZEB1</i> and <i>ZEB2</i> in HMEC-hTERT cells ectopically expressing either <i>SNAI1</i>, <i>SNAI2</i> or <i>SNAI3</i>. Levels expressed relatively to the housekeeping <i>HPRT1</i> gene transcripts were normalized with respect to HMEC-hTERT cells ±SD of triplicates.</p