Whole-mount three-dimensional imaging of internally localized immunostained cells within mouse embryos

We describe a three-dimensional (3D) confocal imaging technique to characterize and enumerate rare, newly emerging hematopoietic cells located within the vasculature of whole-mount preparations of mouse embryos. However, the methodology is broadly applicable for examining the development and 3D architecture of other tissues. Previously, direct whole-mount imaging has been limited to external tissue layers owing to poor laser penetration of dense, opaque tissue. Our whole-embryo imaging method enables detailed quantitative and qualitative analysis of cells within the dorsal aorta of embryonic day (E) 10.5-11.5 embryos after the removal of only the head and body walls. In this protocol we describe the whole-mount fixation and multimarker staining procedure, the tissue transparency treatment, microscopy and the analysis of resulting images. A typical two-color staining experiment can be performed and analyzed in ∼6 d

Infrared Spectral Energy Distributions of Seyfert Galaxies: Spitzer Space Telescope Observations of the 12 micron Sample of Active Galaxies

The mid-far-infrared spectral energy distributions (SEDs) of 83 active galaxies, mostly Seyfert galaxies, selected from the extended 12 micron sample are presented. The data were collected using all three instruments, IRAC, IRS, and MIPS, aboard the Spitzer Space Telescope. The IRS data were obtained in spectral mapping mode, and the photometric data from IRAC and IRS were extracted from matched, 20 arcsec diameter circular apertures. The MIPS data were obtained in SED mode, providing very low resolution spectroscopy (R ~ 20) between ~ 55 and 90 microns in a larger, 20 by 30 arcsec synthetic aperture. We further present the data from a spectral decomposition of the SEDs, including equivalent widths and fluxes of key emission lines; silicate 10 and 18 micron emission and absorption strengths; IRAC magnitudes; and mid-far infrared spectral indices. Finally, we examine the SEDs averaged within optical classifications of activity. We find that the infrared SEDs of Seyfert 1s and Seyfert 2s with hidden broad line regions (HBLR, as revealed by spectropolarimetry or other technique) are qualitatively similar, except that Seyfert 1s show silicate emission and HBLR Seyfert 2s show silicate absorption. The infrared SEDs of other classes with the 12 micron sample, including Seyfert 1.8-1.9, non-HBLR Seyfert 2 (not yet shown to hide a type 1 nucleus), LINER and HII galaxies, appear to be dominated by star-formation, as evidenced by blue IRAC colors, strong PAH emission, and strong far-infrared continuum emission, measured relative to mid-infrared continuum emission.Comment: 78 pages, 13 figure

Inflammatory signaling regulates embryonic hematopoietic stem and progenitor cell production

Identifying signaling pathways that regulate hematopoietic stem and progenitor cell (HSPC) formation in the embryo will guide efforts to produce and expand HSPCs ex vivo. Here we show that sterile tonic inflammatory signaling regulates embryonic HSPC formation. Expression profiling of progenitors with lymphoid potential and hematopoietic stem cells (HSCs) from aorta/gonad/mesonephros (AGM) regions of midgestation mouse embryos revealed a robust innate immune/inflammatory signature. Mouse embryos lacking interferon γ (IFN-γ) or IFN-α signaling and zebrafish morphants lacking IFN-γ and IFN-ϕ activity had significantly fewer AGM HSPCs. Conversely, knockdown of IFN regulatory factor 2 (IRF2), a negative regulator of IFN signaling, increased expression of IFN target genes and HSPC production in zebrafish. Chromatin immunoprecipitation (ChIP) combined with sequencing (ChIP-seq) and expression analyses demonstrated that IRF2-occupied genes identified in human fetal liver CD34(+) HSPCs are actively transcribed in human and mouse HSPCs. Furthermore, we demonstrate that the primitive myeloid population contributes to the local inflammatory response to impact the scale of HSPC production in the AGM region. Thus, sterile inflammatory signaling is an evolutionarily conserved pathway regulating the production of HSPCs during embryonic development

Erythroid/myeloid progenitors and hematopoietic stem cells originate from distinct populations of endothelial cells

SummaryHematopoietic stem cells (HSCs) and an earlier wave of definitive erythroid/myeloid progenitors (EMPs) differentiate from hemogenic endothelial cells in the conceptus. EMPs can be generated in vitro from embryonic or induced pluripotent stem cells, but efforts to produce HSCs have largely failed. The formation of both EMPs and HSCs requires the transcription factor Runx1 and its non-DNA binding partner core binding factor β (CBFβ). Here we show that the requirements for CBFβ in EMP and HSC formation in the conceptus are temporally and spatially distinct. Panendothelial expression of CBFβ in Tek-expressing cells was sufficient for EMP formation, but was not adequate for HSC formation. Expression of CBFβ in Ly6a-expressing cells, on the other hand, was sufficient for HSC, but not EMP, formation. The data indicate that EMPs and HSCs differentiate from distinct populations of hemogenic endothelial cells, with Ly6a expression specifically marking the HSC-generating hemogenic endothelium

Information transmission around block trades on the Spanish stock exchange

This study investigates the informational effects of large transactions, or Block Trades (BT), in the Spanish Stock Exchange (SSE). In the open market period, this topic was not facilitated in the SSE as it was in other markets until 1998. The SSE thus provides a special environment for analysing the information transmission of these specific transactions. It is assumed that information can be better reflected by changes in true asset value, proxied by the midpoint of bid-ask best quotes. Therefore, we will look at changing true asset value orders instead of trades. Three different effects are studied around BTs: price, liquidity and information transmission. To capture them, three different endogenous variables are considered: true asset returns, relative spreads and adverse selection spread component. With this approach, no clear effects of BTs are found. The main result of the study is that there seems to be an increase in information asymmetries when one looks at the adverse selection spread component in some of the different subsample classifications (buyer, seller and sweeping BT), but there is no significant permanent effect on returns. This result could be related to insiders trading in the market. In sharp contrast with adverse selection evidence, a temporary decrease in bid/ask spread around BTs is also observed. These changes reflect temporary liquidity effects related to other spread components (order processing costs and inventory costs).

Semaphorin 3d signaling defects are associated with anomalous pulmonary venous connections

10.1038/nm.3185NATURE MEDICINE196760-

Lithium Directs Embryonic Stem Cell Differentiation Into Hemangioblast‐Like Cells

[EN] Definitive hematopoietic stem cells (HSCs) derive from specialized regions of the endothelium known as the hemogenic endothelium (HE) during embryonic developmental processes. This knowledge opens up new possibilities for designing new strategies to obtain HSCs in vitro from pluripotent stem cells (PSCs). Previous advances in this field show that the Wnt/beta-catenin signaling pathway plays a crucial role in PSC-derived HSC formation. In this work, lithium, a GSK3 inhibitor, is identified as an element capable of stabilizing beta-catenin and inducing embryonic stem cells (ESCs) differentiation in hemangioblast-like cells, highly consistent with the role of Wnt agonists on ESC differentiation. ESCs treated with 10 mm lithium express CD31+, SCA-1+, Nkx2-5+, CD34+, and FLK1+ cells characteristic of the hemangioblast cells that precede HE development. However, 10 mm Li treated cells remain arrested in a hemangioblast-like phase, which switched into the expression of HE markers after stimulation with maturation medium. The ability of lithium-treated ESCs to further derive into HE is confirmed after defined maturation, resulting in a rapid increase in cells positive for the HE markers RUNX1 and SOX17. The results represent a novel strategy for generating HSC precursors in vitro as a multipotent source of stem cells for blood disease therapies.P.R. acknowledges support from the Spanish Ministry of Science, Innovation, and Universities (RTI2018-096794), and Fondo Europeo de Desarrollo Regional (FEDER). CIBER-BBN was an initiative funded by the VI National R&D&I Plan 2008-2011, Iniciativa Ingenio 2010, Consolider Program, CIBER Actions and financed by the Instituto de Salud Carlos III with assistance from the European Regional Development Fund. M.S.-S. acknowledges support from the UK Engineering and Physical Sciences Research Council (EPSRC - EP/P001114/1Mnatsakanyan, H.; Salmerón Sánchez, M.; Rico Tortosa, PM. (2021). Lithium Directs Embryonic Stem Cell Differentiation Into Hemangioblast-Like Cells. Advanced Biosystems. 5(8):1-10. https://doi.org/10.1002/adbi.2020005691105