Stiffness of primordial germ cells is required for their extravasation in avian embryos

細胞の血行性転移の新たな仕組みを発見 --世界初、新たながん転移抑止戦略の開発にも期待--. 京都大学プレスリリース. 2022-12-13.Unlike mammals, primordial germ cells (PGCs) in avian early embryos exploit blood circulation to translocate to the somatic gonadal primordium, but how circulating PGCs undergo extravasation remains elusive. We demonstrate with single-cell level live-imaging analyses that the PGCs are arrested at a specific site in the capillary plexus, which is predominantly governed by occlusion at a narrow path in the vasculature. The occlusion is enabled by a heightened stiffness of the PGCs mediated by actin polymerization. Following the occlusion, PGCs reset their stiffness to soften in order to squeeze through the endothelial lining as they transmigrate. Our discovery also provides a model for the understanding of metastasizing cancer extravasation occurring mainly by occlusion

N-acetylglucosamine suppresses osteoclastogenesis in part through the promotion of O-GlcNAcylation

Osteoclasts are the only cells in an organism capable of resorbing bone. These cells differentiate from monocyte/macrophage lineage cells upon stimulation by receptor activator of NF-κB ligand (RANKL). On the other hand, osteoclastogenesis is reportedly suppressed by glucose via the downregulation of NF-κB activity through suppression of reactive oxygen species generation. To examine whether other sugars might also affect osteoclast development, we compared the effects of monomeric sugars (glucose, galactose, N-acetylglucosamine (GlcNAc), and N-acetylgalactosamine (GalNAc)) on the osteoclastogenesis of murine RAW264 cells. Our results demonstrated that, in addition to glucose, both GlcNAc and GalNAc, which each have little effect on the generation of reactive oxygen species, suppress osteoclastogenesis. We hypothesized that GlcNAc might affect osteoclastogenesis through the upregulation of O-GlcNAcylation and showed that GlcNAc increases global O-GlcNAcylation, thereby suppressing the RANKL-dependent phosphorylation of NF-κB p65. Furthermore, an inhibitor of N-acetyl-β-D-glucosaminidase, O-(2-acetamido-2-deoxy-D-glucopyranosylidene) amino N-phenylcarbamate (PUGNAc), which also increases O-GlcNAcylation, suppressed the osteoclastogenesis of RAW264 cells and that of human peripheral blood mononuclear cells. Together, these data suggest that GlcNAc suppresses osteoclast differentiation in part through the promotion of O-GlcNAcylation. Keywords: Osteoclast, N-acetylglucosamine, GlcNAc, O-GlcNAcylation, NF-κ

Spatiotemporal Gene Expression Regions along the Anterior–Posterior Axis in Mouse Embryos before and after Palatal Elevation

The mammalian secondary palate is formed through complex developmental processes: growth, elevation, and fusion. Although it is known that the palatal elevation pattern changes along the anterior–posterior axis, it is unclear what molecules are expressed and whether their locations change before and after elevation. We examined the expression regions of molecules associated with palatal shelf elevation (Pax9, Osr2, and Tgfβ3) and tissue deformation (F-actin, E-cadherin, and Ki67) using immunohistochemistry and RT–PCR in mouse embryos at E13.5 (before elevation) and E14.5 (after elevation). Pax9 was expressed at significantly higher levels in the lingual/nasal region in the anterior and middle parts, as well as in the buccal/oral region in the posterior part at E13.5. At E14.5, Pax9 was expressed at significantly higher levels in both the lingual/nasal and buccal/oral regions in the anterior and middle parts and the buccal/oral regions in the posterior part. Osr2 was expressed at significantly higher levels in the buccal/oral region in all parts at E13.5 and was more strongly expressed at E13.5 than at E14.5 in all regions. No spatiotemporal changes were found in the other molecules. These results suggested that Pax9 and Osr2 are critical molecules leading to differences in the elevation pattern in palatogenesis

Heterogeneous Precipitation and Mechanical Property Change by Heat Treatments for the Laser Weldments of V-4Cr-4Ti Alloy

Bead-on-plate welds were produced on the 4 mm-thick V-4Cr-4Ti alloy (NIFS-HEAT-2), using a 2.0 kW YAG laser. The post-weld heat treatments (PWHT) were carried out in various conditions. Microstructures, Vickers hardness and Charpy impact properties were obtained for the weld metal after the PWHT. After PWHT for one hour, the hardness increased and after the peak declined with temperature. At 873 K, the hardness increased and after the peak declined with the time of PWHT. Microstructural observation showed that high density of fine precipitates formed homogeneously when the hardness increased, but the precipitate distribution changed into heterogeneous forming islands of developed precipitate aggregates, coincident with decrease in hardness and recovery of impact properties. Optical microscope observations suggested that a cellular structure of precipitate aggregate region was formed by PWHT. Microchemical analysis showed that Ti was enriched in the precipitate aggregate region. Therefore the areal oscillation of Ti concentration with cellular structures formed by melting and resolidification during the welding resulted in the heterogeneous precipitation by the following PWHT. The precipitation in the Ti-rich area purified the matrix of the weld metal and induced the recovery of hardening and impact property degradation. Optimum PWHT conditions were discussed according to the present results

Lzts1 controls both neuronal delamination and outer radial glial-like cell generation during mammalian cerebral development

発生期の大脳で分化細胞と未分化細胞の移動開始をもたらす共通の分子を解明 --「大脳のシワ」形成に貢献する外側放射状グリアはどのように誕生するか--. 京都大学プレスリリース. 2019-07-08.In the developing central nervous system, cell departure from the apical surface is the initial and fundamental step to form the 3D, organized architecture. Both delamination of differentiating cells and repositioning of progenitors to generate outer radial glial cells (oRGs) contribute to mammalian neocortical expansion; however, a comprehensive understanding of their mechanisms is lacking. Here, we demonstrate that Lzts1, a molecule associated with microtubule components, promotes both cell departure events. In neuronally committed cells, Lzts1 functions in apical delamination by altering apical junctional organization. In apical RGs (aRGs), Lzts1 expression is variable, depending on Hes1 expression levels. According to its differential levels, Lzts1 induces diverse RG behaviors: planar division, oblique divisions of aRGs that generate oRGs, and their mitotic somal translocation. Loss-of-function of lzts1 impairs all these cell departure processes. Thus, Lzts1 functions as a master modulator of cellular dynamics, contributing to increasing complexity of the cerebral architecture during evolution

Differences in the mechanical properties of the developing cerebral cortical proliferative zone between mice and ferrets at both the tissue and single-cell levels

Cell-producing events in developing tissues are mechanically dynamic throughout the cell cycle. In many epithelial systems, cells are apicobasally tall, with nuclei and somata that adopt different apicobasal positions because nuclei and somata move in a cell cycle–dependent manner. This movement is apical during G2 phase and basal during G1 phase, whereas mitosis occurs at the apical surface. These movements are collectively referred to as interkinetic nuclear migration, and such epithelia are called pseudostratified. The embryonic mammalian cerebral cortical neuroepithelium is a good model for highly pseudostratified epithelia, and we previously found differences between mice and ferrets in both horizontal cellular density (greater in ferrets) and nuclear/somal movements (slower during G2 and faster during G1 in ferrets). These differences suggest that neuroepithelial cells alter their nucleokinetic behavior in response to physical factors that they encounter, which may form the basis for evolutionary transitions towards more abundant brain-cell production from mice to ferrets and primates. To address how mouse and ferret neuroepithelia may differ physically in a quantitative manner, we used atomic force microscopy to determine that the vertical stiffness of their apical surface is greater in ferrets (Young’s modulus = 1700 Pa) than in mice (1400 Pa). We systematically analyzed factors underlying the apical-surface stiffness through experiments to pharmacologically inhibit actomyosin or microtubules and to examine recoiling behaviors of the apical surface upon laser ablation and also through electron microscopy to observe adherens junction. We found that although both actomyosin and microtubules are partly responsible for the apical-surface stiffness, the mouse<ferret relationship in the apical-surface stiffness was maintained even in the presence of inhibitors. We also found that the stiffness of single, dissociated neuroepithelial cells is actually greater in mice (720 Pa) than in ferrets (450 Pa). Adherens junction was ultrastructurally comparable between mice and ferrets. These results show that the horizontally denser packing of neuroepithelial cell processes is a major contributor to the increased tissue-level apical stiffness in ferrets, and suggest that tissue-level mechanical properties may be achieved by balancing cellular densification and the physical properties of single cells