roper coolsnap ccd camera  (Roper Scientific Inc)

Journal: Developmental Cell

Article Title: Force-generating apoptotic cells orchestrate avian neural tube bending

doi: 10.1016/j.devcel.2022.02.020

Figure Lengend Snippet: Apoptosis is highly present at the DLHP and is required for the bending of DLHP (see also ) (A) Cas3 staining in the trunk region of whole-mount embryo (HH9) indicating cells in the process of dying (arrows) and dead cells (bracket). Dotted lines outline the neural folds. (B) Schematics of neuroepithelium before (left) and after (right) bending at DLHP. Apoptotic cells (green) along DV axis (black arrow) were quantified between the ventral midline and dorsal fusion points of the neuroepithelium (dark gray). VLR = ventrolateral region. (C) 100-μm depth projections of transversal sections of Cas3 immunostained embryos at 3–8 ss. (D) Distribution of average number of dying cells per transversal section along DV axis. (E) Quantification of dying cells (black) and the angle measured at the DLHP (purple) over time. (F) Schematics of experimental design for QVD-OPh drug treatment. (G) Schematics showing the 5th somite pair at different developmental stages. Gray = neural epithelium, black rectangles = somites, and red dotted line = level of section. (H and I) Transversal sections at the level of the 5th somite pair of control (H) or QVD-OPh-treated (I) embryos at the 5–8 somite stage and Cas3 staining. Arrowheads and blue/orange lines indicate the site of angle measurement. (J and K) Outline of apical curvature (blue/orange lines) of the neuroepithelium of control (J) or QVD-OPh-treated (K) embryos shown in (H) and (I). Red arrows indicate angle at DLHP. (L) Boxplot quantification of angle measured at the (predicted) DLHP at the level of the 5th somite pair in embryos at 5–8 somite stage.

Article Snippet: Antero-posterior distribution of apoptosis in the trunk of the embryo was acquired on a Leica DM6000 B microscope (light source Lumencor SOLA light, Roper Scientific CoolSNAP HQ2 CCD camera, MetaMorph 7.8.10 software) using a 2.5x objective or a Zeiss Axio-Imager type 2 (Colibri 8 multi-diode light source) with a 10x objective.

Techniques: Staining, Control

Journal: Developmental Cell

Article Title: Force-generating apoptotic cells orchestrate avian neural tube bending

doi: 10.1016/j.devcel.2022.02.020

Figure Lengend Snippet: Apoptotic cells mechanically impact the neighboring tissue (see also ) (A) Measurements of the apical deformation of cells undergoing apoptosis over time. Horizontal black dotted line: average level of apical side 25 min after pulling peak (−2.13 μm below initial level). Red dotted line: peak of apical pulling = t0. Blue dotted line: fragmentation time (average of 4–6 ss embryos). Schematics of apoptotic cell before pulling, at pulling peak, and at fragmentation. The apical surface recovers its initial position after 25′ at early stages (1–3 ss), whereas it does not at later stages (4–6 ss). (B) Dot plot quantification of the angle of apical deformation of apoptotic cells before pulling, at the peak of apical pulling and at fragmentation. (C) Dot plot quantification of the speed of apical pulling or release. (D) Dot plot quantification of the time between the start and the peak of pulling (start peak) and the time between the peak of pulling and the fragmentation (peak fragmentation). (E and F) Cell orientation changes at the vicinity of control cells (E) or constricting apoptotic cells (F) in quail embryos transgenic for cytoskeletal markers. Raw images and cross sections are shown in Figure S4 . Outline of the first row of neighbors in representative examples (upper panel); scheme showing the control or apoptotic reference cells in green and the orientation of neighboring cells by the black lines at maximal apical constriction (T0), 15′ before (−T15′) and 15′ after (+T15′) (middle panel). Quantification of the orientation of the 1st row of neighbors (lower panel). 0° indicates a perfect radial orientation. (G) Distribution of apical myosin II along the DV axis of the neural tube. (H) Model of the apoptotic force impact in vertebrates neuroepithelium. Actomyosin accumulates underneath a basally located nucleus. An apico-basal cable is formed and is linked to the nucleus. The apoptotic force drives apical and the basal deformations of the epithelium and basal deformation of the nucleus. The nucleus blebs during its upward movement and fragments when it reaches the apical side, which coincides with basal detachment and apical release. Apoptotic cell neighbors keep a topological memory of the apoptotic force exerted, which, cumulatively, impacts tissue dynamics and participates in the active bending of the dorsal part of the neural tube.

Article Snippet: Antero-posterior distribution of apoptosis in the trunk of the embryo was acquired on a Leica DM6000 B microscope (light source Lumencor SOLA light, Roper Scientific CoolSNAP HQ2 CCD camera, MetaMorph 7.8.10 software) using a 2.5x objective or a Zeiss Axio-Imager type 2 (Colibri 8 multi-diode light source) with a 10x objective.

Techniques: Control, Transgenic Assay