Journal: Oncotarget

Article Title: Progerin impairs chromosome maintenance by depleting CENP-F from metaphase kinetochores in Hutchinson-Gilford progeria fibroblasts

doi: 10.18632/oncotarget.8267

Figure Lengend Snippet: A. HGPS (HGADFN127) fibroblasts were labeled with DAPI (DNA, blue), and anti-progerin antibody (red). Representative images exhibiting chromatin lagging during the different stages of mitosis are shown. Arrowheads indicate the position of chromatin lagging. B. Binucleated cells were labeled with DAPI (blue) and calnexin (green); a representative image is shown. Double merged signals for progerin with DNA or calnexin with DNA are shown. Scale bar, 10 μm. C. The numbers of cells exhibiting lagging chromatin at different stages of mitosis were scored as described in the Materials and Methods in control (GMO3349C, GMO1651C) and HGPS cells (HGADFN127, HGADFN003). The average mitotic index was on average 1.40% in control cultures and 0.94% in HGPS cultures. Quantification of mitotic cells exhibiting lagging chromatin was scored for each stage of mitosis and the percentage relative to the total number of mitotic cells present in control or HGPS cultures was determined as indicated. A total number of 5217 cells were screened in control and 4976 in HGPS cultures out of 6 independent cultures for each control and HGPS cell lines. The values are reported as means ± SD and evaluated using Student's t-test. D. The percentage of binucleated cells and cells with giant nuclei were scored by direct count of a total of 1559 control (GMO3349C, GMO1651C) cells and 2130 HGPS (HGADFN127, HGADFN003) cells derived from 3 independent cultures. Data are presented as means ± SD and evaluated using Student's t-test. E. In parallel to the evaluation described in (C), the numbers of cells found at a specific mitotic stage were scored and determined as described in (C). Mitotic HGPS cells were found to be increased during anaphase.

Article Snippet: DNA was stained with DAPI (Vector Inc., VEC-H-1200).

Techniques: Labeling, Derivative Assay

Journal: PLoS ONE

Article Title: The Effect of New Zealand Kanuka, Manuka and Clover Honeys on Bacterial Growth Dynamics and Cellular Morphology Varies According to the Species

doi: 10.1371/journal.pone.0055898

Figure Lengend Snippet: The effects of 4% (w/v) of a high-MGO honey (M3) and a high-hydrogen peroxide honey (MK1) on bacterial cellular morphology were examined. Overnight cultures of B. subtilis, E. coli, S. aureus and P. aeruginosa were treated with these honeys, cells collected at both lag and log phases of growth as indicated in , fixed with glutaraldehyde, stained with DAPI and imaged using fluorescence microscopy. All images are overlays of the phase-contrast image and the DAPI-stained (red) fluorescence image. The two left-hand panels show the no-honey treated control cells, the two middle panels M3 honey-treated cells, and the two right-hand panels show the MK1 honey-treated cells. In all images, condensed DNA is shown by green arrows; and dispersed DNA in B. subtilis cells is shown by blue arrows. An asterisk indicates lysed cells for B. subitlis (MK1, lag-phase cells). The scale bar represents 2 µm, except for S. aureus images, where it represents 1 µm.

Article Snippet: Harvested cells were treated for microscopy as described previously but with the following modifications: 20 μL of fixed cells were diluted 1∶1 with the DNA staining agent DAPI (4',6-diamidino-2-phenylindole; Life Technologies), to give a final DAPI concentration of 0.4 μg/mL for E. coli , B. subtilis and P. aeruginosa , and 0.8 μg/mL for S. aureus .

Techniques: Staining, Fluorescence, Microscopy, Control

Journal: Nature communications

Article Title: FOXO1 Inhibition Yields Functional Insulin-Producing Cells In Human Gut Organoid Cultures

doi: 10.1038/ncomms5242

Figure Lengend Snippet: Insulin-positive cells in 184-day-old human gut organoids. a–b , qPCR analysis of different markers in gut organoids transduced with control (empty bars) or HA-Δ256 FOXO1 adenovirus (black bars). c , Quantification of insulin- and GLP1-positive cells in gut organoids transduced with control (empty bars) or HA-Δ256 FOXO1 adenovirus (black bars). d–e , Immunohistochemistry with insulin (green), C-peptide (red), and CGA (magenta). f , Magnification of a typical flask-shaped insulin-positive cell from panel e . g–h , Co-immunohistochemistry with insulin (green), HA (to detect HA-Δ256 Foxo1 adenovirus) (red), and CGA (magenta). i–j , Co-immunohistochemistry with insulin (green) and FOXO1 (red), or ( k, l ) insulin (green), α-SMA (red) and CDX2 (magenta). Insets in h, j, and l show magnifications of individual cells. DAPI (blue) was used throughout to visualize DNA. Scale bars: 50 µm in a–e ; 10 µm in f (n=3–6 for qPCR and 3 for histology) (* p < 0.05). We present quantitative data as means ± SEM.

Article Snippet: We counterstained DNA with DAPI (Cell Signaling).

Techniques: Transduction, Control, Immunohistochemistry