Journal: PLoS ONE

Article Title: Tumor Necrosis Factor Receptor Associated Factor 6 Is Not Required for Atherogenesis in Mice and Does Not Associate with Atherosclerosis in Humans

doi: 10.1371/journal.pone.0011589

Figure Lengend Snippet: Lethally irradiated 6 week old TRAF6 +/+ /LDLR −/− mice received TRAF6-deficient (hatched bars, N = 21) or competent fetal liver cells (white bars, N = 21), TRAF6 +/− /LDLR −/− mice received TRAF6-deficient fetal liver cells (black bars, N = 22) only. Subsequently, all groups consumed high cholesterol diet (HCD) for 18 weeks. Sections of the aortic roots were analyzed for macrophage- (A), lipid- (B), smooth muscle cell- (C), collagen (D), and T cell-content (E). Mac-3-, oil-red-O-, α-actin-, picrosirius red, and CD4-positive staining in per cent of total wall area is displayed as mean±SEM.

Article Snippet: Air dried and formalin-fixed frozen sections were incubated for 3 h in 0.1% solution of picrosirius red (Polysciences) in saturated aqueous picric acid (Sigma-Aldrich).

Techniques: Irradiation, Staining

Journal: Signal Transduction and Targeted Therapy

Article Title: The β-catenin/YAP signaling axis is a key regulator of melanoma-associated fibroblasts

doi: 10.1038/s41392-019-0100-7

Figure Lengend Snippet: SK-MEL-24; GFP/Fb spheroid and SK-MEL-24; bcat-GFP/Fb spheroid cultures were grown for 96 h. Representative bright-field images of SK-MEL-24; GFP/Fb spheroid a – d and SK-MEL-24; bcat-GFP/Fb spheroid cultures e – h were taken using a Carl Zeiss Axiovert 100 TV inverted microscope at ×5 magnification at 24, 48, 72, and 96 h. i Ki67 immunofluorescence staining (red) of SK-MEL-24; GFP/Fb and SK-MEL-24; bcat-GFP/Fb spheroids embedded in paraffin. The nuclei were stained with DAPI (blue). Scale bar: 200 μm. j Quantification of Ki67-positive cells per square millimeter of SK-MEL-24; GFP/Fb spheroid sections and SK-MEL-24; bcat-GFP/Fb spheroid sections. A minimum of ten randomly selected fields was counted for each spheroid type. k Flow cytometry was used to analyze apoptosis and death of SK-MEL-24 melanoma cells in both SK-MEL-24; GFP/Fb spheroids and SK-MEL-24; bcat-GFP/Fb spheroids using annexin V and PI staining. l Percentages of SK-MEL-24 cell subpopulations based on PI and annexin V staining in SK-MEL-24; GFP/Fb spheroids and SK-MEL-24; bcat-GFP/Fb spheroids are shown. m – p α-SMA m–n and FSP-1 o – p immunostaining was performed on 5-μm-thick paraffin sections of SK-MEL-24; GFP/Fb and SK-MEL-24; bcat-GFP/Fb spheroids. Pictures were taken using a Nikon microscope at ×20 magnification. Scale bar: 200 μm. q Collagen content in SK-MEL-24; GFP/Fb spheroids and SK-MEL-24; bcat-GFP/Fb spheroids was visualized by picrosirius red staining. Pictures were taken using an inverted light microscope at ×20 magnification. r Quantification of collagen content was performed by collagen extraction and colorimetric measurement. Collagen content was normalized to total protein content for each sample. s – v Fibronectin s – t and vimentin u – v immunostaining was performed on 5-μm-thick paraffin sections of SK-MEL-24; GFP/Fb and SK-MEL-24; bcat-GFP/Fb spheroids. Pictures were taken using a Nikon microscope at ×20 magnification. Scale bar: 200 μm. w – z Photographs were taken at ×40 magnification. Scale bar: 100 μm.

Article Snippet: Picrosirius red staining was performed according to the manufacturer’s instructions (American MasterTech, Lodi, CA).

Techniques: Inverted Microscopy, Immunofluorescence, Staining, Flow Cytometry, Immunostaining, Microscopy, Light Microscopy, Extraction

Journal: Reproductive Sciences

Article Title: Cervix Stromal Cells and the Progesterone Receptor A Isoform Mediate Effects of Progesterone for Prepartum Remodeling

doi: 10.1177/1933719118820462

Figure Lengend Snippet: Top: Photomicrographs of progesterone receptor (PR) stained cells and counterstained cell nuclei (CN) in cervix sections from nonpregnant (NP), pregnant (days 15 and 18 postbreeding), and day of birth postpartum (PP) mice. Scale bar is 25 µm. Middle: Density of PR cells normalized to CN per area to account for variability in cell nuclei density due to heterogeneity of tissue morphology within and among sections in individuals, as well as within and among groups. Data are the mean ± SE (n = 5-21/group). Bottom: As inversely related to cross-linked collagen in the extracellular matrix,2 optical density of birefringence of picrosirius red-stained cervix sections cells was normalized to CN per area. Data are mean ± SE (n = 4-10/group; *P < .05 vs NP mice or **vs NP, day 15 and day 16 postbreeding groups by 1-way ANOVA). See Methods for details about staining and analyses. SE indicates standard error; ANOVA, analysis of variance.

Article Snippet: Additional sections were stained with a picrosirius red stain kit (PK-4000; Polysciences, Warrington, Pennsylvania) to assess extracellular collagen organization and counterstained with hematoxylin (SH26; Fisher Scientific, Asheville, NC) as described previously.

Techniques: Staining