Journal: The American journal of pathology

Article Title: Increased expression of versican in the inflammatory response to UVB- and reactive oxygen species-induced skin tumorigenesis.

doi: 10.1016/j.ajpath.2011.08.042

Figure Lengend Snippet: Figure 3. Expression of continuously up-regulated genes after UVB expo- sure. A: Four genes were significantly up-regulated at both 3 and 24 hours after UVB exposure. Relative expression levels were determined by real-time quantitative PCR. Each of the two groups of RNAs isolated from wild-type and Ogg1 knockout mice was assayed in duplicate. Reactions were normal- ized to GAPDH expression levels. *P 0.05; **P 0.01; and ***P 0.001. B: Immunohistochemical study of versican expression after UVB irradiation. Versican is barely expressed in the wild-type mouse epidermis and dermis in the absence of UVB irradiation. At 24 hours after UVB exposure, versican was expressed in the wild-type epidermis. In Ogg1 knockout mice at 24 hours, versican was strongly expressed in the epidermis (arrowheads), as well as in dermal fibroblasts (arrows). Positive signals are seen as reddish-brown deposits produced on reaction with the 3-amino-9-ethylcarbazole substrate. Scale bar 30 m. C: Versican expression after UVB exposure, as deter- mined by Western blotting. Versican was more strongly up-regulated in Ogg1 knockout than in wild-type mice at 24 and 48 hours after UVB exposure. This up-regulation was time-dependent. The band at approximately 75 kDa indi- cates that the antibody for versican can detect the V1 isoform. /-Tubulin was used as the loading control. Data are representative of three separate determinations.

Article Snippet: Sections were incubated for 16 hours at 4°C with the following primary antibodies: rabbit polyclonal anti-mouse IL-1 (1:1000 dilution; Abcam, Cambridge, MA), rabbit polyclonal anti-mouse versican (1: 100 dilution; LifeSpan Biosciences, Seattle, WA), rabbit polyclonal anti-human versican (1:125 dilution; Atlas Antibodies, Stockholm, Sweden), or rabbit polyclonal antimouse p53 (CM5) (1:500 dilution; Leica Biosystems Newcastle, Newcastle upon Tyne, UK).

Techniques: Expressing, Real-time Polymerase Chain Reaction, Isolation, Knock-Out, Immunohistochemical staining, Irradiation, Produced, Western Blot, Control

Journal: The American journal of pathology

Article Title: Increased expression of versican in the inflammatory response to UVB- and reactive oxygen species-induced skin tumorigenesis.

doi: 10.1016/j.ajpath.2011.08.042

Figure Lengend Snippet: Figure 4. Versican expression in developing skin tumors of chronically UVB-exposed mice. Representative histological sections of SCC tumors from wild-type and Ogg1 knockout mice red-colored staining positively for versi- can are shown, with a summary of versican-positive wild-type and Ogg1 knockout mouse tumors overall. *P 0.05 for the ratio of malignant tumor to total tumors analyzed for each genotype.

Article Snippet: Sections were incubated for 16 hours at 4°C with the following primary antibodies: rabbit polyclonal anti-mouse IL-1 (1:1000 dilution; Abcam, Cambridge, MA), rabbit polyclonal anti-mouse versican (1: 100 dilution; LifeSpan Biosciences, Seattle, WA), rabbit polyclonal anti-human versican (1:125 dilution; Atlas Antibodies, Stockholm, Sweden), or rabbit polyclonal antimouse p53 (CM5) (1:500 dilution; Leica Biosystems Newcastle, Newcastle upon Tyne, UK).

Techniques: Expressing, Knock-Out, Staining

Journal: The American journal of pathology

Article Title: Increased expression of versican in the inflammatory response to UVB- and reactive oxygen species-induced skin tumorigenesis.

doi: 10.1016/j.ajpath.2011.08.042

Figure Lengend Snippet: Figure 5. Versican expression in human skin tumors. A: Immunohistochemical study of versican expression in malignant (lentigo maligna melanoma, basal cell carcinoma, and squamous cell carcinoma) and benign (seborrheic keratosis and lentigo senilis) tumors. Arrowheads indicate the borders between the seborrheic keratosis tumor and the normal skin. Scale bar 100 m. B: Classification of versican expression in skin tumors, grouped as dermal and stromal versus tumoral.

Article Snippet: Sections were incubated for 16 hours at 4°C with the following primary antibodies: rabbit polyclonal anti-mouse IL-1 (1:1000 dilution; Abcam, Cambridge, MA), rabbit polyclonal anti-mouse versican (1: 100 dilution; LifeSpan Biosciences, Seattle, WA), rabbit polyclonal anti-human versican (1:125 dilution; Atlas Antibodies, Stockholm, Sweden), or rabbit polyclonal antimouse p53 (CM5) (1:500 dilution; Leica Biosystems Newcastle, Newcastle upon Tyne, UK).

Techniques: Expressing, Immunohistochemical staining

Journal: The American journal of pathology

Article Title: Increased expression of versican in the inflammatory response to UVB- and reactive oxygen species-induced skin tumorigenesis.

doi: 10.1016/j.ajpath.2011.08.042

Figure Lengend Snippet: Figure 7. Proposed relationships of versican in the inflammatory response leading to the development of skin tumors in terms of UVB-induced 8-oxoG accumulation. The accumulation of UVB/ROS-induced 8-oxoG in the skin leads to inflammatory reactions. High versican expression is induced by a highly inflammatory microenvironment with high numbers of infiltrated neu- trophils; conversely, neutrophil infiltration induces versican overexpression. More ROS will be generated at the inflammatory sites by neutrophils.

Article Snippet: Sections were incubated for 16 hours at 4°C with the following primary antibodies: rabbit polyclonal anti-mouse IL-1 (1:1000 dilution; Abcam, Cambridge, MA), rabbit polyclonal anti-mouse versican (1: 100 dilution; LifeSpan Biosciences, Seattle, WA), rabbit polyclonal anti-human versican (1:125 dilution; Atlas Antibodies, Stockholm, Sweden), or rabbit polyclonal antimouse p53 (CM5) (1:500 dilution; Leica Biosystems Newcastle, Newcastle upon Tyne, UK).

Techniques: Expressing, Over Expression, Generated

Journal: The American journal of pathology

Article Title: Increased expression of versican in the inflammatory response to UVB- and reactive oxygen species-induced skin tumorigenesis.

doi: 10.1016/j.ajpath.2011.08.042

Figure Lengend Snippet: Figure 6. Versican expression and neutrophil localization in human and murine skin tumors. A: Versican expression and inflammatory cells in human squamous cell carcinoma. Versican is strongly expressed in the dermal components (arrows) and inflammatory cells (arrowheads), seen at low magnification (left) and high magnification (right). Two focused areas are taken from different part of sections. Inflammatory cells, especially seg- mented leukocytes (neutrophils) (arrowheads), were strongly positive for versican. Scale bars 30 m. B: Versican expression and neutrophils in mouse skin. In the merged iamge, neutrophils with versican expression (arrows) appear yellow. Scale bar 30 m. C: Neutrophil infiltration after UVB irradiation in wild-type and Ogg1 knockout mice. Arrows indicate Gr-1-positive cells (green) in the dermis at 24 and 48 hours after UVB exposure in the wild-type and Ogg1 knockout mice. Scale bar 30 m. The accompanying graphs show the average number of neutrophils per 800 m2

Article Snippet: Sections were incubated for 16 hours at 4°C with the following primary antibodies: rabbit polyclonal anti-mouse IL-1 (1:1000 dilution; Abcam, Cambridge, MA), rabbit polyclonal anti-mouse versican (1: 100 dilution; LifeSpan Biosciences, Seattle, WA), rabbit polyclonal anti-human versican (1:125 dilution; Atlas Antibodies, Stockholm, Sweden), or rabbit polyclonal antimouse p53 (CM5) (1:500 dilution; Leica Biosystems Newcastle, Newcastle upon Tyne, UK).

Techniques: Expressing, Irradiation, Knock-Out

Journal: PLoS ONE

Article Title: The Human Neutrophil Subsets Defined by the Presence or Absence of OLFM4 Both Transmigrate into Tissue In Vivo and Give Rise to Distinct NETs In Vitro

doi: 10.1371/journal.pone.0069575

Figure Lengend Snippet: A ) Neutrophils were immunofluorescently stained for OLFM4 (green) together with specific granule marker NGAL or azurophil granule marker CD63 (red), and DNA was labeled with DAPI (blue). Colocalization was analyzed by imaging flow cytometry and images show representative cells from the double positive populations (BF = brightfield). The diagram shows the mean colocalization score for the two fluorophores +SD from at least three experiments. The technical positive control was FITC-conjugated mouse anti-human CD16 antibody followed by Alexa Fluor 647-coupled goat anti-mouse secondary antibody and the technical negative control was FITC-conjugated mouse anti-human CD16 antibody together with DAPI, showing the minimum and maximum values that can be obtained by this analysis. The biological positive control was specific granule marker lactoferrin (LF) together with NGAL, and the biological negative control was lactoferrin together with CD63, showing the resolution of the analysis. B ) Pooled neutrophils from three donors were subjected to subcellular fractionation, and the fractions containing peak content of azurophil granule marker (MPO; fraction 1), specific granule marker (lactoferrin; fractions 10-12), gelatinase granule marker (gelatinase; fractions 13-15) and secretory vesicle marker (latent alkaline phosphatase; fraction 22) were subjected to Western blot with anti-OLFM4 antibody using rOLFM4 as a positive control. One representative blot out of three is shown. Lactoferrin (specific granule marker) and gelatinase (gelatinase granule marker) blots are also shown for fractions 10-15.

Article Snippet: The contents of lactoferrin and gelatinase were analyzed by Western blotting, using rabbit anti-human lactoferrin (DAKO) and rabbit anti-human gelatinase (Merck, Darmstadt, Germany), both at 2 μg/ml.

Techniques: Staining, Marker, Labeling, Imaging, Flow Cytometry, Positive Control, Negative Control, Fractionation, Western Blot