Journal: Cancer Science

Article Title: Synthetic disulfide-bridged cyclic peptides mimic the anti-angiogenic actions of chondromodulin-I

doi: 10.1111/j.1349-7006.2012.02276.x

Figure Lengend Snippet: Effects of the synthetic and tailed chondromodulin-I (ChM-I) cyclic peptide on tumor angiogenesis and growth in an animal model in which OUMS-27 cells (5 × 10 6 cells) were subcutaneously inoculated in the backs of 4 week-old nude mice. (A) Time-course of tumor volume changes. When the tumor volume reached about 45 mm 3 , each mouse was injected around the tumor mass each day for the initial 5 days with PBS (50 μL) alone (♢), PBS containing 4.3 nmol (20 μg) ChM-I cyclic peptide with a tail (○), or PBS containing 0.2 nmol (5 μg) recombinant human ChM-I (rhChM-I) (●) (arrows). The tumor volumes were determined by the width 2 × length × 0.52. Values are the means ± SD from at least six animals per group. (B) Gross appearance of tumors excised on day 21. Bar, 10 mm. (C) Immunohistochemical staining of the CD31-positive vasculature. On day 21, tumor tissues were excised, fixed, and cross-sectioned. The sections were then stained with toluidine blue (left panels), and semi-serial sections were stained with an anti-type II collagen antibody ( green signal in the right panels) and an anti-CD31 antibody ( red signal in the right panels), respectively. Bars, 100 μm. (D) The CD31-positive area was measured as described in the methods section. Values are the means ± SD of five tumors per group. * P < 0.05, ** P < 0.01.

Article Snippet: Anti-CD31 antibody and anti-collagen type II antibody were obtained from BD PharMingen (San Diego, CA, USA) and Rockland (Gilvertsville, PA, USA), respectively.

Techniques: Animal Model, Injection, Recombinant, Immunohistochemical staining, Staining

Journal: Oncotarget

Article Title: Hypoxia-mediated alterations and their role in the HER-2/neuregulated CREB status and localization

doi: 10.18632/oncotarget.10474

Figure Lengend Snippet: A. DBA-1 mice were injected with parental or CREB-deficient HER-2/neu + cells as described in Materials and Methods and tumors were removed after 42 days. Representative photos of parental and CREB-deficient HER-2/neu + tumors are shown. The arrows indicate the blood vessels on the tumor surface. The tumor volume is given. The bar represents 1 cm (left). 5 μm slices of paraffin-embedded tumors were stained with the indicated primary antibody followed by an anti-rabbit secondary antibody. The detection was performed with the peroxidase substrate DAB. Slides were counterstained with methylene blue. The arrow heads indicate the blood vessels. The bar represents 100 μm; Magnification: 40x (right). B. The blood vessel density of the tumors was analysed by counting vessel structures in the anti-CD31 mAb-stained samples (see 1A). Bars represent mean values from four samples/group with four counted fields/sample. C. The necrotic area was analysed in the HE-stained samples. Bars represent mean values from four samples/group with four counted fields/sample. D. The hypoxic area was analysed in the anti-HIF-1α-stained samples. Bars represent mean values from four samples/group with four counted fields/sample. E. 1×10 4 HUVEC/well were seeded in a 96 well plate on polymerized growth factor reduced matrigel. 100 μl/well fresh medium or cell conditioned medium was added and the cells were incubated for 16 h by 37°C. The morphology of the HUVEC under these distinct culture conditions was compared (left) and the mesh-like structures were quantified (right) as described by Zhang . The bar represents 80 μm; Magnification: 10x. F. 1×10 5 parental and CREB-deficient HER-2/neu + cells resuspended in matrigel were injected into the flank of female DBA-1 mice (n = 8). 7 days after injection the mice were killed and the removed matrigel plugs were photographed. The bar represents 1 cm (up). 5 μm slices of the matrigel plugs from parental and CREB-deficient HER-2/neu + cells were stained as indicated. The bar represents 100 μm; Magnification: 10x (down). G. Matrigel plugs from parental and CREB-deficient HER-2/neu + cells were homogenized and their hemoglobin content was analysed as described as in Material and Methods. The bars represent the hemoglobin concentration of each plug from mice injected with the indicated cell line normalized to the weight of the plug. Data demonstrate the results of one out of two independent experiments (with five Matrigel plugs in each experiments) regarding the hemoglobin content/plug from parental and CREB-deficient HER-2/neu + cells.

Article Snippet: Tumor samples were cut into 5 μm slices and then incubated with primary mAb directed against CD31 (Abbiotec) or HIF-1α (Novus) for 24 h. A compatible secondary antibody linked with HRP SignalStain ® Boost IHC Detection Reagent (Cell Signaling) was used for the detection with the substrate DAB.

Techniques: Injection, Staining, Incubation, Concentration Assay

Journal: Oncotarget

Article Title: Hypoxia-mediated alterations and their role in the HER-2/neuregulated CREB status and localization

doi: 10.18632/oncotarget.10474

Figure Lengend Snippet: Primary and labeled secondary antibodies used in this study

Article Snippet: Tumor samples were cut into 5 μm slices and then incubated with primary mAb directed against CD31 (Abbiotec) or HIF-1α (Novus) for 24 h. A compatible secondary antibody linked with HRP SignalStain ® Boost IHC Detection Reagent (Cell Signaling) was used for the detection with the substrate DAB.

Techniques: Labeling, Western Blot, Staining, Flow Cytometry, Ubiquitin Proteomics

Journal: Brain, behavior, and immunity

Article Title: Bone marrow mononuclear cells exert long-term neuroprotection in a rat model of ischemic stroke by promoting arteriogenesis and angiogenesis

doi: 10.1016/j.bbi.2013.07.010

Figure Lengend Snippet: Transplanted BMMNCs incorporate into brain vessels and promote angiogenesis. Representative images of leptomeningeal anastomoses on days 14, 28 and 42 (A–I) and intracranial parenchymal microvessels on day 42 (J–L) stained with anti-BrdU antibody (green) and anti-CD31 antibody (red) (scale bar = 50 μm). (M–N) Quantification showed an increase in the number of BrdU-positive endothelial cells (ECs, M) and smooth muscle cells (SMCs, N). *p<0.05 vs. the number of BrdU-positive ECs or SMCs on day 14 or 42; n=6/time point.

Article Snippet: Mouse anti-BrdU monoclonal antibody (BBI, Shanghai, China), rabbit anti-rat polyclonal antibody against smooth muscle actin (α-SMA; BBI), and rabbit anti-rat polyclonal antibody against CD31 (BBI) were used as primary antibodies at a dilution of 1:250.

Techniques: Staining

Journal: Biomedicines

Article Title: Mechanistic Illustration: How Newly-Formed Blood Vessels Stopped by the Mineral Blocks of Bone Substitutes Can Be Avoided by Using Innovative Combined Therapeutics

doi: 10.3390/biomedicines9080952

Figure Lengend Snippet: Vasculoneogenesis in bone substitutes subcutaneously implanted in nude mice. ( A , B ) H&E staining of bone substitutes (green lines) with either empty (NF-) ( A ) or HEP/VEGF- ( B ) SNCs, at 12 dpi. Yellow arrowheads indicate blood vessels. Scale bars: 200 µm. The images at the top right show enlargements of blood vessels. Scale bars: 20 µm. ( C ) Quantitative analysis of the vessels found in the implanted bone substitutes. The average number and diameter of the vessels found, together with the average surface covered are given at 4 and 12 dpi. Values are expressed as mean ± SD of at least five images/section and four sections/sample. **: p ≤ 0.05. ( D , E ) Ultrastructural view of a blood vessel found in the HEP/VEGF-SNCs bone substitute, shown as a transverse section. Mural cells enveloping the blood vessels (e.g., the pericyte indicated with a red asterisk) were found in the nano-active bone substitute. ( E ) This is the enlargement of the area delimited by the yellow frame in ( D ). No blood cells are in the lumen of the vessel, as a contrast agent was perfused (residues indicated with a blue asterisk). E: endothelial cell; eN: nucleus of the endothelial cell; P: pericyte; pN: nucleus of the pericyte. Scale bars: 5 µm in d, 2 µm in e. ( F ) Endothelial cells of human origin in the bone substitute, as revealed with anti-human (top panels) or anti-human/mouse PECAM1 antibody (mid panel). Mouse control bone was negative to the anti-human PECAM1 antibody (lower panel). Scale bars: 200 µm.

Article Snippet: Rabbit anti-human PECAM1/CD31 (Microm, Brignais, France) and non-species-specific anti-PECAM1/CD31 (Abcam, Paris, France), then secondary antibody raised against rabbit antibodies and coupled with Alexa 594 fluorochrome (Molecular Probes, Life Technologies, Thermo Fisher Scientific, Illkirch-Graffenstaden, France), diluted 1:200 in PBS 1% BSA, were used.

Techniques: Staining

Journal: bioRxiv

Article Title: 3-nitrotyrosine shortens axons of a non-dopaminergic neuron by inhibiting mitochondrial motility

doi: 10.1101/2024.05.09.593299

Figure Lengend Snippet: 3-NT reduced the axonal length in CGNs. (A) CGNs at 4 days in vitro (DIV) were exposed to varying concentrations of 3-NT for 1 day, followed by immunocytochemical analysis utilizing Tau-1 (axonal marker; magenta) and MAP2 (dendritic marker; green) antibodies. Nuclear staining using Hoechst dye (blue) is revealed. (B) CGNs at 5 DIV, transduced with a vector expressing EGFP, were subjected to 3-NT treatment for 1 day. (C, D) Quantification of the length of the longest (axonal) process (C) and the rest of the processes (D) in each neuron was performed. Whiskers denote the range extending 1.5 times the interquartile range above and below the box limits. Data points are depicted as dots. 3-NT led to a reduction in the length of the longest process compared to the control condition. The Wilcoxon rank-sum test with the Benjamini-Hochberg correction was employed for comparison against 0 μM (white boxes) of 3-NT (n > 90 neurons from at least 3 experiments per condition). * p < 0.05, ** p < 0.01, *** p < 0.001. Neurons with processes less than 20 μm were excluded from analysis. Scale bars represent 50 μm.

Article Snippet: Primary antibodies used included mouse monoclonal antibodies against Tau-1 (1:1000; AB5622, Merck) or α-tubulin (1:1000; 12G10, AB_1157911, Developmental Studies Hybridoma Bank, University of Iowa, Iowa City, IA), and rabbit polyclonal antibodies against MAP2 (1:1000; AB5622, Merck) or 3-NT (1:1000; 06-284, Merck).

Techniques: In Vitro, Marker, Staining, Transduction, Plasmid Preparation, Expressing, Control, Comparison