Journal: Frontiers in Immunology

Article Title: Halted Lymphocyte Egress via Efferent Lymph Contributes to Lymph Node Hypertrophy During Hypercholesterolemia

doi: 10.3389/fimmu.2019.00575

Figure Lengend Snippet: Dyslipidemia induces LN lymphangiogenesis (B) LN sections from WT and apoE −/− mice at 22 to 28 weeks of age were stained for B220 and LYVE-1. Images are representative of four independent experiments ( n = 3–4 mice per group). (A) LN cells were stained for CD45, podoplanin and CD31 to identify LECs by FACS analysis. The number of LEC was enumerated in LNs from WT and apoE −/− mice at 22 to 28 weeks of age and expressed as fold change over WT mice. Data is pooled from three independent experiments with 3–5 mice per group in each experiment; * p < 0.05. (C) LYVE-1 + vessels were analyzed for co-expression of the proliferative marker KI67. Arrows indicate co-localization of lymphatics with Ki67. Ki67 bright dividing lymphocytes were also observed. Images are representative of three to four independent experiments ( n = 3–4 mice per group).

Article Snippet: Antibodies used included the following: rat anti-mouse CD31 (Serotec) detected with anti-rat-APC, hamster anti-mouse podoplanin (clone 8.1.1; Developmental Studies Hybridoma Bank, University of Iowa, Iowa City, IA) detected with anti-hamster PE and PerCPcy5.5—conjugated anti-mouse CD45.2 (BD Biosciences).

Techniques: Staining, Expressing, Marker

Journal: Frontiers in Immunology

Article Title: Halted Lymphocyte Egress via Efferent Lymph Contributes to Lymph Node Hypertrophy During Hypercholesterolemia

doi: 10.3389/fimmu.2019.00575

Figure Lengend Snippet: The extended lymphatic network in apoE −/− mice LNs exhibits structural abnormalities. (A) Immunoreactivity for LYVE-1 was examined in LN sections from WT and apoE −/− mice at 22–28 weeks of age. (B) Lumen area of cortical and medullary sinuses was determined on LN sections from 22–28 week-old WT and apoE −/− mice. n = 4 mice per group; *** p < 0.0005 (C) Immunoreactivity for LYVE-1 and B220, (D) LYVE-1 and collagen (Col) type IV or (E) LYVE-1 and CD31 was examined in LN sections from apoE −/− mice at 22–28 weeks of age. Images are representative of 3–4 independent experiments ( n = 3–5 mice per group). Arrows indicate loss of LYVE-1 expression.

Article Snippet: Antibodies used included the following: rat anti-mouse CD31 (Serotec) detected with anti-rat-APC, hamster anti-mouse podoplanin (clone 8.1.1; Developmental Studies Hybridoma Bank, University of Iowa, Iowa City, IA) detected with anti-hamster PE and PerCPcy5.5—conjugated anti-mouse CD45.2 (BD Biosciences).

Techniques: Expressing

Journal: The American Journal of Pathology

Article Title: Inhibition of Vascular Endothelial Growth Factor (VEGF) Signaling in Cancer Causes Loss of Endothelial Fenestrations, Regression of Tumor Vessels, and Appearance of Basement Membrane Ghosts

doi: 10.1016/s0002-9440(10)63273-7

Figure Lengend Snippet: Figure 1. Loss of vessel patency in simple vascular network of trachea after inhibition of VEGF signaling. Fluorescence micrographs showing vasculature of mouse tracheas stained by intravenous injection of lectin (A–C) and same vessels stained for CD31 immunoreactivity (D and E). Three conditions are compared: A and D, normal state; B and E, after 2 days of treatment with AG013736; and C and F, after 10 days of treatment with AG013736. After 2 days of treatment the lectin stains fewer vessels than CD31 because vessel patency is lost and blood flow stops before the CD31 disappears as endothelial cells degenerate. At 10 days, fewer vessels are evident by either method. Arrows mark regions where the lectin staining and CD31 immunoreactivity do not match. Scale bar, 30 m.

Article Snippet: Vascular basement membrane was examined with rabbit polyclonal anti-type IV collagen antibody (1:10,000; Cosmo Bio Co., Tokyo, Japan).12 Secondary antibodies were goat anti-rat IgG labeled with fluorescein isothiocyanate or Cy3 for rat CD31, CD105, 5 integrin, and PDGFR- antibodies; goat anti-hamster IgG labeled with fluorescein isothiocyanate for hamster CD31 antibody; donkey anti-rabbit IgG labeled with Cy3 for rabbit anti-VEGFR-2 antibody; donkey anti-goat IgG labeled with Cy3 for goat anti-VEGFR-3 antibody; and goat anti-rabbit IgG labeled with Cy3 or Cy5 for rabbit anti-type IV collagen antibody (1:400; Jackson ImmunoResearch, West Grove, PA).

Techniques: Inhibition, Fluorescence, Staining, Injection

Journal: The American Journal of Pathology

Article Title: Inhibition of Vascular Endothelial Growth Factor (VEGF) Signaling in Cancer Causes Loss of Endothelial Fenestrations, Regression of Tumor Vessels, and Appearance of Basement Membrane Ghosts

doi: 10.1016/s0002-9440(10)63273-7

Figure Lengend Snippet: Figure 2. Reduction in tumor vessel patency and blood flow after inhibition of VEGF signaling. Confocal microscopic images of tumor vessels in RIP-Tag2 mice treated with vehicle (A, C, E) or AG013736 (B, D, F) for 1 day and injected with lectin before fixation by vascular perfusion. Lectin staining (A, green) of blood vessels in vehicle-treated tumor closely matches CD31 immunoreactivity (C, red). However, after treatment with AG013736, multiple vessels (arrows) lack lectin staining (B, green) but have CD31 immunoreactivity (D, red), indicating loss of vessel patency. Vessels without flow appear red in merged images (E, F). Bar graphs show area density (%) of lectin staining and CD31 immunoreactivity in vessels of RIP-Tag2 tumors after treatment for 1, 2, or 7 days with AG013736 (G) or VEGF-Trap (H) or their respective vehicle. After 1 or 2 days of treatment by either agent, the area density of lectin staining is significantly less than the amount of CD31 immunoreactivity. By 7 days, this discrepancy no longer exists, suggesting that the surviving vessels have blood flow. *, Different from vehicle (P 0.05). †, Different from CD31 (P 0.05). Scale bar, 25 m.

Article Snippet: Vascular basement membrane was examined with rabbit polyclonal anti-type IV collagen antibody (1:10,000; Cosmo Bio Co., Tokyo, Japan).12 Secondary antibodies were goat anti-rat IgG labeled with fluorescein isothiocyanate or Cy3 for rat CD31, CD105, 5 integrin, and PDGFR- antibodies; goat anti-hamster IgG labeled with fluorescein isothiocyanate for hamster CD31 antibody; donkey anti-rabbit IgG labeled with Cy3 for rabbit anti-VEGFR-2 antibody; donkey anti-goat IgG labeled with Cy3 for goat anti-VEGFR-3 antibody; and goat anti-rabbit IgG labeled with Cy3 or Cy5 for rabbit anti-type IV collagen antibody (1:400; Jackson ImmunoResearch, West Grove, PA).

Techniques: Inhibition, Injection, Staining