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. 2014 Oct 4;16:425. doi: 10.1186/s13058-014-0425-7

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© Novitskiy et al.; licensee BioMed Central Ltd. 2014

This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.

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Increased angiogenesis and proliferation in mammary carcinomas of mouse mammary tumor virus (MMTV)-Neu mice with dominant-negative (DN) transforming growth factor β (TGFβ) receptor II (DNIIR). (A) Autopsy of mice with spontaneous tumor formation of the mammary gland, which indicates increased tumor vasculature in mice with DNIIR expression. (B) Isolated tumor tissue on 12^th week after implanted carcinoma cells via collagen plugs to the mammary fat pad. (C) H&E staining indicated morphological changes in control (NK1Mul) and experimental mice (NK1Mul/DN). (D) Immunohistochemistry (IHC) staining of frozen sections for endothelial cell marker CD31. Staining was performed with hematoxylin (blue) to counterstain all nuclei. (E) Immunofluoresence staining of frozen sections for the endothelial cells CD31 (green), pericyte marker NG2 (red) and nuclei by 4',6-diamidino-2-phenylindole (DAPI) (blue). (F) Representative fluorescence-activated cell sorting plots and quantitative analysis of pericyte percentage (PDGFRβ⁺PDGFRα⁻) in tumor tissue. Cells were gated as non-immune, non-epithelial (CD45⁻CD326⁻). (G) Formalin-fixed paraffin-embedded sections of fluorescein isothiocyanate (FITC-)dextran (green)-perfused tumor tissue for indication of vessel leakage. DAPI (blue) was used to counterstain all nuclei. (H) IHC for 5-bromo-2'-deoxyuridine (BrdU) on primary tumors, which indicate newly synthesized DNA in proliferating cells. Quantification of IHC (right) BrdU-positive cells indicates 2.5 times more positive cells in mice with DNIIR than control tissue section. (Scale bars: C, D, E, J, K, 500 μM; F, 1,000 μM).