Transgenic model of cardiac rhabdomyosarcoma formation

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Transgenic model of cardiac rhabdomyosarcoma formation Christiane Köbbert, PhD, Christa Möllmann, Michael Schäfers, MD, Sven Hermann, MD, Hideo A. Baba, MD, Andreas Hoffmeier, MD, Günter Breithardt, MD, Hans H. Scheld, MD, Gabriele Weissen–Plenz, PhD, Jürgen R. Sindermann, MD The Journal of Thoracic and Cardiovascular Surgery Volume 136, Issue 5, Pages 1178-1186 (November 2008) DOI: 10.1016/j.jtcvs.2008.04.022 Copyright © 2008 The American Association for Thoracic Surgery Terms and Conditions

Figure 1 Panel A shows comparative displays of the heart of an approximately 12-week-old mouse and an age-matched control animal. The heart of the transgenic animal presents a giant tumor formation in the left ventricle, which is associated with an overtly enlarged right and left atrium. Panel B shows a cross-section of the same transgenic heart (arrow) stained with hematoxylin and eosin. The heart was cut at the ventricular level below the heart basis. The spherical tumor occupies about 55% of the cross-sectional luminal area of the left ventricle. Panel C displays tumor tissue and adjacent regular cardiac muscle of the same specimen stained with hematoxylin and eosin. A small blood-filled area is surrounded by the tumor tissue on one side and regular cardiac muscle on the other side. The tumor infiltrates between struts of regular cardiac muscle. Tissues were prepared as described in the Methods section. Bars: A, 5 mm; B, 1 mm; C, 100 μm. The Journal of Thoracic and Cardiovascular Surgery 2008 136, 1178-1186DOI: (10.1016/j.jtcvs.2008.04.022) Copyright © 2008 The American Association for Thoracic Surgery Terms and Conditions

Figure 2 High-resolution positron emission tomographic (PET) scan 1 hour after injection of 10 MBq of 2-[18F]fluoro-2-deoxy-D-glucose (FDG) of the transgenic mouse displayed in Figure 1 and an age-matched control animal. Left panel, Whole-body slice through the heart; right panel, short axis (SA), horizontal long axis (HLA), and vertical long axis (VLA) of the heart. Images display enhanced myocardial glucose uptake in the enlarged atria and the right ventricle of the transgenic mouse compared with normal glucose uptake in the control mouse. The left ventricular tumor (arrows) displays a positive FDG-PET signal. However, this signal is relatively low compared with the strong FDG uptake in the myocardium of mice. The Journal of Thoracic and Cardiovascular Surgery 2008 136, 1178-1186DOI: (10.1016/j.jtcvs.2008.04.022) Copyright © 2008 The American Association for Thoracic Surgery Terms and Conditions

Figure 3 Histologic displays of a left ventricular tumor. The tumor of an approximately 9-week-old transgenic mouse is based at the transition between the wall and the interventricular septum. Panels A through C depict hematoxylin and eosin staining in various magnifications to illustrate the microscopic characteristics of the tumor. Panel D displays immunohistochemical staining for TAg (brown nuclei), which is expressed in the tumor but not in the adjacent regular cardiac muscle (TAg antibody Clone PAb 101; PharMingen, San Diego, Calif; color development was performed with diaminobenzidine and hematoxylin counterstain). Bars: A, 600 μm; B, 120 μm; C and D, 30 μm. The Journal of Thoracic and Cardiovascular Surgery 2008 136, 1178-1186DOI: (10.1016/j.jtcvs.2008.04.022) Copyright © 2008 The American Association for Thoracic Surgery Terms and Conditions

Figure 4 Immunohistochemical staining of a left ventricular tumor of an approximately 9-week old transgenic mouse. Staining was performed as explained in the Methods section. The tumor revealed positive staining for smooth muscle α-actin (A), desmin (B), and troponin C (C) and positive staining of single nuclei for Myo D1 (D). In addition, panel E displays immunohistochemical staining for proliferative cell nuclear antigen. Panel F shows a negative control receiving a secondary antibody but no primary antibody. Color development was performed with diaminobenzidine and hematoxylin counterstaining. Bar: 50 μm. The Journal of Thoracic and Cardiovascular Surgery 2008 136, 1178-1186DOI: (10.1016/j.jtcvs.2008.04.022) Copyright © 2008 The American Association for Thoracic Surgery Terms and Conditions

Figure 5 Immunohistochemical staining for TAg in various smooth muscle–containing tissues of transgenic mice at the age of approximately 9 to 11 weeks. Panel A displays positive TAg staining and increased cellular density, especially in the inner lamellar unit of the thoracic aorta. TAg expression is also shown for the common carotid artery (B). In addition, TAg expression is also displayed for the smooth muscle of the colon (C), urinary bladder (D), and uterus (E). Panel F shows a negative control (urinary bladder) specimen that received a secondary antibody but no primary antibody. Color development was performed with diaminobenzidine and hematoxylin counterstaining. Bar: 50 μm. The Journal of Thoracic and Cardiovascular Surgery 2008 136, 1178-1186DOI: (10.1016/j.jtcvs.2008.04.022) Copyright © 2008 The American Association for Thoracic Surgery Terms and Conditions

Figure 6 Quantitative analysis of the cellular density of the heart, colon, urinary bladder, and thoracic aorta of transgenic mice and control animals. Cell nuclei of cross-sections were visualized with hematoxylin. Cellular cross-sectional density was determined by counting area-defined representative microscopic fields and relating the cell number to the areas for transgenic animals (n = 8; bladder, n = 7) and age-matched control animals (n = 8). Cellular density was most pronounced in the cardiac tumors and was significantly higher than in the regular heart tissue of the same transgenic mice (nontumor cardiac tissue) and significantly higher than the cellular density of the hearts of control animals. Data are presented as means ± standard deviation. The Journal of Thoracic and Cardiovascular Surgery 2008 136, 1178-1186DOI: (10.1016/j.jtcvs.2008.04.022) Copyright © 2008 The American Association for Thoracic Surgery Terms and Conditions

Figure 7 Quantitative analysis of the fraction of TAg-positive cells in the heart tumors (n = 8), colon (n = 8), urinary bladder (n = 7), uterus (n = 7), and thoracic aorta (n = 8) of transgenic mice. TAg was detected by means of immunohistochemistry with antibody Clone PAb 101 (PharMingen, San Diego, Calif), as described in the Methods section. Color development was performed with diaminobenzidine. Hematoxylin counterstaining was used for the visualization of all cell nuclei. Fractions of TAg-positive cells were determined by counting area-defined representative microscopic fields. Data are presented as means ± standard deviation. The fraction of TAg-positive cells in the heart tumors was significantly higher than the fraction of TAg-positive cells in other tissues studied (P < .05). The Journal of Thoracic and Cardiovascular Surgery 2008 136, 1178-1186DOI: (10.1016/j.jtcvs.2008.04.022) Copyright © 2008 The American Association for Thoracic Surgery Terms and Conditions