The mouse aorta model: Influence of genetic background and aging on bFGF- and VEGF-induced angiogenic sprouting Wen-Hui Zhu 1 , Monica Iurlaro 1 , Angela MacIntyre 2 , Eric Fogel 3 & Roberto F. Nicosia 1,3 1 Department of Pathology, University of Washington, Seattle, Washington, USA; 2 Department of Pathology, Microbiology, and Immunology, Philadelphia College of Osteopathic Medicine, Philadelphia, Pennsylvania, USA; 3 Division of Pathology and Laboratory Medicine, Veterans Administration Puget Sound Health Care System, Seattle, Washington, USA Received 20 October 2003; accepted in revised form 10 November 2003 Key words: angiogenesis, aorta, assay, bFGF, endothelium, mouse Abstract Angiogenesis can be studied ex vivo by culturing rat or mouse aortic rings in collagen gels. Unlike rat aorta explants, unstimulated mouse aortic rings were unable to spontaneously produce an angiogenic response under serum-free conditions. They, however, responded to bFGF and VEGF, generating networks of branching neovessels. Aortic rings from GFP-Tie2-transgenic mice generated GFP-labeled neovessels that could be easily identified by their distinctly green fluorescence. Aortic rings from 1- to 2-month-old mice produced microvessels faster, more uniformly and in greater number than aortic rings from 6- to 10-month-old mice, particularly in VEGF-treated cultures. Aortic rings from 129/SVJ mice were capable of a much stronger and sustained angiogenic response to bFGF than those of C57BL/6 or BALB/c mice, which were in turn more angiogenic than aortic rings from FVB mice. The same strains of mice responded differently to VEGF, as C57BL/6 mouse aortic rings produced more microvessels than those of BALB/c, FVB, and 129/SVJ mice, which were capable of only a limited response. The significant impact that aging and genetic background have on mouse aortic angiogenesis should be taken into account when the aortic-ring assay is used to evaluate function of genes that have been deleted or overexpressed in genetically modified mice. Abbreviations: bFGF – basic fibroblast growth factor; GFP – green fluorescent protein; VEGF – vascular endo- thelial growth factor Introduction The rat aorta model of angiogenesis, also known as the aortic ring assay, has gained wide acceptance as a tool to study angiogenesis and its mechanisms [1–6]. In this system, aortic rings cultured in collagen or fibrin gels generate a self-limited angiogenic response, which, under appropriate culture conditions, can be monitored and quantified for a variety of applications [1, 7]. This approach offers many advantages, including the possi- bility of generating many assays per animal and the lack of inflammatory complications. Microvessels have his- totypic features and develop from native angioformative cells that have not been modified by repeated passages in culture. The assay can be used to evaluate the ability of growth factors to induce angiogenesis from quies- cent vascular explants or to study how modifications in the extracellular matrix affect the angiogenic process [8–10]. The mouse, whose genome has been sequenced in parallel with the human genome, is gradually becoming the animal of choice for molecular biologists, develop- mental biologists and experimental oncologists. Gene manipulation technology has greatly enhanced the experimental usefulness of this animal, which has now become a powerful model to study protein function in angiogenesis [11, 12]. Mice are also commonly utilized to evaluate the anti-tumor efficacy of anti-angiogenic drugs, prior to human testing in clinical trials [13–16]. Recently, the aortic ring model has been applied to mice. Methodological modifications introduced in the mouse aortic ring model [17, 18], however, suggest that there might be significant differences in the way the aortic wall of different animal species responds to angio- Correspondence to: Roberto F. Nicosia, MD, PhD, Division of Pathology and Laboratory Medicine (S-113-Lab), VA Puget Sound Health Care System, 1660 South Columbian Way, Seattle, WA 98108, USA. Tel: +1-206-764-2284; Fax: +1-206-764-2001; E-mail: roberto.nicosia@med.va.gov Angiogenesis 6: 193–199, 2003. 193 Ó 2004 Kluwer Academic Publishers. Printed in the Netherlands.