Development of immortalized mouse aortic endothelial cell lines

• Type of Material: Article
• Authors: Ni Chih-Wen, Emory University; Sandeep Kumar, Emory University; Casey J Ankeny, Emory University; Hanjoong Jo, Emory University
• Language: English
• Date: 2014-04-01
• Publisher: BioMed Central Ltd.
• Publication Version: Final Published Version
• Copyright Statement: © 2014 Ni et al.
• License: Creative Commons Attribution 2.0 Generic
• Final Published Version (URL): http://dx.doi.org/10.1186/2045-824X-6-7
• Title of Journal or Parent Work: Vascular Cell
• Volume: 6
• Issue: 1
• Start Page: 7
• End Page: 7
• Grant/Funding Information: This work was supported by funding from NIH grants HL095070, HL114772, HL113451 to HJ. This work was also supported by the National Heart Lung and Blood Institute of the NIH as a Program of Excellence in Nanotechnology award HHSN268201000043C to HJ. SK is an American Heart Association Postdoctoral fellow.
• Keywords: MAEC; Shear stress; p47phox; eNOS; cav1; Endothelial cells
• Research Categories: Health Sciences, General; Engineering, Biomedical

Abstract

Background: The understanding of endothelial cell biology has been facilitated by the availability of primary endothelial cell cultures from a variety of sites and species; however, the isolation and maintenance of primary mouse aortic endothelial cells (MAECs) remain a formidable challenge. Culturing MAECs is difficult as they are prone to phenotypic drift during culture. Therefore, there is a need to have a dependable in vitro culture system, wherein the primary endothelial cells retain their properties and phenotypes. Methods. Here, we developed an effective method to prepare immortalized MAEC (iMAEC) lines. Primary MAECs, initially isolated from aortic explants, were immortalized using a retrovirus expressing polyoma middle T-antigen. Immortalized cells were then incubated with DiI-acetylated-low density lipoprotein and sorted via flow cytometry to isolate iMAECs. Results: iMAECs expressed common markers of endothelial cells, including PECAM1, eNOS, VE-cadherin, and von Willebrand Factor. iMAECs aligned in the direction of imposed laminar shear and retained the ability to form tubes. Using this method, we have generated iMAEC lines from wild-type and various genetically modified mice such as p47phox-/-, eNOS-/-, and caveolin-1-/-. Conclusion: In summary, generation of iMAEC lines from various genetically modified mouse lines provides an invaluable tool to study vascular biology and pathophysiology. © 2014 Ni et al.; licensee BioMed Central Ltd.