In vitro culture and characterization of gene targeted mouse endothelium

Endothelial cells play a crucial role in maintaining cardiovascular homeostasis. Although many cardiovascular disorders involve endothelial cell dysfunction, the specific cellular and molecular mechanisms involved are not well known. We sought to establish a reproducible method of endothelial cell isolation from gene targeted mice to specifically examine endothelial pathophysiological mechanisms. Primary aortic endothelial cell cultures were established from wild type and intercellular adhesion molecule‐1 (ICAM‐1) deficient mice. Isolation of mouse aortic endothelial cells (MAEC) by fluorescent activated cell sorting routinely resulted in pure, homogenous, primary cultures. Wild type and ICAM‐1 deficient endothelial cell morphology was similar, with both cultures showing cobblestone morphology and DiI‐Ac‐LDL staining. Monocyte adhesion to ICAM‐1 deficient aortic endothelial cells was decreased by 86% as compared with wild type MAEC. Monocyte adhesion was also determined using YN‐1, an ICAM‐1 blocking antibody. YN‐1 decreased monocyte adhesion to wild type aortic endothelial cells by 25%, whereas YN‐1 did not further decrease monocyte adhesion to ICAM‐1 deficient MAEC. These data demonstrate that gene targeted endothelial cell cultures are an effective means of identifying specific cellular and molecular mechanisms involved in endothelial cell physiology and dysfunction.