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(Arteriosclerosis, Thrombosis, and Vascular Biology. 2004;24:1836.)
© 2004 American Heart Association, Inc.

Vascular Biology

In Vivo Transcriptional Response of Cardiac Endothelium to Lipopolysaccharide

J. Gregory Maresh; Huaxia Xu; Nan Jiang; Ralph V. Shohet

From Internal Medicine–Cardiology, University of Texas Southwestern Medical Center, Dallas, Tex.

Correspondence to Ralph V. Shohet, MD, Internal Medicine–Cardiology, University of Texas Southwestern Medical Center at Dallas, 5323 Harry Hines Blvd, Dallas, TX 75390-8573. E-mail ralph.shohet{at}utsouthwestern.edu

Objective— Vascular endothelial cells must integrate stimuli from multiple sources, including plasma, leukocytes, and neighboring components of the vessel. These stimuli are difficult to recapitulate in vitro. We have developed a method to examine the in vivo regulation of gene expression in endothelial cells and have applied it to a model of sepsis.

Methods and Results— We used fluorescent-activated cell sorting to isolate highly purified endothelial cells from the hearts of transgenic mice that express green fluorescent protein driven by the endothelial-specific promoter Tie2. We treated these mice with intraperitoneal lipopolysaccharide and identified those genes within cardiac endothelium that were >3-fold dysregulated 4 and 24 hours later by microarray analysis. These findings were confirmed by real-time polymerase chain reaction and compared with in vitro regulation in a murine endothelial cell line.

Conclusions— The in vivo regulation was distinct and, in general, more robust than that seen in vitro. We identified endothelial-expressed genes not previously recognized to be regulated in response to lipopolysaccharide. This approach provides insight into the cardiac-specific responses of the endothelium that contribute to the specific responses of the heart to sepsis, and can be generalized to the exploration of endothelial responses in any organ.

Fluorescent cell sorting was used to isolate green fluorescent protein expressing endothelial cells from the hearts of mice exposed to lipopolysaccharide. In vivo transcriptional changes were distinct compared with in vitro experiments and identified novel regulated genes. This method can be generalized to the exploration of in vivo endothelial responses in any organ.

Key Words: lipopolysaccharide • microarray • gene expression • endothelial • sepsis

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