Autocrine Regulation of Endothelial Cell Function by CAP37

Abstract

Objective Cationic antimicrobial peptide (CAP37, azurocidin) has been identified as the primary soluble mediator through which neutrophils (PMN) compromise endothelial cell (EC) barrier function at sites of inflammation. Findings herein show barrier function and viability of activated EC are altered by neutralization of CAP37 produced in the absence of neutrophils, suggesting that CAP37 regulates EC function through an autocrine pathway. Methods Human umbilical vein endothelial cells (HUVEC) were grown to confluence on electrode arrays. TNF alpha was added, or not, and transendothelial electrical impedance (an index of EC barrier function) was measured using an electric cell-substrate impedance sensor system (ECIS). Rabbit anti-CAP37 antibody or Normal Rabbit Serum (NRS) was added 12h after activation. HUVEC were also plated on 96-well plates, treated as above, and viability assessed via MTT. Results Cells treated with anti-CAP37 antibody initially showed an increase in barrier function followed by a dose dependent loss of cell resistance. NRS controls showed no change in barrier function, and thrombin controls verified cell functionality. Blocking CAP37 also led to loss of cell viability compared to controls. Conclusions CAP37 neutralization demonstrates that endothelial cells contribute to regulation of their own barrier function. In addition, endogenous CAP37 appears to be an important endothelial cell survival factor. NIH066194, AI28018