Inhibition of glycan shedding and leukocyte-endothelial adhesion in postcapillary venules by suppression of matrixmetalloprotease activity with doxycycline

Abstract

Objective: The aims of this study were to examine the role of matrixmetalloproteinases (MMPs) in causing shedding of glycan components of the endothelial glycocalyx and delineate the efficacy of doxycycline as an inhibitor of white blood cell-endothelial cell (WBC-EC) adhesion and glycan shedding in postcapillary venules.

Methods: WBC-EC adhesion in postcapillary venules of mesentery (rat) was examined in response to superfusion with the chemoattractant, f-Met-Leu-Phe (fMLP). Glycan shedding was delineated by using fluorescently labeled microspheres (FLMs; 0.1 microm in diameter) coated with lectins and infused into the systemic circulation. The shedding of FLMs in response to fMLP was examined during superfusion with graded concentrations of doxycycline and the zinc chelator, ilomastat.

Results: Superfusion of mesentery with 10(-7) M of fMLP caused a reduction in FLM adhesion due to shedding of the glycocalyx and a rise in WBC-EC adhesion. WBC-EC adhesion and FLM shedding were reduced with subantimicrobial concentrations of doxycycline equal to or greater than 0.5 muM with an EC(50) value of 0.15 microM. MMP activation was verified by inhibition of shedding and attenuation of circulating MMP substrate cleavage at the venular wall with the zinc chelator, ilomastat (GM6001, 2.6 microM; US Biological, Swampscott, Massachusetts, USA).

Conclusions: MMPs play a significant role in glycan shedding and WBC-EC adhesion, and doxycycline may stabilize the endothelial glycocalyx by inhibition of MMP activation.