P63 Hydrogen sulfide – a novel guardian of the endothelial glycocalyx?

Abstract

The endothelial glycocalyx occupies substantial space at the luminal surface of the endothelium and serves as protective layer that regulates adhesion of leukocytes to the endothelium. Shedding of the endothelial glycocalyx and increased leukocyte adhesion are common features in inflammatory pathologies pathologies (e.g. ischaemia–reperfusion injury, diabetes) and are mediated in part by ’reactive oxygen species’ (ROS). It has been suggested that H 2 S down-regulates leukocyte adherence via K ATP channels activation on leukocytes and the endothelium as a novel mechanism to limit inflammation. Blood levels of H 2 S are substantially lower in diabetic patients compared to controls suggesting ROS consumption of H 2 S in the diabetic vasculature. However, the role H 2 S in leukocyte adhesion is not understood. We have investigated the effects of GYY4137 in hyperglycaemia (HG) induced endothelial glycocalyx dysfunction and the mechanisms involved in modulation of leukocyte adhesion. Primary bovine retinal endothelial cells (BREC) were exposed to 5.6 mM glucose (NG) and 25 mM glucose (HG) ± GYY4137 (100 μM) for 24 h. Cell based fluorescence assay (CBF) with WGA-FITC was used to study EG changes. Glibenclamide, gliclazide and cromakalim (10 μM) were used to test the role of endothelial K ATP channels in leukostesis. Leukocyte adhesion to BREC was measured in Ibidi perfusion slides (shear stress 1 dyn/cm 2 ). HG-induced endothelial glycocalyx degradation was significantly inhibited by GYY4137 [HG 93 ± 5% vs HG + GYY 107 ± 8%, p < 0.001, n = 12]. Cromakalim, glibenclamide or gliclazide alone had no effect on glycocalyx changes. HG increased leukocyte adhesion was attenuated by GYY4137 [HG 275 ± 1% vs HG GYY 105 ± 11%]. H 2 S driven decrease in leukocyte adhesion was not inhibited by glibenclamide or gliclazide ruling out endothelial K ATP channel activation. H 2 S added simultaneously with HG to BREC pretreated with cromakalim did not affect leukostasis. GYY4137 inhibited HG induced endothelial glycocalyx degradation and leukocyte adhesion highlighting the therapeutic potential for H 2 S donors or the modulation of endogenous H 2 S in preserving endothelial integrity under hyperglycaemic conditions.