Abstract

ObjectiveThe endothelial glycocalyx and the regulation of its shedding are important to vascular health. Endo-{beta}-D-glucuronidase heparanase-1 (HPSE1) is the only enzyme that can shed heparan sulfate. However, the mechanisms are not well understood.\n\nApproach and resultsTo investigate HPSE1 and its endogenous inhibitor, heparanase-2 (HPSE2), we used cell culture, lentiviral protein overexpression, a microfluidic chip model of cell culture under shear stress conditions, and lipopolysaccharide (LPS) injections in mice. We show that HPSE1 activity aggravated Toll-like receptor 4 (TLR4)-mediated response of endothelial cells to LPS. On the contrary, HPSE2 overexpression was protective. The microfluidic chip flow model confirmed that HPSE2 prevented heparan sulfate shedding by HPSE1. Furthermore, heparan sulfate did not interfere with cluster of differentiation-14 (CD14)-dependent LPS binding, but instead reduced the presentation of the LPS-CD14 complex to TLR4. HPSE2 reduced LPS-mediated TLR4 activation by LPS, subsequent cell signaling, and cytokine expression. Moreover, HPSE2-overexpressing endothelial cells remained protected against LPS-mediated loss of cell-cell contacts. In vivo, expression of HPSE2 in plasma and kidney medullary capillaries was decreased in mouse sepsis model. We next applied purified HPSE2 in mice and observed decreases in TNF and IL-6 plasma concentrations after intravenous LPS injections.\n\nConclusionsOur data demonstrate the important role of heparan sulfate and the glycocalyx in endothelial cell activation and suggest a protective role of HPSE2 in microvascular inflammation. HPSE2 offers new options for protection against HPSE1-mediated endothelial damage and preventing microvascular disease.\n\nO_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=179 SRC=\"FIGDIR/small/705939v1_ufig1.gif\" ALT=\"Figure 1\">\nView larger version (50K):\norg.highwire.dtl.DTLVardef@1bae526org.highwire.dtl.DTLVardef@11a6039org.highwire.dtl.DTLVardef@a36deforg.highwire.dtl.DTLVardef@1869ea8_HPS_FORMAT_FIGEXP M_FIG Graphical abstract\n\nC_FIG O_TBL View this table:\norg.highwire.dtl.DTLVardef@11a3deborg.highwire.dtl.DTLVardef@1d71919org.highwire.dtl.DTLVardef@b0c8a3org.highwire.dtl.DTLVardef@4477f3org.highwire.dtl.DTLVardef@11f1588_HPS_FORMAT_FIGEXP M_TBL O_TABLECAPTIONNon standard abbreviations\n\nC_TABLECAPTION C_TBL