Abstract: FR-PO683

Heparanase II Inhibits Heparanase I-Mediated Cleavage of Endothelial Glycocalyx, Preserves Glomerular Integrity, and Prevents Microvascular Injury

Session Information

Category: Glomerular

  • 1001 Glomerular: Basic/Experimental Immunology and Inflammation

Authors

  • Kiyan, Yulia, Hannover Medical School, Hannover, Germany
  • Stahl, Klaus, MHH, Hannover, Germany
  • Tkachuk, Sergey, Medizinische Hochschule Hannover, Hannover, Germany
  • Schroder, Patricia Ann, Mount Desert Island Biological Laboratory, Salisbury Cove, Maine, United States
  • Beverly-Staggs, Laura L., Mount Desert Island Biological Lab, Salisbury Harbor, Maine, United States
  • Schiffer, Mario, Hannover Medical School, Hannover, Germany
  • Kiyan, Roman, Laser Zentrum Hannover e.V., Hannover, Germany
  • Chichkov, Boris, Laser Zentrum Hannover e.V., Hannover, Germany
  • Haller, Hermann G., Hannover Medical School, Hannover, Germany
Background

Regulation of heparan sulfate (HS) chains of the glycocalyx is an important pathomechanism of vascular and renal diseases. Heparanase 1 (HPSE1) is the only known glucuronidase capable of degrading HS chains. Recently cloned heparanase 2 (HPSE2) is catalytically inactive and its functions are yet unclear. We have tested the hypothesis that HPSE2 antagonizes HPSE1, prevents HS chain cleavage and protects endothelial cell function.

Methods

In vitro study of endothelial cells (EC) were carried out (1) in cell culture and (2) in a microfluidic chip under flow conditions. We developed a lentiviral construct and upregulated HPSE2 expression in EC. To assess HSPE2 function in vivo we used a transgenic zebrafish model (Tg(I-fabp:GFP-DBP) and measured loss of fluorescent protein form the circulation.

Results

Addition of active HPSE1 led to the shedding of HS layer from endothelial cell surface. Overexpressing HPSE2 protected against HPSE1-induced glycocalyx shedding and damage of VE-cadherin junctions, cytoskeletal rearrangements and from glucose-induced ICAM expression and adhesion of AM-labelled monocytes. LPS stimulation with increased expression of IL-6, IL-1β, and RANTES, as well as phosphorylation of p65 subunit of NFκB, p38, and MEK kinases was diminished by HPSE2 overexpression. HPSE2 knockdown in zebrafish showed a phenotype characterized by general body edema and albuminuria. Diffuse leakage of from the intravascular compartment into the interstitial tissue of the fish tail was visualized. Simultaneous injection of human HPSE2 full length mRNA we partially rescued the proteinuric phenotype of HPSE2-KD fish.

Conclusion

HPSE2 is important for endothelial development and function in zebrafish model and in vitro under flow conditions. Our results suggest that HPSE2, expressed locally by EC or delivered with blood, fulfills protective role in microvasculature via several distinct mechanisms. First, it binds to and protects the HS glycocalyx from enzymatic shedding by HPSE1. In addition, HPSE2 binding to HS diminishes HS involvement in receptor-ligand interaction and is anti-inflammatory. We suggest that the C-terminal part of the HPSE2 protein containing HS-binding motif might be critical for endothelial function.

Funding

  • Government Support - Non-U.S.