Heparan Sulfate Proteoglycan Is a Mechanosensor on Endothelial Cells

Abstract

The objective of this study was to test whether a glycosaminoglycan component of the surface glycocalyx layer is a fluid shear stress sensor on endothelial cells (ECs). Because enhanced nitric oxide (NO) production in response to fluid shear stress is a characteristic and physiologically important response of ECs, we evaluated NO_x (NO₂⁻ and NO₃⁻) production in response to fluid shear stress after enzymatic removal of heparan sulfate, the dominant glycosaminoglycan of the EC glycocalyx, from cultured ECs. The significant NO_x production induced by steady shear stress (20 dyne/cm²) was inhibited completely by pretreatment with 15 mU/mL heparinase III (E.C.4.2.2.8) for 2 hours. Oscillatory shear stress (10±15 dyne/cm²) induced an even greater NO_x production than steady shear stress that was completely inhibited by pretreatment with heparinase III. Addition of bradykinin (BK) induced significant NO_x production that was not inhibited by heparinase pretreatment, demonstrating that the cells were still able to produce abundant NO after heparinase treatment. Fluorescent imaging with a heparan sulfate antibody revealed that heparinase III treatments removed a substantial fraction of the heparan sulfate bound to the surfaces of ECs. In summary, these experiments demonstrate that a heparan sulfate component of the EC glycocalyx participates in mechanosensing that mediates NO production in response to shear stress. The full text of this article is available online at http://www.circresaha.org.