Shear-Induced Increase in Hydraulic Conductivity in Endothelial Cells Is Mediated by a Nitric Oxide–Dependent Mechanism

Abstract

Abstract-This study addresses the role of nitric oxide (NO) and its downstream mechanism in mediating the shear-induced increase in hydraulic conductivity (L p ) of bovine aortic endothelial cell monolayers grown on porous polycarbonate filters. Direct exposure of endothelial monolayers to 20-dyne/cm 2 shear stress induced a 4.70Ϯ0.20-fold increase in L p at the end of 3 hours. Shear stress (20 dyne/cm 2 ) also elicited a multiphasic NO production pattern in which a rapid initial production was followed by a less rapid, sustained production. In the absence of shear stress, an exogenous NO donor, S-nitroso-N-acetylpenicillamine, increased endothelial L p 2.23Ϯ0.14-fold (100 mol/L) and 4.8Ϯ0.66-fold (500 mol/L) at the end of 3 hours. In separate experiments, bovine aortic endothelial cells exposed to NO synthase inhibitors, N G -monomethyl-L-arginine and N G -nitro-L-arginine methyl ester, exhibited significant attenuation of shear-induced increase in L p in a dose-dependent manner. Inhibition of guanylate cyclase (GC) with 583 (1 mol/L) or protein kinase G (PKG) with KT5823 (1 mol/L) failed to attenuate the shear-induced increase in L p .Furthermore, direct addition of a stable cGMP analogue, 8-bromo-cGMP, had no effect in altering baseline L p , indicating that the GC/cGMP/PKG pathway is not involved in shear stress-NO-L p response. Incubation with iodoacetate (IAA), a putative inhibitor of glycolysis, dose-dependently increased L p . Addition of IAA at levels that did not affect baseline L p greatly potentiated the response of L p to 20-dyne/cm 2 shear stress. Key Words: shear stress Ⅲ nitric oxide Ⅲ hydraulic conductivity Ⅲ endothelial cells E ndothelial cells provide the principal barrier to transport of fluids and solutes between blood and underlying tissue. In arteries, shear-dependent permeability has been hypothesized to play a role in the localization of atherosclerotic lesions in regions of vessel branching and curvatures. 1 The first clear demonstration of a direct effect of shear stress on endothelial permeability was reported by Jo et al. 2 Other investigators 3-5 have provided supporting evidence for flowor shear-dependent endothelial solute permeability. More recently, Sill et al, 6 using the same in vitro model as Jo et al, demonstrated that endothelial monolayer hydraulic conductivity (L p ) is also sensitive to acute changes in shear stress level. This finding is consistent with an earlier whole-artery study reported by Lever et al 7 and with a study involving frog mesenteric venular capillary measurements performed by Williams and Huxley. 4 The only insight into the cellsignaling mechanism underlying the shear stress response of L p was provided by Sill et al, who demonstrated that the response could be blocked by addition of a cAMP analogue or a phosphodiesterase inhibitor, suggesting a role for cAMP in the pathway.It is well established that shear stress stimulates the release of nitric oxide (NO) from cultured endothelial cells. 8,9 Direct infusion of NO synthase (NOS) inhibitors into inta...

Regulation of capillary hydraulic conductivity in response to an acute change in shear

“…Therefore, in the present study, the measurements with 50 M L-NAME treatment were conducted within 5-15 min to attenuate the possible role of leukocyte adhesion in J v /S measurements. Our in vivo observations of an attenuation in the L p response with L-NAME in the presence of an increase in shear also support the previous in vitro findings (3,37) that a change in fluid shear stress may regulate L p by releasing NO.…”

“…However, occlusion of the upstream arteriole also induces an increase in pressure upstream of the occluder and can lead to an increase in capillary pressure. In our previous measurements of A 3 arteriolar pressure (21) in the Wistar rat mesentery (same model as the current study), the mean value was 43 mmHg and the pressure increase due to occlusion was 7.4 mmHg. Therefore, it was expected that the pressure increase in the capillary due to arteriolar occlusion would be ϳ7.4 mmHg with increasing shear.…”

“…Several studies have shown that transvascular exchange can be affected by a change in perfusion rate (14,17,19,32,33,43). A small-pore model has been proposed as a possible mechanism to account for flow-dependent transport of small solutes in cannulated vessels (32); moreover, there have been several recent studies of shearinduced increases in fluid filtration performed in vitro (3,6,37) and in vivo (27,40,41). The mechanism by which shear forces could increase L p appear to be dependent on nitric oxide (NO) (3,15,24).…”

“…A small-pore model has been proposed as a possible mechanism to account for flow-dependent transport of small solutes in cannulated vessels (32); moreover, there have been several recent studies of shearinduced increases in fluid filtration performed in vitro (3,6,37) and in vivo (27,40,41). The mechanism by which shear forces could increase L p appear to be dependent on nitric oxide (NO) (3,15,24). Therefore, when a capillary stops flowing, it might be expected that L p could decline in a matter of seconds as shear-induced production of NO is attenuated.…”

Shear stress inhibits smooth muscle cell migration via nitric oxide-mediated downregulation of matrix metalloproteinase-2 activity

“…1,6,8,16,36) because, under normal physiological conditions, they are directly exposed to the SS of flowing blood, having a magnitude on the order of 10 dyn/cm 2 (26). It had long been thought that the underlying SMCs were shielded from blood flow and were only subjected to SS when the EC layer was denuded, as in balloon angioplasty, directly exposing SMCs to blood flow SS.…”