Abstract
PurposeAcute increases in hydrostatic pressure activate endothelial signaling pathways that modulate barrier function and vascular permeability. We investigated the role the glycocalyx and established mechanotransduction pathways in pressure-induced albumin transport across rat lung microvascular endothelial cells.MethodsRat lung microvascular endothelial cells (RLMEC) were cultured on Costar Snapwell chambers. Cell morphology was assessed using silver nitrate staining. RLMEC were exposed to zero pressure (Control) or 30 cmH2O (Pressure) for 30 or 60 min. Intracellular albumin uptake and transcellular albumin transport was quantified. Transcellular transport was reported as solute flux (Js) and an effective permeability coefficient (Pe). The removal of cell surface heparan sulfates (heparinase), inhibition of NOS (L-NAME) and reactive oxygen species (apocynin, Apo) was investigated.ResultsAcute increase in hydrostatic pressure augmented albumin uptake by 30–40% at 60 min and Js and Pe both increased significantly. Heparinase increased albumin uptake but attenuated transcellular transport while L-NAME attenuated both pressure-dependent albumin uptake and transport. Apo interrupted albumin uptake under both control and pressure conditions, leading to a near total lack of transcellular transport, suggesting a different mechanism and/or site of action.ConclusionPressure-dependent albumin uptake and transcellular transport is another component of endothelial mechanotransduction and associated regulation of solute flux. This novel albumin uptake and transport pathway is regulated by heparan sulfates and eNOS. Albumin uptake is sensitive to ROS. The physiological and clinical implications of this albumin transport are discussed.
Highlights
In 1991 Dull et al.[6] published a report characterizing hydraulic conductivity (Lp), solute flux (Js) and the effective permeability coefficient (Pe) of albumin across cultured aortic endothelial monolayers during step increases in hydrostatic pressures
It was shown that pressure-dependent endothelial nitric oxide synthase (eNOS) activation and the increase in Lp was mediated by heparan sulfate proteoglycans
At 60 min, Bovine serum albumin (BSA) uptake increased with pressure as the control monolayers had 607.1 ± 76.1 lg/mL and pressure-treated monolayer increased to 914.9 ± 105.2 lg/mL (p
Summary
In 1991 Dull et al.[6] published a report characterizing hydraulic conductivity (Lp), solute flux (Js) and the effective permeability coefficient (Pe) of albumin across cultured aortic endothelial monolayers during step increases in hydrostatic pressures. Z. CHIGNALIA quently showed that pressure activated NO production in lung capillary endothelial cells grown on a rigid support where stretch was virtually absent. CHIGNALIA quently showed that pressure activated NO production in lung capillary endothelial cells grown on a rigid support where stretch was virtually absent In those studies, it was shown that pressure-dependent eNOS activation and the increase in Lp was mediated by heparan sulfate proteoglycans. Inhibition of eNOS mitigates both pressure and shear-induced changes in permeability as demonstrated using a variety of endothelial cell types.[5,11,15–17] Dull et al.[7] extend these studies to a perfused lung model and demonstrated that acute increase in pulmonary capillary pressure resulted in an increase in lung filtration coefficient (Kf) and this was mediated by a heparan sulfate-nitric oxide mechanism. We demonstrated that inhibition of lung NOS, prevented pulmonary edema in a rat model of acute hypertensive heart failure, despite persistently elevated pulmonary capillary pressure.[4]
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