Constitutive NOS expression in cultured endothelial cells is elevated by fluid shear stress

Abstract

The role of chronic fluid shear stress on endothelial constitutive nitric oxide synthase (cNOS) levels may have an important role in vessel diameter control. We subjected primary human umbilical vein endothelial cells (HUVEC) or bovine aortic endothelial cells (BAEC, passages 2-14) to steady laminar shear stress. In both cell types, the intracellular level of cNOS was elevated within 3 h of flow exposure at 25 dyn/cm2 and remained elevated at 6 and 12 h of flow exposure, compared with stationary controls, as indicated by digital immunofluorescence microscopy. Shear stress exposure for 6 h caused a 2.2 +/- 0.3- and 2.8 +/- 0.3-fold elevation of cNOS protein levels in BAEC (n = 3, P < 0.01) and HUVEC (n = 3, P < 0.01), respectively, in the presence or absence of 1 microM dexamethasone. Dexamethasone suppresses induction of the inducible NOS gene, indicating that cNOS was elevated by fluid shear stress. Flow exposure at 4 dyn/cm2 caused no enhancement of cNOS levels in either cell type. The flow induction of the cNOS protein levels was not blocked by preincubation of BAEC with 100-400 microM of NG-nitro-L-arginine methyl ester, indicating that flow-induced NO (or guanosine 3',5'-cyclic monophosphate) was not involved in the elevation of cNOS levels. Protein kinase C inhibitor H-7 (10 microM) had no effect on induction of NOS protein in BAEC exposed to 25 dyn/cm2. The cNOS mRNA levels were found to be elevated by two- to threefold in BAEC after 6 or 12 h of flow exposure at either 4 or 25 dyn/cm2, and this induction of NOS mRNA occurred in the presence of dexamethasone. The elevation of cNOS levels by chronic flow exposure may provide a mechanism for chronic regulation of vessel diameter by endothelial response to prevailing blood flow.

Superoxide as a Messenger of Endothelial Function

“…Another reason for increased eNOS expression might be hemodynamic forces such as shear stress, which have been shown to upregulate eNOS in vitro (Ranjan et al, 1995). However, neither blood pressure, heart rate, nor hematocrit value changed with age in these animals.…”

Ventilation and oxygenation induce endothelial nitric oxide synthase gene expression in the lungs of fetal lambs.

“…In vivo, endothelial cells are exposed to fluid mechanical forces, including hydrostatic pressure, strain, and shear stress (25,26,38,39). Shear stress is proportional to the velocity of blood and its viscosity and inversely proportional to the internal radius of the blood vessel to the third power (38,39).…”

“…Initially, NOS activity and NO production are increased when endothelial cells are exposed to increased shear stress (12,14,15,(20)(21)(22)(23). If the shear is maintained there are then subsequent increases in eNOS mRNA and protein expression (24)(25)(26). The immediate increase in pulmonary blood flow at birth may increase shear forces on the pulmonary vascular endothelium inducing eNOS mRNA and protein expression.…”

Nitric oxide synthase and NAD(P)H oxidase modulate coronary endothelial cell growth

“…Although eNOS is constitutively expressed, its expression and activity are mediated by many physio-pathological processes e.g. while shear stress increases its expression in cultured endothelial cells, hypoxia elicits a decrease in eNOS expression/activity [6,7]. NO plays significant roles in the regulation of vascular tone, in the modulation of endothelial cell permeability and in the inhibition of vascular smooth muscle cell (VSMC) proliferation and migration [1].…”