An in vitro bioassay system was developed to study endothelium-mediated, shear stressinduced, or flow-dependent generation of endothelium-derived relaxing factor (EDRF). Monolayers of aortic endothelial cells were grown on a rigid and large surface area of microcarrier beads and were packed in a small column perfused with Krebs bicarbonate solution. The perfusate was allowed to superfuse three endothelium-denuded target pulmonary arterial strips arranged in a cascade. T here are two components that make up the total force applied to the intimal surface of blood vessels during blood flow. One is a perpendicular pressure component and the other is a tangential component called the wall shear stress, a frictional force produced when the blood flows across the endothelial surface. The perpendicular pressure component is not appreciable at the endothelial surface because the force is borne primarily by structural proteins in the blood vessel wall. However, wall shear stress falls entirely on the endothelial cell layer. Thus, fluid shear stress refers to the mechanical forces generated at the endothelial cell surface by blood flowing under pressure.The vascular endothelium is capable of modulating the tone of the underlying smooth muscle in conduit and resistance vessels of the arterial bed in response to local changes in shear stress, pressure, and other mechanical factors.

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A bioassay cascade superfusion technique was utilized to study the properties of endothelium derived relaxing factor (EDRF) from human umbilical vein (HUV) and compare its actions on umbilical, chorionic plate and bovine pulmonary arterial strips. Histamine (1 μm), bradykinin (1 μm) and A‐23187 (0.3 μm, 1 μm) but not acetylcholine (1 μm) released EDRF. The non‐innervated human foetoplacental vessels, i.e., umbilical and chorionic plate arteries, do relax to EDRF by a guanosine 3′: 5′‐cyclic monophosphate (cyclic GMP)‐mediated mechanism. The sensitivity of the human umbilical arterial strips to EDRF was less than that of the chorionic plate arterial strips. Bovine pulmonary arterial strips were the most sensitive to the relaxant actions of human umbilical EDRF.

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The objective of this study was to determine whether the vascular smooth muscle contractile effect of NG_ methyl-L-arginine (NMA) is endothelium dependent and attributed to a decline in smooth muscle levels of cyclic GMP. Vascular smooth muscle levels of cyclic GMP are severalfold greater in endothelium-intact than in endothelium-denuded preparations because of the continuous formation and release of a lipophilic endothelium-derived chemical factor that diffuses into the underlying smooth muscle and activates cytosolic guanylate cyclase. This chemical substance, believed to be nitric oxide (NO) or a labile nitroso precursor, appears to account for the biological actions of endothelium-derived relaxing factor. NMA inhibits the formation of NO from endogenous L-arginine in endothelial cells. In the present study, NMA caused marked endothelium-dependent contraction of isolated rings of bovine pulmonary artery and vein, and this was similar to the contraction elicited by hemoglobin, an inhibitor of the relaxant action of NO. Both NMA and hemoglobin caused endothelium-dependent potentiation of contractile responses to phenylephrine in artery and vein. NMA caused endothelium-dependent decreases in the resting or basal levels of cyclic GMP in artery and vein to levels that were characteristic of those in endothelium-denuded vessels. Finally, NMA inhibited endothelium-dependent relaxant responses and cyclic GMP formation stimulated by acetylcholine and bradykinin.These observations reveal that interference with the continuous or basal generation of endothelium-derived NO in artery and vein can cause marked increases in vascular smooth muscle tone as a result of inhibition of cyclic GMP formation.Endothelium-derived NO is believed to account for the biological actions of an endothelium-derived relaxing factor (EDRF) discovered almost 10 years ago (1). These actions include vascular smooth muscle relaxation (2-4) and inhibition of platelet function (5, 6), which were originally demonstrated for authentic NO also about 10 years ago (7,8 (23,26,27). Indeed, the observation that NMA causes an increase in systemic blood pressure after intravenous injection into guinea pigs or rabbits suggests that basal NO generation is sufficient to influence peripheral vascular resistance (21,22). The contractile effects of hemoglobin, methylene blue, and pyrogallol are accompanied by decreases in vascular levels of cyclic GMP (26). This is attributed to prevention or inhibition of EDRFelicited activation of cytosolic guanylate cyclase. Increases in vascular cyclic GMP levels are generally associated with smooth muscle relaxation (28), and this is consistent with the hypothesis that cyclic GMP mediates NO-elicited vascular smooth muscle relaxation (7,29). The objective of this study was to elucidate the mechanism by which NMA causes arterial and venous contraction. The hypothesis tested was that NMA causes endothelium-dependent contraction by decreasing the resting levels ofcyclic GMP in artery and vein.MATERIALS AND METHODS Reagent...