Aortic fibronectin (FN) expression is augmented in hypertension. Increasing evidence suggests that both angiotensin II (Ang II) and mechanical factors may induce vascular remodeling in response to hypertension. We have previously shown that, in vitro, increased transmural pressure enhances FN expression in rabbit aortic media. To investigate the existence of a link between the effects of pressure and Ang II and to explore the mechanisms underlying such a relationship, we quantified the effect of Ang II and Ang II inhibitors on the pressure-dependent FN expression in a 3-day organ culture model of rabbit aorta using immunolabeling analysis and detected FN mRNAs by in situ hybridization. A dose-dependent effect of Ang II on FN expression was observed at both 80 and 150 mm Hg but not at 0 mm Hg (relaxed vessels). One mumol/L Ang II increased the media cross-sectional surface, showing FN expression from 7.9 +/- 0.7% (n = 9) to 18.9 +/- 1.1% (n = 4) at 80 mm Hg (P < .01) and from 17.4 +/- 1.8% (n = 9) to 56.6% +/- 3.6 (n = 4) at 150 mm Hg (P < .001). In situ hybridization revealed that Ang II and pressure upregulated FN mRNA expression. Losartan, an AT1 antagonist, not only blocked the Ang II effect but also inhibited the transmural pressure effect. Angiotensin-converting enzyme inhibition abolished the pressure-dependent FN expression and significantly diminished the effect of pressure in the presence of Ang II. The effect of renin-angiotensin system inhibitors was specific for FN, since neither bFGF nor laminin expression was affected by these agents. Taken together, the results demonstrate that (1) the effect of transmural pressure is mediated by the stimulation of a local renin-angiotensin system, resulting in a net Ang II production in the culture medium, (2) transmural pressure and Ang II act synergistically to enhance vascular FN expression, (3) AT1 receptors mediate both the effects of pressure and of exogenous Ang II, and (4) the effect of Ang II on FN expression is regulated at a pretranslational level.

Structural adaptation of the blood vessel wall occurs in response to mechanical factors related to blood pressure and flow. To elucidate the relative roles of pressure, flow, and medium composition, we have developed a novel organ culture system in which rabbit thoracic aorta, held at in vivo length, can be perfused and pressurized at independently varied flow and pressure for several days. Histology and histomorphometry, as well as scanning electron microscopy, revealed a well-preserved wall structure. In arteries perfused and pressurized at 80 mm Hg, endothelial injury led to a 2-fold increase in [3H]thymidine incorporation in the media, which peaked at 3 to 5 days and returned to baseline level at 6 to 8 days. In intact endothelialized vessels cultured for 3 days under no-flow conditions, pressure per se had no effect on DNA synthesis. In contrast, in the presence of serum, total protein synthesis, as assessed by [35S]methionine incorporation into the media, was enhanced 6-fold at 150 mm Hg compared with vessels pressurized at 0 or 80 mm Hg. In intact vessels perfused at a constant flow of 40 mL/min for 3 days, DNA synthesis was unchanged regardless of the pressure level when vessels were cultured in the presence of serum but increased 8-fold at both 80 and 150 mm Hg in the absence of serum. Unlike DNA synthesis, total protein synthesis was enhanced 12-fold by flow regardless of the presence or absence of serum. Expression of fibronectin was markedly enhanced at high transmural pressure, and serum potentiated its expression in the arterial wall. This novel organ culture system of perfused and pressurized vessels allowed identification of differential effects of pressure, flow, and serum on DNA and total protein synthesis, including cellular fibronectin expression.

Abstract-Different forms of mechanical stimulation are among the physiological factors constantly acting on the vessel wall. We previously demonstrated that subjecting vascular smooth muscle cells (VSMCs) in culture to cyclic stretch increased the expression of high-molecular-weight caldesmon, a marker protein of a differentiated, contractile, VSMC phenotype. In the present work the effects of mechanical factors, in the form of circumferential stress and shear stress, on the characteristics of SM contractile phenotype were studied in an organ culture of rabbit aorta. Application of an intralumininal pressure of 80 mm Hg to aortic segments cultured in Dulbecco's modified Eagle's medium containing 20% fetal calf serum for 3 days prevented the decrease in high-molecular-weight caldesmon content (70Ϯ4% of initial level in nonpressurized vessel, 116Ϯ17% at 80 mm Hg) and filamin content (80Ϯ5% in nonpressurized vessel, 100Ϯ2% at 80 mm Hg). SM myosin and low-molecular-weight caldesmon contents showed no dependence on vessel pressurization. Neither endothelial denudation nor alteration of intraluminal flow rates affected marker protein content in 3-day vessel culture, thus excluding the possibility of any shear or endothelial effects. Maintenance of high high-molecular-weight caldesmon and filamin levels in the organ cultures of pressurized and stretched vessels demonstrates the positive role of mechanical factors in the control of the VSMC differentiated phenotype. (Arterioscler Thromb Vasc Biol. 1998;18:922-927.)Key Words: stretch Ⅲ aorta Ⅲ marker protein Ⅲ caldesmon Ⅲ filamin T he arterial wall is continuously subjected to the action of mechanical forces in the form of tensile stress and shear stress. In addition, the pulsatile nature of blood pressure imposes cyclic stretch on the wall. In vivo studies and clinical observations suggest that decreased values of tensile and shear stress in surgically injured vessels are correlated with activation of cell proliferation and extracellular matrix production, which may result in vessel occlusion.1 In contrast, reestablishment of baseline tensile stress and shear stress levels is correlated with inhibition of cell proliferation and restoration of vessel wall morphology and cellular ultrastructure characteristic of the differentiated VSMC phenotype. 1,2These features and morphology indicate that VSMCs undergo phenotypic changes in regions of altered hemodynamic patterns. Experiments on isolated vessels suggest that appropriate mechanical stimulation is essential for the maintenance of VSMC contractile function and sensitivity to vasoconstrictors, 3 as well as morphology of the vessel wall. 4,5 However, the role of mechanical stimulation in the maintenance of SM marker protein content is still unclear.Changes in the expression of VSMC marker proteins are usually coordinated during VSMC transition to the synthetic phenotype in primary culture or at the loci of vascular injury.6,7 However, under certain conditions, expression of some marker proteins may be regulated independent...