Jo G. R. De Mey

Structural properties of rat mesenteric small arteries after 4-wk exposure to elevated or reduced blood flow

We determined the structure of mesenteric small arteries after chronic elevation and chronic reduction of blood flow. In 6-wk-old rats, we ligated second-order side branches of every other first-order side branch of the superior mesenteric artery. This persistently reduced blood flow (-90%) in the vessels feeding into the ligated trees and elevated blood flow (+80%) in the nonligated mesenteric artery side branches. Four weeks after surgery, vessels that had been exposed to high blood flow (HF) or low blood flow (LF) and vessels from sham-operated rats (Sham) were isolated and mounted in a pressure myograph system. At an intraluminal pressure of 100 mmHg, the internal diameter at rest was larger in HF (533 +/- 23 microm) and smaller in LF (262 +/- 14 microm) than in Sham vessels (427 +/- 15 microm). Also, wall and media cross-sectional areas were larger in HF and smaller in LF than in Sham vessels (media: 22 +/- 1, 11 +/- 2, and 16 +/- 1 x 10(3) microm2, respectively), but circumferential wall stress did not differ among groups. DNA content was significantly increased in HF vessels (+100%) and was not modified in LF vessels. Maximal vasoconstrictions elicited by high potassium or norepinephrine were slightly increased in HF vessels but were reduced by 50% in LF vessels. Thus chronic changes in blood flow give rise to structural changes that normalize circumferential wall stress. Elevated blood flow resulted in outward hypertrophic remodeling involving hyperplasia. Reduced blood flow resulted in inward hypotrophic remodeling accompanied by hyporeactivity of the arterial smooth muscle.We determined the structure of mesenteric small arteries after chronic elevation and chronic reduction of blood flow. In 6-wk-old rats, we ligated second-order side branches of every other first-order side branch of the superior mesenteric artery. This persistently reduced blood flow (-90%) in the vessels feeding into the ligated trees and elevated blood flow (+80%) in the nonligated mesenteric artery side branches. Four weeks after surgery, vessels that had been exposed to high blood flow (HF) or low blood flow (LF) and vessels from sham-operated rats (Sham) were isolated and mounted in a pressure myograph system. At an intraluminal pressure of 100 mmHg, the internal diameter at rest was larger in HF (533 ± 23 μm) and smaller in LF (262 ± 14 μm) than in Sham vessels (427 ± 15 μm). Also, wall and media cross-sectional areas were larger in HF and smaller in LF than in Sham vessels (media: 22 ± 1, 11 ± 2, and 16 ± 1 × 103μm2, respectively), but circumferential wall stress did not differ among groups. DNA content was significantly increased in HF vessels (+100%) and was not modified in LF vessels. Maximal vasoconstrictions elicited by high potassium or norepinephrine were slightly increased in HF vessels but were reduced by 50% in LF vessels. Thus chronic changes in blood flow give rise to structural changes that normalize circumferential wall stress. Elevated blood flow resulted in outward hypertrophic remodeling involving hyperplasia. Reduced blood flow resulted in inward hypotrophic remodeling accompanied by hyporeactivity of the arterial smooth muscle.

Uterine Artery Remodeling and Reproductive Performance Are Impaired in Endothelial Nitric Oxide Synthase-Deficient Mice

Abstract The progressive rise in uterine blood flow during pregnancy is accompanied by outward hypertrophic remodeling of the uterine artery (UA). This process involves changes of the arterial smooth muscle cells and extracellular matrix. Acute increases in blood flow stimulate endothelial production of nitric oxide (NO). It remains to be established whether endothelial NO synthase (eNOS) is involved in pregnancy-related arterial remodeling. We tested the hypothesis that absence of eNOS results in a reduced remodeling capacity of the UA during pregnancy leading to a decline in neonatal outcome. UA of nonpregnant and pregnant wild-type (Nos3+/+) and eNOS-deficient (Nos3−/−) mice were collected and processed for standard morphometrical analyses. In addition, cross sections of UA were processed for cytological (smoothelin, smooth muscle α-actin) and proliferation (Ki-67) immunostaining. We compared the pregnancy-related changes longitudinally and, together with the data on pregnancy outcome, transversally by analysis of variance with Bonferroni correction. During pregnancy, the increases in radius and medial cross sectional area of Nos3−/− UA was significantly less than those of Nos3+/+ UA. Smooth muscle cell dedifferentiation and proliferation were impaired in gravid Nos3−/− mice as deduced from the lack of change in the expression of smoothelin and smooth muscle α-actin, and the reduced Ki-67 expression. Until 17 days of gestation, litter size did not differ between both genotypes, but at birth the number of viable newborn pups and their weights were smaller in Nos3−/− than in Nos3+/+ mice. We conclude that absence of eNOS adversely affects UA remodeling in pregnancy, which may explain the impaired pregnancy outcome observed in these mice.

Hypertension Is Associated with Marked Alterations in Sphingolipid Biology: A Potential Role for Ceramide

Background Hypertension is, amongst others, characterized by endothelial dysfunction and vascular remodeling. As sphingolipids have been implicated in both the regulation of vascular contractility and growth, we investigated whether sphingolipid biology is altered in hypertension and whether this is reflected in altered vascular function. Methods and Findings In isolated carotid arteries from spontaneously hypertensive rats (SHR) and normotensive Wistar-Kyoto (WKY) rats, shifting the ceramide/S1P ratio towards ceramide dominance by administration of a sphingosine kinase inhibitor (dimethylsphingosine) or exogenous application of sphingomyelinase, induced marked endothelium-dependent contractions in SHR vessels (DMS: 1.4±0.4 and SMase: 2.1±0.1 mN/mm; n = 10), that were virtually absent in WKY vessels (DMS: 0.0±0.0 and SMase: 0.6±0.1 mN/mm; n = 9, p<0.05). Imaging mass spectrometry and immunohistochemistry indicated that these contractions were most likely mediated by ceramide and dependent on iPLA2, cyclooxygenase-1 and thromboxane synthase. Expression levels of these enzymes were higher in SHR vessels. In concurrence, infusion of dimethylsphingosine caused a marked rise in blood pressure in anesthetized SHR (42±4%; n = 7), but not in WKY (−12±10%; n = 6). Lipidomics analysis by mass spectrometry, revealed elevated levels of ceramide in arterial tissue of SHR compared to WKY (691±42 vs. 419±27 pmol, n = 3–5 respectively, p<0.05). These pronounced alterations in SHR sphingolipid biology are also reflected in increased plasma ceramide levels (513±19 pmol WKY vs. 645±25 pmol SHR, n = 6–12, p<0.05). Interestingly, we observed similar increases in ceramide levels (correlating with hypertension grade) in plasma from humans with essential hypertension (185±8 pmol vs. 252±23 pmol; n = 18 normotensive vs. n = 19 hypertensive patients, p<0.05). Conclusions Hypertension is associated with marked alterations in vascular sphingolipid biology such as elevated ceramide levels and signaling, that contribute to increased vascular tone.

Regional Heterogeneity of Arterial Structural Changes

Arterial structural changes in experimental models of hypertension and restenosis differ between vessel types and within vessels. Inspired by the diversity of short-term functional responses to vasoactive agents, hypotheses are presented with respect to the heterogeneity of structural alterations. Considered are the multifactorial nature of smooth muscle cell growth control and the possibility that vascular smooth muscle is not homogeneous but composed of different smooth muscle cell populations. These hypotheses may help explain the origin of both intervascular and intravascular heterogeneity of vascular structural responses.

Aortic Wall Properties in Normotensive and Hypertensive Rats of Various Ages In Vivo

The distensibility of the arterial system, which is partly determined by arterial wall structure, smooth muscle tone, and actual pressure level, decreases with aging and hypertension. Our aim was to compare aortic wall properties in 3- and 6-month-old normotensive Wistar-Kyoto rats (WKY) and spontaneously hypertensive rats (SHR) at comparable blood pressures in vivo. During ketamine/xylazine anesthesia in rats we performed ultrasound arterial wall tracking and invasive pressure measurements to determine, at the level of the thoracic aorta, diastolic pressure, diastolic lumen area, changes in pressure and lumen area during the cardiac cycle, and indexes of compliance and distensibility. These observations were combined with histological measurements for determination of media cross-sectional area and thickness and the incremental elastic modulus under conditions as expected in situ. Anesthesia abolished the difference in diastolic pressure between SHR and WKY. Between 3 and 6 months of age in WKY, diastolic area and incremental elastic modulus increased significantly, distensibility decreased, and all other recorded variables were not modified. Between 3 and 6 months of age in SHR, diastolic area and incremental elastic modulus increased, distensibility of the aortic wall decreased, and all other mechanical and structural properties did not change significantly. At both ages, diastolic area and compliance were significantly smaller in SHR than WKY. The other mechanical and structural properties measured or calculated at comparable pressure did not differ between strains. Differences between the aorta of 3- and 6-month-old rats and between strains observed in vivo at comparable pressures can largely be attributed to differences in lumen caliber.(ABSTRACT TRUNCATED AT 250 WORDS)

Chronic Moderate Hypoxia and Protein Malnutrition Both Induce Growth Retardation, But Have Distinct Effects on Arterial Endothelium-Dependent Reactivity in the Chicken Embryo

Deviations in the rate of intrauterine growth may change organ system development, resulting in cardiovascular disease in adult life. Arterial endothelial dysfunction often plays an important role in these diseases. The effects of two interventions that reduce fetal growth, chronic hypoxia and protein malnutrition, on arterial endothelial function were investigated. Eggs of White Leghorn chickens were incubated either in room air or in 15% O2 from d 6 until d 19 of the 21-d incubation. Protein malnutrition was induced by removal of 10% of the total albumen content at d 0. In vitro reactivity of the femoral artery in response to vasodilators was measured at d 19. Both chronic hypoxia and protein malnutrition reduced embryonic body weight at d 19 by 14% without affecting relative brain weight. Chronic hypoxia or protein malnutrition did not change sensitivity to the exogenous nitric oxide donor, sodium nitroprusside (5.74 ± 0.15 versus 5.85 ± 0.23 and 6.05 ± 0.18 versus 6.01 ± 0.34, respectively). Whereas protein malnutrition did not modify arterial sensitivity to acetylcholine (7.00 ± 0.10 versus 7.12 ± 0.05), chronic hypoxia reduced sensitivity to this endothelium-dependent vasodilator (6.57 ± 0.07 versus 7.02 ± 0.06). In the presence of Nω-nitro-l-arginine methyl ester, this difference in sensitivity to acetylcholine was no longer apparent (6.31 ± 0.13 versus 6.27 ± 0.06), indicating that chronic exposure to hypoxia reduced sensitivity to acetylcholine by lowering nitric oxide release. In additional experiments, a decrease in basal nitric oxide release in arteries of 3- to 4-wk-old chickens that had been exposed to in ovo chronic hypoxia was observed (increase in K+ contraction: −0.16 ± 0.33 N/m versus 0.68 ± 020 N/m). Protein malnutrition and chronic hypoxia both induce disproportionate growth retardation, but only the latter impairs arterial endothelial function. Intrauterine exposure to chronic hypoxia induces changes in arterial endothelial properties that may play a role in the development of cardiovascular disease in adult life.

Vasomotor responses in chronically hyperperfused and hypoperfused rat mesenteric arteries

We evaluated the reactivity of small arteries after remodeling induced by elevated or reduced blood flow. In 6-wk-old rats, every other first-order side branch of the superior mesenteric artery was ligated near the bifurcation of second-order branches. Four weeks after surgery, vessels that had been exposed to high flow (HF) or low flow (LF) were isolated and mounted in a pressure myograph at 100 mmHg and were compared with vessels from sham-operated rats (Sham). In HF: 1) basal lumen diameter was increased; 2) sensitivity to norepinephrine, arginine vasopressin, and perivascular nerve stimulation was not modified; 3) maximal constrictor responses (Δ diameter) to these stimuli and 125 mM K+ were increased; and 4) sensitivity and maximal dilator responses to sodium nitroprusside, acetylcholine, and flow were not modified. In LF: 1) basal diameter was reduced; 2) sensitivity to constrictor stimuli was not altered; 3) maximal responses to all vasoconstrictors except arginine vasopressin were reduced; and 4) sensitivity but not maximal dilator responses to sodium nitroprusside and acetylcholine was reduced. During acute flow-induced dilatations, lower shear stress was maintained in HF (48 ± 7 dyn/cm2) than in Sham (63 ± 10 dyn/cm2), but no shear stress regulation was observed in LF. These observations indicate that arterial structural responses to altered blood flow are accompanied by modified reactivity of the arterial smooth muscle, which entails changes in responsiveness to neurogenic and endothelium-dependent stimuli.We evaluated the reactivity of small arteries after remodeling induced by elevated or reduced blood flow. In 6-wk-old rats, every other first-order side branch of the superior mesenteric artery was ligated near the bifurcation of second-order branches. Four weeks after surgery, vessels that had been exposed to high flow (HF) or low flow (LF) were isolated and mounted in a pressure myograph at 100 mmHg and were compared with vessels from sham-operated rats (Sham). In HF: 1) basal lumen diameter was increased; 2) sensitivity to norepinephrine, arginine vasopressin, and perivascular nerve stimulation was not modified; 3) maximal constrictor responses (delta diameter) to these stimuli and 125 mM K+ were increased; and 4) sensitivity and maximal dilator responses to sodium nitroprusside, acetylcholine, and flow were not modified. In LF: 1) basal diameter was reduced; 2) sensitivity to constrictor stimuli was not altered; 3) maximal responses to all vasoconstrictors except arginine vasopressin were reduced; and 4) sensitivity but not maximal dilator responses to sodium nitroprusside and acetylcholine was reduced. During acute flow-induced dilatations, lower shear stress was maintained in HF (48 +/- 7 dyn/cm2) than in Sham (63 +/- 10 dyn/cm2), but no shear stress regulation was observed in LF. These observations indicate that arterial structural responses to altered blood flow are accompanied by modified reactivity of the arterial smooth muscle, which entails changes in responsiveness to neurogenic and endothelium-dependent stimuli.

Nitric oxide and potassium channels are involved in brain natriuretic peptide induced vasodilatation in man.

Objective Brain natriuretic peptide (BNP) causes vasodilatation but the mechanisms by which this is accomplished are not fully known. The aim of the present study was to determine whether, besides K+Ca2+-channels, nitric oxide (NO) is involved in BNP-induced vasodilatation. Methods We studied 10 healthy males twice, in random order, at an interval of 2 weeks. Experiments always started with infusion of BNP (8–16–32–64 pmol/dl per min) into the brachial artery. On the first day this infusion was followed by a second BNP infusion combined with the K+Ca2+-channel-blocker, tetraethylammonium (TEA, 0.1 mg/dl per min), and on the other day by BNP infusion combined with the NO-synthase inhibitor, l-NG-monomethyl arginine (l-NMMA, 0.8 μmol/dl per min). The latter was then followed by a combined infusion of BNP, l-NMMA and TEA. All infusions were separated by a 1 h washout period. Forearm blood flow (FBF) was determined by venous occlusion plethysmography. Results Mean arterial pressure and heart rate did not change during any of the experiments. BNP alone induced a dose-dependent dilatation, which was similar on both days. TEA, l-NMMA, and their combination all reduced the BNP-induced dilatation (P < 0.05). The combined infusion had a significantly greater effect than TEA alone (P = 0.005). BNP infusions were associated with a significant increase in plasma cyclic guanosine monophosphate (cGMP) and C-type natriuretic peptide (CNP) (P < 0.05). Conclusions BNP induces arterial vasodilatation not only by opening K+Ca2+-channels, but also via stimulation of NO production. In addition, BNP stimulates net CNP increase.

High Sodium Intake Increases Blood Pressure and Alters Renal Function in Intrauterine Growth–Retarded Rats

A suboptimal fetal environment increases the risk to develop cardiovascular disease in the adult. We reported previously that intrauterine stress in response to reduced uteroplacental blood flow in the pregnant rat limits fetal growth and compromises renal development, leading to an altered renal function in the adult offspring. Here we tested the hypothesis that high dietary sodium intake in rats with impaired renal development attributable to intrauterine stress, results in increased blood pressure, altered renal function, and organ damage. In rats, intrauterine stress was induced by bilateral ligation of the uterine arteries at day 17 of pregnancy. At the age of 12 weeks, the offspring was given high-sodium drinking water (2% sodium chloride). At the age of 16 weeks, rats were instrumented for monitoring of blood pressure and renal function. After intrauterine stress, litter size and birth weight were reduced, whereas hematocrit at birth was increased. Renal blood flow, glomerular filtration rate, and the glomerular filtration fraction were increased significantly after intrauterine stress. High sodium intake did not change renal function and blood pressure in control animals. However, during high sodium intake in intrauterine stress offspring, renal blood flow, glomerular filtration rate, and the filtration fraction were decreased, and blood pressure was increased. In addition, these animals developed severe albuminuria, an important sign of renal dysfunction. Thus, a suboptimal fetal microenvironment, which impairs renal development, results in sodium-dependent hypertension and albuminuria.

Chronic NG-Nitro-l-Arginine Methyl Ester Treatment Does Not Prevent Flow-Induced Remodeling in Mesenteric Feed Arteries and Arcading Arterioles

Although endothelium-derived NO is an important mediator in acute flow-induced changes in arterial tone, the role of NO in chronic flow-induced changes in the resistance artery and arteriolar structure remains largely unresolved. We investigated the effects of chronic inhibition of NO synthase on arterial and arteriolar remodeling in a rat mesenteric model in which flow changes were induced. Alternating first-order mesenteric arteries were ligated to shunt blood flow through the intermittent patent arteries. Animals received no treatment (NT) or a continuous infusion of NG-nitro-l-arginine methyl ester (L-NAME, 25 mg/kg SC per day). After 2 weeks, local in vivo blood flow and in vitro arterial pressure-diameter relationships were assessed, as were the in situ diameters of arcading arterioles. Medial cross-sectional areas (CSAs) were measured histologically. In both groups of animals, blood flow was significantly increased in patent arteries and decreased in ligated arteries compared with control vessels. Nonetheless, in L-NAME–treated rats, patent artery flow was increased to a lesser extent, although control flow was not significantly reduced (0.18±0.05 versus 0.26±0.05 mL/min). In NT rats, the diameter of patent arteries was significantly larger and the diameter of ligated arteries was significantly smaller than that of control arteries. CSAs displayed the same pattern of change (11.9±0.6×103, 6.1±0.7×103, and 8.2±1.0×103 &mgr;m2 for patent, ligated, and control arteries, respectively). Arterioles in the NT collateral pathway (218±15 &mgr;m) had diameters similar to control arteriole diameters (201±15 &mgr;m) but had a significantly larger CSA (6.2±0.6×103 versus 4.2±0.4×103 &mgr;m2). In L-NAME–treated rats, the flow-induced changes of the diameter and CSA in patent arteries, ligated arteries, and arcading arterioles mimicked those in NT rats. Nonetheless, control feed arteries (430±21 versus 497±16 &mgr;m) and arcading arterioles (156±21 &mgr;m) were significantly narrower after L-NAME treatment. Thus, chronic blockade of NO oxide synthase (1) tended to reduce arterial blood flow and resulted in inward remodeling of mesenteric arteries and arterioles and (2) did not prevent arterial and arteriolar remodeling in response to imposed changes in blood flow. Endothelium-derived mediators other than NO can play a major role in flow-induced arterial remodeling.