Livius V. d'Uscio

Angiotensin II Increases Tissue Endothelin and Induces Vascular Hypertrophy

BACKGROUNDnIn vitro studies on vascular smooth muscle cells suggest that endothelin has a stimulating effect on cellular proliferation. This study was designed to determine the endogenous effect of endothelin on angiotensin II-induced hypertrophy of small arteries in vivo.nnnMETHODS AND RESULTSnTwo weeks of angiotensin II administration (200 ng x kg[-1] x min[-1]) increased media thickness, media/lumen ratio, and cross-sectional area of basilar and small mesenteric arteries, confirming the proliferative properties of angiotensin II. The tissue levels of endothelin-1 were elevated in mesenteric arteries after angiotensin II administration. The administration of the selective and specific ET(A)-receptor antagonist LU135252 (50 mg x kg[-1] x d[-1]) in combination with angiotensin II prevented the changes of vascular geometry and partially reduced the increase in blood pressure induced by angiotensin II. Indeed, part of the effect on the vascular structure of the endothelin-receptor antagonist seemed pressure-independent.nnnCONCLUSIONSnOur results therefore demonstrate that angiotensin II increases the production of endothelin in the blood vessel wall that, via ET(A) receptors, mediates changes in vascular structure of the cerebral and mesenteric circulation. Endothelin antagonists may therefore be of value to reduce blood pressure and to prevent vascular structural changes in conditions of increased activity of the renin-angiotensin system.

ETA Receptor Blockade Prevents Increased Tissue Endothelin-1, Vascular Hypertrophy, and Endothelial Dysfunction in Salt-Sensitive Hypertension

Sodium plays an important role in the pathogenesis and therapy of hypertension, a major risk factor for cardiovascular disease. This study investigated the involvement of endothelin in vascular alterations in salt-induced Dahl hypertension. Salt-sensitive (DS) and salt-resistant (DR) Dahl rats were treated with a high-sodium diet (NaCl 4%) with or without ET A receptor antagonist LU135252 for two months, and effects of treatments on systolic blood pressure, vascular endothelin-1 (ET-1) protein content, aortic hypertrophy, and vascular reactivity of isolated aortic rings were studied. In DS rats, a high-sodium diet increased systolic pressure (190±4 versus 152±2 mm Hg, P P P P P =.0011). ET-1 tissue levels were highly and inversely correlated with endothelium-dependent relaxations ( r =0.931, P r =0.77, P =.0007). LU135252 treatment reduced systolic blood pressure only in part (168±3 versus 190±4 mm Hg. P P A receptor antagonism may have therapeutic potential for lowering vascular ET-1 content, improving endothelial function, and preventing structural changes in salt-sensitive hypertension.

Effects of Chronic ETA-Receptor Blockade in Angiotensin II-Induced Hypertension

Angiotensin II, a constrictor and mitogen of vascular smooth muscle cells, affects the release of endothelium-derived factors such as nitric oxide or endothelin-1. This study investigated the influence of endothelin-1, using the selective endothelin A receptor antagonist LU135252, on blood pressure and endothelial function in angiotensin II-induced hypertension in the rat. Two weeks of angiotensin II administration (200 ng/kg per minute) increased systolic blood pressure (+35 +/- 5 mm Hg; tail-cuff method) compared with placebo (P < .05). LU135252 alone did not affect systolic pressure but lowered the angiotensin II-induced pressure increase (P < .05). In isolated aortic rings, endothelium-dependent relaxations to acetylcholine were reduced in the angiotensin II group (P < .05 versus placebo) and improved by concomitant chronic LU135252 treatment (P < .05 versus angiotensin II). Blood pressure elevation strongly correlated with impaired endothelium-dependent relaxations to acetylcholine (r = -.967). LU135252 did not affect endothelium-independent relaxations to sodium nitroprusside, which were diminished after angiotensin II treatment (P < .05). In quiescent rings, chronic angiotensin II administration enhanced endothelium-dependent contractions to acetylcholine, which were reduced by LU135252 (P < .05). Impaired contractions to endothelin-1 and norepinephrine in the angiotensin II group were normalized after treatment with LU135252 (P < .05). Thus, chronic therapy with LU135252 partially prevents angiotensin II-induced hypertension and the alternations of the endothelial function observed in this experimental model.

Obesity Is Associated With Tissue-Specific Activation of Renal Angiotensin-Converting Enzyme In Vivo: Evidence for a Regulatory Role of Endothelin

In the C57BL/6J mice model, we investigated whether obesity affects the function or expression of components of the tissue renin-angiotensin system and whether endothelin (ET)-1 contributes to these changes. ACE activity (nmol. L His-Leu. mg protein(-1)) was measured in lung, kidney, and liver in control (receiving standard chow) and obese animals treated for 30 weeks with a high-fat, low cholesterol diet alone or in combination with LU135252, an orally active ET(A) receptor antagonist. ACE mRNA expression was measured in the kidney, and the effects of LU135252 on purified human ACE were determined. Aortic and renal tissue ET-1 protein content was measured, and the vascular contractility to angiotensin II was assessed. Obesity was associated with a tissue-specific increase in ACE activity in the kidney (55+/-4 versus 33+/-3 nmol/L) but not in the lung (34+/-2 versus 32+/-2 nmol/L). Long-term LU135252 treatment completely prevented this activation (13.3+/-0.3 versus 55+/-4 nmol/L, P<0.05) independent of ACE mRNA expression, body weight, or renal ET-1 protein but did not affect pulmonary or hepatic ACE activity. Obesity potentiated contractions in response to angiotensin II in the aorta (from 6+/-2% to 33+/-5% KCl) but not in the carotid artery (4+/-1% to 3.6+/-1% KCl), an effect that was completely prevented with LU135252 treatment (6+/-0.4% versus 33+/-5% KCl). No effect of LU135252 on purified ACE was observed. Thus, obesity is associated with the activation of renal ACE in vivo independent of its mRNA expression and enhanced vascular contractility to angiotensin II. These effects are regulated by ET in an organ-specific manner, providing novel mechanisms by which ET antagonists may exert organ protection.

Structure and Function of Small Arteries in Salt-Induced Hypertension: Effects of Chronic Endothelin-Subtype-A–Receptor Blockade

The involvement of endothelin in salt-induced hypertension is unclear. In the Dahl rat model, we studied the effects of a selective endothelin-subtype A (ET[A]) receptor antagonist, LU135252, on blood pressure, vascular structure, and function. Dahl salt-sensitive and salt-resistant rats were treated for 8 weeks with 4% NaCl alone or in combination with LU135252 taken orally (60 mg/kg per day). The geometry and reactivity of basilar and mesenteric arteries were studied in vitro under perfused and pressurized conditions using a video dimension analyzer. Chronic salt administration increased systolic blood pressure by 37 +/- 3 mm Hg and media-lumen ratio of the basilar and mesenteric arteries in salt-sensitive rats (P<.05). These structural changes were caused by eutrophic remodeling in basilar and hypertrophic remodeling in mesenteric arteries. Endothelium-dependent relaxations to acetylcholine and contractions to endothelin-1 were impaired in mesenteric arteries of salt-sensitive rats on a high NaCl diet. LU135252 prevented part of the increase in systolic blood pressure and structural and functional alterations but increased plasma endothelin 1 levels (P<.05 versus salt-treated, saltsensitive rats). LU135252 had no effect on these parameters in salt-resistant rats. These findings suggest that the long-term pressor effect of salt administration is mediated in part by the action of endogenous endothelin acting via ET(A) receptors. Thus, chronic ET(A) receptor blockade may be useful therapeutically to lower arterial pressure and prevent endothelial dysfunction and hypertrophic remodeling of resistance arteries in salt-sensitive forms of hypertension.

Losartan but Not Verapamil Inhibits Angiotensin II–Induced Tissue Endothelin-1 Increase: Role of Blood Pressure and Endothelial Function

Endothelin partially mediates angiotensin (Ang) II-induced vascular changes in vivo. This study investigated the effects of the angiotensin type 1 receptor antagonist losartan and the calcium channel blocker verapamil on vascular reactivity and tissue endothelin-1 levels in aortas of Wistar-Kyoto rats treated for 2 weeks with Ang II (200 ng x kg(-1) x min(-1)). Ang II increased systolic blood pressure (39+/-4 mm Hg, P<0.05). Concomitant treatment with losartan abolished the Ang II-induced pressure increase (P<0.05), whereas verapamil reduced it only partially (P<0.05). In the aortas of rats with Ang II-induced hypertension, tissue endothelin-1 content was increased threefold and contractions to endothelin-1 were impaired (P<0.05). Interestingly, these alterations were normalized by losartan (P<0.05) but not by verapamil. Hence, there was a strong, negative correlation between contractions to endothelin-1 and tissue endothelin-1 content (r=-0.733, P<0.0001). In contrast, both antihypertensive drugs normalized impaired endothelium-dependent relaxations to acetylcholine and reduced the sensitivity of vascular smooth muscle to sodium nitroprusside compared with Ang II-treated rats (P<0.05). Ang II-induced hypertension enhanced endothelium-dependent contractions to acetylcholine, and these were normalized by either drug. In conclusion, these findings suggest that long-term treatment with Ang II modulates endothelin-1 protein expression in the rat aorta. Although both antihypertensive agents lowered blood pressure and normalized endothelial function, only losartan prevented the increase in tissue endothelin-1 content, suggesting that angiotensin type 1 receptor antagonists but not calcium antagonists modulate tissue endothelin-1 in vivo.

Structure and reactivity of small arteries in aging

OBJECTIVEnIncreased pulse pressure has been observed in aging subjects, but the impact on the structure and reactivity of small arteries has been scarcely evaluated.nnnMETHODSnThis study presents the modifications of vascular structure and function observed in female rats of 5, 18 and 32 months of age, and their relation to the prevailing hemodynamic status. Geometry and reactivity of perfused and pressurized basilar and mesenteric small arteries were analyzed in vitro using a video dimension analyzer.nnnRESULTSnMean arterial pressure was similar in the three age groups, and only pulse pressure was increased in the oldest group. Media thickness and cross sectional area increased in basilar and mesenteric arteries of the oldest rats and these structural abnormalities were positively related to pulse pressure but not to mean, systolic or diastolic arterial pressure. Only minor changes of vascular reactivity were noted with age: there was a decreased contraction to angiotensin II in mesenteric arteries and an enhanced contraction to endothelin-1 in the basilar arteries.nnnCONCLUSIONnIn conclusion, aging is associated with increased pulse pressure and hypertrophy of basilar and mesenteric resistance arteries, suggesting that this hemodynamic variable may influence cerebral and peripheral vascular structure in aging.

Endothelium-independent relaxation and hyperpolarization to C-type natriuretic peptide in porcine coronary arteries

Endothelial cells produce C-type natriuretic peptide (CNP), which has been proposed as an endothelium-derived hyperpolarizing factor. In porcine coronary arteries, we investigated the vasodilatory effects of CNP and compared them with endothelium-dependent relaxations and hyperpolarizations to bradykinin. Isolated epicardial porcine coronary arteries were studied in organ chambers, and concentration-response curves to CNP and bradykinin were obtained. Membrane potential was measured in endothelial cells and smooth muscle of intact porcine coronary arteries during stimulation with CNP or bradykinin. In precontracted porcine coronary arteries with or without endothelium, CNP (10[-10]-10[-6] M) evoked relaxations (maximum, 42 +/- 4%) smaller than those evoked by bradykinin (100 +/- 1%), blunted in preparations contracted by KCl instead of U46619 (9,11-dideoxy-11a,9a-epoxymethano-prostaglandin F2alpha; p < 0.05) and unaffected by inhibition of NO synthase (NS). CNP evoked hyperpolarization of vascular smooth muscle of similar magnitude in endothelium-intact (-4.4 +/- 1 mV) and endothelium-denuded (-4.6 +/- 1 mV) porcine coronary arteries. Bradykinin (10[-10]-10[-6] M) evoked concentration-dependent relaxations in preparations with endothelium only. Although atrial natriuretic peptide-receptor antagonist HS-142-1 (25 microM) slightly reduced the sensitivity to bradykinin (log shift at IC50, twofold; p < 0.05), it had no effect on the maximal response to bradykinin. Inhibition of NO synthase partially attenuated, whereas high potassium chloride (30 mM) markedly inhibited relaxations to bradykinin (p < 0.05). Hyperpolarization to bradykinin was much more pronounced than that to CNP (-17 +/- 3 mV; p < 0.05 vs. CNP) and was observed in endothelium-intact preparations only and unaffected by HS-142-1. In conclusion, in contrast to bradykinin, CNP induces endothelium-independent and weaker relaxation and hyperpolarization of coronary artery vascular smooth muscle, suggesting that CNP is an unlikely mediator of endothelium-dependent hyperpolarization of porcine coronary arteries.

Vasopeptidase Inhibition Exhibits Endothelial Protection in Salt-Induced Hypertension

Omapatrilat represents a new class of drugs capable of inhibiting both ACE and neutral endopeptidase 24.11, the so-called vasopeptidase inhibitors. It therefore contributes to neurohumoral modulation, which might improve endothelial function in cardiovascular diseases. This study investigated the effect of omapatrilat in comparison to the ACE inhibitor captopril on systolic blood pressure and endothelial function in salt-induced hypertension. Dahl salt-sensitive rats (n=6/group) on standard or salt-enriched (4% NaCl) chow were treated for 8 weeks with either omapatrilat (36±4 mg/kg per day), captopril (94±2 mg/kg per day), or placebo. Aortic rings were then isolated and suspended in organ chambers for isometric tension recording. Systolic blood pressure of salt-fed, placebo-treated animals increased to 196±6 mm Hg, which was prevented by omapatrilat (162±5 mm Hg, P <0.05) and captopril (164±7 mm Hg, P <0.05) to a comparable degree. In control rats, acetylcholine (10−10 to 10−5 mol/L) induced endothelium-dependent relaxation (97±4%), which was reduced by high-salt diet to 30±5% (P <0.005; n=6). Omapatrilat improved relaxation to a greater extent (86±5%) than did captopril (57±6%;P <0.05). eNOS protein expression and aortic nitrite/nitrate content were reduced in hypertensive rats and improved by both omapatrilat and captopril. Aortic endothelin-1 levels were increased in salt-fed animals and unaffected by omapatrilat or captopril. These data suggest that despite comparable blood pressure, omapatrilat is superior to captopril in improving endothelium-dependent relaxation in salt-sensitive hypertension.

Chronic ETA receptor blockade prevents endothelial dysfunction of small arteries in apolipoprotein E-deficient mice

Objective: This study investigated whether endothelial dysfunction occurs in mesenteric arteries of apoE-deficient mice and determined the role of endothelin (ET)-1, which is increased in human atherosclerosis, using an orally active endothelin ETA receptor antagonist. Methods: ApoE-deficient and C57BL/6J control mice were fed for 30 weeks with normal chow or high-fat Western-type diet alone or in combination with darusentan (LU135252; 50 mg/kg/day). Vasomotor reactivity of isolated small mesenteric arteries (I.D. 200–250 μm) was studied in vitro under perfused and pressurized conditions. Results: In both mouse strains, about one fourth of the endothelium-dependent relaxant response to acetylcholine was insensitive to inhibition by l-NAME and indomethacin. In mesenteric arteries of apoE-deficient mice on Western-type diet, increased intima-media thickness and levels of endothelin-1 protein were observed. In addition, NO-mediated endothelium-dependent relaxation to acetylcholine was reduced without affecting l-NAME/indomethacin insensitive relaxation and contractions to endothelin-1 and serotonin were enhanced. Treatment with darusentan normalized vascular structure, NO-mediated relaxation to acetylcholine and contractions to endothelin-1 and serotonin without affecting blood pressure or plasma cholesterol levels. Conclusions: Severe hypercholesterolemia in apoE-deficient mice is associated with attenuation of NO-mediated relaxation to acetylcholine and increased vascular endothelin-1 content. Chronic ETA receptor blockade may provide a new therapeutic approach to improve NO-mediated endothelium-dependent vasomotion in small arteries.