Obesity and diabetes in humans are associated with hypertrophic remodeling and increased media:lumen ratio of small resistance arteries, which is an independent predictor of cardiovascular events. In order to minimize increases in media:lumen ratio, hypertrophic remodeling should be accompanied by outward remodeling. We aimed to investigate the mechanisms of structural remodeling in small pial arteries (PAs) and terminal mesenteric arteries (TMAs) from obese Göttingen Minipigs with or without diabetes. Göttingen Minipigs received either control diet (lean control (LC)), high fat/high fructose/high cholesterol diet (FFC), or FFC diet with streptozotocin (STZ)-induced diabetes (FFC/STZ) for 13 months. At the end of the study (20 months), we assessed body weight, fasting plasma biochemistry, passive vessel dimensions, mRNA expression (matrix metallopeptidases 2/9 ( MMP2, MMP9 ), tissue inhibitor of metallopeptidase 1 ( TIMP1 ), transglutaminase 2 ( TGM2 ), Rho-kinase 1 ( ROCK1 ), TGFβ-receptor 2 ( TGFBR2 ), and IGF1-receptor ( IGFR1 ) genes), and immunofluorescence in PAs and TMAs. We performed multiple linear correlation analyses using plasma values, structural data, and gene expression data. We detected outward hypertrophic remodeling in TMAs and hypertrophic remodeling in PAs from FFC/STZ animals. ROCK1 and TGM2 genes were up-regulated in PAs and TMAs from the FFC/STZ group. Passive lumen diameter (PLD) of TMAs was correlated with plasma values of glucose (GLU), fructosamine (FRA), total cholesterol (TC), and triglycerides (TGs). ROCK1 and TGM2 expressions in TMAs were correlated with PLD, plasma GLU, fructosamine, and TC. ROCK1 and TGM2 proteins were immunolocalized in the media of PAs and TMAs, and their fluorescence levels were increased in the FFC/STZ group. Hyperglycemia/hyperlipidemia is involved in regulation of ROCK1 and TGM2 expression leading to outward remodeling of small resistance arteries in obese diabetic Göttingen Minipigs.

Endothelial dysfunction is a common problem associated with hypertension and is considered a precursor to the development of micro- and macro-vascular complications. The present study investigated the involvement of nNOS (neuronal nitric oxide synthase) and H 2 O 2 (hydrogen peroxide) in the impaired endothelium-dependent vasodilation of the mesenteric arteries of DOCA (deoxycorticosterone acetate)-salt-hypertensive mice. Myograph studies were used to investigate the endothelium-dependent vasodilator effect of ACh (acetylcholine). The expression and phosphorylation of nNOS and eNOS (endothelial nitric oxide synthase) were studied by Western blot analysis. Immunofluorescence was used to examine the localization of nNOS and eNOS in the endothelial layer of the mesenteric artery. The vasodilator effect of ACh is strongly impaired in mesenteric arteries of DOCA-salt-hypertensive mice. Non-selective inhibition of NOS sharply reduced the effect of ACh in both DOCA-salt-hypertensive and sham mice. Selective inhibition of nNOS and catalase led to a higher reduction in the effect of ACh in sham than in DOCA-salt-hypertensive mice. Production of H 2 O 2 induced by ACh was significantly reduced in vessels from DOCA-salt-hypertensive mice, and it was blunted after nNOS inhibition. The expression of both eNOS and nNOS was considerably lower in DOCA-salt-hypertensive mice, whereas phosphorylation of their inhibitory sites was increased. The presence of nNOS was confirmed in the endothelial layer of mesenteric arteries from both sham and DOCA-salt-hypertensive mice. These results demonstrate that endothelial dysfunction in the mesenteric arteries of DOCA-salt-hypertensive mice is associated with reduced expression and functioning of nNOS and impaired production of nNOS-derived H 2 O 2 . Such findings offer a new perspective for the understanding of endothelial dysfunction in hypertension.

In the present in vivo study, we have investigated whether inhibitors of the Na + /Mg 2+ exchanger quinidine and imipramine influence the development of hypertension and whether this is associated with modulation of mitogen-activated protein (MAP) kinase activation in arteries and kidneys of hypertensive rats. Sprague—Dawley rats were divided into four groups ( n =6/group): control (vehicle), angiotensin II (Ang II; 150 ng/kg of body weight per min subcutaneously), quinidine [Ang II (150 ng/kg of body weight per min)+quinidine (5 mg/kg of body weight per day in food)] and imipramine groups [Ang II (150 ng/kg of body weight per min)+imipramine (5 mg/kg/day in food)]. Rats were studied for 3 weeks. Phosphorylation of vascular and renal extracellular-signal-regulated protein kinase 1/2 (ERK1/2), p38MAP kinase and c-Jun N-terminal kinase (JNK) were assessed using phospho-specific antibodies. Ang II increased systolic blood pressure from 112±5 mmHg to 215±9 mmHg ( P <0.01). Development of hypertension was attenuated in Ang II-infused rats treated with quinidine (173±6 mmHg) and imipramine (152±6 mmHg) ( P <0.01). Phosphorylation of ERK1/2, p38MAP kinase and JNK, which were increased 2–3-fold in arteries of the Ang II group, were reduced by quinidine and imipramine ( P <0.05). Activation of renal MAP kinases was also increased in the Ang II group ( P <0.05). Quinidine and imipramine reduced the phosphorylation of renal ERK1/2, but did not modify renal p38MAP kinase or JNK. Our data demonstrate that Ang II induces severe hypertension in Sprague—Dawley rats and this is associated with increased phosphorylation of vascular and renal MAP kinases. Quinidine and imipramine attenuated the development of hypertension and normalized MAP kinase activity. The findings from this study suggest a possible role for the Na + /Mg 2+ exchanger in vascular signalling events associated with blood pressure elevation in Ang II-dependent hypertension.

Endothelin-1 (ET-1) and adrenomedullin (ADM) are both produced in the arterial wall, but have opposing biological actions. Evidence from experimental animals suggests a functional interaction between ET-1 and ADM. We have tested this in humans. Small resistance arteries were obtained from gluteal biopsies taken from patients with chronic heart failure (CHF) due to coronary heart disease (CHD), or with CHD and preserved ventricular function. The contractile responses to big ET-1 and to ET-1 in both sets of vessels were studied in the absence (control) and presence of ADM at 20 pmol/l (low ADM) or 200 pmol/l (high ADM), using wire myography. ADM did not affect the conversion of big ET-1 into ET-1 in vessels from patients with either CHD or CHF. Low ADM did not alter the contractile response to ET-1 in vessels from patients with CHF. Low ADM was not tested in vessels from patients with CHD, but high ADM did not affect this response in arteries from these patients. High ADM did, however, significantly reduce the vasoconstrictor effect of ET-1 in vessels from patients with CHF. The maximum response, as a percentage of the response to high potassium, was 199% (S.E.M. 25%) in the control experiments ( n = 14), 205% (27%) in the low-ADM ( n = 7) studies and 150% (17%) in the high-ADM ( n = 6) experiments ( P < 0.001). Furthermore, the Hill coefficient increased from 0.57±0.05 in the absence of ADM to 1.16±0.15 in the high-ADM experiments, indicating that ADM at 200 pmol/l specifically antagonized one receptor type in vessels from patients with CHF. We conclude that there is a one-site receptor interaction between ADM and ET-1 that is specific for vessels from patients with CHF. This functional interaction between ADM and ET-1 in resistance arteries may be of pathophysiological importance in CHF.

Resveratrol has been shown to induce vasorelaxation. In this study, we investigated the mechanism(s) of resveratrol-induced vasorelaxation in resistance mesenteric arteries from male lean and dietary-induced obese rats. Compared with lean rats, arteries from dietary-obese rats showed significant ( P < 0.001) endothelial dysfunction, as indicated by a decrease (> 20%) in maximal acetylcholine-induced vasorelaxation. Resveratrol (5–35 µ mol/l) induced concentration-dependent relaxation of mesenteric arteries preconstricted with noradrenaline (8 µ mol/l) or KCl (125 mmol/l) from both lean and dietary-obese rats. There were no significant differences between the two groups, achieving a maximum relaxation of > 95% at a concentration of 35 µ mol/l. In noradrenaline-preconstricted arteries from lean rats, N G -nitro- L -arginine methyl ester ( L -NAME; 100 and 300 µ mol/l) caused a significant ( P < 0.01) concentration-dependent rightward shift in reseveratrol activity, with no effect on maximal responses. However, L -NAME (100 and 300 µ mol/l) did not alter the effects of reseveratrol on arteries from dietary-obese rats, giving superimposed concentration–responses curves. Indomethacin was also ineffective in altering resveratrol activity in arteries from both lean and dietary-obese rats. In noradrenaline-precontracted arteries from dietary-obese rats, responses to resveratrol were not attenuated by endothelial denudation, indicating an action independent of the endothelium. This study indicates that: (a) the maximal effects of resveratrol on resistance arteries from lean and dietary-obese rats are not effected by endothelial dysfunction, and (b) the effects of resveratrol in lean animals (where endothelial function is not impaired), but not in dietary-obese rats, are mediated via NO.

Naturally occurring hydroxystilbenes have been shown to induce vasorelaxation. Here, we studied the mechanism of resveratrol-induced vasorelaxation in different types of blood vessels, namely mesenteric (resistance) and main uterine (conductance) arteries, from female guinea-pigs on day 7 and day 15 of the oestrous cycle. Resveratrol (5–70 µ mol/l) induced concentration-dependent relaxation of both mesenteric and uterine arteries preconstricted with either noradrenaline (NA; 10 µ mol/l) or KCl (125 mmol/l). Resveratrol was 2-fold more potent in inducing relaxation of mesenteric arteries than of uterine arteries. Its effects on uterine arteries from both day-7 and day-15 guinea-pigs were similar, irrespective of the constrictor used, but it was significantly ( P < 0.01) more potent in inducing relaxation of mesenteric arteries contracted with NA compared with those constricted with KCl. In day-7 arteries precontracted with NA, N G -nitro- l -arginine methyl ester ( l -NAME; 10 µ mol/l) had no effects on the time course of resveratrol-induced vasorelaxation in either mesenteric or uterine arteries. However, indomethacin (50 µ mol/l) significantly ( P < 0.05) potentiated resveratrol's effect on mesenteric, but not uterine, arteries. Indomethacin had no effect on resveratrol-induced vasorelaxation of arteries contracted with KCl, whereas l -NAME significantly ( P < 0.05) reduced the effects of resveratrol on uterine, but not on mesenteric, arteries. In day-15 arteries, l -NAME significantly ( P < 0.01) attenuated the effects of resveratrol on mesenteric arteries contracted with NA. Indomethacin had no effect on resveratrol activity. This study indicates that: (a) the effect of resveratrol on resistance arteries is greater than that on conductance arteries; (b) the effects of resveratrol are not mediated via prostanoids, but NO may play a role; and (c) the stage of the oestrous cycle has no influence on resveratrol-induced vasorelaxation.

1. Previous studies have indicated that younger hypertensive subjects may have abnormal endothelium-dependent relaxation, which could contribute to the elevated peripheral resistance seen in established hypertension. This study was designed to examine the functional behaviour of the endothelium of small arteries from elderly hypertensive and normotensive subjects. 2. Resistance arteries were obtained from gluteal biopsies taken under local anaesthesia in 28 subjects of mean age 70 (range 60–76) years, and studied in an isometric myograph. Eighteen subjects had untreated essential hypertension, and 10 were normotensive. 3. After measurement of the contractile response to noradrenaline, relaxation responses to a variety of endothelium-dependent (acetylcholine and bradykinin) and endothelium-independent (iloprost and sodium nitroprusside) mechanisms were assessed in vessels precontracted with noradrenaline. Endothelium-dependent responses were also studied after incubation with N G -nitro- l -arginine to inhibit nitric oxide synthase. 4. There were no significant differences in the contraction or relaxation responses between elderly subjects with or without high blood pressure. Inhibition of nitric oxide synthase prevented any relaxation with acetylcholine and significantly attenuated the relaxation with bradykinin. Near-complete relaxation was however achieved with the endothelium-independent vasodilator sodium nitroprusside. 5. Hypertension in elderly subjects is not associated with a reduction in endothelial vasodilating function in the subcutaneous vessels of the gluteal region compared with age-matched normotensive controls. The results of this study do not support the hypothesis of a defect of resistance artery endothelium-dependent relaxation in the pathophysiology of hypertension in the elderly.

1. Impaired endothelium-dependent relaxation has been previously demonstrated in blood vessels of hypertensive rats and in humans with essential hypertension. Arteries from spontaneously hypertensive rats have been shown to produce, in response to high concentrations of acetylcholine, a vasoconstrictor substance called endothelium-derived contracting factor, the production of which can be inhibited by indomethacin or other cyclo-oxygenase inhibitors, suggesting that it is a prostanoid. The mechanisms involved in endothelium-dependent relaxation of human arteries are unclear, and the potential generation of endothelium-derived contracting factor by endothelium in human hypertension has not been established. 2. We investigated the effects of acetylcholine on precontracted small arteries dissected from gluteal subcutaneous fat biopsies from normotensive subjects and subjects with borderline and mild essential hypertension. Vessels from normotensive subjects and those from borderline hypertensive patients, precontracted by noradrenaline, were relaxed completely by acetylcholine, whereas those from patients with mild essential hypertension relaxed slightly but significantly less, indicating that generation of endothelium-derived relaxing factor (endothelium-derived nitric oxide) was only minimally reduced or that production of minor amounts of endothelium-derived contracting factor occurred in small arteries from these hypertensive subjects. This impairment of endothelium-dependent relaxation was not corrected by indomethacin, which indicated that the contribution of endothelium-derived contracting factor, if any, was minimal in this subset of essential hypertensive patients. In contrast, mesenteric small arteries of adult spontaneously hypertensive rats presented strong contractions in response to the higher concentrations of acetylcholine, which were abolished by exposure to indomethacin. 3. The relaxation induced by acetylcholine in arteries from both hypertensive and normotensive humans was partially blunted (by 30%) by pretreatment with 0.1 mmol/l N G -nitro-l-arginine methyl ester or N G -nitro-monomethyl-l-arginine (inhibitors of nitric oxide synthase) and by 10 μmol/l Methylene Blue (a blocker of soluble guanylate cyclase), indicating the role of endothelium-derived nitric oxide and the generation of its intracellular second messenger cyclic guanosine monophosphate in acetylcholine-induced relaxation. The remaining relaxation elicited by acetylcholine could be blocked with 30 mmol/l KCl or with 10 μmol/l ouabain (inhibitor of Na + , K + -ATPase), and, when combined with N G -nitro-l-arginine methyl ester, these interventions abolished acetylcholine-induced relaxation. Tolbutamide at 2 mmol/l or 10 μmol/l glyburide (blockers of ATP-sensitive potassium channels) partially inhibited N G -nitro-l-arginine methyl ester-resistant endothelium-dependent relaxation. Apamin (a blocker of small-conductance calcium-activated potassium channels), which has been shown to block N G -nitro-l-arginine methyl ester-resistant endothelium-dependent relaxation in rat arteries, was without effect. Charybdotoxin (blocker of large-conductance calcium-activated potassium channels) displaced to the right the responses to acetylcholine, in the absence and presence of N G -nitro-l-arginine methyl ester and/or tolbutamide. 4. In conclusion, in contrast to mesenteric small arteries from spontaneously hypertensive rats, which produce endothelium-derived contracting factor, subcutaneous small arteries from subjects with mild essential hypertension appear not to do so in significant amounts. Thirty per cent of the relaxation induced by acetylcholine in human small arteries is mediated by release of endothelium-derived nitric oxide, whereas more than 60% is the result of release of another agent, perhaps the putative endothelium-derived hyperpolarizing factor, which may act in part by opening ATP-sensitive potassium channels and large-conductance calcium-activated potassium channels. Endothelium-dependent relaxation induced by acetylcholine was normal in borderline hypertensive subjects and only slightly abnormal in the mildly hypertensive patients studied.

1. Using a Mulvany-Halpern myograph to measure changes in isometric tension, we have investigated the effect of ouabain on noradrenaline-induced contraction of human subcutaneous resistance arteries. 2. Low concentrations of ouabain (10 nmol/l or less) were shown not to alter vascular smooth muscle contractility or sensitivity to noradrenaline. 3. In contrast, higher concentrations of ouabain (100 nmol/l or more) were found to depress vascular smooth muscle contractility and to reduce the sensitivity of the noradrenaline concentration-response relationship. 4. These findings may have implications regarding the presence of an endogenous inhibitor of the sodium pump in essential hypertension and in pregnancy-associated hypertension.

1. In order to examine the effects of experimental hypertension on intracellular pH in mesenteric resistance arteries, intracellular pH was measured in mesenteric resistance arteries from rats with coarctation 72 h, 9 days and 28 days after the aorta was partially constricted between the origins of the renal arteries. Carotid arterial pressure was significantly raised at all time points. 2. Second-order mesenteric resistance arteries were mounted in a myograph and were loaded with the acetoxymethyl ester of the pH-sensitive dye 2′,7′-bis(carboxyethyl)-5,6-carboxyfluorescein. Morphological measurements demonstrated that arteries from rats with coarctation had an increased media volume at 9 days and at 28 days compared with vessels from sham-operated control animals, but this was only statistically significant at 28 days. 3. Resting intracellular pH was not significantly different at any time point in arteries from rats with coarctation compared with control animals, although there was a rise in intracellular pH in both groups of rats between 72 h and 9 days. The application of 4,4-di-isothiocyanatostilbene-2,2′-disulphonic acid produced a fall in intracellular pH which was significantly greater in the sham-operated rats at 9 days; this difference was not found at 28 days. Blockade of Na + /H + exchange with 60 μmol/l ethylisopropylamiloride led to a similar fall in intracellular pH in both groups of rats at 9 days but a significantly greater fall in intracellular pH in arteries of rats with coarctation at 28 days. Activation with noradrenaline (10 μmol/l) induced acid changes in intracellular pH that were similar in both groups of rats. 4. It is concluded that induction of coarctation of the aorta was associated with the development of significant growth in mesenteric resistance arteries, but that this structural change was not associated with an intracellular pH that was significantly different from that seen in sham-operated control rats at any time. Increased regulation of intracellular pH by Na + /H + exchange was observed only at 28 days, after medial growth had occurred, which argues against a fundamental role of this exchange system as a mediator of cell growth.

1. Using a myograph to measure isometric tension, we have looked at the action of N G -monomethyl-l-arginine on the endothelium-dependent relaxation of human subcutaneous resistance arteries. 2. N G -Monomethyl-l-arginine, the novel inhibitor of endothelium-derived relaxing factor synthesis, caused concentration-dependent but only partial inhibition of maximal relaxation induced by acetylcholine in human subcutaneous resistance arteries. 3. The inhibitory action of N G -monomethyl-l-arginine on acetylcholine-induced maximal relaxation was partially reversed by incubation of the arteries in equimolar concentrations of l-arginine and N G -monomethyl-l-arginine. Subsequent incubation in l-arginine led to further reversal, but this was no greater than with incubation in physiological saline. 4. A component of acetylcholine-induced relaxation was sensitive to indomethacin, suggesting that this response is mediated by prostanoids as well as by endothelium-derived relaxing factor. 5. N G -Monomethyl-l-arginine did not increase the tension of resting human subcutaneous resistance arteries. N G -Monomethyl-l-arginine did enhance the contractile response to noradrenaline, possibly due to inhibition of release of endothelium-derived relaxing factor resulting from stimulation of α 2 -adrenoreceptors on the endothelial cells.