Free-radical scavengers, thiol-containing reagents and endothelium-dependent relaxation in isolated rat and human resistance arteries

1. Small arteries were isolated from either rat mesentery or human subcutaneous fat, and mounted in a myograph for the measurement of isometric force. 2. Superoxide dismutase, either in the presence or absence of catalase, relaxed noradrenaline-induced tone. This effect was abolished by removal of the endothelium or incubation with an inhibitor of NO synthase, N-omega-nitro-L-arginine methyl ester. Catalase alone had a negligible effect on noradrenaline-induced tone. 3. Captopril, an angiotensin-converting enzyme inhibitor and putative free-radical scavenger, did not relax pre-contracted isolated vessels. N-Acetylcysteine caused an endothelium-independent relaxation of rat vessels. Similar effects were observed in human vessels. 4. Acetylcholine induced a concentration-dependent relaxation of isolated resistance arteries, which was inhibited by removal of the endothelium or N-omega-nitro-L-arginine methyl ester, but unaffected by indomethacin. Preincubation with captopril, N-acetylcysteine or catalase alone did not alter the acetylcholine concentration-response relationship, but superoxide dismutase in combination with catalase enhanced responses to acetylcholine, causing a six-fold increase in potency. 5. Superoxide dismutase causes endothelium-dependent relaxation of resistance arteries and potentiates responses to acetylcholine. This action is probably due to the ability of the enzyme to scavenge superoxide anions which inhibit endothelium-dependent relaxation. 6. N-Acetylcysteine causes an endothelium-independent relaxation of resistance arteries which is probably unrelated to the putative ability of this compound to scavenge superoxide radicals and may reflect a direct action on vascular smooth muscle.     

Impaired endothelium-dependent relaxation in isolated resistance arteries of spontaneously diabetic rats

1. Previous studies have shown that endothelium-dependent relaxation in the aorta of spontaneously diabetic bio bred rats (BB) is impaired. 2. We have investigated noradrenaline (NA) contractility, endothelium-dependent acetylcholine (ACh) and bradykinin (BK) relaxation, and endothelium-independent sodium nitroprusside (SNP) relaxation in mesenteric resistance arteries of recent onset BB rats and established insulin treated BB rats, compared to their age-matched non diabetic controls. 3. There was no significant difference in the maximum contractile response or sensitivity to noradrenaline in either of the diabetic groups compared to their age-matched controls. 4. Incubation with the nitric oxide synthetase inhibitor NG-nitro-L-arginine (L-NOARG) resulted in a significant increase in maximum contractile response to noradrenaline in the recent onset age-matched control group (P < 0.05). Analysis of the whole dose-response curve (using ANOVA for repeated measures with paired t test) showed a significant left-ward shift following the addition of L-NOARG (P < 0.001). A similar but less marked shift (P < 0.01) was evident in vessels from recent onset diabetics. An overall shift in both sensitivity and maximum response was also evident in the age-matched non diabetic controls of the insulin-treated group (P < 0.05). However, by contrast, there was no significant change in sensitivity in the insulin-treated diabetic rats. 5. ACh-induced endothelium-dependent relaxation was significantly impaired in the recent onset diabetic rats compared to their age-matched controls (47 +/- 11% versus 92 +/- 2%, P < 0.05, n = 6), and in the insulin treated diabetic rats (34 +/- 5% versus 75 +/- 6%, P < 0.05, n = 6). The relaxation responses to BK also were significantly impaired in the diabetic rats compared to their age-matched controls (recent onset: 20 +/- 3% versus 72 +/- 7%, P < 0.05, n = 6; insulin treated: 12 +/- 9% versus 68 +/- 7%, P < 0.05, n = 7). 6. Incubation with either the nitric oxide synthetase substrate, U-arginine, or the free radical scavenging enzyme superoxide dismutase (150 mu ml-1) failed to improve the attenuated response of acetylcholine-induced relaxation in the diabetic vessels. 7. Endothelium-dependent relaxation mediated by ACh and BK was significantly attenuated in both the diabetic and control vessels after incubation with L-NOARG. 8. Pretreatment with a cyclo-oxygenase inhibitor, indomethacin, significantly enhanced the relaxation to ACh in both the recent onset and insulin treated diabetic rats (42 +/- 10%, n = 7 versus 64 +/- 7%, n = 7, P < 0.05, and 40 +/- 5%, n = 7 versus 65 +/- 9%, n = 6, P < 0.05). 9. Following endothelium removal, there was a marked impairment in endothelium-dependent relaxation responses to ACh and BK in both the diabetic and control vessels. 10. Incubation with the thromboxane A2 receptor antagonist SQ29548, did not significantly improve the ACh endothelium-dependent relaxation response in the diabetic vessels. 11. Endothelium-independent relaxation to sodium nitroprusside was significantly impaired in the first group of diabetic vessels studied; however, subsequent studies showed no impairment of the sodium nitroprusside response in the diabetic vessels. 12. In conclusion, the ability of the endothelium to regulate vascular contractility is reduced in recent onset diabetic vessels, and significantly impaired in established insulin treated diabetics. Relaxation to the endothelium-dependent vasodilators ACh and BK was impaired in both the recent onset and the established insulin treated diabetics, and the ACh response was significantly improved following pretreatment with indomethacin, suggesting a role for a cyclo-oxygenase-derived vasoconstrictor. Preliminary studies with a thromboxane A2, receptor antagonist, SQ29548 did not significantly improve the impaired relaxation to ACh, indicating that the vasoconstrictor prostanoid is not thromboxane A2.     

Evaluation of a new selective medium for the isolation of Corynebacterium urealyticum

1. The objective was to determine whether nitric oxide participates in stress adaptive responses. Acute stress (AS) decreased endothelium-dependent vasoconstriction to noradrenaline (NA) in rat aorta [control rat (CR) 3.90 +/- 0.18, n = 22; AS 2.76 +/- 0.20, n = 13; P < 0.05]. 2. Chronic stress exposure previous to AS (CS) potentiated this effect [CS 1.93 +/- 0.19; n = 9; P < 0.05 related to CR, P < 0.05 related to AS]. 3. Methylene blue and NG-monomethyl-L-arginine, but not indomethacin, restored the decreased aorta reactivity to NA. 4. No reactivity alteration was observed in aorta without endothelium either in both stress conditions or in the presence of inhibitors. These data show that the nitric oxide participates in stress responses.     

Effect of elevated glucose on contractile and relaxant response of aortic strips in Indian bonnet monkey (Macaca radiata radiata)

Vascular contractile response to phenylephrine and potassium chloride were examined in strips of isolated thoracic aorta from non-diabetic monkeys with and without intact endothelium exposed to glucose (5.5 mM; control) and (11 mM; high) concentration. Acetylcholine causes relaxation in isolated aortic strips with intact endothelial cells while it causes contraction in aortic strips with damaged endothelial cells. In preparations with intact and damaged endothelium, there was a significant increase in the maximal contractile response to potassium chloride when added cumulatively, on exposure to elevated glucose (11 mM) concentration as compared to control. It was also observed that relaxant response to acetylcholine and sodium nitroprusside in control (5.5 mM) and high glucose (44 mM) concentration. Endothelium-dependent relaxation to acetylcholine decreased significantly in the presence of 44 mM glucose. In preparation without endothelium, contraction caused by acetylcholine increased in the presence of glucose (44 mM). Direct smooth muscle relaxation to sodium nitroprusside remained unchanged in aortic strips with and without endothelium. Relaxation response to sodium nitroprusside decreased in strips with damaged endothelium on exposure to high glucose when compared to control glucose.     

UFT and leucovorin: a review of its clinical development and therapeutic potential in the oral treatment of cancer

Different receptors mediating the release of endothelium-derived nitric oxide (EDNO) have been identified at endothelial level. In the present study we aimed to characterise, on rabbit aorta by means of pharmacological tools, the generation of EDNO by receptors located on endothelial cell membrane (M3, P2u, P2y) and by direct activation of Ca2+ entry into the endothelial cell. Four vasodilating drugs were tested (acetylcholine, UTP, A23187 and 2-methyl-thio-ATP); they were active only if the endothelial layer was intact, suggesting that they act through endothelial receptors. The effect of different nitric oxide synthase (NOS) inhibitors (0.1 mM: L- and D-NAME, L-NMMA, L-NIO and 7-NI) was investigated on NO-mediated relaxation induced by the relaxants in vessels with intact endothelium. NOS inhibitors differently affected relaxation mediated by the vasoactive drugs in isolated rabbit aorta. Reversibility of the inhibition by using a fixed concentration of L-arginine (0.1 mM) was different depending on the relaxing drug and NOS-inhibitor. The data obtained support the coexistence in aortic vessel of more than one endothelial cell NOS isoform, each provided with different receptor coupling.     

Endothelium-dependent modulation of resistance vessel contraction: studies with NG-nitro-L-arginine methyl ester and NG-nitro-L-arginine

1. The effect of NG-nitro-L-arginine methyl ester (L-NAME) and NG-nitro-L-arginine (L-NOARG) on noradrenaline (NA)-induced contractility and acetylcholine (ACh)-induced endothelium-dependent relaxation was studied in rat mesenteric resistance arteries. 2. Third order branches of mesenteric arteries were dissected and mounted on two forty micron wires in a Mulvany myograph. 3. Incubation with L-NAME and L-NOARG (10 microM) caused a time-dependent shift in the 50% response to NA (ED50) (0.01 microM-10 microM) but was not associated with an increase in the maximum contractile response. 4. L-NAME and L-NOARG (10 microM) caused a time-dependent inhibition of ACh (1 microM)-induced relaxation with a maximum effect after 120 min. 5. Following endothelium removal, incubation with either L-NAME or L-NOARG caused no significant shift in the ED50, although the residual relaxation response to ACh (1 microM) was further attenuated. 6. Incubation with the cyclo-oxygenase inhibitor, indomethacin, enhanced the relaxation to ACh and reduced the inhibitory effects of L-NAME and L-NOARG. 7. In conclusion, L-NAME and L-NOARG are potent inhibitors of acetylcholine-induced endothelium-dependent relaxation in mesenteric resistance arteries. The shift in ED50 associated with these inhibitors suggests a probable role for the endothelium in modulating the contractility of the resistance vasculature.     

Reactivity of small intramyocardial arteries from atherosclerotic and non-atherosclerotic human hearts

Little is known about how the vascular reactivity of the coronary microcirculation is affected by upstream atherosclerotic disease. We have examined, with a wire myograph, the responses of intramyocardial arteries from hearts in which the epicardial vessels were either free of atherosclerotic lesions (non-diseased group) or were affected by atherosclerosis (diseased group). Vasodilator responses of preconstricted vessels to substance P (84.1 +/- 12.6 compared to 42.0 +/- 19.7%) were less in vessels from the diseased group (p < 0.05). In contrast, the relaxation to bradykinin (70.2 +/- 21.2 compared to 100.6 +/- 7.9%) was increased in vessels from the diseased group (p < 0.05). The dilator responses to acetylcholine, adenosine diphosphate, histamine and sodium nitroprusside showed no significant differences between arteries from each group. 5-Hydroxytryptamine was without any significant vasodilator effect in arteries from either group. Assessment of contractile function revealed that the responses to 5-hydroxytryptamine, acetylcholine, U46619, endothelin-1 and L-N(G)-monomethylarginine in each group were not significantly different. Histamine, noradrenaline and dopamine were without any significant contractile response. These results demonstrate that upstream atherosclerosis does not confer any global impairment of endothelium-dependent vasorelaxant responses or smooth muscle hyperreactivity to vasoconstrictors in the arteries that penetrate the myocardium.     

The endothelium-dependent response of human hepatic artery after preservation with the UW solution

The University of Wisconsin's (UW) solution has been used commonly for current liver transplantation. However, its effect on the vascular endothelium remains unclear. Experiments were designed to study the effects. Human hepatic arteries harvested from patients with hepatocellular carcinoma undergoing liver resection were preserved in 4 degree C physiological solution (group 1, the content showed on the text) and UW solution (group 2) for 1 hr. Segments of preserved and control (group 3) hepatic arteries were suspended in organ chamber to measure the isometric force. The relaxations to acetylcholine (ACH) and adenosine diphosphate in segments of hepatic artery with endothelium were significantly greater than those segments without endothelium. The maximal relaxation to ACH in arterial segments with endothelium of group 2 was significantly different from those of group 1 and 3 (group 1 to group 3, 82 +/- 2%, 57 +/- 6%, and 83 +/- 4% of the initial tension contracted by neoepinephrine (3 X 10-7 mole/l, P < 0.05). The maximal relaxation to adenosine diphosphate was similar to the response to ACH. Perfusate hypoxia (oxygen tension 30 +/- 5 mmHG) caused endothelium-dependent contraction of the arterial segments (group 1 to group 3, 233 +/- 32%, 276 +/- 35%, and 251 +/- 40% of the initial tension, P < 0.05). Endothelium-independent relaxation and contraction of human hepatic artery to sodium nitroprusside and norepinephrine were not altered by UW solution. In summary, the impaired endothelium-dependent relaxation by UW solution and prominent endothelium-dependent contraction to hypoxia of human hepatic artery would favor vasospasm and thrombus formation after liver transplantation.     

The response of the isolated ductus arteriosus to transmural stimulation and drugs

1 Responses of isolated ductus arteriosus preparations from near term guinea-pigs and lambs to transmural electrical stimulation and drugs were studied in a low oxygen medium (Po(2) 19 to 28 mmHg).2 Acetylcholine and noradrenaline contracted both vessels in a dose-dependent manner, their threshold being between 10(-8) and 10(-7) M. Transmural stimulation (pulse width 0.2 to 0.6 ms, typically 20 Hz) also contracted the vessels.3 Atropine and phentolamine or dibenzyline selectively blocked responses to acetylcholine and noradrenaline, respectively.4 In the guinea-pig ductus, part of the response to transmural stimulation was due to activation of intrinsic adrenergic nerves since the responses were reduced by alpha-adrenoceptor antagonists, bretylium or prior reserpine treatment, but not by atropine. The response of the lamp ductus to transmural stimulation varied greatly in magnitude and was inconsistently affected by alpha-adrenoceptor blocking drugs.5 There was no evidence that transmural stimulation activated cholinergic nerves in either species.6 After inactivation of alpha-adrenoceptors with dibenzyline, noradrenaline caused a beta-adrenoceptor-mediated relaxation. Both this effect and isoprenaline-mediated relaxation were blocked by propranolol. beta-Adrenoceptor activity was more prominent in the ductus of the guinea-pig than of the lamb.7 Raising the Po(2) from 19-28 to 92-98 mmHg increased the response of the guinea-pig ductus to transmural stimulation suggesting that, in this species, physiological elevation of oxygen tension at birth may increase transmitter release from intrinsic adrenergic nerves. Whether this mechanism would contribute to ductus closure remains an open question.8 We postulate that beta-adrenoceptor-mediated relaxation has a role in maintaining ductus patency in the guinea-pig foetus.     

Perfadex is superior to Euro-Collins solution regarding 24-hour preservation of vascular function

BACKGROUND: The aim of this study was to compare Perfadex with Euro-Collins solution regarding 24-hour preservation of endothelium-dependent relaxation and vascular smooth muscle function. METHODS: The infrarenal aorta of 72 isogenic rats was studied in organ baths as fresh controls, after 24 hours of cold (4 degrees C) storage, and after 24-hour storage followed by transplantation and examination after 7 or 30 days. The thromboxane A2 analogue U-46619 was used to test contractility. Acetylcholine chloride was used to elicit endothelium-dependent relaxation and papaverine hydrochloride, to elicit endothelium-independent relaxation. RESULTS: With both solutions, all grafts were patent after 7 and 30 days. Vessels preserved in Euro-Collins solution for 24 hours lost 95% (p < 0.001) of their contractility compared with fresh controls; 7 days after transplantation, they had regained 40% of initial contractility, and after 30 days, there was no significant decrease in contractility. Vessels preserved in Perfadex manifested no significant decrease in contractility at any time. Endothelium-dependent relaxation could not be evaluated in vessels stored for 24 hours in Euro-Collins solution because they had lost almost all contractility; 7 days after transplantation, endothelium-dependent relaxation was reduced by 65% (p < 0.001), but at 30 days after transplantation, there was no significant decrease in endothelium-dependent relaxation. Vessels preserved in Perfadex for 24 hours lost 17% (p < 0.05) of endothelium-dependent relaxation, but 7 and 30 days after transplantation, there was no significant decrease in endothelium-dependent relaxation. CONCLUSIONS: Perfadex, but not Euro-Collins solution, has the capacity to preserve vascular function after 24 hours of storage followed by in vivo reperfusion.     

Functional and morphologic characterization of thoracic aorta in heritable hyperlipidemic Yoshida rats of different ages

We investigated serum and aortic tissue lipid content, in vitro aortic response to drugs, and morphology of thoracic aorta in Pittsburgh Yoshida rats (YOS), a new animal model of endogenous hyperlipidemia. Experiments were performed on 2-, 6-, and 18-month-old rats. Normolipidemic Brown Norway rats (BN) were used as controls. Both serum cholesterol and triglycerides increased significantly with age in YOS rats, but remained constantly low in the control group. In YOS rats, absolute serum concentration of high density lipoprotein (HDL)-cholesterol increased significantly with age, although HDL-cholesterol/total-cholesterol ratio decreased. In contrast, no difference in cholesterol content in aortic tissue was detected between the two animal strains or among different age groups. The contractile force generation of thoracic aorta to norepinephrine (NE) and serotonin increased with age in both strains of animals. The endothelium-dependent relaxation induced by acetylcholine (ACh) was significantly reduced in 6- and 18-month-old YOS as compared with 2-month-old YOS but not in BN. ATP-induced relaxation was significantly impaired in YOS thoracic aorta. In contrast, the relaxation induced by NaNO2 acting in smooth muscle did not vary with age in either YOS or BN. Only alterations in endothelial cells, not typical atheromatous injuries in thoracic aorta wall were detected in YOS even at age 18 months. Our data indicate that despite high serum lipid levels, YOS do not develop typical atheromatous lesions or functional and morphologic damage of smooth muscle cells in thoracic aorta, whereas YOS show decreased endothelium-dependent relaxation and morphologic alteration of endothelial cells.     

Botulinolysin, a thiol-activated hemolysin produced by Clostridium botulinum, inhibits endothelium-dependent relaxation of rat aortic ring

The effects of botulinolysin (Blyn), a thiol-activated hemolysin produced by Clostridium botulinum, on contractility of rat aortic ring were studied in order to clarify an underlying mechanism of vasoconstriction by the toxin observed previously as an increase in perfusion pressure in isolated rat organs. Blyn (30 hemolytic units/ml; HU/ml) itself did not elicit any apparent change in resting tension of the ring. Contractile tension elicited by a high concentration of phenylephrine in endothelium-intact rings increased significantly after treatment with Blyn (30 HU/ml), while phenylephrine-induced contraction of endothelium-denuded rings was not influenced by toxin treatment. In rings with intact endothelium, acetylcholine (ACh)-induced relaxation was significantly inhibited after treatment with Blyn (30, 10, 1 HU/ml). In contrast, relaxation of denuded rings by sodium nitroprusside was not affected by toxin treatment (30 HU/ml). Arginine (10(-4) M) partly reversed the inhibition of ACh-induced relaxation by the toxin (1 HU/ml). Endothelium-dependent relaxation by histamine or adenosine triphosphate was also inhibited by Blyn (1 HU/ml), but the relaxation elicited by calcium ionophore A23187 was not influenced by the toxin. The results indicate that Blyn acts on endothelium and inhibits agonist-induced endothelium-dependent relaxation of blood vessels.     

Impairment of endothelium-dependent vasorelaxation in experimental atherosclerosis is dependent on gender

OBJECTIVE: Nitric oxide (NO) has been suggested to have antiatherosclerotic effects. It has also been demonstrated that there is a greater basal release of endothelium derived relaxing factor (EDRF) in female as compared to male rabbit aorta, which also might have beneficial effects in atherosclerosis. We thus sought to determine if gender influences the severity of atherosclerosis. METHODS: We studied 18 female and 18 male New Zealand White rabbits that were randomly divided in two groups of 9 animals each and fed either a standard or a cholesterol diet (0.75%) for 15 weeks. RESULTS: In cholesterol-fed rabbits the percentage of atherosclerotic lesions in the aorta was identical in females and males and was inversely correlated with the maximal aortic relaxation to acetylcholine as assessed in organ chamber experiments (females: P < 0.0008, males: P < 0.0002). Importantly, the cholesterol diet induced a significantly (P < 0.025) more severe impairment of maximal vasorelaxation to acetylcholine in males from 78.4 +/- 1.2% to 29.4 +/- 10.2%) compared to females (from 84.4 +/- 1.2% to 60.7 +/- 8.5%). Both male gender (P < 0.0001) and the extent of impairment of endothelium-dependent relaxation (P < 0.0002) were associated with a reduced aortic sensitivity to S-nitroso-N-acetyl-D,L-penicillamine, which releases NO into the organ bath. In contrast, the aortic sensitivity to the organic nitrates pentaerythritol tetranitrate and isosorbide 5-nitrate, which release NO after enzymatic metabolization within the smooth muscle, was not reduced. CONCLUSIONS: These results suggest that the impairment of endothelium-dependent vasorelaxation induced by atherosclerosis is dependent on gender. This may be due to a greater degradation of extracellular NO in the vessel wall of males.     

Liver granulomas in schistosomiasis: mast cell-dependent induction of SCF expression in hepatic stellate cells is mediated by TNF-alpha

Abnormalities of vasomotor tone are characteristic of heart failure. This study was designed to assess the effects of chronic heart failure on endothelium-dependent relaxation in both large conduit arteries and small resistance vessels and to determine whether or not impaired nitric oxide (NO) production is involved. Segments of pulmonary artery (PA), abdominal aorta (AA), and small mesenteric artery (MA) were harvested from rats with heart failure resulting from coronary artery ligation and from sham-operated controls. Organ-bath experiments done in the presence of indomethacin to avoid the influence of vasodilatory prostanoids demonstrated that relaxation to acetylcholine (ACh) was impaired in the PA but not the AA or MA of the group with heart failure. Endothelium-independent relaxation to nitroglycerin was not significantly affected by the development of heart failure. Constriction to prostaglandin (PG) F(2alpha) was enhanced in PA but not in AA or MA segments. Preincubation with N(omega)-nitro-L-arginine (NNA) to inhibit the production of NO increased baseline force in vessels from all three beds, but the effect was greatest in the PA. Although relaxation to ACh was significantly diminished by NNA in the PA, it was not completely abolished. Furthermore, ACh-mediated relaxation in the presence of NAA was still impaired in the group with heart failure compared with the sham-operated control group. NNA had only mild effects on ACh-mediated relaxation in MA. These results demonstrate that (a) the mediators of endothelium-dependent relaxation may vary throughout the arterial circulation, (b) the contribution of NO to endothelium-dependent relaxation is substantial in PA and minimal in mesenteric resistance vessels, (c) endothelium-dependent relaxation is not uniformly impaired throughout the arterial bed by the development of heart failure, and (d) although a defect in NO production may account for enchanced vasoconstriction seen in response to PGF(2alpha), it does not account for the diminished vasodilatory response to ACh in this experimental model of heart failure.     

Exposure to oxidized low-density lipoprotein in vivo enhances intimal thickening and selectively impairs endothelium-dependent dilation in the rabbit

OBJECTIVES: Based on in vitro studies, oxidized low-density lipoprotein (oxLDL) has been implicated in atherogenesis and the associated deficiency in endothelium-dependent relaxation. The aim of this study was to investigate the effects of in vivo exposure to oxLDL on intimal thickening and relaxing behaviour. METHODS: Intimal thickening was evoked by the placement of silicone collars around the carotid arteries of the rabbit for 3 or 14 days. OxLDL (Cu(2+)-oxidized, 7 micron/h) or the vehicle phosphate-buffered saline (PBS) was infused in the collars via subdermally implanted osmotic minipumps. RESULTS: The collared vessels receiving PBS developed discrete intimal thickening after 14 days (intima/media (I/M) ratio 11 +/- 2%). OxLDL infusion resulted in intimal thickening after 3 days and significantly enhanced the intimal thickness by 14 days (I/M ratio 98 +/- 16%). Collaring alone for 3 or 14 days and 3 days exposure to oxLDL did not impair the endothelium-dependent relaxations to acetylcholine or calcium ionophore, nor to the NO donors glyceryl trinitrate (GTN) and S-nitroso-N-acetylpenicillamine (SNAP). However, the sensitivity to acetylcholine was decreased after exposure to oxLDL for 14 days (-logEC50 oxLDL 6.95 +/- 0.11 vs. 7.52 +/- 0.11 collar alone) and the maximal relaxation to the endothelium-dependent agonist was reduced by 50%, this in the presence of a virtually intact endothelium. Complete relaxation was still obtained with the nitric oxide donors. CONCLUSION: Our results show for the first time that local vascular exposure to oxLDL in vivo promotes intimal thickening and inhibits endothelium-dependent dilation, thereby supporting an active role for oxLDL in the morphological and functional changes observed in atherosclerotic blood vessels.     

15-Hydroxyeicosatetraenoic acid and diabetic endothelial dysfunction in rabbit aorta

We examined the effects of diabetes on eicosanoid metabolism and endothelium-dependent relaxation in isolated aorta from alloxan-induced diabetic rabbits and that from normal rabbits incubated in increased concentrations (44 mM) of glucose in vitro for 6 h. Immunoreactive 15-hydroxyeicosatetraenoic acid (HETE) was assayed in the incubation media of isolated aortic segments. Basal and acetylcholine (ACh)-stimulated release of 15-HETE was significantly greater in aorta of diabetic animals as compared with those of normal rabbits. Incubation of aortic segments from normal rabbits in increased concentrations of glucose caused a significant increase in basal and ACh-stimulated release of 15-HETE; and the release was significantly greater in aortic segments with endothelium than in segments without endothelium. Basal and ACh-stimulated release of 15-HETE was inhibited by indomethacin, a cyclooxygenase inhibitor. 15-HETE caused contractions of aortic rings that were inhibited by the prostaglandin H2 (PGH2) thromboxane A2 (TXA2) receptor blocker SQ-29548, but not by the TXA2 synthase inhibitor carbethoxyhexyl imidazole or indomethacin. Treatment of aortic rings with subthreshold concentrations of 15-HETE impaired ACh-induced relaxation; this was prevented by treatment with SQ-29548. Thus, abnormal release of endothelium-derived 15-HETE may play a role in endothelial cell dysfunction and increased vasoconstriction in diabetes by a mechanism that involves interaction with PGH2/TXA2 receptors.     

In vitro perfusion studies of resistance artery function in genetic hypertension

To examine the function of resistance-sized arteries in hypertension under in vitro conditions that approximate in vivo conditions as much as possible, we mounted segments of second-order mesenteric resistance arteries from spontaneously hypertensive rats (SHR) and Wistar-Kyoto normotensive control rats aged 12 to 13 weeks in a perfusion myograph and exposed them to conditions of constant flow and pressure. The endothelial integrity was validated both functionally and histologically. Vascular sensitivity to norepinephrine was examined when the hormone was applied either intraluminally or extraluminally and before and after removal of the endothelium. Both endothelium-dependent and -independent dilatation was assessed by the intraluminal application of acetylcholine and sodium nitroprusside, respectively. Sodium nitroprusside was applied to arteries after endothelium removal. Arterial responses were measured by changes in intraluminal diameter recorded with a video camera and imaging system. Vessels from SHR demonstrated depressed endothelium-dependent relaxation but similar endothelium-independent relaxation and greater sensitivity to norepinephrine with both intraluminal and extraluminal application. Removal of the endothelium abolished the differences in sensitivity to norepinephrine between the two strains. The results demonstrate that resistance arteries from SHR when examined under in vitro perfusion display enhanced sensitivity to norepinephrine due to depressed endothelium-dependent dilatation, and the data suggest that functional modifications in the endothelium may play an important role in hypertensive vascular disease.     

Some pharmacological and elastic characteristics of isolated subcutaneous small arteries from patients with essential hypertension

OBJECTIVE: To investigate the elastic characteristics of the wall of isolated subcutaneous resistance arteries from patients with essential hypertension, the response of the vessels to endothelium-dependent and -independent vasodilators and the dependence on calcium. METHODS: Subcutaneous resistance arteries were isolated from 16 patients with never-treated essential hypertension and from 16 normotensive controls matched for age and sex. The vessels were mounted in a myograph for isometric force development. The passive elastic characteristics were determined and then the response to acetylcholine, nitroprusside, felodipine, caffeine and calcium (in the presence of noradrenaline and prazosin or yohimbine) were determined. RESULTS: Young's elastic modulus as a function of wall stress was similar in the two groups of vessels. The relaxation of vessels from hypertensive and normotensive in response to acetylcholine, nitroprusside and felodipine was also similar. However, the response to caffeine was increased in vessels from the hypertensive patients, although the relationship between the dependence on the effect of calcium on the behaviour of arteries from hypertensives and controls was similar in the presence of prazosin and yohimbine. CONCLUSIONS: The altered morphology of subcutaneous resistance arteries from hypertensives is not caused by a change in the elastic characteristics of the wall material. The data support our previous observation of abnormal calcium handling in vessels from hypertensives, although they do not support the hypothesis that a generalized abnormality in endothelium-dependent or endothelium-independent relaxation is of importance in essential hypertension.     

Comparison of the effects of hypercholesterolaemia on relaxation responses in aortas of spontaneously hypertensive rats and Dahl salt-sensitive rats

1. We investigated the effects of hypercholesterolaemia on relaxation responses in thoracic aortas isolated from two different types of hypertensive rats; spontaneously hypertensive rats (SHR) and Dahl salt-sensitive rats (DSR). 2. All rats fed the high cholesterol diet for 8 weeks showed a significant increase in the serum cholesterol level. The high cholesterol diet did not change the blood pressure of SHR, but increased that of hypertensive DSR fed a high-salt diet. 3. In aortas of SHR, the high-cholesterol diet did not change the endothelium-dependent and -independent relaxation induced by acetylcholine (ACh) and sodium nitroprusside (SNP), respectively. 4. In aortas of hypertensive DSR, the high-cholesterol diet notably reduced the ACh-induced relaxations and slightly reduced SNP-induced relaxation. 5. These results suggest that hypercholesterolaemia causes greater impairment of endothelium-dependent relaxation in rat aorta with salt-induced hypertension than genetic hypertension.     

Relaxing actions of corticotropin-releasing factor on rat resistance arteries

1. Although it well established that corticotropin-releasing factor (CRF) injected i.v. can cause hypotension and vasodilatation, there is no in vitro evidence that CRF acts as a vasodilator. We have therefore tested the hypothesis that the hypotensive effect of i.v. CRF is due to a direct vasodilator action by carrying out experiments in vitro on rat resistance arteries (i.d. 150-300 microns). 2. Initial in vivo experiments confirmed that CRF (1.5 nmol.kg-1) injected i.v. caused hypotension in rats, this being partially antagonized by the CRF analogue CRF9-41. 3. For the in vitro experiments, vessels were taken from the mesenteric, cerebral and femoral vascular beds, and mounted as ring preparations in an isometric myograph. The vessels were pre-contracted with one of 3 agonists (prostaglandin F2 alpha, arginine vasopressin or noradrenaline) or with a high-potassium solution (K+). 4. With maximal concentrations of the agonists, CRF caused relaxation of mesenteric and cerebral vessels with 10 nM, and near complete relaxation with 100 nM. Femoral vessels pre-constricted with agonists and all vessels pre-constricted with K+ were less affected by CRF. In the mesenteric vessels, with sub-maximal levels of pre-constriction, CRF caused substantial relaxation at 1 nM and could cause complete relaxation at 10 nM. 5. The relaxant effect of CRF on contractions of mesenteric vessels was antagonized by 100 nM CRF9-41. Neither tetraethyl ammonium (30 mM) nor glibenclamide (3 microM) antagonized the relaxant effect of CRF. 6. The relaxant effect of CRF on mesenteric small arteries was found to be unaffected by removal of the endothelium. 7. The results indicate that CRF causes an endothelial-independent vasodilatation of rat resistance arteries under in vitro conditions at concentrations which are consistent with this being an important cause of the hypotension observed with i.v. injection of CRF.