Nitric oxide and vascular responses in Type I diabetes

Vascular complications are major causes of morbidity and mortality in patients with diabetes mellitus. The mechanisms underlying the development of microvascular and macrovascular angiopathy in Type I (insulin-dependent) diabetes mellitus are complex and incompletely understood. The discovery of endothelium-derived nitric oxide has greatly improved our understanding of vascular biology. Nitric oxide has an important role in the regulation of vascular tone and impaired nitric oxide activity could be implicated in the development of diabetic vasculopathy. Vascular studies of endothelial function in Type I diabetes have produced conflicting results. The role of nitric oxide in diabetic vasculopathy is still not clear. [Diabetologia (2000) 43: 137–147]     

Apelin对两肾一夹高血压大鼠离体主动脉环的舒张作用

目的：探讨小分子活性肽Apelin—13对两肾一夹（2K1C）高血压大鼠离体主动脉环的舒张作用及意义。方法：①清洁级雄性SD大鼠27只，随机分为正常对照组和2K1C高血压模型组。测定平均颈动脉压（mCAV）、左心重与体重比值（LVM／BW）以及右肾重／左肾重（RK／LK），以评价2K1C模型。②采用累计加药法，观察Apelin-13分别对用去甲肾上腺素（NE）预收缩的对照组大鼠和2K1C高血压组大鼠离体胸主动脉环张力的作用。结果：①模型组大鼠mCAP、LVM／BW和RK／LK分别较对照组高50．38％（P〈0．01）、32．16％（P〈0．01）和51．02％（P〈0．01）。②Apenn—13在（10^-11～10^-7）mol·L^-1浓度范围内对内皮完整的两组大鼠胸主动脉环具有浓度依赖性的舒张作用，且模型组舒张率高于对照组（P〈0．05），而模型组与对照．EC50相比差异无显著性（P〉0．05）。结论：Apelin-13具有显著的舒张主动脉的作用，并且对高血压大鼠血管环的舒张作用更强，提示Apelin—13在血管的保护作用中扮演了重要角色。     

Coronary dilatation reserve in experimental hypertension and chronic heart failure: effects of blockade of the renin-angiotensin system

1. The aim of the present study was to investigate left and right ventricular (LV and RV, respectively) coronary vasodilatation reserve (CVR; fluorescent microsphere technique) in rats with hypertension (spontaneously hypertensive rats (SHR)) or congestive heart failure (CHF) and the effects of early and chronic renin-angiotensin system (RAS) blockade thereupon. 2. In adult SHR, both LV and RV CVR were impaired, especially in the non-hypertrophied RV, the main factor involved being coronary vascular remodelling. Blockade of the RAS normalized both LV and RV CVR, mainly through the prevention of hypertension and suppression of the resulting pericoronary fibrosis. 3. In postischaemic CHF rats, there was an early and severe degradation of LV and RV CVR that developed before any significant vascular remodelling and appeared to be linked to the deterioration of cardiac hypertrophy and haemodynamics. This degradation in CVR further worsened over the longer term due to late-developing pericoronary fibrosis and endothelial dysfunction. Blockade of the RAS had no early effects on LV and RV CVR, but improved RV CVR over the long term, mainly by limiting RV hypertrophy and by preventing the development of pericoronary fibrosis and coronary endothelial dysfunction. 4. In kallikrein-kinin system-deficient mice, CVR was not different from that of wild-type mice, suggesting that this system is not implicated in normal CVR regulation.     

Corrigendum

1. Epidemiological and experimental data have shown that homocysteine may provoke vascular lesions and that moderate homocysteinaemia may constitute an independent risk factor for vascular disease. It is now documented that homocysteine damages human endothelial cells in culture, possibly by producing hydrogen peroxide in an oxygen-dependent reaction.
2. In this study, we have examined the direct effect of this sulphur amino acid on pancreatic vascular resistance. Experiments were performed on the vascular bed of the rat isolated pancreas perfused at constant pressure; thus, any change in pancreatic vascular resistance resulted in a change in the flow rate. D,L-Homocysteine perfused for one hour at three different concentrations (200 μM, 2 mM, 20 mM) did not induce any significant change in the flow rate per se. Homocysteine infusion for 30 min at a concentration of 200 μM or 2 mM abolished the endothelium-dependent vasodilatation induced by acetylcholine (0.05 μM), but did not modify adenosine (1.5 μM)-induced vasodilatation.
3. The effect of D,L-homocysteine (200 μM or 2 mM) cannot be ascribed to a direct antimuscarinic effect since 30 min pretreatment of rat ileum with these concentrations did not significantly change the contractile effect of increasing concentrations of acetylcholine (0.015–15 μM).
4. Preincubation of human umbilical vein endothelial cells with D,L-homocysteine (0.2–5.0 mM) had no significant effect on overall cell number or viability during 18 h of incubation; the endothelial cells exposed to concentrations up to 5 mM exhibited a spindle-shaped, whirled pattern. This pattern was reversed 48 h after the removal of homocysteine. A cytotoxic effect was seen after 18 h incubation in 10 mM D,L-homocysteine.
5. In conclusion, an acute infusion of homocysteine altered acetylcholine endothelium-induced vasodilatation, whereas the adenosine vasodilatator effect was insensitive to the deleterious action of homocysteine in vitro.

     

Endothelium-independent relaxation of aortic rings by the nitric oxide synthase inhibitor diphenyleneiodonium

1. The flavoprotein binder diphenyleneiodonium (DPI) is a potent, irreversible inhibitor of nitric oxide synthase (NOS), but produces only a transient pressor response following systemic administration to animals, despite evidence of persistent NOS inhibition. To characterize further the effects of DPI on vascular tone, isometric tension was recorded from rat isolated aortic rings mounted between steel wires in an organ bath.
2. The NOS inhibitor N^G-nitro-L-arginine methyl ester (L-NAME, 1 mM) initiated an additional contraction of prostaglandin F_2α-preconstricted rings with endothelium which was sustained throughout the period of L-NAME exposure (+234±39% at 15 min). In contrast, addition of DPI (5 μM) to rings with endothelium produced a transient initial contraction (+111±27% at 2 min) followed by a more sustained relaxation (−27±19% at 15 min, P<0.001 vs L-NAME).
3. The contraction to DPI was also observed in rings without endothelium, was abolished by L-NAME pretreatment, and was unaffected by the α-adrenoreceptor inhibitor prazosin. Relaxation in response to DPI was not inhibited by endothelium removal or by pretreatment with either L-NAME or with the ATP-sensitive potassium channel blocker glibenclamide.
4. The endothelium-independent relaxation to DPI was inhibited at 23°C and its time course was delayed by pretreatment with the guanylate cyclase inhibitor methylene blue.
5. Thus, in addition to a transient initial contraction due to NOS inhibition, DPI produces an endothelium-independent, temperature-dependent relaxation which appears in part due to activation of guanylate cyclase. This relaxant effect of DPI may explain the transient nature of its pressor effect in vivo despite sustained NOS inhibition.

     

Pharmacological characterization of endothelin-induced rat pulmonary arterial dilatation

1. The aim of study was to characterize endothelin (ET)-induced vasodilatation in isolated extrapulmonary rat arteries (EPA) and in intrapulmonary arteries (IPA) preconstricted with 1 μM phenylephrine.
2. The ET-3 (1 nM–100 nM)- and ET-1 (10 nM–100 nM)-induced transient vasodilatations in EPA were more potent than those in IPA. The vasodilatation induced by ET-3 (100 nM) was larger than that induced by ET-1 (100 nM).
3. Both the ET_B antagonist, BQ788 (3 μM) and or endothelium denudation, but not the ET_A antagonist, BQ123 (3 μM), abolished the vasodilatation induced by ET-1 or ET-3 (100 nM each) in EPA and in IPA. The ATP-sensitive K+channel blocker, glibenclamide (20 μM) and the nitric oxide synthase inhibitor, N^G-monomethyl-L-arginine (L-NMMA, 1 mM) suppressed the ET-induced vasodilatation in EPA and in IPA.
4. We conclude that the vasodilatation induced by endothelins is markedly reduced in rat isolated IPA, and suggest that the endothelial ET_B-mediated vasodilatation varies depending on rat pulmonary arterial regions. Furthermore, ET_B-mediated vasodilatation involves activation of ATP-sensitive K⁺ channels and of nitric oxide synthase in rat isolated EPA and IPA.

     

Role of Na+-K+ ATPase in cyclic GMP-mediated relaxation of canine pulmonary artery smooth muscle cells

1. Sodium-potassium adenosine triphosphatase (Na⁺-K⁺ ATPase) plays a role in the regulation of vascular tone, but contribution of this enzyme to nitrovasodilator-induced pulmonary vasodilatation remains uncertain. We thus studied the interaction between guanosine 3′:5′-cyclic monophosphate (cyclic GMP) and Na⁺-K⁺ ATPase in smooth muscle cells isolated from canine pulmonary artery.
2. To assess the contractile properties, changes in smooth muscle cell length were determined microscopically. Application of potassium chloride (KCl) shortened the cell length, an effect which was reduced by sodium nitroprusside and 8-bromo-cyclic GMP in a concentration-dependent manner. Pretreatment of cells with the cyclic GMP-dependent kinase inhibitor KT 5823 (2 μM) abolished the effects of sodium nitroprusside and 8-bromo-cyclic GMP.
3. Ouabain (0.3 μM) did not alter the KCl-induced muscle shortening, but inhibited the relaxant responses to sodium nitroprusside and 8-bromo-cyclic GMP.
4. Incubation of smooth muscle cells with sodium nitroprusside concentration-dependently increased intracellular cyclic GMP levels and ouabain-sensitive ⁸⁶Rb uptake, and these values were significantly correlated. In the presence of KT 5823, sodium nitroprusside increased cyclic GMP levels but did not alter ouabain-sensitive ⁸⁶Rb uptake.
5. These results suggest that there is a link between accumulation of intracellular cyclic GMP and activation of sarcolemmal Na⁺-K⁺ ATPase in pulmonary artery smooth muscle cells and that this link may be involved in the sodium nitroprusside-induced pulmonary vasodilatation.

     

P2Y receptor‐induced EDHF vasodilatation is of primary importance for the regulation of perfusion pressure in the peripheral circulation of the rat

Extracellular nucleotides have been shown to induce vasodilatation of conductance arteries by release of the endothelium‐derived hyperpolarizing factor (EDHF). As small resistance arteries are of greater importance for blood pressure regulation, a whole rat mesenteric arterial bed preparation was used in the present study when evaluating the physiological relevance for EDHF in mediating nucleotide dilatation. Dilatory responses were examined after pre‐contraction with noradrenaline in the presence of 10 mM indomethacin. Adenosine‐5′‐O‐thiodiphosphate (ADPβS), adenosine triphosphate (ATP) and uridine triphosphate (UTP) induced vasodilatation (pEC₅₀=6.5–7 and E_max=40–70%), while uridine diphosphate (UDP) was ineffective. Endothelium‐derived hyperpolarizing factor was studied in the presence of 0.5 mM N?‐nitro‐L ‐arginine (L ‐NOARG). ADPβS and UTP induced strong and potent EDHF‐dilatations, while ATP only had a weak effect (E_max=25%). After P2X₁ receptor desensitization with 10 μM αβ‐methylene‐adenosine triphosphate, the ATP response was significantly increased (E_max=65%). Putatively, this could be because of simultaneous activation of both endothelial P2Y receptors and P2X₁ receptors on smooth muscle cells, which resulted in the release of EDHF and subsequent hyperpolarization, and depolarization, respectively. Nitric oxide (NO) was studied in the presence of 60 mM K⁺. ADPβS, ATP and UTP induced weak NO dilatations, suggesting a minor role for NO as compared with EDHF. In conclusion, extracellular nucleotides stimulate dilatation by activating P2Y₁ and P2Y₂/P2Y₄ receptors, but not P2Y₆ receptors. The dilatory responses are mediated primarily by EDHF in the peripheral vascular bed.     

Sympathetic stimulation induced by hand cooling alters cold-induced vasodilatation in humans

Hand cooling is a cold pressor test, which induces general sympathetic stimulation. This cooling procedure is often performed to investigate cold induced vasodilatation (CIVD) in one finger. To investigate the effects of this sympathetic stimulation on local CIVD, 12 subjects immersed either the right index finger (T1), right hand (T2) or left hand and right index finger (T3) for 30 min in water at 5°C followed by 15-min recovery. Skin temperature and skin blood flow (Q˙ _sk) measured by laser Doppler flowmetry on the right index finger, as well as heart rate (f _c) and mean arterial blood pressure (), were continuously monitored during the three tests. Cutaneous vascular conductance was calculated as Q˙ _sk/. Concentrations of plasma noradrenaline (NA) and adrenaline (AD) were measured at different times during the tests. The results showed no cardiovascular change in T1, whereas f _c and increased significantly at the beginning of both T2 and T3. Similarly, sympathetic stimulation was reflected in the NA concentrations, which increased significantly (P < 0.01) during T2 and T3 after 5 min of immersion, and remained elevated until the recovery period. The AD concentration did not change during the three tests. During T2, the CIVD appeared later and slower in comparison with T1 [CIVD onset: 12.81 (SEM 2.30) min in T2 and 5.62 (SEM 0.33) min in T1] . During T3, the CIVD onset was not delayed compared to T1 [6.38 (SEM 0.67) min], but the rewarming was lower [+5.40 (SEM 0.86)°C in T3 and +9.10 (SEM 1.31)°C in T1]. These results showed that CIVD could be altered by sympathetic stimulation but it also appeared that the onset of CIVD could be influenced by local cooling, independently of the general sympathetic stimulation. Accepted: 23 September 1999