Chak Leung Au

Cyclooxygenase-2–Derived Prostaglandin F2α Mediates Endothelium-Dependent Contractions in the Aortae of Hamsters With Increased Impact During Aging

Hypertension and vascular dysfunction result in the increased release of endothelium-derived contracting factors (EDCFs), whose identity is poorly defined. We tested the hypothesis that endothelial cyclooxygenase (COX)-2 can generate EDCFs and identified the possible EDCF candidate. Changes in isometric tension of aortae of young and aged hamsters were recorded on myograph. Real-time changes in intracellular calcium concentrations ([Ca2+]i) in native aortic endothelial cells were measured by imaging. Endothelium-dependent contractions were triggered by acetylcholine (ACh) after inhibition of nitric oxide production and they were abolished by COX-2 but not COX-1 inhibitors or by thromboxane–prostanoid receptor antagonists. 2-Aminoethoxydiphenyl borate (cation channel blocker) eliminated endothelium-dependent contractions and ACh-stimulated rises in endothelial cell [Ca2+]i. RT-PCR and Western blotting showed COX-2 expression mainly in the endothelium. Enzyme immunoassay and high-performance liquid chromatography-coupled mass spectrometry showed release of prostaglandin (PG)F2&agr; and prostacyclin (PGI2) increased by ACh; only PGF2&agr; caused contraction at relevant concentrations. COX-2 expression, ACh-stimulated contractions, and vascular sensitivity to PGF2&agr; were augmented in aortae from aged hamsters. Human renal arteries also showed thromboxane–prostanoid receptor–mediated ACh- or PGF2&agr;-induced contractions and COX-2–dependent release of PGF2&agr;. The present study demonstrates that PGF2&agr;, derived from COX-2, which is localized primarily in the endothelium, is the most likely EDCF underlying endothelium-dependent, thromboxane–prostanoid receptor–mediated contractions to ACh in hamster aortae. These contractions involved increases in endothelial cell [Ca2+]i. The results support a critical role of COX-2 in endothelium-dependent contractions in this species with an increased importance during aging and, possibly, a similar relevance in humans.

Angiotensin II Type 1 Receptor-Dependent Oxidative Stress Mediates Endothelial Dysfunction in Type 2 Diabetic Mice

The mechanisms underlying the effect of the renin-angiotensin-aldosterone system (RAAS) inhibition on endothelial dysfunction in type 2 diabetes are incompletely understood. This study explored a causal relationship between RAAS activation and oxidative stress involved in diabetes-associated endothelial dysfunction. Daily oral administration of valsartan or enalapril at 10 mg/kg/day to db/db mice for 6 weeks reversed the blunted acetylcholine-induced endothelium-dependent dilatations, suppressed the upregulated expression of angiotensin II type 1 receptor (AT(1)R) and NAD(P)H oxidase subunits (p22(phox) and p47(phox)), and reduced reactive oxygen species (ROS) production. Acute exposure to AT(1)R blocker losartan restored the impaired endothelium-dependent dilatations in aortas of db/db mice and also in renal arteries of diabetic patients (fasting plasma glucose level > or =7.0 mmol/l). Similar observations were also made with apocynin, diphenyliodonium, or tempol treatment in db/db mouse aortas. DHE fluorescence revealed an overproduction of ROS in db/db aortas which was sensitive to inhibition by losartan or ROS scavengers. Losartan also prevented the impairment of endothelium-dependent dilatations under hyperglycemic conditions that were accompanied by high ROS production. The present study has identified an initiative role of AT(1)R activation in mediating endothelial dysfunction of arteries from db/db mice and diabetic patients.

Raloxifene protects endothelial cell function against oxidative stress

Maintaining a delicate balance between the generation of nitric oxide (NO) and removal of reactive oxygen species (ROS) within the vascular wall is crucial to the physiological regulation of vascular tone. Increased production of ROS reduces the effect and/or bioavailability of NO, leading to an impaired endothelial function. This study tested the hypothesis that raloxifene, a selective oestrogen receptor modulator, can prevent endothelial dysfunction under oxidative stress.

Contribution of K+ channels to relaxation induced by 17beta-estradiol but not by progesterone in isolated rat mesenteric artery rings.

17&bgr;-Estradiol and progesterone were found to relax various vascular beds through multiple mechanisms. However, the exact ionic mechanisms underlying the acute relaxant responses to both hormones are incompletely understood. This study was aimed to examine the possible role of K+ channel activation in the relaxation induced by both hormones in isolated rat mesenteric artery rings. Isometric tension of each ring was measured with Grass force displacement transducers. In rat endothelium–denuded rings preconstricted by 9,11-dideoxy-11&agr;,9&agr;-epoxy-methanoprostaglandin F2&agr; (U46619), the relaxation induced by 17&bgr;-estradiol was partially inhibited by tetrapentylammonium, 4-aminopyridine, iberiotoxin, BaCl2, and tertiapin-Q but not by tetraethylammonium, charybdotoxin, apamin, or glibenclamide. In contrast, these putative K+ channel blockers, except for glibenclamide, did not affect the relaxant response to progesterone. In 4 × 10−2M K+-preconstricted rings, the K+ channel blockers lost their inhibitory effects on 17&bgr;-estradiol–induced relaxation. Endothelium did not seem to be involved in the effects of K+ channel blockers on 17&bgr;-estradiol–mediated relaxation. Nifedipine-induced relaxation was not inhibited but was instead enhanced by tetrapentylammonium, iberiotoxin, 4-aminopyridine, and BaCl2. The above results indicate that in rat mesenteric artery rings, nonselective activation of K+ channels contributes partially to the relaxation induced by 17&bgr;-estradiol. These K+ channels involved in the estrogen response appeared to be sensitive to inhibition by KCa, KV, and KIR channel blockers. Lack of effect of K+ channel blockers on progesterone-induced relaxation suggests that these K+ channels play little or no role. The present findings provide pharmacological evidence for an additional mechanism contributing to acute vasorelaxation induced by 17&bgr;-estradiol.

Pivotal Role of Protein Kinase Cδ in Angiotensin II–Induced Endothelial Cyclooxygenase-2 Expression: A Link to Vascular Inflammation

Objective—The purpose of this study was to examine the hypothesis that angiotensin II (Ang II) induced endothelial cyclooxygenase-2 (COX-2) expression, which in turn mediated the generation of proinflammatory cytokines. Methods and Results—Western blot analysis on primary rat endothelial cells showed Ang II induced COX-2 expression, which was abolished by cotreatment of p38 mitogen-activated protein kinase (SB 202190) and extracellular signal–regulated kinase 1/2 (PD 98059) inhibitors. Protein kinase C&dgr; (PKC&dgr;) inhibitor (rottlerin) prevented extracellular signal–regulated kinase 1/2 phosphorylation and COX-2 expression. The pivotal role of PKC&dgr; was further supported by a similar stimulatory effect of the PKC activator on COX-2 expression, signified by Ang II–stimulated translocation of PKC&dgr; to the plasma membrane, and confirmed by PKC&dgr; phosphorylation at Tyr311. Small interfering RNA targeting PKC&dgr; diminished COX-2 expression, which was further abrogated by SB 202190. Human mesenteric arteries incubated with Ang II showed increased levels of endothelial COX-2 and monocyte chemoattractant protein-1; the former was inhibited by SB 202190 plus rottlerin, whereas the latter was prevented by COX-2 inhibitor. Conclusion—The present study pinpoints a novel role of PKC&dgr; in Ang II–induced endothelial COX-2 upregulation and identifies a COX-2-dependent proatherosclerotic cytokine monocyte chemoattractant protein-1. The findings raise the possibility of curtailing endothelial COX-2 expression as a means of limiting or preventing vascular inflammation.

Therapeutic concentrations of raloxifene augment nitric oxide-dependent coronary artery dilatation in vitro

Raloxifene improves cardiovascular function. This study examines the hypothesis that therapeutic concentrations of raloxifene augment endothelium‐dependent relaxation via up‐regulation of eNOS expression and activity in porcine coronary arteries.

Contribution of Na+‐Ca2+ exchanger to pinacidil‐induced relaxation in the rat mesenteric artery

Pinacidil relaxes blood vessels through opening the KATP channels with a resultant membrane hyperpolarization and inhibition of Ca2+ influx. The aim of this study was to examine the mechanisms thereby pinacidil induces K+ channel‐independent relaxation in isolated endothelium‐denuded rat mesenteric artery. Pinacidil‐induced relaxation was inhibited by glibenclamide (1–10 μM) in phenylephrine‐preconstricted rings, but was unaffected by glibenclamide after inhibition of K+ channels and VGCCs. Pinacidil‐induced K+ channel‐independent relaxation remained unchanged after treatment with cyclopiazonic acid (10 μM), thapsigargin (1 μM), ouabain (100 μM), propranolol (10 μM), Rp‐cAMPS triethylamine (30 μM), L‐NNA (100 μM), or ODQ (10 μM). Pinacidil induced more relaxant effect in the presence of nifedipine than in the presence of 60 mM K+ plus nifedipine. Pretreatment with Na+‐Ca2+ exchanger inhibitors, nickel (30–300 μM) or benzamil (20 μM) attenuated pinacidil‐induced relaxation in normal or in nifedipine‐containing solution. Pinacidil (1 μM) produced less relaxant effect with decreasing extracellular Na+ concentration. Na+‐free condition abolished the inhibitory effect of benzamil. Both nickel and benzamil inhibited pinacidil‐induced relaxation in the presence of glibenclamide (10 μM). Nickel (300 μM) did not affect the relaxant response to sodium nitroprusside. Pinacidil relaxed the rings preconstricted by active phorbol and U46619 with similar potency. The present results indicate that stimulation of the forward mode Na+‐Ca2+ exchange pathway is in part responsible for pinacidil‐induced K+ channel‐independent vasorelaxation. Pinacidil also induces K+ channel‐dependent but VGCCs‐independent relaxation. The PKC‐mediated cellular pathway may be a target site for pinacidil only in higher concentrations.

Role of inducible nitric oxide synthase in endothelium‐independent relaxation to raloxifene in rat aorta

Raloxifene can induce both endothelium‐dependent and ‐independent relaxation in different arteries. However, the underlying mechanisms by which raloxifene triggers endothelium‐independent relaxation are still incompletely understood. The purpose of present study was to examine the roles of NOSs and Ca2+ channels in the relaxant response to raloxifene in the rat isolated, endothelium‐denuded aorta.