Raloxifene, tamoxifen and vascular tone.

1. Oestrogen deficiency causes progressive reduction in endothelial function. Despite the benefits of hormone-replacement therapy (HRT) evident in earlier epidemiological studies, recent randomized trials of HRT for the prevention of heart disease found no overall benefit. Instead, HRT users had higher incidences of stroke and heart attack. Most women discontinue HRT because of its many side-effects and/or the increased risk of breast and uterine cancer. This has contributed to the development of selective oestrogen receptor modulators (SERMs), such as tamoxifen and raloxifene, as alternative oestrogenic agents. 2. A SERM is a molecule that binds with high affinity to oestrogen receptors but has tissue-specific effects distinct from oestrogen, acting as an oestrogen agonist in some tissues and as an antagonist in others. Clinical and animal studies suggest multiple cardiovascular effects of SERMs. For example, raloxifene lowers serum levels of cholesterol and homocysteine, attenuates oxidation of low-density lipoprotein, inhibits endothelial-leucocyte interaction, improves endothelial function and reduces vascular smooth muscle tone. 3. Available evidence suggests that raloxifene and tamoxifen are capable of acting directly on both endothelial cells and the underlying vascular smooth muscle cells and cause a multitude of favourable modifications of the vascular wall, which jointly contribute to improved local blood flow. The outcome of the Raloxifene Use for the Heart (RUTH) trial will determine whether raloxifene, currently approved for the treatment of post-menopausal osteoporosis, could substitute for HRT in alleviating cardiovascular symptoms in post-menopausal women.

Endothelium-independent relaxation to raloxifene in porcine coronary artery.

Although the vascular action of raloxifene has been studied in several vascular beds, the underlying mechanisms are still incompletely understood. The role of endothelium in raloxifene-induced vascular responses was controversial. The present study was designed to examine endothelium-independent effects of raloxifene in isolated porcine left circumflex coronary arteries. Arterial rings were suspended in organ baths and changes in isometric tension were measured. The large-conductance Ca2+-activated K+(BK(Ca)) currents were recorded using a whole-cell patch-clamp technique. Treatment with raloxifene (1-10 micromol/l) reduced the contractions to 9,11-dideoxy-11alpha,9alpha-epoxy-methanoprostaglandin F2alpha (U46619), serotonin (5-HT), endothelin-1 in normal Krebs solution and to CaCl2 in a Ca2+-free, high K+-containing solution. In endothelin-1-contracted rings, raloxifene (0.3 to 50 micromol/l) caused relaxations which were comparable in rings with and without endothelium. The raloxifene-induced relaxation was reduced by putative K+ channel blockers, iberiotoxin and tetraethyl ammonium chloride (TEA+) in rings with and without endothelium, or by elevated extracellular K+ ions (30 mmol/l K+ and 60 mmol/l K+). 13-methyl-7-[9-(4,4,5,5,5-pentafluoropentylsulfinyl)nonyl]-7,8,9,11,12,13,14,15,16, 17-decahydro-6H-cyclopenta[a] phenanthrene-3,17-diol (ICI 182,780) did not affect raloxifene-induced relaxation. Raloxifene enhanced the outward BK(Ca) currents, which were sensitive to inhibition by iberiotoxin. In summary, the present study shows that raloxifene acutely relaxes porcine coronary arteries via an endothelium-independent mechanism without involving the ICI 182,780-sensitive estrogen receptors. Raloxifene mainly acts on the vascular smooth muscle cells to induce vasorelaxation by the inhibition of Ca2+ channels and the activation of BK(Ca) channels. The former mechanism appears to play a more significant role.

Endothelial mediators of the acetylcholine-induced relaxation of the rat femoral artery.

This study examined endothelium-derived mediators of acetylcholine-induced relaxation in male rat femoral arteries. Arterial rings were suspended in a myograph for the measurement of isometric force. The generation of hydrogen peroxide (H2O2) in endothelial cells was detected using the fluorescent probe, 5-(and-6)-chloromethyl-2',7'-dichlorodihydrofluorescein diacetate acetyl ester. N(G)-nitro-L-arginine methyl ester (L-NAME, NOS inhibitor) and 1H-[1,2,4]oxadiazolo[4,2-alpha]quinoxalin-1-one (ODQ, guanylate cyclase inhibitor) alone or in combination with indomethacin (cycloxygenase inhibitor) diminished acetylcholine-induced endothelium-dependent relaxation to a similar extent. A small relaxation to acetylcholine in 60 mM KCl-constricted rings was abolished by L-NAME. Acetylcholine-induced relaxation was reduced by charybdotoxin plus apamin (intermediate- and small-conductance Ca2+-activated K+ channel blockers, respectively) or by 30 mM KCl. Both ouabain (Na+/K+ ATPase inhibitor) and BaCl2 (K(IR) channel blocker) also inhibited the relaxation albeit to a lesser degree. In the presence of L-NAME, ODQ plus indomethacin, charybdotoxin plus apamin or ouabain plus BaCl2 produced further inhibition. Catalase attenuated acetylcholine-induced relaxations and this attenuation was prevented by 3-amino-1,2,4-triazole (catalase inhibitor). Catalase did not affect acetylcholine-induced relaxations in rings treated with L-NAME or ODQ. Acetylcholine increased the dichlorofluorescein fluorescence intensity in native endothelial cells and this effect was abolished by catalase and by L-NAME. Exogenous H2O2 caused endothelium-independent relaxation that was slightly inhibited by iberiotoxin, ODQ or significantly reduced by elevated KCl, and abolished by catalase. The present results indicate that in addition to nitric oxide (NO) and endothelium-derived hyperpolarizing factor (EDHF, sensitive to charybdotoxin plus apamin, ouabain, and BaCl2), the endothelium of rat femoral artery can release H2O2 in response to acetylcholine, which was sensitive to L-NAME. Thus, the eNOS-dependent H2O2 is likely to be the third mediator of acetylcholine-mediated relaxations in rat femoral arteries.

Cilnidipine, a slow-acting Ca2+ channel blocker, induces relaxation in porcine coronary artery: role of endothelial nitric oxide and [Ca2+]i.

Cilnidipine is a dual blocker of L-type voltage-gated Ca(2+) channels in vascular smooth muscle and N-type Ca(2+) channels in sympathetic nerve terminals that supply blood vessels. However, the clinical benefits of cilnidipine and underlying mechanisms are incompletely understood. This study was designed to compare the time course of relaxant responses to cilnidipine and nifedipine, and to examine the role of endothelial NO and [Ca(2+)](i) in the vasorelaxation. Porcine left circumflex coronary arteries were isolated and isometric tension was measured with Grass force transducers. Endothelial [Ca(2+)](i) in intact arteries was determined by a calcium fluorescence imaging technique. The free radical scavenging capacity was also assayed. Cilnidipine and nifedipine induced concentration-dependent relaxations in high KCl-precontracted artery rings, while the former-induced relaxation was slower as compared to the latter. Treatment with L-NAME or ODQ reduced relaxations to cilnidipine or nifedipine to the same extent as in rings without endothelium. Indomethacin or omega-conotoxin had no effects. L-Arginine antagonized the effect of L-NAME on cilnidipine-induced relaxations. Cilnidipine did not affect sodium nitroprusside-induced relaxation in rings with and without endothelium. Cilnidipine and nifedipine caused extracellular Ca(2+)-dependent increases in endothelial [Ca(2+)](i) in intact arteries and cilnidipine's action had a slower onset, similar to that of cilnidipine-induced relaxation. Neither cilnidipine nor nifedipine exhibited a free radical scavenging property. The present results demonstrate that cilnidipine can produce endothelium-dependent relaxation in porcine coronary arteries in vitro in addition to blocking Ca(2+) channels. Like short-acting nifedipine, cilnidipine-dependent relaxation, albeit to a slower onset, is partly mediated by endothelial NO but not by prostacyclin. The increased release or bioavailability of NO may causally result from elevated endothelial [Ca(2+)](i) in arteries. The Ca(2+) channel-independent effect suggests the usefulness of cilnidipine in the treatment of cardiovascular diseases associated with diminished NO release, such as atherosclerosis.