Cationic derivatives of dextran and hydroxypropylcellulose as novel potential heparin antagonists.

Cationic derivatives of dextran (Dex) and hydroxypropylcellulose (HPC) were studied as potential alternatives of protamine sulfate (PS) used in the reversal of anticoagulant activity of heparin. The modification was performed by the attachment of cationic groups to the Dex main chain or by grafting short side chains of a polycation onto HPC. The cationic derivatives of these polysaccharides were found to bind heparin with the efficiency increasing with growing degree of cationic modification. The degree of cationic modification and consequently the ζ potential of the polymers do not have to be high to achieve effective heparin binding. The size of the complexes of cationic Dex with unfractionated heparin (UFH) is a few micrometers. For complexes of cationic HPC and UFH the size is much below 1 µm, both below and above the lower critical solution temperature of HPC. None of the cationic polysaccharides studied caused hemolysis. The concentrations of the polymers inducing the aggregation of human erythrocytes in vitro were determined.

Compensation of endothelium-dependent responses in coronary circulation of eNOS-deficient mice.

Nitric oxide plays a fundamental role in the regulation of blood flow. Here we analyzed compensatory mechanisms for the genetic eNOS deficiency in aorta and in coronary circulation. Vasodilation induced by acetylcholine, bradykinin, adenosine, and ADP as well as by S-nitroso-penicillamine (SNAP) was assessed in isolated aorta and in isolated mouse hearts from eNOS-/- and age-matched eNOS+/+ mice. In aorta from eNOS+/+ mice acetylcholine-induced vasodilation was entirely dependent on NO, and this response was absent in aorta from eNOS-/- mice. In eNOS+/+ mouse hearts responses induced by bradykinin, adenosine and ADP were partially dependent on NO, but not on PGI2, cytochrome P450-dependent metabolites, or H2O2. On the other hand, vasodilation induced by acetylcholine involved NO, but not PGI2, in its immediate, short-lasting phase, whereas PGI2 and NO mediated delayed, longer-lasting phase of this response. In eNOS-/- mouse hearts coronary vasodilator function was compensated. Responses induced by acetylcholine and adenosine, but not by bradykinin or ADP, were in part compensated by NO, most likely derived from nNOS. However, the major mechanisms compensating for the loss of eNOS in the coronary circulation did not rely on NO, PGI2, cytochrome P450-derived metabolites of arachidonic acid or on H2O2. Deficiency of eNOS is largely compensated in coronary circulation but not in aorta.

Statins as coronary vasodilators in isolated bovine coronary arteries--involvement of PGI2 and NO.

Here we studied direct vasodilation induced by statins in isolated bovine coronary arteries. In rings of coronary bovine arteries preconstricted with prostaglandin F(2 alpha) (3 x 10(-8) - 10(-5)), lovastatin, simvastatin, atorvastatin and cerivastatin (3-30 microM) but not pravastatin induced concentration-dependent vasodilation. Removal of endothelium diminished response to simvastatin, cerivastatin and atorvastatin (30 microM) (67.4+/-4.56 vs. 22.7+/-8.14%, 96.9+/-2.27% vs. 54.5+/-6.86%, 67.4+/-4.01% vs. 34.6+/-5.66%, respectively). In presence of L-NAME (300 microM) or indomethacin (5 microM) responses to simvastatin, atorvastatin and cerivastatin, were also partially diminished. In contrast, lovastatin-induced vasorelaxation was not significantly affected by removal of endothelium (35.6+/-4.19% vs. 28.8+/-5.24%) or by pretreatment with L-NAME or indomethacin. In summary, with the exception of pravastatin, statins act as coronary vasodilators. Simvastatin, cerivastatin and atorvastatin but not lovastatin induced vasodilation displayed endothelium dependent- and endothelium-independent components. The endothelium-dependent effect of statins was mediated by NO and PGI(2), while the mechanism of smooth muscle cells-dependent component remains to be determined.

Hypertriglyceridemia but not hypercholesterolemia induces endothelial dysfunction in the rat.

In humans, hypercholesterolemia and hypertriglyceridemia induce endothelial dysfunction and therefore lead to atherosclerosis. In contrast, rats are resistant to atherosclerosis. Here we analyze whether rats respond to hypercholesterolemia and hypertriglyceridemia by developing of endothelial dysfunction. To induce hypercholesterolemiaWistar-Kyoto (WKY) and spontaneous hypertensive (SHR) rats were fed for 12 weeks with AIN93 diet supplemented with cholesterol (1%) and butter (20%). To induce hypertriglyceridemiaWistar were fed for 8 weeks with AIN93 diet supplemented with 60% fructose. In all experimental groups nitric oxide (NO)-dependent and prostacyclin (PGI(2))-dependent function was assessed in the isolated aorta. Additionally in hypertriglyceridemic rats endothelial function in the isolated mesenteric resistance artery was analyzed. NO-dependent vasodilation induced by acetylcholine or histamine in aorta of SHR and WKY rats was modestly impaired. Hypercholesterolemic diet fed to WKY and SHR rats induced a rise in total cholesterol and low-density lipoproteins (LDL) cholesterol by 2.5 and 4.5 fold, respectively, but did not further impair NO-dependent vasodilation. Although basal production of PGI(2) in aortic rings from SHR rats was five fold higher than in aortic rings from WKY rats, the hypercholesterolemic diet did not further affect aortic PGI(2) production in either rat strain. Endothelium-independent vasodilation induced by SNAP remained also unchanged. On the other hand, the hypertriglyceridemic diet given to Wistar rats led to a selective 1.5-2 fold elevation of triglycerides that was associated with the impairment of NO-dependent relaxation in aorta as well as in the mesenteric resistance artery. Interestingly, the basal production PGI(2) by aortic rings was not modified by hypertriglyceridemic diet. Again endothelium-independent relaxation induced by S-nitroso-N-acetyl-penicilamine (SNAP) was not affected. In summary, although in humans both hypercholesterolemia and hypertriglyceridemia are associated with endothelial dysfunction, in rats hypertriglyceridemia only led to the impairment of NO-dependent vasodilation. Hypercholesterolemia did not modify endothelial function even in hypertensive rats that display pre-existing alterations in vasodilator function.

Statins rise cytoplasmic calcium level [Ca2+]i in cultured endothelial cells.

Recently, we have shown that some HMG-CoA reductase inhibitors (statins) induce immediate pleiotropic effects in vascular endothelium both in vivo and in vitro, to mention only PGI2-mediated thrombolysis in rats and NO-mediated endothelium-dependent vasodilation in guinea pig coronary circulation. Here we look whether immediate endothelial effect of statins is associated with mobilization of intracellular calcium ions [Ca2+]i in cultured bovine aortic endothelial cells (BAEC). We analyzed the effects of various statins (atorvastatin, cerivastatin, simvastatin, lovastatin and pravastatin at concentration of 10-30 microM) on [Ca2+]i in BAEC in comparison to responses induced by bradykinin (Bk) (10 nM), adenosine diphosphate (1 microM), acetylcholine (100 nM), adrenaline (10 microM), serotonin (10 microM) or calcium ionophore A 23187 (0.1 microM) using FURA-2 according to fluorimetric method of Grynkiewicz et al. Basal [Ca2+]i level in BAEC was between 60 and 100 nM. Bk was the most potent to induce [Ca2+]i response. Delta[Ca2+]i induced by Bk was 331.9 +/- 19.49 nM (n = 36). Delta[Ca2+]i induced by statins (30 microM), i.e. atorvastatin, cerivastatin, simvastatin, lovastatin and pravastatin were 66.4 +/- 7.38% (n = 6), 54.8 +/- 10.12% (n = 5), 58.8 +/- 13.9% (n = 8), 27.7 +/- 7.19% (n = 5) and 0% (n = 5) of the response induced by Bk (10 nM), respectively. In summary, all statins tested, except pravastatin, induce immediate increase in [Ca2+]i in endothelium. This pleiotropic activity of statins in endothelium, most likely not related to the inhibition of HMG-CoA reductase, may represent an intracellular correlate for the immediate release of NO and PGI2 by these drugs that was reported by us previously.