Endothelium-injured Coronary Arteries Research Articles

Intraplatelet tetrahydrobiopterin plays an important role in regulating canine coronary arterial thrombosis by modulating intraplatelet nitric oxide and superoxide generation.

Platelet-derived nitric oxide inhibits platelet aggregation via constitutive NO synthase (NOS). Tetrahydrobiopterin (BH(4)), a cofactor of NOS, augments NO formation, whereas its deficiency decreases NO bioactivity and increases superoxide generation by NOS. The roles of intraplatelet BH(4) in platelet aggregation and thrombus formation, however, are unknown. Accordingly, we investigated whether intraplatelet BH(4) is involved in regulating cyclic flow variations (CFVs) and platelet aggregation in a canine model with stenosed and endothelium-injured coronary arteries that mimics acute coronary syndromes in humans. After developing CFVs, dogs received saline or BH(4) (10 or 30 mg/kg) intravenously. Intraplatelet BH(4) and cGMP levels were decreased and intraplatelet nitrotyrosine production was increased during CFVs. ADP- and U46619-induced ex vivo platelet aggregation and platelet P-selectin expression were augmented during CFVs. BH(4) administration restored intraplatelet BH(4) and cGMP levels and decreased intraplatelet nitrotyrosine production, resulting in reduced CFVs and inhibited ex vivo platelet aggregation and platelet P-selectin expression. CFVs again developed after N(G)-monomethyl-L-arginine, an inhibitor of NOS, in BH(4)-treated dogs. Ex vivo platelet NOS activity at baseline, during CFVs, and after BH(4) administration did not differ. Intraplatelet BH(4) may play an important role in regulating thrombus formation by modulating platelet-derived nitric oxide and superoxide generation by platelet NOS.

Abolition of cyclic flow variations in stenosed, endothelium-injured coronary arteries in nonhuman primates with a peptide fragment (VCL) derived from human plasma von Willebrand factor-glycoprotein Ib binding domain.

Platelets play an important role in the pathophysiology of acute coronary syndromes. The interaction between the platelet glycoprotein Ib receptor and von Willebrand factor is a critical event allowing platelet adhesion and aggregation and subsequent thrombus formation in vessels with high shear rates and damaged endothelium. Therefore, we tested the hypotheses that VCL, an antagonist of von Willebrand-glycoprotein Ib binding domain, (1) attenuates/abolishes cyclic flow variations in stenosed, endothelium-injured coronary arteries in nonhuman primates and (2) reduces botrocetin-induced platelet aggregation in vitro after intravenous in vivo administration. Cyclic flow variations were established in anesthetized, open-chest baboons (n = 18). The baboons were divided into three groups. One group (n = 8) received a bolus of VCL (4 mg/kg IV) followed by an infusion (6 mg.kg-1.h-1) for 90 minutes (schedule A). Another group (n = 6) received a 2-mg/kg bolus followed by an infusion of 3 mg.kg-1.h-1 for 90 minutes (schedule B). The third group received a placebo infusion of normal saline. Under dosing schedule A, cyclic flow variations were abolished in 7 of 8 baboons after 33 +/- 18 minutes and markedly attenuated in 1. The frequency of cyclic flow variations fell from 18 +/- 9.4 per hour during the control period to 1 +/- 2.5 per hour after VCL infusion, P < .002. After cessation of infusion, cyclic flow variations remained abolished in 5 of 7 animals for > 3 hours and returned in 2 of 7 after 2 to 2.5 hours. Under schedule B, cyclic flow variations were abolished in 3 of 6 baboons and markedly reduced in the remainder. The number of cyclic flow variations fell from 17 +/- 4.8 per hour during the control period to 5 +/- 4.9 per hour after the VCL infusion, P < .001. The cyclic flow variations returned spontaneously at 38 +/- 40 minutes under this dosing schedule. Placebo infusion of saline had no effect on cyclic flow frequency or severity. VCL administration was associated with slight prolongation in bleeding time and a reduction in botrocetin-induced platelet aggregation. The bleeding time increased from a control time of 88 +/- 32 to 276 +/- 204 seconds, P < .03, and from 142 +/- 28 to 176 +/- 36 seconds, P = .056, for schedules A and B, respectively. VCL decreased platelet aggregation in response to botrocetin (20 micrograms/mL), from a control value of 66 +/- 30.3 to 33 +/- 31.3 omega, P < .05, and from 64 +/- 23.5 to 46 +/- 15.8 omega, P = .006, for dosing schedules A and B, respectively. Therefore, administration of a peptide fragment corresponding to von Willebrand-glycoprotein Ib binding domain (1) is effective in abolishing cyclic flow variations in stenosed, endothelium-injured coronary arteries and (2) reduces platelet aggregation in vivo in response to botrocetin in nonhuman primates.

Liposome-bound prostaglandin E1 often prevents cyclic flow variations in stenosed and endothelium-injured canine coronary arteries.

Prostaglandin E1 is a potent vasodilator with anti-inflammatory and antiplatelet effects. We tested the hypothesis that prostaglandin E1 attenuates or prevents platelet aggregation-associated cyclic flow variations (CFVs) in severely stenosed and endothelium-injured coronary arteries. We induced CFVs in 21 dogs by placing an external constrictor around the left anterior descending coronary artery at the site where the endothelium had been mechanically injured. The blood flow velocity in the artery was monitored by a pulsed Doppler flow probe. Sixty minutes after CFVs were established, liposome-bound prostaglandin E1, a stable formulation, was administered intravenously to 12 dogs at incremental doses of 0.25, 0.5, 1, and 2 micrograms/kg body wt; it abolished CFVs in 8 of the 12 dogs (67%). After CFVs were eliminated, epinephrine was infused intravenously, and at a dose of 6.6 +/- 1.6 micrograms/min, it restored CFVs in 7 of 7 dogs. Control dogs (n = 9) were treated with free prostaglandin E1, which did not abolish CFVs in any dog. Liposome-bound but not free prostaglandin E1 effectively diminishes CFVs in severely stenosed and endothelium-injured canine coronary arteries.

Active oxygen species play a role in mediating platelet aggregation and cyclic flow variations in severely stenosed and endothelium-injured coronary arteries.

A canine model with cyclic flow variations (CFVs) in stenosed and endothelium-injured coronary arteries was used to examine the role of active oxygen species in platelet aggregation in vivo. We studied 90 anesthetized dogs in which the pericardial cavity was opened and the heart was exposed. The velocity of blood flow in the left anterior descending coronary artery (LAD) was monitored by a pulsed Doppler flow probe. In 67 dogs, the LADs were stenosed by applying external constrictors at the site where the endothelium was mechanically injured. CFVs developed in all 67 dogs. Treatment with the antioxidants recombinant human copper-zinc superoxide dismutase (r-h-CuZnSOD), recombinant human manganese superoxide dismutase (r-h-MnSOD), and catalase eliminated platelet aggregation-associated coronary CFVs in 63%, 62%, and 64% of animals, respectively. Intravenous infusion of epinephrine restored CFVs in most dogs. Ketanserin, a serotonin (5-hydroxytryptamine2) receptor antagonist, abolished epinephrine-restored CFVs and eliminated CFVs in dogs in which CFVs had not been eliminated by free radical scavengers. In an additional 23 dogs, the LADs were stenosed but not mechanically injured. For control studies, saline was infused into the LADs of 5 dogs. Xanthine/xanthine oxidase was infused into the LADs of 8 dogs and induced CFVs in 4. Hydrogen peroxide was infused into the other 10 dogs and induced CFVs in 9. Histological analysis of the coronary artery revealed that the intima was significantly injured by the infusion. In ex vivo platelet aggregation studies, the in vivo treatment with r-h-CuZnSOD, r-h-MnSOD, and catalase significantly inhibited platelet aggregation induced by platelet-activating factor. Thus, active oxygen species are involved in mediating platelet aggregation and cyclic flow variations in stenosed and endothelium-injured canine coronary arteries in vivo.

ADP plays an important role in mediating platelet aggregation and cyclic flow variations in vivo in stenosed and endothelium-injured canine coronary arteries.

The goal of this study was to test the hypotheses that endogenous ADP plays an important role in vivo in mediating platelet aggregation and cyclic coronary artery blood flow variations (CFVs) in stenosed and endothelium-injured coronary arteries in an experimental canine model. Anesthetized animals were studied and coronary blood flow velocities monitored by a pulsed Doppler flow probe positioned around the left anterior descending coronary artery. CFVs were established by an external constrictor positioned at sites with injured endothelium. Apyrase, an ADP-removing enzyme, was infused into the left anterior descending coronary artery (0.3-1.8 units/min) 30 minutes or 2 hours after the establishment of CFVs. Complete abolition of CFVs was achieved in 81% (13/16) of dogs with 30-minute CFVs and in 83% (five of six) of dogs with 2-hour CFVs. In other dogs, a potent inhibitor of ADP-induced platelet aggregation, clopidogrel, was administered as a 10 mg/kg i.v. bolus and a 2.5 mg/kg/hr infusion 30 minutes and 3 hours after the establishment of CFVs. This treatment resulted in complete abolition of CFVs in 14 dogs (100%) with either 30-minute or 3-hour CFVs. Epinephrine was infused into some dogs after CFVs had ceased as a result of either apyrase or clopidogrel administration and into some dogs in whom SQ29548, a thromboxane A2 receptor antagonist, had been given when apyrase failed to abolish CFVs. Epinephrine restored CFVs in all dogs treated with apyrase alone, 67% (four of six) of dogs treated with the combination of apyrase and SQ29548, and 29% (two of seven) of dogs treated with clopidogrel. The plasma epinephrine levels required for CFV restoration were 20 times higher than baseline values in dogs receiving apyrase alone, 100 times higher when a combination of apyrase and SQ29548 had been given, and more than 5,000 times higher in dogs receiving clopidogrel. In vitro studies showed that apyrase only inhibited ADP-induced platelet aggregation, whereas clopidogrel not only inhibited ADP-induced platelet aggregation, but also reduced platelet aggregation induced by the thromboxane mimetic U46619 and serotonin. These data suggest that 1) ADP is an important mediator of platelet aggregation and CFVs in vivo and 2) combined inhibition of thromboxane A2 and ADP's effects provides marked protection against CFVs in experimentally stenosed and endothelium-injured canine coronary arteries. These data and our previous observations are consistent with the possibility that specific antagonists of thromboxane A2, serotonin, and ADP, alone and together, may provide substantial protection against platelet aggregation leading to CFVs at sites of endothelial injury and coronary artery stenosis.

Role of thromboxane and serotonin as mediators in the development of spontaneous alterations in coronary blood flow and neointimal proliferation in canine models with chronic coronary artery stenoses and endothelial injury

Platelet-mediated obstruction of stenotic and endothelium-injured coronary arteries may be important in the abrupt progression from chronic stable to unstable coronary heart disease syndromes in patients. Transcardiac accumulation of thromboxane A2and serotonin has been demonstrated in patients as chronic stable angina is converted to unstable angina. In this study in anesthetized open chest dogs with coronary artery stenosis and endothelial injury, thromboxane A2and serotonin were shown to be important mediators of intermittent coronary obstruction caused by platelet aggregation and dynamic vasoconstriction. Furthermore, thromboxane A2synthesis inhibitors and receptor antagonists and serotonin receptor antagonists, singly and together, provided substantial protection against repetitive platelet aggregation and dislodgment in canine models with coronary artery stenosis and endothelial injury even when systemic catecholamine concentrations were markedly elevated. These same observations apply in chronically instrumented, awake, unsedated dogs with coronary artery stenosis and endothelial injury in which recurrent platelet attachment and dislodgment cause cyclic flow alterations that may be prevented by thromboxane A2synthesis inhibitors and receptor antagonists and serotonin receptor antagonists. Chronically instrumented dogs with coronary stenosis and endothelial injury in which recurrent platelet attachment and dislodgment occurred also developed neointimal proliferation of varying severity within 19 days to 3 weeks; the morphologic appearance of the neointimal proliferation was identical to that found in patients who develop restenosis after coronary angioplasty.