[Establishment of a canine slow transit constipation model and evalution of defecation, gastrointestinal transit and pathological sections].

Objective: To establish a canine model of slow transit constipation (STC), and to test the changes in defecation, gastrointestinal transit time and pathology sections. Methods: Baseline information was measured in 8 beagle dogs, and these dogs were randomly divided into the control group and the model group. The dogs in model group were given a diet of canned meat, as well as a combination of compound diphenoxylate and alosetron hydrochloride for 5 weeks. Dogs in control group were given normal diet with no special intervention. Stool frequency and consistency were observed and recorded daily, and the gastrointestinal transit time (GITT) were measured every week. All animals underwent the midline laparotomy and the colonic tissues were taken from the rectosigmoid colon, then investigated by light microscopy, electron microscopy, and immunohistochemistry to evaluate changes of protein gene product 9.5(PGP9.5), synaptophysin and c-kit between two groups. Results: 8 beagle dogs underwent all experiment items successfully.Both of the stool frequency and scores of stool consistency decreased in model group(F=6.568, P=0.043; F=25.954, P=0.002). GITT delayed in model group(F=42.573, P=0.001). After 5 weeks of intervention, in the model group, the myenteric neurons and interstitial cells of Cajal showed damage such as swelling of mitochondria under electron microscopy, and both of the PGP9.5 and synaptophysin integrated option density of rectosigmoid colon were decreased (t=3.471, P=0.013; t=2.506, P=0.046)under immunohistochemistry. The c-kit integrated option density showed no statistically significant differences between two groups(t=1.709, P=0.138). Conclusions: The canine model of STC which was consistent with clinical symptoms and pathological changes was successfully established, and it can be used to observe and evaluate the therapeutic effect of electrical stimulation, surgery and so on.

Impaired vagal activity to meal in patients with functional dyspepsia and delayed gastric emptying.

Objective This study was performed to investigate impaired vagal activity to meal in patients with functional dyspepsia (FD) with delayed gastric emptying (GE). Methods Eighty-five patients were studied. GE parameters, including those in the overall and proximal stomach, were measured by GE functional tests at the Department of Nuclear Medicine. Autonomic nervous function was tested by spectral analysis of heart rate variability (HRV). The vagal activity and sympathetic activity were analyzed by recording the power in the high-frequency component (HF), low-frequency component (LF), and LF/HF ratio. Results Overall and proximal GE were delayed in 47.2% and 50.9% of the patients, respectively. Spectral analysis of HRV showed that the HF in patients with delayed proximal GE was significantly lower and that the LF/HF ratio was significantly higher than those in patients with normal proximal GE after a meal. Conclusion Delayed proximal GE might be caused by disrupted sympathovagal balance as a result of decreased vagal activity after a meal. Improvement in vagal activity may constitute an effective treatment method for patients with FD.

Increased expression of nerve growth factor correlates with visceral hypersensitivity and impaired gut barrier function in diarrhoea-predominant irritable bowel syndrome: a preliminary explorative study.

BACKGROUND: Neural-immune-endocrine network mechanism has attracted increased attention in diarrhoea-predominant irritable bowel syndrome (IBS-D). Pre-clinical evidence indicates that nerve growth factor (NGF) mediates visceral hypersensitivity and gut barrier dysfunction, via interactions with mast cells and sensory nerve fibres. AIM: To explore the role of nerve growth factor, as well as mast cell-nerve growth factor-nerve interaction in IBS-D pathophysiology. METHODS: In this cross-sectional study, IBS-D patients and healthy controls first underwent clinical and psychological assessments. Visceral sensitivity to rectal distension was tested. As gut barrier function markers, serum diamine oxidase and d-lactate were detected. Rectosigmoid biopsies were taken for the analyses of nerve growth factor expression, mast cell count and activation, and sensory nerve fibres expressing transient receptor potential vanilloid 1 and calcitonin gene-related peptide. Correlations between these parameters were examined in patients. RESULTS: Thirty-eight IBS-D patients (28 males, 10 females; average age 30.2 years) and 20 healthy controls (12 males, 8 females; average age 26.8 years) participated in the study. The patients presented increased psychological symptoms, visceral hypersensitivity and impaired gut barrier function. NGF gene expression, mast cell count and sensory nerve fibres were significantly increased in the patients (P < 0.05). In correlation analysis, NGF expression was positively correlated with the disease severity, anxiety and serum diamine oxidase; visceral sensitivity thresholds were negatively associated with NGF expression (Bonferroni corrected P < 0.0029). CONCLUSIONS: Elevated mucosal NGF may interact with mast cells and sensory nerve fibres, contributing to visceral hypersensitivity and impaired gut barrier function in IBS-D.

Relationship between symptoms and gastric emptying of solids in functional dyspepsia.

OBJECTIVE: Functional dyspepsia is a heterogeneous disorder and different pathophysiological mechanisms underlie its symptom patterns. This study investigated the relationship between dyspepsia symptoms and overall and proximal gastric emptying in patients with functional dyspepsia. METHODS: A total of 93 patients with functional dyspepsia and 32 healthy subjects were enrolled in this cross-sectional study. Prevalence and severity of eight dyspepsia symptoms were recorded. Gastric emptying was measured using single photon emission computed tomography scanning. RESULTS: Overall and proximal gastric emptying were delayed in 47.3% (44/93) and 46.2% (43/93) of the patients, respectively. Logistic regression analyses showed that presence of nausea was associated with delayed proximal gastric emptying (odds ratio 4.951; 95% confidence interval 1.321, 18.558). There were no significant differences between normal and delayed overall gastric emptying according to presence of symptoms. CONCLUSIONS: Presence of nausea might indicate delayed gastric emptying of the proximal stomach. Promotion of proximal gastric emptying may constitute an effective therapy for patients with functional dyspepsia who report nausea as the dominant symptom.

Localized administration of sodium nitroprusside enhances its protection against platelet aggregation in stenosed and injured coronary arteries.

Sodium nitroprusside, a potent vasodilator, was evaluated for its effect on platelet aggregation in stenosed and endothelium-injured coronary arteries in a canine model. Twenty-five anesthetized dogs were studied; coronary blood flow velocity was continuously monitored. Recurrent intracoronary platelet aggregation and dislodgment (indicated by cyclic variations in coronary blood flow) were induced by mechanically injuring and stenosing the left anterior descending coronary artery. Sodium nitroprusside was administered either intrapericardially or intravenously 30 min after cyclic flow variations were established. Intrapericardial administration of saline (control) did not affect cyclic flow variations in any of 6 tested dogs. Sodium nitroprusside abolished cyclic flow variations in all 7 dogs (100%) when given intrapericardially and in 5 to 7 dogs (71%) when given intravenously (compared to intrapericardial salines, p < 0.01). A smaller dose of sodium nitroprusside was required to abolish cyclic flow variations when given intrapericardially than when given intravenously (1.6 +/- 0.5 micrograms.kg-1.min-1 vs 4.8 +/- 0.8 micrograms.kg-1.min-1, p < 0.01). The mean aortic pressure was reduced by 10 to 20 mmHg after intrapericardial sodium nitroprusside administration and by 30 to 40 mmHg after intravenous sodium nitroprusside administration. To investigate the mechanism of protection by sodium nitroprusside, NG-monomethyl-L-arginine, an inhibitor of nitric oxide synthetase, was used to induce cyclic flow variations in mildly injured and stenosed left anterior descending coronary arteries in 5 dogs. Intrapericardial sodium nitroprusside abolished the cyclic flow variations in all 5 dogs. Then oxyhemoglobin, an inhibitor of nitric oxide, was administered into the left anterior descending coronary arteries of these dogs, and it restored the sodium nitroprusside-abolished cyclic flow variations in all 5 dogs. Thus, sodium nitroprusside protects against platelet aggregation and cyclic flow variations in stenosed and endothelium-injured canine coronary arteries, probably by the action of nitric oxide, and it is more effective and hemodynamically safer when administered intrapericardially than when administered intravenously.

Endogenous and exogenous nitric oxide protect against intracoronary thrombosis and reocclusion after thrombolysis.

BACKGROUND: Nitric oxide (NO), an endothelium-derived relaxing factor, plays an important role in regulating platelet activation. We evaluated the effect of NO in a canine model of intracoronary thrombosis, thrombolysis, and reocclusion. METHODS AND RESULTS: Before thrombosis was induced, 34 anesthetized dogs were treated with a continuous intracoronary infusion of saline (n = 8); NG-nitro-L-arginine (L-NNA, n = 8), an inhibitor of NO synthetase; L-arginine (n = 7), the precursor for NO; or sodium nitroprusside (SNP, n = 11), an NO donor. Ten minutes after the infusion was begun, an electric current of 150 microA was applied to the endothelium of coronary arteries to induce thrombosis. Occlusive thrombi developed in all dogs in the saline group (38 +/- 4 minutes) and the L-NNA group (30 +/- 6 minutes), in 6 of 7 dogs in the L-arginine group (81 +/- 18 minutes), and in 6 of 11 dogs in the SNP group (102 +/- 21 minutes) (P < .01). The time to thrombus was prolonged by L-arginine (P < .05) and SNP (P < .01). After 3 hours of thrombus formation in coronary arteries, tissue plasminogen activator and heparin were administered intravenously. Thrombi were lysed in 4 (of 8) dogs in the saline group (71 +/- 8 minutes), in 4 (of 8) dogs in the L-NNA group (72 +/- 8 minutes), in 4 (of 6) dogs in the L-arginine group (50 +/- 14 minutes), and in 4 (of 6) dogs in the SNP group (49 +/- 11 minutes) (P > .05). After thrombolysis, coronary artery reocclusion developed in all reperfused dogs in the saline group (30 +/- 8 minutes) and in the L-NNA group (48 +/- 12 minutes), in 3 (of 4) reperfused dogs in the L-arginine group (123 +/- 26 minutes), and in 3 (of 4) reperfused dogs in the SNP group (128 +/- 19 minutes) (P < .01). The ex vivo platelet aggregation induced by collagen was inhibited after in vivo treatment with L-arginine or SNP. CONCLUSIONS: Increasing NO production or giving an NO donor may inhibit platelet aggregation and delay intracoronary thrombus formation and reocclusion after thrombolysis.

Clopidogrel is more effective than aspirin as adjuvant treatment to prevent reocclusion after thrombolysis.

Clopidogrel is a thienopyridine derivative and a potent inhibitor of ADP-induced platelet aggregation. We compared clopidogrel with aspirin as an adjunctive treatment to tissue-type plasminogen activator (t-PA) for thrombolysis and reocclusion. Thrombosis was induced in coronary arteries of 32 dogs by injuring the endothelium with an electric charge. Coronary blood flow velocity was monitored by a pulsed Doppler flow probe placed around the artery. After the artery had been occluded by a thrombus for 3 continuous hours, each animal was given one of the following intravenous treatments: 1) t-PA (80 micrograms/kg + 8 micrograms.kg-1.h-1) and heparin (200 U/kg) (group 1, n = 7); 2) t-PA, heparin, and aspirin (5 mg/kg) (group 2, n = 8); 3) t-PA, heparin, and clopidogrel (5 mg/kg) (group 3, n = 9); and 4) t-PA, heparin, and clopidogrel (10 mg/kg + 2.5 mg.kg-1.h-1) (group 4, n = 8). After treatment, thrombolysis developed in 45 +/- 12 min in group 1, 39 +/- 10 min in group 2, 39 +/- 10 min in group 3, and 27 +/- 10 min in group 4 (compared with group 1, P > 0.05). After thrombolysis, reocclusion occurred in 5 of 5 dogs in group 1 and 7 of 7 in group 2, but only 2 of 7 in group 3 and none of 7 in group 4 (compared with groups 1 and 2, P < 0.001).(ABSTRACT TRUNCATED AT 250 WORDS)

Blockade of platelet membrane glycoprotein Ib receptors delays intracoronary thrombogenesis, enhances thrombolysis, and delays coronary artery reocclusion in dogs.

Von Willebrand factor and platelet membrane glycoprotein Ib receptors interact to mediate platelet adhesion and thrombogenesis in stenosed and endothelium-injured arteries. We wished to determine whether blocking glycoprotein Ib receptors with a recombinant von Willibrand factor binding domain (VCL) increases the time required for thrombus formation after injury to the coronary arteries. We also wished to determine whether, after thrombolysis with tissue plasminogen activator (TPA), VCL delays or protects against coronary artery reocclusion. Twenty-seven dogs were treated with either saline, VCL, or aspirin before thrombosis was induced in their coronary arteries by electrical injury. The time from injury to the formation of occlusive thrombi was significantly greater with VCL (70 +/- 10 minutes) and aspirin (69 +/- 20 minutes) than with saline (18 +/- 3 minutes, P < .001 and P < .05). Thrombosis was induced in 30 other dogs that then received thrombolytic treatment in four groups. Our major finding was that coronary artery reocclusion occurred in 72 +/- 11 minutes after treatment with TPA (80 micrograms/kg + 8 micrograms.kg-1.min-1) and heparin (200 U/kg) (n = 7); in 142 +/- 24 minutes after TPA, heparin, and VCL (4 mg/kg + 2 mg.kg-1.h-1) (n = 7) (compared with TPA and heparin, P < .05); in 74 +/- 13 minutes after TPA, heparin, and aspirin (5 mg/kg) (n = 8); and in 173 +/- 8 minutes after TPA, heparin, VCL, and aspirin (n = 8) (compared with TPA and heparin, P < .001). Thus, VCL increases the length of time required for thrombus formation in coronary arteries, and, when given with TPA and heparin, delays coronary artery reocclusion more effectively than aspirin.

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

BACKGROUND: 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. METHODS AND RESULTS: 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. CONCLUSIONS: Liposome-bound but not free prostaglandin E1 effectively diminishes CFVs in severely stenosed and endothelium-injured canine coronary arteries.

Combined ADP and thromboxane A2 antagonism prevents cyclic flow variations in stenosed and endothelium-injured arteries in nonhuman primates.

BACKGROUND: This study was designed to test the hypothesis that clopidogrel, a potent inhibitor of platelet aggregation, can eliminate cyclic flow variations in stenosed and endothelium-injured coronary and femoral arteries in nonhuman primates. METHODS AND RESULTS: We studied five anesthetized, open-chest baboons. Blood flow velocity in the coronary and femoral arteries was monitored by pulsed Doppler flow probes placed around the arteries. Cyclic flow variations were established by mechanically injuring the endothelium of the arteries and by narrowing the arteries with external constrictors. Clopidogrel (10 to 20 mg/kg i.v. bolus plus 2.5 mg.kg-1 x h-1 continuous infusion) was administered 60 minutes after cyclic flow variations were established. Clopidogrel abolished cyclic flow variations in the coronary and femoral arteries of all five baboons (frequency of cyclic flow variations, 0/h versus 14/h at baseline, P < .001). Then epinephrine was infused (maximum average dose, 2.2 micrograms.kg-1 x min-1 i.v.). Epinephrine did not restore cyclic flow variations in the coronary or femoral arteries of any baboon. Before treatment with clopidogrel, ADP, collagen, and U46619, a thromboxane A2 mimetic, induced dose-dependent platelet aggregation in vitro. Serotonin, however, did not induce platelet aggregation in vitro. Clopidogrel given in vivo completely inhibited ADP-induced platelet aggregation and significantly diminished collagen- and U46619-induced platelet aggregation in vitro. CONCLUSIONS: Clopidogrel eliminates cyclic flow variations in stenosed and endothelium-injured coronary and femoral arteries of nonhuman primates at least in part by antagonizing the platelet proaggregatory effects of ADP and thromboxane A2.

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.

Combination of inhibition of thrombin and blockade of thromboxane A2 synthetase and receptors enhances thrombolysis and delays reocclusion in canine coronary arteries.

BACKGROUND: The efficacy of thrombolytic therapy in treating patients with acute myocardial infarction is limited by failure to achieve reperfusion in some patients, by the prolonged time required to achieve reperfusion, and by reocclusion of some coronary arteries. We designed this study to examine the effect of combined inhibition of thrombin and thromboxane synthesis and blockade of thromboxane A2 receptors in addition to tissue-type plasminogen activator (t-PA) on thrombolysis and reocclusion in an experimental canine model with coronary thrombosis. METHODS AND RESULTS: Blood flow velocity in the left anterior descending coronary artery (LAD) of 32 anesthetized mongrel dogs was monitored by a pulsed Doppler flow probe. Coronary thrombosis was induced by applying electrical stimulation to the LAD at the site where an external constrictor was used to narrow the artery. Three hours after the formation of occlusive thrombus, animals were randomly assigned to receive one of the following: 1) t-PA (80 micrograms/kg + 8 micrograms.kg-1.min-1 i.v.) and saline; 2) t-PA and hirulog, a hirudin-based synthetic peptide and specific thrombin inhibitor (2 mg/kg + 2 mg.kg-1.hr-1 i.v.); 3) t-PA and ridogrel, a combined thromboxane A2 synthetase inhibitor and receptor antagonist (5 mg/kg + 2.5 mg.kg-1.hr-1 i.v.); or 4) t-PA, hirulog, and ridogrel. Reperfusion developed in 14% (one of seven) of dogs treated with t-PA alone at an average of 86 +/- 4 minutes after treatment, in 78% (seven of nine) of dogs treated with t-PA plus hirulog at 53 +/- 11 minutes, in 13% (one of eight) of dogs treated with t-PA plus ridogrel at 85 +/- 5 minutes, and in 88% (seven of eight) of dogs treated with t-PA, hirulog, and ridogrel at 37 +/- 10 minutes (comparison of the frequency of and the time to reperfusion, both p < 0.01). Among the dogs with reestablished coronary blood flow, reocclusion developed in the one treated with t-PA alone at 36 minutes after reperfusion, in seven of the seven treated with t-PA plus hirulog at 66 +/- 15 minutes, and in two of the seven treated with t-PA, hirulog, and ridogrel at 151 +/- 21 minutes (comparison of the frequency of and time to reocclusion, both p < 0.05). Reocclusion was not detected in the one dog treated with t-PA and ridogrel or in the other five dogs treated with t-PA, hirulog, and ridogrel within 180 minutes after reperfusion. Hirulog prolonged and maintained activated clotting times at a level twice that of baseline values. Hirulog inhibited ex vivo platelet aggregation induced by thrombin, and ridogrel inhibited platelet aggregation induced by U46619, a thromboxane mimetic. CONCLUSIONS: Inhibition of thrombin in addition to treatment with t-PA enhances thrombolysis. A combination of inhibition of thrombin and thromboxane synthetase and blockade of thromboxane A2 receptor enhances thrombolysis and delays or may prevent reocclusion of the recanalized coronary arteries.

Endogenous nitric oxide protects against platelet aggregation and cyclic flow variations in stenosed and endothelium-injured arteries.

BACKGROUND: This study was designed to test the hypothesis that endogenously produced nitric oxide protects against platelet aggregation and cyclic flow variations in stenosed and endothelium-injured arteries of mongrel dogs. METHODS AND RESULTS: NG-Monomethyl-L-arginine (L-NMMA), an inhibitor of nitric oxide formation, was administered at 5 mg/kg to 15 dogs after the left anterior descending coronary artery was mechanically injured and narrowed by external constrictors and to nine dogs before endothelial injury of the femoral artery and after injury and moderate arterial constriction. Treatment with L-NMMA resulted in cyclic flow variations (as detected by external Doppler flow probes) in the left anterior descending artery of seven of 15 dogs and in the femoral artery of four of nine dogs after endothelial injury. L-Arginine, the precursor for nitric oxide synthesis, was administered at 60 mg/kg and abolished cyclic flow variations in each of the 11 dogs. D-Arginine did not change the L-NMMA-induced cyclic flow variations. Saline infusion did not induce or change cyclic flow variations in any of the animals. Acetylcholine (1, 10, and 100 micrograms/min; n = 9) was administered in the femoral artery of nine additional dogs before and after endothelial injury in moderately stenosed femoral arteries. Acetylcholine did not induce cyclic flow variations in any animal; however, it did increase the severity of cyclic flow variations that developed in severely stenosed arteries. The diameter of the femoral artery was measured by intravascular ultrasound imaging. L-NMMA caused vasoconstriction of normal arteries, but no change was detected in endothelium-injured arteries. In contrast, L-arginine caused vasodilation of normal arteries, but, again, no change was noted in endothelium-injured arteries. Acetylcholine dilated normal femoral arteries but constricted arteries with endothelial injury. In both in vitro and ex vivo platelet studies, L-NMMA enhanced platelet aggregation, whereas L-arginine significantly reduced platelet aggregation. D-Arginine and acetylcholine showed no effect on platelet aggregation. CONCLUSIONS: Promotion of nitric oxide production decreases platelet aggregation and may eliminate cyclic flow variations, whereas a reduction in nitric oxide formation enhances platelet aggregation and may induce cyclic flow variations. Acetylcholine causes vasoconstriction at the femoral arterial site of endothelial injury and may increase the severity of cyclic flow variations.

Thrombin inhibition enhances tissue-type plasminogen activator-induced thrombolysis and delays reocclusion.

The objectives of this study were to test the hypotheses that thrombin inhibitors 1) enhance tissue-type plasminogen activator (t-PA)-induced coronary thrombolysis and 2) prevent or delay coronary artery reocclusion. Seventy-one dogs developed occlusive coronary thrombi after introducing a copper coil into the left anterior descending coronary artery (LAD). Coronary blood flows were monitored by an externally positioned pulsed Doppler flow probe. t-PA was given with or without heparin at different times after LAD occlusions. In some experiments, hirugen, a synthetic hirudin-based peptide and specific thrombin inhibitor, was given as 4 mg/kg i.v. bolus and 3 mg.kg-1.h-1 i.v. infusion at 30 min after LAD occlusion with t-PA and a bolus of heparin. Thrombolysis times were significantly shorter in t-PA- and heparin-treated dogs than in dogs treated with t-PA alone. Reocclusion times were significantly longer in t-PA- and heparin-treated dogs than in dogs treated with t-PA alone. Continuous heparin infusions prolonged reocclusion times to greater than 180 min in all treated dogs. The addition of hirugen to t-PA plus one bolus heparin prolonged reocclusion times to 90 +/- 6 min in dogs with 30-min thrombi. Thus heparin enhances t-PA-induced thrombolysis and delays reocclusion. Addition of a specific thrombin inhibitor, such as hirugen, to heparin enhances its effect on delaying reocclusion.

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.

Frequency and severity of cyclic flow alternations and platelet aggregation predict the severity of neointimal proliferation following experimental coronary stenosis and endothelial injury.

The role of recurrent platelet aggregation in the development of neointimal proliferation of coronary arteries was explored in this study, and the hypothesis was evaluated that recurrent platelet aggregation and the consequent frequency and severity of cyclic coronary blood flow variations are important pathophysiologic factors in the subsequent development of neointimal proliferation. In 24 chronically instrumented dogs, variable degrees of coronary artery neointimal proliferation were observed 3 weeks after mechanical injury of the arterial endothelium and the placement of an external coronary artery constrictor. The severity of neointimal proliferation at 21 days was closely related to the frequency and severity of cyclic coronary blood flow variations during the initial 7 days after instrumentation of the animals, itself a manifestation of recurrent platelet aggregation and dislodgement. Pharmacological therapy with a dual thromboxane A2 synthetase inhibitor and receptor antagonist and with a serotonin S2 receptor antagonist frequently was successful in abolishing cyclic blood flow variations and in retarding neointimal proliferation.

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 A2 and 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 A2 and serotonin were shown to be important mediators of intermittent coronary obstruction caused by platelet aggregation and dynamic vasoconstriction. Furthermore, thromboxane A2 synthesis 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 A2 synthesis 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 10 days to 3 weeks; the morphologic appearance of the neointimal proliferation was identical to that found in patients who develop restenosis after coronary angioplasty.