Heat shock protein 90 mediates the balance of nitric oxide and superoxide anion from endothelial nitric-oxide synthase.

The balance of nitric oxide (.NO) and superoxide anion (O(2)) plays an important role in vascular biology. The association of heat shock protein 90 (Hsp90) with endothelial nitric-oxide synthase (eNOS) is a critical step in the mechanisms by which eNOS generates.NO. As eNOS is capable of generating both.NO and O(2), we hypothesized that Hsp90 might also mediate eNOS-dependent O(2) production. To test this hypothesis, bovine coronary endothelial cells (BCEC) were pretreated with geldanamycin (GA, 10 microg/ml; 17.8 microm) and then stimulated with the calcium ionophore, (5 microm). GA significantly decreased -stimulated eNOS-dependent nitrite production (p < 0.001, n = 4) and significantly increased -stimulated eNOS-dependent O(2) production (p < 0.001, n = 8). increased phospho-eNOS(Ser-1179) levels by >1.6-fold over vehicle (V)-treated levels. Pretreatment with GA by itself or with increased phospho-eNOS levels. In unstimulated V-treated BCEC cultures low amounts of Hsp90 were found to associate with eNOS. Pretreatment with GA and/or increased the association of Hsp90 with eNOS. These data show that Hsp90 is essential for eNOS-dependent.NO production and that inhibition of ATP-dependent conformational changes in Hsp90 uncouples eNOS activity and increases eNOS-dependent O(2) production.

Diabetes and hyperglycemia impair activation of mitochondrial K(ATP) channels.

Hyperglycemia is an important predictor of cardiovascular mortality in patients with diabetes. We investigated the hypothesis that diabetes or acute hyperglycemia attenuates the reduction of myocardial infarct size produced by activation of mitochondrial ATP-regulated potassium (K(ATP)) channels. Acutely instrumented barbiturate-anesthetized dogs were subjected to a 60-min period of coronary artery occlusion and 3 h of reperfusion. Myocardial infarct size (triphenyltetrazolium chloride staining) was 25 +/- 1, 28 +/- 3, and 25 +/- 1% of the area at risk (AAR) for infarction in control, diabetic (3 wk after streptozotocin-alloxan), and hyperglycemic (15% intravenous dextrose) dogs, respectively. Diazoxide (2.5 mg/kg iv) significantly decreased infarct size (10 +/- 1% of AAR, P < 0.05) but did not produce protection in the presence of diabetes (28 +/- 5%) or moderate hyperglycemia (blood glucose 310 +/- 10 mg/dl; 23 +/- 2%). The dose of diazoxide and the degree of hyperglycemia were interactive. Profound (blood glucose 574 +/- 23 mg/dl) but not moderate hyperglycemia blocked the effects of high-dose (5.0 mg/kg) diazoxide [26 +/- 3, 15 +/- 3 (P < 0.05), and 11 +/- 2% (P < 0.05), respectively]. There were no differences in systemic hemodynamics, AAR, or coronary collateral blood flow (by radioactive microspheres) between groups. The results indicate that diabetes or hyperglycemia impairs activation of mitochondrial K(ATP) channels.

Ethanol enhances the functional recovery of stunned myocardium independent of K(ATP) channels in dogs.

Chronic, intermittent exposure to small amounts of ethanol reduces myocardial infarct size in vivo. We tested the hypothesis that acute administration of ethanol enhances the functional recovery of stunned myocardium and that adenosine triphosphate-dependent potassium (K(ATP)) channels mediate this beneficial effect. Barbiturate-anesthetized dogs were instrumented for measurement of aortic and left ventricular pressure, +dP/dt(max), and subendocardial segment shortening (%SS) and were subjected to five 5-min periods of coronary artery occlusion, each separated by 5 min of reperfusion followed by a 3-h final reperfusion. In four groups (n = 7 each), dogs received 0.9% saline or ethanol (0.25, 0.5, or 1.0 g/kg over 30 min) in a random manner before occlusions and reperfusions. In other groups (n = 7 each), dogs received the K(ATP) channel antagonist glyburide (0.3 mg/kg, IV) 30 min before saline or ethanol (0.25 g/kg) was administered. Dogs receiving saline or glyburide alone demonstrated poor recovery of contractile function during reperfusion (%SS = 0.9% +/- 2.0% and 1.6% +/- 1.2% at 3 h, respectively). Recovery of %SS was enhanced in dogs receiving the 0.25- and 0.5-g/kg doses of ethanol (10.0% +/- 1.8% and 8.6% +/- 2.2% at 3 h, respectively) independent of alterations in hemodynamics or coronary collateral blood flow (radioactive microspheres). Glyburide did not affect improvement of recovery of stunned myocardium produced by ethanol (11.8% +/- 2.2% at 3 h). The results indicate that ethanol enhances the functional recovery of stunned myocardium independent of K(ATP) channels in vivo.

K(ATP) channels mediate the beneficial effects of chronic ethanol ingestion.

Chronic ingestion of low doses of ethanol protects the myocardium from ischemic injury by activating adenosine receptors and protein kinase C. We tested the hypothesis that ATP-dependent potassium (K(ATP)) channels mediate these beneficial effects. Dogs were fed with ethanol (1.5 g/kg) or water mixed with dry food twice per day for 12 wk. After they were acutely instrumented for measurement of hemodynamics, dogs received saline (vehicle) or glyburide (0.1 mg/kg iv) and were subjected to 60 min of coronary artery occlusion followed by 3 h of reperfusion. Infarct size (through triphenyltetrazolium chloride staining) was significantly (P < 0.05) reduced to 14 +/- 1% of the left ventricular area at risk in ethanol-pretreated dogs compared with controls (25 +/- 2%). Glyburide alone did not affect infarct size (25 +/- 3%) but abolished the protective effects of ethanol pretreatment (28 +/- 3%). No differences in hemodynamics or coronary collateral blood flow (through radioactive microspheres) were observed among groups. The results indicate that K(ATP) channels mediate the protective effects of chronic consumption of ethanol.

Sarcolemmal and mitochondrial adenosine triphosphate- dependent potassium channels: mechanism of desflurane-induced cardioprotection.

BACKGROUND: Volatile anesthetic-induced preconditioning is mediated by adenosine triphosphate-dependent potassium (KATP) channels; however, the subcellular location of these channels is unknown. The authors tested the hypothesis that desflurane reduces experimental myocardial infarct size by activation of specific sarcolemmal and mitochondrial KATP channels. METHODS: Barbiturate-anesthetized dogs (n = 88) were acutely instrumented for measurement of aortic and left ventricular pressures. All dogs were subjected to a 60-min left anterior descending coronary artery occlusion followed by 3-h reperfusion. In four separate groups, dogs received vehicle (0.9% saline) or the nonselective KATP channel antagonist glyburide (0.1 mg/kg intravenously) in the presence or absence of 1 minimum alveolar concentration desflurane. In four additional groups, dogs received 45-min intracoronary infusions of the selective sarcolemmal (HMR 1098; 1 microg. kg-1. min-1) or mitochondrial (5-hydroxydecanoate [5-HD]; 150 microg. kg-1. min-1) KATP channel antagonists in the presence or absence of desflurane. Myocardial perfusion and infarct size were measured with radioactive microspheres and triphenyltetrazolium staining, respectively. RESULTS: Desflurane significantly (P < 0.05) decreased infarct size to 10 +/- 2% (mean +/- SEM) of the area at risk as compared with control experiments (25 +/- 3% of area at risk). This beneficial effect of desflurane was abolished by glyburide (25 +/- 2% of area at risk). Glyburide (24 +/- 2%), HMR 1098 (21 +/- 4%), and 5-HD (24 +/- 2% of area at risk) alone had no effects on myocardial infarct size. HMR 1098 and 5-HD abolished the protective effects of desflurane (19 +/- 3% and 22 +/- 2% of area at risk, respectively). CONCLUSION: Desflurane reduces myocardial infarct size in vivo, and the results further suggest that both sarcolemmal and mitochondrial KATP channels could be involved.

Isoflurane preconditions myocardium against infarction via activation of inhibitory guanine nucleotide binding proteins.

BACKGROUND: Isoflurane-induced myocardial protection during ischemia is mediated by adenosine triphosphate-regulated potassium (KATP) channels; however, the intracellular signal transduction cascade responsible for this process has been incompletely evaluated. The authors tested the hypothesis that isoflurane reduces myocardial infarct size through a Gi protein-mediated process. METHODS: Forty-eight hours after pretreatment with vehicle (0.9% saline) or the Gi protein inhibitor pertussis toxin (10 microg/kg intravenously), barbiturate-anesthetized dogs (n = 43) were instrumented for measurement of aortic and left ventricular pressures and maximum rate of increase of left ventricular pressure. All dogs were subjected to a 60-min left anterior descending coronary artery occlusion followed by 3-h reperfusion. In four separate groups, vehicle- or pertussis toxin-pretreated dogs were studied with or without administration of 1 minimum alveolar concentration isoflurane. In two additional groups, dogs received the direct KATP channel agonist nicorandil (100 microg/kg bolus and 10 microg x kg-1 x min-1 intravenous infusion) in the presence or absence of pertussis toxin pretreatment. Myocardial perfusion and infarct size were measured with radioactive microspheres and triphenyltetrazolium staining, respectively. RESULTS: Isoflurane significantly (P < 0.05) decreased infarct size to 7 +/- 2% of the area at risk compared with control experiments (26 +/- 2%). Pertussis toxin pretreatment alone had no effects on myocardial infarct size (31 +/- 4%) but blocked the beneficial effects of isoflurane (21 +/- 3%). Nicorandil decreased infarct size (11 +/- 2%), but, in contrast to isoflurane, this effect was independent of pertussis toxin pretreatment (11 +/- 1%). CONCLUSION: Isoflurane reduces myocardial infarct size by a Gi protein-mediated mechanism in vivo.

Diabetes abolishes ischemic preconditioning: role of glucose, insulin, and osmolality.

Recent evidence indicates that hyperglycemia is an important risk factor for the development of cardiovascular disease. We tested the hypothesis that myocardial infarct size is related to blood glucose concentration in the presence or absence of ischemic preconditioning (PC) stimuli in canine models of diabetes mellitus and acute hyperglycemia. Barbiturate-anesthetized dogs were subjected to a 60-min period of coronary artery occlusion and 3-h reperfusion. Infarct size was 24 +/- 2% of the area at risk (AAR) for infarction in control dogs. PC significantly (P < 0.05) decreased the extent of infarction in normal (8 +/- 2% of AAR), but not diabetic (22 +/- 4% of AAR), dogs. Infarct size was linearly related to blood glucose concentration during acute hyperglycemia (r = 0.96; P < 0.001) and during diabetes (r = 0.74; P < 0.002) in the presence or absence of PC stimuli. Increases in serum osmolality caused by administration of raffinose (300 g) did not increase infarct size (11 +/- 3% of AAR) or interfere with the ability of PC to protect against infarction (2 +/- 1% of AAR). The results indicate that hyperglycemia is a major determinant of the extent of myocardial infarction in the dog.

Regional hyperthermia for deep-seated malignancies using the BSD annular array.

Forty-four patients were treated using the BSD-1000 Annular Phased Array between April 1983 and December 1986. There were 32 pelvic, nine abdominal, two extremity, and one thoracic sites treated. Mean tumour volume was 646 cc. Thirty-nine patients had concurrent radiation therapy, receiving a mean dose of 38 Gy. Mean average temperature was 41.0 +/- 1.4 degrees C. Most patients experienced local or systemic toxicity, requiring temporary treatment interruption in 33 patients, and termination of treatment in eight. Chronic complications were seen in four, but these were in patients receiving high total radiation doses as well. There were six complete and five partial responses. Among the 32 patients with pelvic tumours, mean tumour volume was 317 cc, mean radiation dose was 42 Gy, and mean average temperature was 41.3 +/- 1.2 degrees C. There were five complete and four partial responses. Achieving tumour temperatures greater than or equal to 42 degrees C with the annular array is difficult, due to both systemic and local toxicity. To improve clinical hyperthermia for thoracic, abdominal, and pelvic tumours, new technologies such as steerable phased array microwave systems; scanned, focused ultrasound; and permanently implantable thermoregulating ferromagnetic seeds, or new approaches such as using drugs to alter blood flow, or combining hyperthermia with antineoplastic drugs or biological agents, will be necessary.

Effects of dihydrocodeine, alcohol, and caffeine on breathlessness and exercise tolerance in patients with chronic obstructive lung disease and normal blood gases.

We measured breathlessness and exercise tolerance in 12 patients with chronic airways obstruction, moderate or severe breathlessness, and low or normal arterial carbon dioxide tension, after the patients received dihydrocodeine, alcohol, caffeine, or placebo (through double-blind administration). Forty-five minutes after ingestion, dihydrocodeine had reduced breathlessness by 20 per cent and increased exercise tolerance by 18 per cent, with a reduction in ventilation and oxygen consumption at submaximal work loads but with no change in spirometric volumes. Oxygen also reduced breathlessness and provided additional benefit to that achieved with dihydrocodeine (at three hours after ingestion) when the two were given together: the reduction of breathlessness was 18 per cent with dihydrocodeine; 22 per cent with oxygen; and 32 per cent with dihydrocodeine plus oxygen. Alcohol increased forced vital capacity by 9 per cent, and exercise tolerance by 7 per cent. Caffeine had no deleterious effect on breathlessness or exercise tolerance, despite increasing ventilation during rest and exercise. We conclude that opiates may be valuable for the treatment of breathlessness in selected patients; further evaluation is needed, particularly of the long-term benefits and safety.

Drug treatment of breathlessness: contrasting effects of diazepam and promethazine in pink puffers.

Fifteen out of 18 "pink and puffing" patients completed a double-blind, placebo-controlled cross-over trial of diazepam and promethazine for breathlessness and reduced exercise tolerance. Dosages were 25 mg and 125 mg daily, respectively, and each course lasted two weeks. Patients with psychiatric or other major medical histories were excluded. Of the three patients who did not complete the trial, one died during an exacerbation of breathlessness while taking diazepam, one was withdrawn because of mild hypercapnia while taking placebo, and one suffered intolerable drowsiness while taking diazepam. Of the remaining 15 patients, six needed a reduction in dosage because of drowsiness: one of these was taking promethazine and five diazepam. Diazepam had no effect on breathlessness and noticeably reduced exercise tolerance. Promethazine reduced breathlessness and improved exercise tolerance without altering lung function. From these results diazepam is contraindicated for breathlessness and reduced exercise tolerance in fixed airways obstruction, but promethazine may be beneficial.

Oxygen relieves breathlessness in "pink puffers".

The effect of oxygen on breathlessness and exercise tolerance was examined in "pink and puffing" patients with fixed airways obstruction. When breathing oxygen, patients were less breathless and walked further. This was true whether the cylinder was carried by the patient or by an assistant. It was not possible to identify those patients who would benefit most. The best method of assessing improvement was by comparing breathlessness on a standardised progressive exercise test on a treadmill. Four patients had a greater than 30% reduction in breathlessness on submaximum exercise when breathing oxygen. Breathing oxygen for five or fifteen minutes before exercise but not during exercise (predose) resulted in a similar improvement in exercise tolerance. For short periods of exercise predosing with oxygen provides a convenient alternative to continuous oxygen. For longer periods of exercise the benefits of portable oxygen in selected patients have been previously underestimated.