Anaerobic co-digestion of sewage sludge and primary clarifier skimmings for increased biogas production.

The objective of the study was to identify the impact of co-digesting clarifier skimmings on the overall methane generation from the treatment plant and additional energy value of the increased methane production. Biogas production from co-digesting clarifier skimmings and sewage sludge in pilot-scale fed-batch mesophilic anaerobic digesters has been evaluated. The digester was fed with increasing quantities of clarifier skimmings loads: 1.5, 2.6, 3.5 and 7.0 g COD equivalent/(L·d) (COD: chemical oxygen demand). Average volatile solids reduction of 65% was achieved in the scum-fed digester, compared with 51% in the control digester. Average 69% COD removal was achieved at highest scum loading (7 g COD eq/(L·d)) with approximate methane yield of 250 L CH(4)/kg COD fed (4 ft(3)/lb COD fed). The results show that scum as co-substrate in anaerobic digestion systems improves biogas yields while a 29% increase in specific CH(4) yield could be achieved when scum load is 7 g COD eq/(L·d). Based on the pilot-scale study results and full-scale data from South East Water Pollution Control Plant and Northeast Water Pollution Control Plant the expected annual energy recovery would be approximately 1.7 billion BTUs or nearly 0.5 million kWh.

Emergency and out of hours care of patients with inherited bleeding disorders.

Recently, the United Kingdom Haemophilia Centre Doctors Organisation published recommendations for the standard of care for assessment and treatment of patients with bleeding disorders in the emergency department (A&E). An audit was undertaken to compare the level of care to the acceptable standards in a tertiary hospital A&E, attached to a haemophilia comprehensive care centre. A&E attendances were found by cross referencing all patients with known bleeding disorders against the EDMS attendance system. Visits from the past 3 years were identified to produce sufficient data and electronic notes from these visits were then accessed, and marked against the proforma. Data were available from 45 of a total of 54 patients, who had a total of 75 emergency visits documented. In all aspects of care, the standards were not adequately met including the average length of time between booking and clinical assessment, early initiation of specific haemostatic treatment, seeking haematology advice and arrangement of follow-up. Also no specialist clotting investigations were done with only 9/11 patients admitted having their haematological diagnosis recorded. In addition, only very few patients had the severity of bleeding disorder noted and less than half their first line treatment documented. There were significant differences in the standard of care for haemophilia patients provided by the A&E department when compared with acceptable standards. Measures have been put in place and policies have been drafted to improve this situation and provide the best possible care to persons with haemophilia.

Healthcare-acquired clusters of COVID-19 across multiple wards in a Scottish health board.

BACKGROUND: Healthcare-acquired COVID-19 has been an additional burden on hospitals managing increasing numbers of patients with SARS-CoV-2. One acute hospital (W) among three in a Scottish healthboard experienced an unexpected surge of COVID-19 clusters. AIM: To investigate possible causes of COVID-19 clusters at Hospital W. METHODS: Daily surveillance provided total numbers of patients and staff involved in clusters in three acute hospitals (H, M and W) and care homes across the healthboard. All clusters were investigated and documented, along with patient boarding, community infection rates and outdoor temperatures from October 2020 to March 2021. Selected SARS-CoV-2 strains were genotyped. FINDINGS: There were 19 COVID-19 clusters on 14 wards at Hospital W during the six-month study period, lasting from two to 42 days (average, five days; median, 14 days) and involving an average of nine patients (range 1-24) and seven staff (range 0-17). COVID-19 clusters in Hospitals H and M reflected community infection rates. An outbreak management team implemented a control package including daily surveillance; ward closures; universal masking; screening; restricting staff and patient movement; enhanced cleaning; and improved ventilation. Forty clusters occurred across all three hospitals before a January window-opening policy, after which there were three during the remainder of the study. CONCLUSION: The winter surge of COVID-19 clusters was multi-factorial, but clearly exacerbated by moving trauma patients around the hospital. An extended infection prevention and control package including enhanced natural ventilation helped reduce COVID-19 clusters in acute hospitals.

Acetoclastic methanogens in an anaerobic digester could be susceptible to trace metal supplementation.

The objective of this study was to investigate the effects of nutrient supplementation on anaerobic biomass. While many studies emphasized the importance of supplementing trace metals such as iron, cobalt, and nickel for maximum methanogenic activity, there is no evidence whether such supplements, even at relatively low concentration, could perturb anaerobic biomass. Effects of supplementing nutrients, including yeast extract, on anaerobic biomass from two full-scale mesophilic digesters, operating under different conditions, at the North East Water Pollution Control Plant in Philadelphia, Pennsylvania, USA, were assessed using biochemical methane potential tests. The results show that acetoclastic methanogens from a recently cleaned digester was not stimulated by nutrient supplementation at relatively low concentrations and a slight perturbation was observed when supplementation was at a relatively high concentration. Furthermore, greater degree of susceptibility to the trace metal supplementation was observed for biomass from another digester that had not been cleaned for over 10 years, thus it had reduced active volume due to grit accumulation. For instance, supplementation of 200 mg/L of iron as FeCl(2)·4H(2)O to the biomass from the reduced-active-volume digester caused 17% reduction in CH(4) production, as compared to a control which did not receive any supplements, while the same concentration had no effect on the biomass from full-active-volume digester. Results strongly suggest that acetoclastic methanogens stressed due to reduced hydraulic/solids retention time may be susceptible to trace metal addition. Therefore, trace metal supplementation for anaerobic digesters should be considered on a case by case basis.

Clinical and genetic correlates of soluble P-selectin in the community.

BACKGROUND: P-selectin is a cell adhesion molecule that is involved in atherogenesis, and soluble concentrations of this biomarker reflect cardiovascular risk. However, the clinical correlates and genetic characterization of soluble P-selectin have not been clearly elucidated. OBJECTIVE: To describe clinical and genetic correlates of circulating P-selectin in the community. METHODS: In Framingham Heart Study Offspring (European descent) and Omni (ethnic/racial minority) participants, we examined the association of cardiovascular risk factors with soluble P-selectin concentrations. In Offspring participants, we evaluated heritability, linkage and association of 29 SELP single-nucleotide polymorphisms (SNPs) with adjusted P-selectin concentrations. RESULTS: In multivariable analysis of 3,690 participants (54% women, mean age 60 +/- 10 years), higher log-transformed P-selectin concentrations were inversely associated with female sex and hormone replacement therapy, and positively associated with age, ethnic/racial minority status, cigarette smoking, waist circumference, systolic blood pressure, fasting glucose, and total/high-density lipoprotein cholesterol and triglyceride concentrations. Clinical factors explained 10.4% of the interindividual variability in P-selectin concentrations. In 571 extended pedigrees (n = 1,841) with >or= 2 phenotyped members per family, multivariable-adjusted heritability was 45.4 +/- 5.8%. Among the SELP SNPs examined, a non-synonymous SNP (rs6136) encoding a threonine-to-proline substitution at position 715 was highly significantly associated with decreased P-selectin concentrations (P = 5.2 x 10(-39)), explaining 9.7% of variation after adjustment for clinical factors. CONCLUSIONS: Multiple clinical factors and an SNP in the SELP gene were significantly associated with circulating P-selectin concentrations. One SNP in SELP explained significant variation in circulating P-selectin concentrations, even after accounting for known clinical correlates.

Nitric oxide released from activated platelets inhibits platelet recruitment.

Vessel injury and thrombus formation are the cause of most ischemic coronary syndromes and, in this setting, activated platelets stimulate platelet recruitment to the growing thrombus. Recently, a constitutive nitric oxide synthase (NOS) has been identified in human platelets. To further define the capacity of platelets to produce nitric oxide (NO), as well as to study the role of this NO in platelet recruitment, we adapted a NO-selective microelectrode for use in a standard platelet aggregometer, thereby permitting simultaneous measurement of platelet aggregation and NO production. Treatment of platelets with the NO synthase inhibitor -NG-nitroarginine methyl ester (L-NAME), reduced NO production by 92+/-8% in response to 5 microM ADP compared to control but increased aggregation by only 15+/-2%. In contrast, L-NAME had a more pronounced effect on platelet recruitment as evidenced by a 35+/-5% increase in the extent of aggregation, a 33+/-3% decrease in cyclic GMP content, and a 31+/-5% increase in serotonin release from a second recruitable population of platelets added to stimulated platelets at the peak of NO production. To study platelet recruitment accurately, we developed an assay that monitors two platelet populations simultaneously. Nonbiotinylated platelets were incubated with L-NAME or vehicle and activated with ADP. At peak NO production, biotinylated platelets were added. As measured by three-color flow cytometry, there was a 56+/-11% increase in the number of P selectin- positive platelets in the nonbiotinylated population treated with L-NAME as compared to control. When biotinylated platelets were added to the L-NAME-treated nonbiotinylated population, the number of P selectin positive biotinylated plate-lets increased by 180+/-32% as compared to biotinylated platelets added to the control. In summary, stimulated platelets produce NO that modestly inhibits platelet activation but markedly inhibits additional platelet recruitment. These data suggest that platelet-derived NO may regulate platelet recruitment to a growing thrombus.

Vascular incorporation of alpha-tocopherol prevents endothelial dysfunction due to oxidized LDL by inhibiting protein kinase C stimulation.

Excess vascular oxidative stress has been linked to impaired endothelium-dependent arterial relaxation in hypercholesterolemia. alpha-Tocopherol (AT) preserves endothelial function in hypercholesterolemia although the mechanism(s) for this protective effect is (are) not known. We examined the tissue-specific effects of AT on oxidized LDL (ox-LDL)-mediated endothelial dysfunction in male New Zealand White rabbits. Animals consumed chow deficient in (< 10 IU/kg) or supplemented with (1,000 IU/kg) AT for 28 d. Exposure of thoracic aortae from AT-deficient animals to ox-LDL (0-500 microg/ml) for 4 h produced dose-dependent inhibition of acetylcholine-mediated relaxation (P < 0.05) while vessels derived from animals consuming AT were resistant to ox-LDL-mediated endothelial dysfunction. Animals consuming AT demonstrated a 100-fold increase in vascular AT content and this was strongly correlated with vessel resistance to endothelial dysfunction from ox-LDL (R = 0.67; P = 0.0014). These results were not explained by an effect of AT on ox-LDL-mediated cytotoxicity by LDH assay or scanning electron microscopy. Vascular incorporation of AT did produce resistance to endothelial dysfunction from protein kinase C stimulation, an event that has been implicated in the vascular response to ox-LDL. Human aortic endothelial cells loaded with AT also demonstrated resistance to protein kinase C stimulation by both phorbol ester and ox-LDL. Thus, these data indicate that enrichment of vascular tissue with AT protects the vascular endothelium from ox-LDL-mediated dysfunction, at least in part, through the inhibition of protein kinase C stimulation. These findings suggest one potential mechanism for the observed beneficial effect of AT in preventing the clinical expression of coronary artery disease that is distinct from the antioxidant protection of LDL.

Role of nitric oxide in parasympathetic modulation of beta-adrenergic myocardial contractility in normal dogs.

In vitro studies indicate that muscarinic cholinergic inhibition of beta-adrenergic cardiac responses may be modulated in part by nitric oxide (NO). To evaluate the role of NO in parasympathetic inhibition of the beta-adrenergic contractile response in vivo, we assessed the inotropic response to dobutamine before and during bilateral vagus nerve stimulation in closed-chest dogs. Dobutamine administration and vagal stimulation were repeated during intracoronary infusion of the NO synthase inhibitor NG-monomethyl-L-arginine (L-NMMA, 10 mumol/min) and again following infusion of L-arginine (100 mg/kg). In eight dogs, intracoronary dobutamine infusion at rates of 25 and 50 micrograms/min increased peak +dP/dt by 131 +/- 24 and 168 +/- 22%, respectively (P < 0.0001). Vagal stimulation (2.5 Hz) attenuated the responses to dobutamine (25 and 50 micrograms/min) by 23 +/- 4 and 21 +/- 4%, respectively (P < 0.001). L-NMMA reduced (by 44-62%; P < 0.001) and L-arginine restored vagal inhibition of the dobutamine-stimulated inotropic response. In a second group of nine dogs, dobutamine was administered systemically to assure a constant concentration in the coronary circulation. Vagal stimulation (2.5 Hz) attenuated the dobutamine-stimulated inotropic response (2.5 and 5.0 micrograms/kg per min) by 40 +/- 12% and 57 +/- 8%, respectively (P < 0.004). As with intracoronary dobutamine, L-NMMA diminished and L-arginine restored vagal inhibition of the inotropic response to dobutamine. Intracoronary infusion of atropine (12 micrograms/min) abolished the vagal inhibitory effect, and intracoronary infusion of 8-bromo-cyclic GMP (1 and 10 mM) caused a dose-dependent attenuation of the dobutamine-stimulated increase in +dP/dt. These data suggest that NO mediates, at least in part, vagal inhibition of the inotropic response to beta-adrenergic stimulation by dobutamine, and thus may play a role in normal physiologic regulation of myocardial autonomic responses.

In vivo transfer of nitric oxide between a plasma protein-bound reservoir and low molecular weight thiols.

Plasma albumin reacts with nitric oxide (NO) to form the bioactive adduct, S-nitroso-albumin (S-NO-albumin). The limited intracellular access of S-NO-albumin suggests the need for a vascular transfer mechanism of NO from a large plasma S-NO-albumin pool to effect biologic function. To study the role of low molecular weight (LMW) thiols in NO transfer in vivo, we administered intravenous S-NO-albumin (1-300 nmol/kg) to rabbits before and after an intravenous infusion of L-cysteine or N-acetyl-L-cysteine. S-NO-albumin produced dose-dependent hypotension that was significantly augmented by prior infusion of either LMW thiol. LMW thiol infusion significantly accelerated the rate of onset and reduced the duration of action of the hypotension induced by S-NO-albumin. The hemodynamic effects of S-NO-albumin after pretreatment with LMW thiols were mimicked by administration of the corresponding LMW S-nitrosothiol. The transfer of NO from albumin to L-cysteine was directly measured in rabbit plasma using a novel technique that couples high performance liquid chromatography to electrochemical detection. These data demonstrate that NO exchange between plasma protein thiol-bound NO and available LMW thiol pools (transnitrosation) occurs in vivo.

Inhibition of neointimal proliferation in rabbits after vascular injury by a single treatment with a protein adduct of nitric oxide.

Endothelium-derived relaxing factor is important for vascular homeostasis and possesses qualities that may modulate vascular injury, including vasodilation, platelet inhibition, and inhibition of smooth muscle proliferation. S-nitrososerum albumin is a naturally occurring adduct of nitric oxide (NO) with a prolonged biologic half-life and is a potent vasodilator and platelet inhibitor. Given the avidity of serum albumin for subendothelial matrix and the antiproliferative effects of NO, we investigated the effects of locally delivered S-nitroso-bovine serum albumin (S-NO-BSA) and a polythiolated form of bovine serum albumin (pS-BSA) modified to carry several S-nitrosothiol groups (pS-NO-BSA) on neointimal responses in an animal model of vascular injury. Locally delivered S-NO-BSA bound preferentially to denuded rabbit femoral vessels producing a 26-fold increase in local concentration compared with uninjured vessels (P = 0.029). pS-NO-BSA significantly reduced the intimal/medial ratio (P = 0.038) and did so in conjunction with elevations in platelet (P < 0.001) and vascular cGMP content (P < or = 0.001). pS-NO-BSA treatment also inhibited platelet deposition (P = 0.031) after denuding injury. Comparison of BSA, S-NO-BSA, pS-NO-BSA, and control revealed a dose-response relationship between the amount of displaceable NO delivered and the extent of inhibition of neointimal proliferation at 2 wk (P < or = 0.001). Local administration of a stable protein S-nitrosothiol inhibits intimal proliferation and platelet deposition after vascular arterial balloon injury. This strategy for the local delivery of a long-lived NO adduct has potential for preventing restenosis after angioplasty.

Dietary probucol preserves endothelial function in cholesterol-fed rabbits by limiting vascular oxidative stress and superoxide generation.

Excess vascular oxidative stress and the local formation of oxidized LDL (ox-LDL) have been implicated in the development of impaired endothelium-dependent arterial relaxation in hypercholesterolemia and atherosclerosis. Dietary antioxidants limit LDL oxidation in vitro and treatment of cholesterol-fed rabbits with dietary antioxidants preserves endothelium-derived relaxing factor (EDRF) action. To investigate the mechanism(s) responsible for these observations, we examined EDRF action, vascular oxidative stress, and antioxidant protection in male New Zealand White rabbits using four dietary treatments. Animals consumed standard chow (chow group) or chow supplemented with: (a) 0.5% cholesterol (0.5% cholesterol group); (b) 1% cholesterol (1% cholesterol group); or (c) 1% cholesterol and 1% probucol (probucol group). After 28 d of dietary treatment, segments of thoracic aorta from the 0.5 and 1% cholesterol groups demonstrated impairment of acetylcholine-mediated endothelium-dependent arterial relaxation compared to chow-fed animals (57 +/- 11% and 45 +/- 9% vs 78 +/- 3%, respectively; P < 0.05). In contrast, vessels from the probucol group demonstrated normal relaxation to acetylcholine (83 +/- 5%). Plasma cholesterol levels and the extent of atherosclerosis were similar among all cholesterol-fed groups. Probucol treatment was associated a threefold increase in LDL resistance to copper-induced oxidative modification (P < 0.05) and a reduction in tissue lipid peroxidation (as assessed by thiobarbituric acid-reactive substances; P < 0.05) compared to animals fed cholesterol alone. Most importantly, both of these changes were strongly correlated with preserved EDRF action. Moreover, cholesterol feeding was associated with a dose-dependent increase in vascular superoxide generation and lysophosphatidylcholine content, both of which were prevented by probucol treatment. From these findings, we conclude that probucol, a lipid-soluble antioxidant, preserves EDRF action in cholesterol-fed rabbits in association with limiting vascular oxidative stress and superoxide generation.

NO forms an adduct with serum albumin that has endothelium-derived relaxing factor-like properties.

Recent evidence suggests that sulfhydryl species can react with oxides of nitrogen under physiologic conditions and thereby stabilize endothelium-derived relaxing factor (EDRF) activity, but the presence of a specific in vivo thiol carrier for nitric oxide (NO) remains controversial. The single free sulfhydryl of serum albumin is the most abundant thiol species in plasma (approximately 0.5 mM) and is particularly reactive towards NO. To examine the potential role of serum albumin in endogenous nitric oxide metabolism, we synthesized S-nitroso-BSA (S-NO-BSA), a model S-nitroso-protein, and examined its effects on platelet function and coronary and systemic vascular tone in 16 mongrel dogs. Intravenous bolus S-NO-BSA markedly reduced mean arterial pressure in a dose-dependent manner and proved seven and a half-fold less potent than intravenous nitroglycerin and 10-fold less potent than intravenous S-nitroso-cysteine (half-maximal response of 75 nmol/kg compared to 10 and 7.5 nmol/kg, respectively; P < 0.05); when given by intravenous infusion (half-maximal response = 10 nmol/kg per min), however, S-NO-BSA and nitroglycerin were equipotent. Intravenous bolus S-NO-BSA had a greater duration of action than either nitroglycerin or S-nitroso-cysteine and produced marked prolongation of the template bleeding time associated with dose-dependent inhibition of ex vivo platelet aggregation (half-maximal response approximately 70 nmol/kg). Intracoronary S-NO-BSA increased coronary blood flow (mean +/- SEM) less effectively than nitroprusside, acetylcholine, or S-nitroso-cysteine (165% +/- 24% vs. 315% +/- 82%, 483% +/- 55%, or 475% +/- 66%, respectively; P < 0.05) although with much longer duration of action. On a molar basis, S-nitroso-cysteine proved more effective than S-nitroso-BSA, nitroprusside, or acetylcholine as an epicardial coronary vasodilator. Thus, serum albumin reacts with oxides of nitrogen to form a stable S-nitroso-thiol with properties reminiscent of authentic EDRF supporting the view that protein associated thiol may participate in the action and metabolism of EDRF.

L-2-Oxothiazolidine-4-carboxylic acid reverses endothelial dysfunction in patients with coronary artery disease.

The effective action of endothelium-derived nitric oxide (EDNO) is impaired in patients with atherosclerosis. This impairment has been attributed in part to increased vascular oxidative stress. EDNO action is improved by administration of ascorbic acid, a water-soluble antioxidant. Ascorbic acid is a potent free-radical scavenger in plasma, and also regulates intracellular redox state in part by sparing cellular glutathione. We specifically investigated the role of intracellular redox state in EDNO action by examining the effect of L-2-oxo-4-thiazolidine carboxylate (OTC) on EDNO-dependent, flow-mediated dilation in a randomized double-blind placebo-controlled study of patients with angiographically proven coronary artery disease. OTC augments intracellular glutathione by providing substrate cysteine for glutathione synthesis. Brachial artery flow-mediated dilation was examined with high-resolution ultrasound before and after oral administration of 4.5 g of OTC or placebo in 48 subjects with angiographically documented coronary artery disease. Placebo treatment produced no change in flow-mediated dilation (7.0+/-3.9% vs. 7.2+/-3.7%), whereas OTC treatment was associated with a significant improvement in flow-mediated dilation (6.6+/-4.4% vs. 11.0+/-6.3%; P = 0.005). OTC had no effect on arterial dilation to nitroglycerin, systemic blood pressure, heart rate, or reactive hyperemia. These data suggest that augmenting cellular glutathione levels improves EDNO action in human atherosclerosis. Cellular redox state may be an important regulator of EDNO action, and is a potential target for therapy in patients with coronary artery disease.

Low-dose alpha-tocopherol improves and high-dose alpha-tocopherol worsens endothelial vasodilator function in cholesterol-fed rabbits.

Abnormalities in endothelium-dependent arterial relaxation develop early in atherosclerosis and may, in part, result from the effects of modified low-density lipoprotein (LDL) on agonist-mediated endothelium-derived relaxing factor (EDRF) release and EDRF degradation. alpha-Tocopherol (AT) is the main lipid-soluble antioxidant in human plasma and lipoproteins, therefore, we investigated the effects of AT on endothelium-dependent arterial relaxation in male New Zealand White rabbits fed diets containing (a) no additive (controls), (b) 1% cholesterol (cholesterol group), or 1% cholesterol with either (c) 1,000 IU/kg chow AT (low-dose AT group) or (d) 10,000 IU/kg chow AT (high-dose AT group). After 28 d, we assayed endothelial function and LDL susceptibility to ex vivo copper-mediated oxidation. Acetylcholine-and A23187-mediated endothelium-dependent relaxations were significantly impaired in the cholesterol group (P < 0.001 vs. control), but preserved in the low-dose AT group (P = NS vs. control). Compared to the control and cholesterol groups, vessels from the high-dose AT group demonstrated profound impairment of arterial relaxation (P < 0.05) and significantly more intimal proliferation than other groups (P < 0.05). In normal vessels, alpha-tocopherol had no effect on endothelial function. LDL derived from both the high- and low-dose AT groups was more resistant to oxidation than LDL from control animals (P < 0.05). These data indicate that modest dietary treatment with AT preserves endothelial vasodilator function in cholesterol-fed rabbits while a higher dose of AT is associated with endothelial dysfunction and enhanced intimal proliferation despite continued LDL resistance to ex vivo copper-mediated oxidation.

Insulin resistance, oxidative stress, hypertension, and leukocyte telomere length in men from the Framingham Heart Study.

Insulin resistance and oxidative stress are associated with accelerated telomere attrition in leukocytes. Both are also implicated in the biology of aging and in aging-related disorders, including hypertension. We explored the relations of leukocyte telomere length, expressed by terminal restriction fragment (TRF) length, with insulin resistance, oxidative stress and hypertension. We measured leukocyte TRF length in 327 Caucasian men with a mean age of 62.2 years (range 40-89 years) from the Offspring cohort of the Framingham Heart Study. TRF length was inversely correlated with age (r = -0.41, P < 0.0001) and age-adjusted TRF length was inversely correlated with the Homeostatic Model Assessment of Insulin Resistance (HOMA-IR) (r =-0.16, P = 0.007) and urinary 8-epi-PGF(2alpha) (r = -0.16, P = 0.005) - an index of systemic oxidative stress. Compared with their normotensive peers, hypertensive subjects exhibited shorter age-adjusted TRF length (hypertensives = 5.93 +/- 0.042 kb, normotensives = 6.07 +/- 0.040 kb, P = 0.025). Collectively, these observations suggest that hypertension, increased insulin resistance and oxidative stress are associated with shorter leukocyte telomere length and that shorter leukocyte telomere length in hypertensives is largely due to insulin resistance.

Chemiosmotic lysis and insulin secretion: studies of isolated granules, intact and permeabilised rat islets of Langerhans.

The possible involvement of chemiosmotic lysis of secretory granules in the exocytosis of insulin from pancreatic beta cells was investigated by comparing insulin release from isolated secretory granules, from intact islets of Langerhans, and from electrically permeabilised islets. Lysis of isolated granules was stimulated by ATP in the presence of Mg2+. ATP-induced granule lysis was pH and temperature dependent and was inhibited by collapsing the pH gradient across the granule membrane by removal of permeant anions, or by increasing the extragranular osmolarity. However, insulin secretion from intact islets in response to glucose, a phosphodiesterase inhibitor or a Ca2+ ionophore was only partially inhibited by anion replacement, while Ca2+ -induced insulin release from electrically permeabilised islets was not affected by altering the extragranular or intragranular pH. These results suggest that studies of the stability of isolated granules in vitro do not necessarily relate to insulin release from whole cells, and do not support a major role for chemiosmotic lysis of secretory granules in the exocytotic release of insulin.

Long-term ascorbic acid administration reverses endothelial vasomotor dysfunction in patients with coronary artery disease.

BACKGROUND: Loss of endothelium-derived nitric oxide (EDNO) contributes to the clinical expression of coronary artery disease (CAD). Increased oxidative stress has been linked to impaired endothelial vasomotor function in atherosclerosis, and recent studies demonstrated that short-term ascorbic acid treatment improves endothelial function. METHODS AND RESULTS: In a randomized, double-blind, placebo-controlled study, we examined the effects of single-dose (2 g PO) and long-term (500 mg/d) ascorbic acid treatment on EDNO-dependent flow-mediated dilation of the brachial artery in patients with angiographically established CAD. Flow-mediated dilation was examined by high-resolution vascular ultrasound at baseline, 2 hours after the single dose, and 30 days after long-term treatment in 46 patients with CAD. Flow-mediated dilation improved from 6.6+/-3.5% to 10.1+/-5.2% after single-dose treatment, and the effect was sustained after long-term treatment (9. 0+/-3.7%), whereas flow-mediated dilation was 8.6+/-4.7% at baseline and remained unchanged after single-dose (7.8+/-4.4%) and long-term (7.9+/-4.5%) treatment with placebo (P=0.005 by repeated-measures ANOVA). Plasma ascorbic acid concentrations increased from 41.4+/-12. 9 to 115.9+/-34.2 micromol/L after single-dose treatment and to 95. 0+/-36.1 micromol/L after long-term treatment (P<0.001). CONCLUSIONS: In patients with CAD, long-term ascorbic acid treatment has a sustained beneficial effect on EDNO action. Because endothelial dysfunction may contribute to the pathogenesis of cardiovascular events, this study indicates that ascorbic acid treatment may benefit patients with CAD.

Iron chelation improves endothelial function in patients with coronary artery disease.

BACKGROUND: Some epidemiological studies have shown that increased iron stores are associated with increased cardiovascular events. Redox-active iron may contribute to lipid peroxidation, endothelial cell activation, and generation of reactive oxygen species (especially hydroxyl radical, via Fenton chemistry). Increased oxidative stress is associated with impaired action of endothelium-derived nitric oxide in patients with atherosclerosis. METHODS AND RESULTS: To test the hypothesis that reducing vascular iron stores would reverse endothelial dysfunction, we examined the effects of the iron chelator deferoxamine (500 mg intra-arterially over 1 hour) on vasomotor function in forearm resistance vessels of patients with coronary artery disease by venous occlusion plethysmography. Patients with coronary artery disease had impaired endothelium-dependent vasodilation in response to methacholine compared with healthy control subjects (P<0.001). Deferoxamine infusion decreased serum iron levels (P<0.001). Deferoxamine improved the blood flow response to methacholine in patients with coronary artery disease (P<0.01 by 2-way repeated-measures ANOVA) but had no effect on the response to sodium nitroprusside. In normal volunteers, deferoxamine had no effect on the response to methacholine. The nitric oxide synthase inhibitor N(G)-monomethyl-L-arginine abolished augmentation of the methacholine response associated with deferoxamine. The hydroxyl radical scavenger mannitol had no effect on the methacholine response. CONCLUSIONS: Deferoxamine improved nitric oxide-mediated, endothelium-dependent vasodilation in patients with coronary artery disease. These results suggest that iron availability contributes to impaired nitric oxide action in atherosclerosis.

Low plasma ascorbic acid independently predicts the presence of an unstable coronary syndrome.

OBJECTIVES: This study sought to investigate the relations between plasma antioxidant status, extent of atherosclerosis and activity of coronary artery disease. BACKGROUND: Previous studies indicate that increased antioxidant intake is associated with decreased coronary disease risk, but the underlying mechanisms remain controversial. METHODS: Plasma samples were obtained from 149 patients undergoing cardiac catheterization (65 with stable angina, 84 with unstable angina or a myocardial infarction within 2 weeks). Twelve plasma antioxidant/oxidant markers were measured and correlated with the extent of atherosclerosis and the presence of an unstable coronary syndrome. RESULTS: By multiple linear regression analysis, age (p < 0.001), diabetes mellitus (p < 0.001), male gender (p < 0.001) and hypercholesterolemia (p = 0.02) were independent predictors of the extent of atherosclerosis. No antioxidant/oxidant marker correlated with the extent of atherosclerosis. However, lower plasma ascorbic acid concentration predicted the presence of an unstable coronary syndrome by multiple logistic regression (odds ratio [OR] 0.59, 95% confidence interval [CI] 0.40 to 0.89, p = 0.01). The severity of atherosclerosis also predicted the presence of an unstable coronary syndrome (OR 1.7, 95% CI 1.14 to 2.47, p = 0.008) when all patients were considered. When only patients with significant coronary disease were considered (at least one stenosis >50%), ascorbic acid concentration (OR 0.56, 95% CI 0.37 to 0.85, p = 0.008) and total plasma thiols (OR 0.52, 95% CI 0.34 to 0.80, p = 0.004) predicted the presence of an unstable coronary syndrome, whereas the extent of atherosclerosis did not. CONCLUSIONS: These data are consistent with the hypothesis that the beneficial effects of antioxidants in coronary artery disease may result, in part, by an influence on lesion activity rather than a reduction in the overall extent of fixed disease.

Coronary artery perforation during excimer laser coronary angioplasty. The percutaneous Excimer Laser Coronary Angioplasty Registry.

OBJECTIVES: The aim of this study was to analyze the risk of vessel perforation during excimer laser angioplasty. BACKGROUND: Vessel perforation is a serious complication of angioplasty. METHODS: A total of 764 patients had 858 stenoses treated with excimer laser angioplasty. Laser catheters had a diameter of 1.4, 1.7 or 2 mm. Laser energy was delivered in pulses of 135 ns, at a frequency of 25 s-1 and at a fluence of 30 to 60 mJ/mm2. Follow-up angiography was requested for all patients who did not require emergency bypass surgery. RESULTS: In the 764 consecutive patients treated with excimer laser coronary angioplasty, vessel perforation occurred in 23 patients (3%). Nine patients had a major complication resulting directly from vessel perforation (cardiac tamponade, myocardial infarction or need for bypass surgery) and 14 had no clinical complications after successful sealing of the puncture site. No patient with a perforation died. Multivariate analysis showed that bifurcation lesions (odds ratio [OR] = 3.5; p = 0.049), diabetes mellitus (OR = 3.15; p = 0.029) and female gender (OR = 2.86; p = 0.013) were associated with an increased risk of vessel perforation. Lesions > 10 mm in length (OR = 0.45; p = 0.206), calcified stenoses (OR = 0.26; p = 0.088) and saphenous vein graft lesions (OR = 0.50; p = 0.295) were not at increased risk. Vessel perforation was seen in 10 (8.3%) of 120 lesions in which the laser catheter was equivalent in diameter to the target vessel (< or = 0.5 mm smaller in size) but in only 8 (1.5%) of 525 lesions in which the laser catheter was > 1 mm smaller than the target vessel (p = 0.001). CONCLUSIONS: Most lesions thought to be suitable for excimer laser treatment are not at increased risk of perforation. The complication may be avoided by improved patient and laser catheter size selection.