Hyperoxaemia does not change concentrations of serotonin and beta-thromboglobulin in blood of healthy humans

BACKGROUND: The mechanisms of oxygen-induced effects on blood vessels (vasoconstriction in hyperoxaemia and vasodilatation during hypoxaemia) are uncertain. Many investigators have suggested that the vasoconstriction seen during hyperoxia/hyperoxaemia is mediated through the endothelium as a result of either increased release or activity of vasoconstrictors (oxygen radicals, endothelin, norepinephrine, angiotensin II, or serotonin (5-HT)), or reduced activity of vasodilators (prostaglandin E2 and nitric oxide). Serotonin has been assumed to have a central role. METHODS: Eight healthy volunteers were exposed to FiO2 of 1.0 for 20 min and serum concentrations of serotonin and activated platelets were measured (indicated by concentrations of beta-thromboglobulin (beta-TG)). RESULTS: During hyperoxaemia in humans, serum concentrations of scrotonin and beta-TG remained unchanged. CONCLUSION: If serotonin is involved in oxygen-induced vasoconstriction, the mechanism is more likely to be either a potentiating effect of serotonin on other vasoconstrictors or increased activity of serotonin on its receptor.     

Effect of the selective serotonin reuptake inhibitor paroxetine on platelet function is modified by a SLC6A4 serotonin transporter polymorphism

Summary. Background: Selective serotonin reuptake inhibitors (SSRIs) have been associated with an increased bleeding tendency. Objectives: To prospectively quantify the dose‐response effects of paroxetine and the influence of the serotonin transporter gene (SLC6A4) promoter polymorphism (5‐HTTLPR) on platelet function. Methods: Nineteen drug‐free psychiatric outpatients (44.5 ± 10.8 years) were tested before and after 6 weeks of paroxetine treatment (20 mg day^?1). Based on clinical symptoms, paroxetine dosages were increased (40–50 mg day^?1) for 6 more weeks in 11 patients. Parameters related to platelet function were assessed by bleeding time, platelet function analyzer (PFA), platelet serotonin, platelet factor 4 (PF4), β‐thromboglobulin (β‐TG), and aggregation tests. Results: Paroxetine 20 mg day^?1 increased mean bleeding time by 1.2 min (95% confidence interval (95% CI) ?0.2–2.7) and reduced median platelet serotonin level (463 ng 10^?9 platelets; inter quartile range (IQR) 361–666), and platelet ß‐TG concentration (3.1 IU 10^?6 platelets; IQR 0.3–6.0). Other platelet parameters did not change significantly. Serial platelet aggregation tests did not become abnormal. Paroxetine dose‐escalation did not further influence platelet function. However, 5‐HTTLPR polymorphisms modified these effects: in L_A/L_A‐carriers, bleeding times did not change (?0.2 min; 95% CI ?0.6 to 0.9), while bleeding times significantly increased in <2L_A‐allele carriers (2.3 min; 95% CI 0.5 to 4.07; P = 0.032). Platelet serotonin decreases were larger in patients without L_A‐alleles (868 ng 10^?9 platelets; IQR 585 to 1213) than in ≥1 L_A‐allele carriers (457 ng 10^?9 platelets; IQR 392 to 598; P = 0.035). PFA closure time and PF4 increased significantly in patients without L_A‐alleles. Conclusions: Paroxetine 20 mg day^?1 does not increase overall bleeding time, but impairs platelet function by decreasing the levels of platelet serotonin and platelet ß‐TG. These paroxetine effects appear to be mediated by 5‐HTTLPR, with most pronounced effects in patients without L_A‐alleles.     

Effect of 12‐month statin therapy on antioxidant potential of LDL and serum antioxidant vitamin concentrations

BACKGROUND. Statins are known to cause short‐term reduction in serum lipid‐soluble antioxidant concentrations, but their long‐term effects are not known.

AIM: We randomised 104 subjects with CHD and hypercholesterolaemia to receive either atorvastatin or simvastatin treatment for 52 weeks and measured the antioxidant potential of LDL and serum antioxidant vitamin concentrations.

METHOD: Initial daily dose for both statins was 20?mg.

RESULTS: LDL antioxidant capacity and serum α‐tocopherol, γ‐tocopherol and β‐carotene concentrations decreased by 22%–35% with both statins during the first 12 weeks' therapy ( P ?<?0.01 for all). After 52 weeks' therapy, the concentrations of serum γ‐tocopherol in the simvastatin group and serum β‐carotene in both treatment groups returned to baseline levels, while the concentrations of serum γ‐tocopherol in the atorvastatin group and LDL antioxidant capacity and serum α‐tocopherol in both treatment groups remained reduced ( P ?<?0.001 for all). The LDL antioxidant capacity:LDL‐cholesterol and the serum α‐tocopherol:LDL‐cholesterol ratios were significantly elevated with both statins after 12 and 52 weeks ( P ?<?0.001 for all). Statistically significant increases were also observed for corresponding ratios of the less abundant vitamins γ‐tocopherol and β‐carotene.

CONLUSIONS: Some of the decreases in serum lipid soluble antioxidant vitamins reported in short‐term statin interventions may become attenuated when therapy continues longer. The relative antioxidant capacity of LDL particles increased during the 52‐week treatment, suggesting that the oxidation resistance of LDL particles did not become impaired and that their atherogenicity did not increase.     

ITAM receptor‐mediated generation of reactive oxygen species in human platelets occurs via Syk‐dependent and Syk‐independent pathways

Background: Ligation of the platelet‐specific collagen receptor, GPVI/FcRγ, causes rapid, transient disulfide‐dependent homodimerization, and the production of intracellular reactive oxygen species (ROS) generated by the NADPH oxidase, linked to GPVI via TRAF4. Objectives: The aim of this study was to evaluate the role of early signaling events in ROS generation following engagement of either GPVI/FcRγ or a second immunoreceptor tyrosine‐based activation motif (ITAM)‐containing receptor on platelets, FcγRIIa. Methods and Results: Using an H₂DCF‐DA‐based flow cytometric assay to measure intracellular ROS, we show that treatment of platelets with either the GPVI agonists, collagen‐related peptide (CRP) or convulxin (Cvx), or the FcγRIIa agonist 14A2, increased intraplatelet ROS; other platelet agonists such as ADP and TRAP did not. Basal ROS in platelet‐rich plasma from 14 healthy donors displayed little inter‐individual variability. CRP, Cvx or 14A2 induced an initial burst of ROS within 2 min followed by additional ROS reaching a plateau after 15–20 min. The Syk inhibitor BAY61‐3606, which blocks ITAM‐dependent signaling, had no effect on the initial ROS burst, but completely inhibited the second phase. Conclusions: Together, these results show for the first time that ROS generation downstream of GPVI or FcγRIIa consists of two distinct phases: an initial Syk‐independent burst followed by additional Syk‐dependent generation.     

MAP Kinase cross talks in oxidative stress‐induced impairment of insulin secretion. Involvement in the protective activity of quercetin

Insulin secretion preservation is a major issue for the prevention or treatment of type 2 diabetes. We previously showed on β‐cells that quercetin (Q), but not resveratrol (R) or N‐acetyl cysteine (NAC), amplified glucose‐induced insulin secretion in a calcium‐ and ERK1/2‐dependent manner. Quercetin, but not resveratrol or NAC, also protected β‐cell function and hyperamplified ERK1/2 phosphorylation in oxidative stress conditions. As quercetin may interfere with other stress‐activated protein kinases (JNK and p38 MAPK), we further explored MAPK cross talks and their relationships with the mechanism of the protective effect of quercetin against oxidative stress. In INS‐1 insulin‐secreting β‐cells, using pharmacological inhibitors of MAPK pathways, we found that under oxidative stress (50 μm H₂O₂) and glucose‐stimulating insulin secretion conditions: (i) p38 MAPK phosphorylation was increased and regulated by ERK1/2 (positively) and JNK (negatively), although p38 MAPK activation did not seem to play any significant role in oxidative stress‐induced insulin secretion impairment; (ii) the JNK pathway appeared to inhibit both ERK1/2 activation and insulin secretion, although JNK phosphorylation was not significantly changed in our experimental conditions; (iii) the functionality of β‐cell in the presence of oxidative stress was closely linked to the level of ERK1/2 activation, (iv) quercetin, resveratrol, or NAC inhibited H₂O₂‐induced p38 MAPK phosphorylation. The preservation of β‐cell function against oxidative stress appears dependent on the balance between ERK1/2 and JNK activation. The protecting effect of quercetin appears due to ERK1/2 hyperactivation, possibly induced by L‐type calcium channel opening as we recently showed (Br. J. Pharmacol. 2013, 169, 1102–1113).     

Hyperosmolar glucose induces vasoconstriction through Rho/Rho‐kinase pathway in the rat aorta

Rho/Rho‐kinase signalling pathway plays a substantial role in vascular contractions. In this study, we investigated any roles of Rho/Rho‐kinase pathway in the vasoconstriction of the rat conductance and capacitance vessels by hyperosmolar glucose solution. Isolated aortic, mesenteric and renal rings were suspended and exposed to hyperosmolar glucose, sucrose and NaCl in the organ chambers filled with Krebs solution gassed with 95% O₂ and 5% CO₂ and maintained at 37 °C. The effect of a Rho‐kinase inhibitor, (+)‐(R)‐trans‐4‐(1‐aminoethyl)‐N‐(4‐pyridyl) cyclohexanecarboxamide dihydrochloride monohydrate (Y‐27632, 10^?5 m ), was tested on the contraction induced by hypertonic solutions. Endothelial integrity was also assessed after hyperosmolar glucose exposure. Moreover, the activity and expression of Rho‐kinase (ROCK‐2) as well as RhoA translocation were detected by Western blotting and enzyme‐linked immunosorbent assay‐based RhoA activity detection method detection kit. The vessels produced substantial contractions in response to hyperosmolar solutions. Y‐27632 significantly reduced hyperosmolarity‐induced vasoconstrictions (P < 0.05). Phosphorylation of myosin‐phosphatase target 1 increased after hyperosmolar glucose exposure, and this phosphorylation was significantly decreased by Y‐27632 (P < 0.05) in the aorta. Furthermore, RhoA translocation but not ROCK‐2 expression markedly increased by hyperosmolar glucose solution. These results may indicate that hyperosmolarity could induce vasoconstriction through Rho/Rho‐kinase signalling.     

Serotonin stimulates platelet receptor shedding by tumor necrosis factor-alpha-converting enzyme (ADAM17)

Summary. Background: Peripheral serotonin (5‐hydroxytryptamine, 5‐HT) is transported by platelets and released upon stimulation. In the platelet cytoplasm, 5‐HT is transamidated to small GTPases, promoting α‐granule release and primary hemostasis. Objective: We hypothesized that 5‐HT could also stimulate platelet receptor shedding after binding to the membrane 5‐HT receptor (5‐HT2AR). Methods: Western blot and flow cytometry were used to determine levels of the adhesion receptor glycoprotein (GP)Ibα on platelets or its shed fragment glycocalicin in plasma and serum from wild‐type mice, Tph1^?/? mice lacking peripheral 5‐HT, and mice lacking functional tumor necrosis factor‐alpha‐converting enzyme (TACE, ADAM17). Flow chamber experiments and intravital microscopy were used to examine the adhesive properties of platelets after stimulation of 5‐HT2AR. Results: Glycocalicin was significantly reduced in Tph1^?/? plasma and serum. In isolated platelets, 5‐HT induced shedding of GPIbα, which was increased to 60% when 5‐HT uptake was inhibited by the selective serotonin reuptake inhibitor fluoxetine. Specific 5‐HT2AR agonism and antagonism suggested activation of this receptor. The shedding could not be induced in TACE^ΔZn/ΔZn platelets, suggesting that activated TACE mediated the shedding of GPIbα. Intracellular signaling involved phosphorylation of p38 mitogen‐activated protein kinase rather than G‐protein signaling. 5‐HT2AR stimulation decreased platelet adhesion to collagen‐bound von Willebrand factor under arterial shear (1500 s^?1) and incorporation into FeCl₃‐induced thrombi in mesenteric arterioles. Conclusions: Stimulation of 5‐HT2AR on platelets induces TACE‐mediated shedding of GPIbα, the key adhesion molecule under high shear conditions. Our observations demonstrate a new pathway through which 5‐HT could modulate cardiovascular disease.     

Effect of Peripheral Vasoconstriction on Pulse Oximetry

Objective. We tested the hypothesis that peripheral vasodilation has an effect on arterial oxygen saturation measurements by pulse oximetry, independent of temperature. Methods. Study 1 compared finger arterial oxygen saturation values (SpO₂), before and after peripheral vasoconstriction while temperature was kept constant. This was achieved by administering dexmedetomidine (peripheral vasoconstrictor) to 16 volunteers given general anesthesia. Study 2 compared SpO₂ before and after peripheral vasodilation (brachial plexus block) in a neurally denervated left hand and a neurally innervated right hand in ten awake volunteers. In both studies measurements were also made of finger blood volume (an indicator of vasoconstriction) by photoplethysmographic determination of light transmission through a finger (LTF), finger temperature and of hemodynamic variables. Results. In Study 1, systolic blood pressure, SpO₂ and LTF values increased (vasoconstriction) during dexmedetomidine infusion, (P<0.0001 for all) while there were no changes in finger temperature. In Study 2, in the left hand (axillary block), temperature increased by 1.9 ± 1.6 °C (P=0.004), SpO₂ decreased by 2.5 ± 1.0 % (P<0.0001) and LTF values decreased (vasodilation) by 42 ± 8 % (P<0.0001) after axillary block. Simultaneously, the axillary block did not induce any changes in temperature, SpO₂ or LTF values in the neurally innervated right hand. Conclusions. Our results demonstrate that finger pulse oximeter SpO₂ measurements can be affected by peripheral vascular tone independent of temperature. The mechanism for this effect remains speculative and unproven.     

The paradoxical stimulation by a reversible thrombin inhibitor of thrombin generation in plasma measured with thrombinography is caused by α2‐macroglobulin‐thrombin

Summary. Background. Thrombin generation (TG) in plasma can be monitored continuously with a fluorogenic thrombin substrate using calibrated automated thrombinography (CAT). In the presence of low concentrations of a reversible direct thrombin inhibitor (DTI), CAT shows an unexpected effect: the endogenous thrombin potential (ETP) increases at low concentrations of the inhibitor to subsequently decrease concentration dependently at higher concentrations (> approximately 100 nm ). Objectives. To find an explanation for this phenomenon, we measured the concentrations of free thrombin and α₂‐macroglobulin‐thrombin complex (α₂MT) with a sub‐sampling technique in the presence of AR‐H067637, a selective DTI. Results. At all concentrations of the DTI there was a gradual dose‐dependent decrease in the concentration of free, not‐inhibited thrombin but a transient increase in free α₂MT due to competition of thrombin and α₂MT for the inhibitor. Because the CAT technique uses an algorithm to subtract α₂MT activity from the total amidolytic activity, this transient increase in α₂MT activity is not subtracted and erroneously attributed to thrombin itself. Conclusions. This study explains the spurious increase in ETP observed at low DTI concentrations. The results obtained in plasma were corroborated by observations in a thrombin generating system reconstituted with purified factors. In practise, the effect of DTIs on TG can be reliably evaluated from the area under the curve till time‐to‐peak.     

Antispasmodic effects of essential oil of Pterodon polygalaeflorus and its main constituent β‐caryophyllene on rat isolated ileum

This study investigates the effects of essential oil of Pterodon polygalaeflorus (EOPP) and β‐caryophyllene (β‐CAR). EOPP and β‐CAR relaxed the basal tone of ileum smooth muscle in a concentration‐dependent manner (IC₅₀s = 394.35 ± 62.12 and 68.65 ± 9.51 μg/mL respectively), an effect that was unaltered by hexamethonium, L ‐nitroarginine methyl ester or indomethacin. Both EOPP and β‐CAR evoked a concentration‐dependent relaxation of ileum pre‐contracted with KCl with an IC₅₀ value of 107.78 ± 10.47 and 17.35 ± 0.75 μg/mL, respectively. EOPP and β‐CAR inhibited the contractions induced by acetylcholine (ACh) and by KCl. In ileal preparations, the CaCl₂‐induced contractions were reduced by EOPP (300 μg/mL) and β‐CAR (100 μg/mL). Furthermore, CaCl₂‐induced contractions were also reduced by EOPP (300 μg/mL) and β‐CAR (100 μg/mL) in ileal preparations pretreated with ACh under Ca²⁺‐free condition and in the presence of verapamil. EOPP (100 and 300 μg/mL) and β‐CAR (30 and 100 μg/mL) reduced the ACh‐induced contractions of isolated rat ileum under Ca²⁺‐free conditions. In the presence of high KCl and Ca²⁺‐free conditions, EOPP (300 μg/mL) and β‐CAR (100 μg/mL) reduced the contractions induced by barium. A similar effect was also observed with verapamil. It is concluded that (i) β‐CAR is an important constituent involved in the myorelaxant and antispasmodic effects induced by EOPP; (ii) the inhibitory effect on intestinal contractility is myogenic and seems mainly mediated through an intracellular mechanism. However, the ability of EOPP and β‐CAR to decrease Ca²⁺ influx through cytoplasmic membrane could not be discounted.     

Influence of long‐term oxygen therapy on heart rate and QT time‐series in hypoxic patients with chronic obstructive pulmonary disease

Background: There is increasing interest in the cardiovascular pathology independently associated with chronic obstructive pulmonary disease (COPD). We examined the influence of long‐term oxygen therapy (LTOT) on heart rate (RR) and QT time‐series in COPD. Methods: Ten hypoxic stable COPD patients underwent Holter ECG monitoring for 24 h and physical activity/energy expenditure monitoring for 5 days before and after LTOT. Variability of RR and QT time‐series was quantified using standard statistics and their structural (correlation/scaling) properties were assessed using multifractal analysis. Pre‐ and post‐LTOT cardiac/activity parameters were compared to examine the influence of oxygen therapy and circadian variation. Results: PaO₂ increased (P = 0·0004) whilst PaCO₂ was unchanged (P = 0·56) following LTOT. Activity/energy expenditure estimates were also unchanged following LTOT (P = 0·64–0·99), but RR variability was increased during the morning (P < 0·05) and night (P < 0·1, trend only). Multifractality of RR and QT time‐series was not significantly changed following LTOT, although QT multifractality showed some time‐dependent fluctuations. Trends in RR and QT time‐series over 24‐h were similar pre‐ and post‐LTOT, indicating a generally normal circadian response. Conclusions: An increase in HRV following LTOT (but notably in the absence of altered activity levels) provides tentative evidence that LTOT has a direct effect on heart rate control in COPD. This beneficial influence was expressed mainly during the morning, and the relevance of this diurnal variation in response requires further investigation. It was also confirmed that both RR and (to a lesser degree) QT time‐series in COPD have a multifractal structure, and this is not affected appreciably by LTOT.     

Vascular hyporesponsiveness to angiotensin II in rats with CCl4-induced liver cirrhosis

Background Portal hypertension is triggered by vasodilation due to impaired contraction of extrahepatic vessels. Angiotensin II type 1 (AT₁) receptor‐induced vasocontraction is mediated by G proteins and may be desensitized by recruitment of β‐arrestin‐2 to the receptor. In this study, we analysed the interaction of AT₁ receptors with β‐arrestin‐2 in the context of vascular hypocontractility in rats with CCl₄‐induced cirrhosis. Methods Micronodular liver cirrhosis in rats (n = 15) was induced by regular CCl₄ exposure. Age‐matched rats (n = 15) served as controls. Contractility of aortic rings was measured by myography. Protein expressions and phosphorylations were assessed by Western blot analysis, and AT₁ receptor interaction with β‐arrestin‐2 by co‐immunoprecipitation. Results Aortic rings from CCl₄ rats were hypocontractile to angiotensin II independent of nitric oxide synthases (Nω‐nitro‐l ‐arginine methyl ester 200 μM). Expression of the AT₁ receptor, Gα_q/11 and the contraction‐mediating effector Rho kinase was similar in aortas from both groups. Expression and AT₁ receptor binding of β‐arrestin‐2 were up‐regulated in aortas from CCl₄ rats. Stimulation of isolated aortas with exogenous angiotensin II caused recruitment of β‐arrestin‐2 in aortas from noncirrhotic rats, but no further interaction of AT₁ receptors with β‐arrestin‐2 was found in aortas from CCl₄ rats. While angiotensin II stimulation resulted in Rho kinase activation in aortas from noncirrhotic rats but not in aortas from CCl₄ rats, extracellular signal‐regulated kinase activation in response to angiotensin II was observed in aortas from both groups. Conclusions Vascular hyporesponsiveness to angiotensin II in CCl₄ rats is due to enhanced interaction of the AT₁ receptor with β‐arrestin‐2 and consecutively changed receptor function.     

Plasma nitrite concentration decreases after hyperoxia-induced oxidative stress in healthy humans

The aim of this study was to measure plasma nitrite, the biochemical marker of endothelial nitric oxide (^?NO) synthesis, before and after hyperoxia, in order to test the hypothesis that hyperoxia‐induced vasoconstriction is a consequence of reduced bioavailability of ^?NO caused by elevated oxidative stress. Ten healthy men breathed 100% normobaric O₂ for 30 min between 15th and 45th min of the 1‐h study protocol. Plasma nitrite and malondialdehyde (MDA), arterial stiffness (indicated by augmentation index, AIx) and arterial oxygen (P_tcO₂) pressure were measured at 1st, 15th, 45th and 60th minute of the study. Breathing of normobaric 100% oxygen during 30 min caused an increase in P_tcO₂ (from 75 ± 2 to 412 ± 25 mm Hg), AIx (from ?63 ± 4 to ?51 ± 3%) and MDA (from 152 ± 13 to 218 ± 15 nm ) values and a decrease in plasma nitrite (from 918 ± 58 to 773 ± 55 nm ). During the 15‐min recovery phase, plasma nitrite, AIx and MDA values remained altered. This study suggests that the underlying mechanism of hyperoxia‐induced vasoconstriction may involve reduced ^?NO bioavailability caused by elevated and sustained oxidative stress.     

The serial effect of iodinated contrast media on renal hemodynamics and oxygenation as evaluated by ASL and BOLD MRI

Contrast‐induced nephropathy is a prevalent cause of renal failure, and the mechanisms underlying this injury are not fully understood. We utilized noninvasive functional MRI in order to determine the serial effect of a single administration of iodinated contrast media (CM) on renal hemodynamics and oxygenation. Fifteen rabbits were randomized to receive an intravenous injection of CM (i.e. iopamidol‐370; 6 ml kg^?1 body weight) or an equivalent amount of 0.9% saline. Both arterial spin‐labeling and blood oxygen level‐dependent imaging sequences were performed at 24 h before and at intervals of 1, 24, 48 and 72 h after injection to obtain serial renal blood flow (RBF) and relative spin–spin relaxation rate (R₂*). Results showed that, in the iopamidol group, the mean cortical RBF decreased at 1 h (p = 0.04 vs baseline), reached its minimum at 24 h (p = 0.01) and gradually returned to baseline by 48 h (p = nonsignificant, NS). The outer medullary RBF decreased to its minimum by 24 h (p = 0.00) and remained less than baseline until 72 h. R₂* in inner stripes was dramatically increased at 1 h (p = 0.00), remained elevated at 24 h (p = 0.05), but returned to baseline by 48 h (p = NS). R₂* values within the cortex and outer stripes and inner medulla were slightly increased, but the changes did not reach a statistical significance (p = NS). Saline did not produce positive change in either RBF or R₂* within different compartments of the kidney. We conclude that iopamidol is associated with a relatively longer‐term hypoperfusion in whole kidney and decreased oxygen level in the inner stripes of the outer medulla. Copyright © 2012 John Wiley & Sons, Ltd.     

Oxygen tension under hyperbaric conditions in healthy pig brain

Objective: To investigate the effect of hyperbaric conditions on brain oxygenation, intracranial pressure and brain glucose/lactate levels in healthy non‐brain‐traumatized animals. Design and setting: Prospective animal study in a hyperbaric chamber. Subjects: Twelve adult Landrace/Yorkshire pigs. Interventions: The animals were normoventilated in a pressure‐controlled mode according to the open lung concept first at normobaric pressures (FiO₂ of 0·4 and 1·0) and subsequently in the hyperbaric chamber at 1·9 and 2·8 bar (both at an FiO₂ of 1·0). Under these conditions brain oxygen tension and intracranial pressure were recorded and brain glucose/lactate levels were measured by microdialysis. Results: At normobaric conditions, increasing the FiO₂ from 0·4 (baseline) to 1·0 resulted in a significant increase in brain oxygen tension from 33 ± 14 to 63 ± 28 mmHg (P<0·05). Compared with baseline, both hyperbaric conditions (at an FiO₂ of 1·0) led to a significant increase in brain oxygen tension to 151 ± 65 mmHg (P<0·001) at 1·9 bar and to 294 ± 134 mmHg (P<0·001) at 2·8 bar. Conclusions: If there is a need for increased oxygenation in the brain, then one way to achieve this is to apply hyperbaric conditions at 100% oxygen. Compared with an atmospheric pressure with a FiO₂ of 0·4, a nine‐fold increase (900%) in PbrO₂ values can be reached by increasing the FiO₂ to 1·0 and the pressure to 2·8 bar. In this study, hyperbaric oxygen pressure in the brain did not lead to changes in intracranial pressure or in brain glucose/lactate levels.     

Antiapoptotic Cardioprotective Effect of Hypothermia Treatment Against Oxidative Stress Injuries

Objectives: The effect of hypothermia on cardiomyocyte injury induced by oxidative stress remains unclear. The authors investigated the effects of hypothermia on apoptosis and mitochondrial dysfunction in cardiomyocytes exposed to oxidative stress. Methods: Cardiomyocytes (H9c2) derived from embryonic rat heart cell culture were exposed to either normothermic (37°C) or hypothermic (31°C) environments before undergoing oxidative stress via treatment with hydrogen peroxide (H₂O₂). The degree of apoptosis was determined by annexin V and terminal deoxynucleotidyl transferase (TUNEL) staining. The amount of reactive oxygen species (ROS) was compared after H₂O₂ exposure between normo‐ and hypothermic‐pretreated groups. Mitochondrial dysfunction in both groups was measured by differential reductase activity and transmembrane potential (ΔΨm). Results: Hydrogen peroxide induced significant apoptosis in both normothermic and hypothermic cardiomyocytes. Hypothermia ameliorated apoptosis as demonstrated by decreased annexin V staining (33 ± 1% vs. 49 ± 4%; p < 0.05) and TUNEL staining (27 ± 17% vs. 80 ±25%; p < 0.01). The amount of intracellular ROS increased after H₂O₂ treatment and was higher in the hypothermic group than that in the normothermic group (237.9 ± 31.0% vs. 146.6 ± 20.6%; p < 0.05). In the hypothermic group, compared with the normothermic group, after H₂O₂ treatment mitochondrial reductase activity was greater (72.0 ± 17.9% vs. 27.0 ± 13.3%; p < 0.01) and the mitochondria ΔΨm was higher (101.0 ± 22.6% vs. 69.7 ± 12.9%; p < 0.05). Pretreatment of cardiomyocytes with the antioxidant ascorbic acid diminished the hypothermia‐induced increase in intracellular ROS and prevented the beneficial effects of hypothermia on apoptosis and mitochondrial function. Conclusions: Hypothermia at 31°C can protect cardiomyocytes against oxidative stress–induced injury by decreasing apoptosis and mitochondrial dysfunction through intracellular ROS‐dependent pathways.     

Insights into the Effects of Contraction Dyssynchrony on Global Left Ventricular Mechano-Energetic Function

Background: The effects of dyssynchrony on global left ventricular (LV) mechanics have been well documented; however, its impact on LV energetics has received less attention. Objective: To assess the effects of LV contraction dyssynchrony on global LV mechano‐energetic function in a pacing‐induced acute model of dyssynchrony. Methods: Using blood‐perfused isolated rabbit heart preparations (n = 11), LV pressure, coronary flow, and arteriovenous oxygen content difference were recorded for isovolumic contractions under right atrial (RA) pacing (control) and simultaneous RA and right ventricular outflow tract (RVOT) pacing (dyssynchrony). LV mechanical function was quantified by the end‐systolic pressure‐volume relationship (ESPVR). Myocardial oxygen consumption‐pressure‐volume area (MVO₂‐PVA) relationship quantified LV energetic function. Internal PVA for MVO_{2 RVOT} was calculated based on the MVO₂‐PVA relationship for RA pacing. Thus, lost PVA (internal PVA–PVA_RVOT) represents the mechanical energy not observable at the global level. Results: Compared to RA pacing, RVOT pacing depressed LV mechanics as indicated by a rightward shift of ESPVR (i.e., increase in V_d from 0.58 ± 0.10 to 0.67 ± 0.10 mL, P < 0.05). Despite depressed mechanics, RVOT pacing was associated with greater MVO₂ such that the MVO₂‐PVA relationship intercept was markedly increased from 0.025 ± 0.003 to 0.029 ± 0.003 mL?O₂/beat/100gLV (P < 0.05). Excess MVO₂ (i.e., MVO_{2 RVOT}– MVO_{2 RA}) significantly correlated with lost PVA (R²= 0.54, P < 0.001). Conclusion: A potential mechanism explaining the observed increase in MVO₂ with dyssynchrony may be that the measured PVA at the global level underestimates the internal PVA at the cellular level, which is likely to be the true determinant of MVO₂.     

Serial oxygen equilibrium and kinetic measurements during RBC storage

Objectives: To contribute to the understanding of the biochemical changes associated with the RBC storage lesion. Aim: To investigate changes in O₂ equilibrium and on/off kinetic rates during routine cold storage. Background: As RBCs are stored between 1 and 6°C numerous biochemical changes occur within the RBCs, including changes in the properties of the haemoglobin itself. This study serially analysed for the first time the O₂ equilibrium and on/off kinetic rates across the RBC membrane during routine storage. Methods/Materials: The oxygen binding (k_on) and offloading (k_off) constants were measured in fresh RBCs and then in AS‐5‐preserved RBCs at weekly intervals, along with oxygen equilibrium curves (OECs), 2,3‐Diphosphoglycerate (2,3‐DPG), p50 and the Hill number (n). Results: The k_on increased slightly as the 2,3‐DPG and p50 decreased during storage, whereas the k_off remained largely unchanged. The OECs demonstrated the expected increase in O₂ affinity, whereas the Hill number was unchanged during storage. Conclusion: In spite of the biochemical, structural and functional changes associated with the storage of RBCs, their in vitro interactions with oxygen were largely preserved through 42 days of storage.     

Cardiorespiratory, tissue oxygen and hepatic NADH responses to graded hypoxia

Objective: To assess cardiorespiratory, tissue oxygen and hepatic nicotine adenine dinucleotide hydride (NADH) responses to graded hypoxia. Design: Prospective, controlled, randomized study. Setting: University laboratory. Animals and interventions: 18 anaesthetised Sprague-Dawley rats spontaneously breathing either 21 % (controls), 12.5 % or 10 % inspired oxygen concentrations (6 rats per group). Measurements and results: All animals in the 21 and 12.5 % O₂ groups survived the 3-h study period, compared to only 1 in the 10 % O₂ group. In this latter group, mean arterial pressure and renal blood flow fell immediately with hypoxaemia, whereas aortic blood flow was maintained until the preterminal stages. Critical cellular hypoxia was suggested by an increasingly severe base deficit, an initial rise then a preterminal fall in hepatic NADH intensity and premature death in all but 1 animal. Hepatic NADH fluorescence intensity was unchanged in control animals but showed a progressive rise in the 12.5 % O₂ group, accompanied by a small though static increase in arterial base deficit. No significant differences were seen in arterial and tissue partial pressure of oxygen between the 12.5 and 10 % O₂ groups. Conclusions: This study demonstrates major differences in cardiorespiratory, hepatic NADH and outcome responses to small variations in the degree of hypoxic hypoxia. The fall in NADH fluorescence intensity presages impending death and is likely to reflect failure of cellular metabolic processes. Received: 2 April 1998 Accepted: 26 August 1998     

Effects of levosimendan on isolated human internal mammary artery and saphenous vein: concurrent use with conventional vasodilators

Graft spasm is a common problem in coronary artery bypass grafting (CABG). In this study, we aimed to investigate the interaction of levosimendan, a novel inodilator, with vasodilator agents that are clinically used for the treatment of graft spasm and with endogenous vasoconstrictors that are thought to play a role in graft vasospasm, in human internal mammary artery (IMA) and saphenous vein (SV). Isolated human IMA and SV segments derived from patients undergoing CABG were suspended in an organ bath. Responses to cumulative concentrations of noradrenaline (NA), serotonin (5‐HT), papaverine, nitroglycerin (NG), and diltiazem were recorded before and after 10^?5 m levosimendan incubation (30 min). In addition, cumulative levosimendan responses were taken in vessels precontracted with NA or 5‐HT. 10^?5 m levosimendan reduced NA E_max and sensitivity in IMA and SV, and 5‐HT E_max responses in IMA. Moreover, levosimendan caused concentration‐dependent relaxation in both grafts. Papaverine E_max or sensitivity was not altered by levosimendan neither in IMA nor in SV. Levosimendan diminished NG sensitivity in IMA and E_max responses in SV and decreased diltiazem E_max responses both in IMA and SV. Our results suggest that levosimendan may be used alone for prevention or treatment of graft spasm in IMA or in combination with papaverine in IMA and SV grafts. However, as concurrent administration with diltiazem or NG causes a reduction in relaxation in vitro, we suggest caution should be exercised when using levosimendan in combination with these agents.