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Background: We aimed to examine whether using a high fraction of inspired oxygen (FIO 2 ) in the context of an individualised intra- and postoperative open-lung ventilation approach could decrease surgical site infection (SSI) in patients scheduled for abdominal surgery., Methods: We performed a multicentre, randomised controlled clinical trial in a network of 21 university hospitals from June 6, 2017 to July 19, 2018. Patients undergoing abdominal surgery were randomly assigned to receive a high (0.80) or conventional (0.3) FIO 2 during the intraoperative period and during the first 3 postoperative hours. All patients were mechanically ventilated with an open-lung strategy, which included recruitment manoeuvres and individualised positive end-expiratory pressure for the best respiratory-system compliance, and individualised continuous postoperative airway pressure for adequate peripheral oxyhaemoglobin saturation. The primary outcome was the prevalence of SSI within the first 7 postoperative days. The secondary outcomes were composites of systemic complications, length of intensive care and hospital stay, and 6-month mortality., Results: We enrolled 740 subjects: 371 in the high FIO 2 group and 369 in the low FIO 2 group. Data from 717 subjects were available for final analysis. The rate of SSI during the first postoperative week did not differ between high (8.9%) and low (9.4%) FIO 2 groups (relative risk [RR]: 0.94; 95% confidence interval [CI]: 0.59-1.50; P=0.90]). Secondary outcomes, such as atelectasis (7.7% vs 9.8%; RR: 0.77; 95% CI: 0.48-1.25; P=0.38) and myocardial ischaemia (0.6% [n=2] vs 0% [n=0]; P=0.47) did not differ between groups., Conclusions: An oxygenation strategy using high FIO 2 compared with conventional FIO 2 did not reduce postoperative SSIs in abdominal surgery. No differences in secondary outcomes or adverse events were found., Clinical Trial Registration: NCT02776046., (Copyright © 2019 British Journal of Anaesthesia. Published by Elsevier Ltd. All rights reserved.)

In atrial-derived HL-1 cells, ryanodine receptor and Na(+)/Ca(2+)-exchanger were altered early by 5 μM doxorubicin. The observed effects were an increase of cytosolic Ca(2+) at rest, ensuing ryanodine receptor phosphorylation, and the slowing of Ca(2+) transient decay after caffeine addition. Doxorubicin triggered a linear rise of reactive oxygen species (ROS) with no early effect on mitochondrial inner membrane potential. Doxorubicin and ROS were both detected in mitochondria by colocalization with fluorescence probes and doxorubicin-induced ROS was totally blocked by mitoTEMPO. The NADPH oxidase activity in the mitochondrial fraction was sensitive to inhibition by GKT137831, and doxorubicin-induced ROS decreased gradually as the GKT137831 concentration added in preincubation was increased. When doxorubicin-induced ROS was prevented by GKT137831, the kinetic response revealed a permanent degree of protection that was consistent with mitochondrial NADPH oxidase inhibition. In contrast, the ROS induction by doxorubicin after melatonin preincubation was totally eliminated at first but the effect was completely reversed with time. Limiting the source of ROS production is a better alternative for dealing with oxidative damage than using ROS scavengers. The short-term effect of doxorubicin on Ca(2+) transporters involved in myocardiac contractility was dependent on oxidative damage, and so the impairment was subsequent to ROS production., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Increase of resting Ca(2+) levels and amplitude of vasopressin-induced Ca(2+) transients were observed when cells in serum-free medium were exposed to 5mM Ca(2+) for 2h. Small effect on cell viability was also observed. A rapid cytotoxic effect was developed in the presence of 10mM Ca(2+) and absence of serum. However, cells exposed to 10mM Ca(2+) in the presence of serum were protected from damage for at least 2days. Resting Ca(2+) levels and cytosolic Ca(2+) transients in serum-containing medium with 10mM Ca(2+) displayed lower increases and a tendency to recover control values. When serum was absent, cells preincubated with 10mM Ca(2+) were more sensitive to thapsigargin-induced damage than cells preincubated with lower Ca(2+). The sensitivity was similar when serum was present. Tolerance to high Ca(2+) in the presence of serum was linked to potentiation of the mitochondrial Ca(2+) entry to decrease the sarcoplasmic reticulum Ca(2+) overload., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Background: The aim of this study was to evaluate the type and incidence of complications during insertion, maintenance, and withdrawal of central arterial catheters used for transpulmonary thermodilution haemodynamic monitoring (PiCCO™)., Methods: We conducted a prospective, observational, multicentre study in 14 European intensive care units (six countries). A total of 514 consecutive patients in whom haemodynamic monitoring by PiCCO™ was indicated were studied., Results: Five hundred and fourteen PiCCO catheters (475 in femoral, 26 in radial, nine in axillary, and four in brachial arteries) were inserted. Arterial access was obtained on the first attempt in 86.4% of the patients. Minor problems such as oozing after insertion (3.3%) or removal of the catheter (3.5%) were observed, but no episodes of serious bleeding (more than 50 ml) were recorded. Small local haematomas were observed after insertion (4.5%) and after removal (1.2%) of the catheter. These complications were not more frequent in patients with coagulation abnormalities. The incidence of site inflammation and catheter-related infection was 2% and 0.78%, respectively. Other complications such as ischaemia (0.4%), pulse loss (0.4%), or femoral artery thrombosis (0.2%) were rare, transient, and all resolved with catheter removal or embolectomy, respectively., Conclusions: In this series of patients, central arterial catheters used for PiCCO™ monitoring were demonstrated to be a safe alternative for advanced haemodynamic monitoring.

The effect of palytoxin was studied in a microsomal fraction enriched in longitudinal tubules of the sarcoplasmic reticulum membrane. Half-maximal effect of palytoxin on Ca(2+)-ATPase activity yielded an apparent inhibition constant of approx. 0.4 microM. The inhibition process exhibited the following characteristics: (i) the degree of inhibition was dependent on membrane protein concentration; (ii) no protection was observed when the ATP concentration was raised; (iii) dependence on Ca(2+) concentration with a decreased maximum catalytic rate; (iv) it occurred in the absence of Ca(2+) ionophoric activity. Likewise, the inhibition mechanism was linked to: (i) rapid enzyme phosphorylation from ATP in the presence of Ca(2+) but lower steady-state levels of phosphoenzyme; (ii) more drastic effect on phosphoenzyme levels when the toxin was added to the enzyme in the absence of Ca(2+); (iii) decreased phosphoenzyme levels at saturating Ca(2+) concentrations; (iv) no effect on kinetics of phosphoenzyme decomposition. The palytoxin effect is related with lock of the enzyme in the Ca(2+)-free conformation so that progression of the catalytic cycle is impeded.

Experimental sarcoplasmic reticulum damage induced by 3 microM thapsigargin or 1 microg/ml tunicamycin provoked viability loss of the cell population in approximately 72 h. Release of cytochrome c from mitochondria was an early event and Bax translocation to the mitochondria preceded or was simultaneous with cytochrome c release. The release of cytochrome c was not related with mitochondria depolarization or caspase activation. Irreversible stress in the sarcoplasmic reticulum, detected by the early activation of caspase 12, was functionally linked to the mitochondrial apoptotic pathway. Caspase 3 processing was blocked by cells preincubation with a selective inhibitor of either caspase 9 or caspase 8 whereas caspase 8 activation was inhibited by a selective caspase 9 inhibitor. This was consistent with the involvement of caspase 8 in a positive feedback loop leading to amplify the caspase cascade. Caspase inhibition did not protect against cell death indicating the existence of alternative caspase-independent mechanisms.

The chemical treatment of sarcoplasmic reticulum vesicles with H2O2 affects both Ca2+ transport and the hydrolytic activity supported by the Ca2+-ATPase protein. Ca2+ transport was much more sensitive to inhibition than ATPase activity and the decrease in Ca2+ transport was not the result of an increase in membrane permeability. The Ca2+/Pi uncoupling can be attributed to the own catalytic mechanism of the enzyme. Under conditions of high uncoupling, Ca2+ binding to the transport sites was barely affected and accumulation of phosphorylated species during the enzyme cycling gave almost maximal levels. These are features defining intramolecular uncoupling mediated by a phosphorylated form of the enzyme. Severe inhibition of the hydrolytic activity was observed when higher peroxide concentrations and leaky vesicles were used. These experimental conditions diminished maximal Ca2+ binding and the steady-state phosphoenzyme level. The low hydrolytic activity can be ascribed to a decrease in the rate of enzyme dephosphorylation.

Tricyclic antidepressants are moderately potent inhibitors of the plasma membrane Ca(2+)-ATPase activity measured in erythrocyte ghosts. For the calmodulin-activated activity, half-maximal inhibition was observed in the presence of 0.25 mM clomipramine. Desipramine, imipramine, and trimipramine show half-maximal inhibition in the range of 0.8 to 1 mM. The inhibition dependence on clomipramine concentration is the same whether the enzyme is activated by exogenous calmodulin or by tryptic digestion. A similar behavior was observed for desipramine. The inhibition mechanisms utilized by clomipramine and desipramine are different. The clomipramine effect is associated with the Ca(2+)-bound enzyme conformation and can be attributed to a decrease in the rate of phosphorylation by ATP. The desipramine effect appears more related to the Ca(2+)-free conformation, since the partial reaction involved in the release of inorganic phosphate is perturbed by this drug. There is also little or no effect of tricyclics on the enzyme's affinity for ligand (Ca(2+) or ATP) binding., (Copyright 1999 Academic Press.)

The Ca(2+)-ATPase activity of human erythrocyte membrane can be inhibited in vitro by ritodrine, a beta 2-adrenergic agonist. The inhibitory profile shows a low-affinity interaction and no competition with the specific transport and catalytic substrates. The activated conformation of the enzyme (in the presence of calmodulin or after trypsin digestion) facilitates the interaction with ritodrine. This suggests that the C-terminal tail of the enzyme plays a protective role. By studying selected partial reactions of the catalytic and transport cycle we found that the inhibition can be basically assigned to a lower rate of phosphorylation by ATP. A minor effect on the phosphorylation level by Pi in the absence of Ca2+ and no effect on the enzyme affinity for Ca2+ or ATP were also observed. The inhibition of the plasma membrane Ca(2+)-ATPase by ritodrine shows a clear similarity with that of the sarcoplasmic/endoplasmic reticulum membrane. The inhibition under study does not foresee a pharmacological effect of ritodrine on the myometrial plasma membrane Ca(2+)-ATPase when administered for the management of preterm labor.

Ritodrine inhibits the steady-state Ca(2+)-ATPase activity of isolated sarcoplasmic reticulum vesicles in a dose-dependent manner. The observed K0.5 value for inhibition (approximately 3 mM) gives proof of a low-affinity interaction. Ritodrine also interferes with the steady-state Ca2+ transport ability decreasing the maximal rate without modification of the Ca2+ or ATP affinity for the enzyme. This is consistent with an absence of competition for the transport and the catalytic sites. Analysis of the catalytic and transport cycle shows that: (i) ritodrine inhibits the phosphorylation partial reaction. This is supported by pre-steady-state kinetic experiments of Ca2+ transport and also by the temperature dependence of the phosphoenzyme level. (ii) At high ritodrine concentrations the dephosphorylation step becomes rate-limiting. This is suggested by the biphasic profile (V-shape) of phosphoenzyme accumulation at different ritodrine concentrations. This was also confirmed by chase experiments of radioactive phosphoenzyme decomposition at steady state. These data reveal a complex pattern of inhibition involving two sites for interaction with low and high ritodrine concentrations. It is envisaged that ritodrine does not exert any direct effect on the smooth muscle sarcoplasmic reticulum Ca(2+)-ATPase when used in the treatment of preterm birth.

Gas chromatographic analysis of whole-cell fatty acids, secondary alcohols and mycolic acid cleavage products could be a useful technique in checking mixed mycobacterial cultures. The mixed cultures were confirmed when species-specific compounds of different mycobacterial species were detected in the same chromatogram.

Ca2+ binding and internalization in sarcoplasmic reticulum ATPase can be investigated by the use of La3+ as a Ca2+ analog. Displacement kinetics of Ca2+ bound by La3+ in native vesicles is a slow biphasic process (k1 = 0.55 s-1 and k2 = 0.05 s-1) that is consistent with the existence of two Ca2+ binding populations whereas in leaky vesicles there appears to be a single population (k = 0.57 s-1). Rapid quench experiments demonstrate that Ca2+ internalization occurs with an initial burst (approximately 8 nmol/mg protein) associated with the presence of a phosphate-donor substrate in the reaction medium. While acid quenching for measurements of phosphoenzyme is instantaneous, La3+ quenching allows completion of one catalytic and transport cycle due to the slow La3+ exchange with Ca2+. This explains the apparent inconsistencies in the kinetics and stoichiometry of phosphoenzyme formation and Ca2+ internalization that are observed under certain experimental conditions.

The rise of intrinsic fluorescence due to calcium binding to sarcoplasmic reticulum ATPase occurs with a kobs of approximately 2 s-1 at pH 6.0, which is much lower than that observed at neutral pH. This is consistent with a H+-Ca2+ competition for the high-affinity sites. An accelerating effect of ATP on the calcium-induced transition can be clearly demonstrated at that pH. Nonhydrolyzable nucleotides, such as AMP-PNP, do not elicit the same response. Acetylphosphate also accelerates the calcium-induced fluorescence rise, demonstrating that this effect is limited to substrates that are able to form the phosphorylated enzyme intermediate. This effect, which is attributed to occupancy of the phosphorylation domain of the catalytic site, is distinct from the known secondary activation of enzyme turnover which is produced by ATP and by inactive nucleotide analogs, but not by acetylphosphate.

The apparent effect of tetraphenylboron on the Ca2+-dependent ATPase activity is rather complex being dependent of the physical state of the membrane. In leaky vesicles, tetraphenylboron decreases the affinity of the enzyme for Ca2+, and high concentrations (100 microM) inhibit the enzyme. These effects are mediated by the membrane structure. In native vesicles, tetraphenylboron induces an apparent stimulation of the maximum hydrolytic activity and a decrease of the Ca2+ gradient formed due to specific Ca2+ releasing activity of the mentioned drug.