Evaluation of a surveillance strategy for early detection of adenovirus by PCR of peripheral blood in hematopoietic SCT recipients: incidence and outcome.

Adenoviruses (AdV) have emerged as important causes of morbidity and mortality in patients after hematopoietic SCT (HSCT). Early diagnosis of the infection by detection of viral DNA may improve the prognosis. A surveillance strategy was evaluated for detection of AdV DNA by PCR in a prospective study of unselected allogeneic HSCT recipients. In parallel with a routine CMV surveillance program, plasma from 20 children and 77 adults was analyzed by quantitative PCR for detection of AdV DNA. In addition, in 12 unselected patients, the presence of AdV-specific T cells were analyzed by enzyme-linked immunosorbent spot (ELISPOT) at 1 to 3 months after transplantation. A total of 5 of 97 (5%) patients had detectable AdV DNA in peripheral blood. Only one patient had high titers and none developed AdV disease. BM as a source of stem cells and myelodysplastic syndrome as the indication for transplantation were independently associated with higher risk of acquiring AdV infection. AdV-specific T cells were detected in 7 (58%) of 12 patients. Although AdV DNA was found in peripheral blood by quantitative PCR in 5% of patients undergoing allogeneic HSCT, the present surveillance program did not have a significant effect on the clinical outcome.

Quantification of adenovirus DNA in unrelated donor hematopoietic stem cell transplant recipients.

BACKGROUND: Adenovirus (AdV) infection is a life threatening condition in immunosuppressed patients. Quantitative AdV assays can improve the clinical management of these patients. OBJECTIVES: To evaluate quantitative measurement of AdV DNA with PCR in blood from hematopoietic stem cell transplant (HSCT) recipients. STUDY DESIGN: Quantitative PCR was used to measure viral DNA levels of AdV in consecutive blood samples from 40 HSCT recipients (27 adults and 13 children) during a 1-year post-engraftment period. All patients received grafts from unrelated donors and were given anti-T-cell antibodies in the conditioning regimen. RESULTS: In the group of 40 patients, six (15%) had detectable AdV DNA in blood for different lengths of time. None of these six patients suffered from severe graft-versus-host disease. In three of the patients a high AdV viral load (>10,000 copies/mL) was detected, one of whom also had high viral load of EBV and CMV and one of EBV only. These three patients died within 2 months after detection of ADV viremia. A low AdV viral load (<500 copies/mL) was detected in three surviving patients and they did not have concomitant high viral load of neither CMV nor EBV. CONCLUSIONS: AdV viremia was present in 15% of the HSCT recipients and a high AdV viral load was associated with fatal outcome. Screening for AdV DNA with quantitative PCR in blood may be of clinical importance in allogeneic HSCT recipients in order to prevent severe clinical virological complications.

Distribution and stability of membrane proteins in lipid membranes on solid supports.

Bacteriorhodopsin and the nicotinic acetylcholine receptor were biotinylated and reconstituted in lipidic membranes on silicon supports by fusion with proteoliposomes. The presence and distribution of the proteins were studied by binding with streptavidin. Radio-labelled streptavidin was employed for quantifying the amounts of protein remaining in the supported membranes after storage in buffer. The proteins within the membranes remained bound to the surface for weeks. The biological activity of reconstituted unlabelled receptor upon storage showed stability in membranes formed on silicon supports and a reduced stability when formed onto lipid monolayer covered supports. Atomic force microscopy studies on preparations in liquid showed bilayer structures but also attached, partly fused liposomes and membrane particles. In air, the surface was smoother and contained less of liposomes and more of stacked lipid layers. Preparations labelled with streptavidin conjugated to colloidal gold and imaged in air showed the proteins individually distributed, with no protein-rich patches or protein aggregates.

[Postoperative laryngospasm may induce pulmonary edema. An unusual or overlooked complication?]. / Postoperativ laryngospasm kan orsaka lungödem. En ovanlig eller förbisedd komplikation?

Young and healthy patients in whom laryngospasm secondary to postoperative extubation is encountered run a risk of developing pulmonary edema. The mechanism behind the edema is thought to be a strongly negative intrathoracic pressure generated by the patient's forced inspiration against a closed glottis. A net flow of fluid occurs to the interstitial space and further to the alveoli. Unusually the pulmonary edema can be noticed within a few minutes after relief of the obstruction, but occasionally it is delayed for several hours. The condition is potentially life-threatening, but usually responds favourably to positive pressure ventilation and diuretics. In anaesthesia and intensive care, it is important to be aware of the complication in order to be ready for adequate therapy when needed. We present three cases from the Central Hospital in Växjö, where pulmonary edema occurred directly following postoperative extubation.

Planar lipid bilayers on solid supports from liposomes--factors of importance for kinetics and stability.

One method to create planar lipid bilayers on solid substrates involves the transfer of lipids from liposomes to the support. We have varied the composition of liposomes systematically using factorial experimental designs and analyzed the adsorption behaviour of lipids from these liposomes onto solid supports. The hydrophilic supports were either used plain or modified with a monolayer of a lipid mixture, exposing hydrophobic groups. The monolayer-covered supports were used to identify factors important for adhesion and stability. Lipid adsorption kinetics was primarily studied on plain silicon supports in an ellipsometric cell or on a silicon nitride surface in a resonant mirror system (IAsys), using the systematic approach. Saturated phospholipids were essential for the required stability. Mixtures of dipalmitoylphosphatidylcholine, dipalmitoylphosphatidylglycerol, dipalmitoylphosphatidylethanolamine and cholesterol in combination with proteins were investigated in further detail as regards kinetics. The propensity to form a supported planar bilayer could be manipulated by the presence of calcium ions.

A comparison of remifentanil and alfentanil in patients undergoing major abdominal surgery.

The efficacy and safety of remifentanil and alfentanil for patients undergoing major abdominal surgery were compared. Premedicated patients received a loading dose of remifentanil (1.0 microgram.kg-1; n = 116) and a continuous infusion of 0.5 microgram.kg-1.min-1, or a loading dose of alfentanil (25 micrograms.kg-1; n = 118) and a continuous infusion of 1.0 microgram.kg-1.min-1. Propofol was administered (10 mg every 10 s) until loss of consciousness. Patients' lungs were ventilated with 66% nitrous oxide and 0.5% (end-tidal) isoflurane in oxygen. The study drug infusion rate was reduced by 50% 5 min after intubation. Alfentanil was discontinued 15 min before the end of surgery, whereas remifentanil was continued in the immediate postoperative period at a reduced dose. Responses to intubation (28%) and skin incision (17%) occurred approximately twice as often in the alfentanil group (15% and 8%; p = 0.014 and p = 0.037, respectively). More patients receiving alfentanil had one or more responses to surgery (72% vs. 57%; p = 0.016). The time to spontaneous respiration, adequate respiration, response to verbal command and time to recovery room discharge were similar. However, owing to decreased variability, the time to extubation was shorter with remifentanil than with alfentanil (p = 0.048). There was a similar overall incidence of adverse events in both groups, 82% and 75% of patients, respectively. Adverse events associated with remifentanil were rapidly controlled by dose reductions. The incidence of intra-operative hypotension and bradycardia was higher in the remifentanil group (p < or = 0.033). An initial remifentanil infusion rate of 0.1 microgram.kg-1.min-1 titrated to individual need provided postoperative pain relief in the presence of adequate respiration in 71% of patients. When using remifentanil in the immediate postoperative setting, rapid administration of bolus doses and infusion rate increases resulted in a relatively high incidence of muscle rigidity, respiratory depression and apnoea. Changing the postoperative regimen to avoid rapid changes in remifentanil blood concentration resulted in more effective analgesia and dramatically reduced the incidence of adverse events during this period. In patients undergoing major abdominal surgery, remifentanil appears to offer superior intra-operative haemodynamic stability during stressful surgical events compared with alfentanil without compromising recovery from anaesthesia. Remifentanil can be administered as a postoperative analgesic agent at a starting dose of 0.1 microgram-.kg-1.min-1; however, it should only be used in the presence of adequate supervision and monitoring of the patient. Administration of bolus doses is not recommended in this setting.

Retained activities of some membrane proteins in stable lipid bilayers on a solid support.

Highly stable lipid bilayers, composed of biologically relevant lipids such as phosphatidylcholine, phosphatidylethanolamine and cholesterol, were formed on platinum surfaces. Bacteriorhodopsin isolated from purple membrane (PM) from Halobacterium halobium, cytochrome oxidase from bovine heart, acetylcholinesterase from bovine brain and the nicotinic acetylcholine receptor from Torpedo electric organ were also incorporated into these reconstituted membranes. The proteins retained their biological activities. Some of them were active several weeks after the reconstitution and after several cycles of assay, washing and storage.

Effects of ischemia on regional ligand binding to adrenoceptors in the rat brain.

Changes in ligand binding to adrenoceptors ([3H]prazosin to alpha 1-receptors, [3H]idazoxan to alpha 2-receptors and [125I]cyanopindolol to beta-receptors) following transient cerebral ischemia were investigated using autoradiographic methods. The binding was quantified in brain sections from control rats, rats subjected to 15 min of 2-vessel occlusion ischemia, and rats with recirculation times of 1 h, 1 week or 4 weeks after ischemia. No significant change in alpha 1-receptor binding was observed during and immediately following ischemia, but a decrease was noted in the vulnerable hippocampal CA1 region following 1 week's survival. In the parietal cortex, the ligand binding to alpha 1-receptors increased at 4 weeks. A reduced [3H]idazoxan binding was observed 1 h after ischemia in the temporal cortex and amygdala. No change in ligand binding to beta-receptors was seen in the early phase postischemia, but a marked increase had occurred in the hippocampal CA1 region at 1 and 4 weeks after ischemia (+163% and +142%, respectively), presumably due to accumulation of macrophages expressing beta-receptors. The early postischemic changes in receptor binding may represent downregulation of the adrenoceptors by processes activated during ischemia, while neuronal degeneration, compensatory mechanisms in surviving neurons and proliferation of non-neuronal cells may account for the subsequent changes.

Brain-oriented intensive care after resuscitation from cardiac arrest.

The 'chain-of-survival' concept has gained general acceptance in the care of cardiac arrest victims. Most standards and guidelines for cardiopulmonary resuscitation, however, focus on the initial links in the chain. We consider appropriate in-hospital care for the survivors a logical extension of the chain of survival. In recent years extensive research activity has probed the pathophysiology and pharmacology of postischemic reperfusion. The present review discusses the current understanding of mechanisms for cerebral damage following global ischemia. Promising pharmacological principles for protection or resuscitation from cerebral ischemia are reviewed. None of them are considered ready for clinical application. Clinical guidelines are proposed, based on the reviewed data and previously published clinical observations. Cornerstones of the proposed brain-oriented intensive care protocol are: (1) hemodynamic monitoring and meticulous treatment of circulatory disturbances, (2) controlled ventilation providing normoventilation and normoxia to all comatose patients, (3) avoiding hyperglycemia and hyperthermia in comatose patients, (4) adequate analgesia and sedation, tempered by the understanding that oversedation impedes neurological evaluation without promoting recovery. An accurate prognosis can usually be made 48-72 h after resuscitation. This permits reevaluation and assignment to an appropriate level of continued hospital care.

Brain cortical tissue levels of noradrenaline and its glycol metabolites: effects of ischemia and postischemic administration of idazoxan.

The brain noradrenaline (NA) system is known to modulate ischemic neuronal damage, and the turnover of NA has been suggested to increase in the early recovery period following cerebral ischemia. Using HPLC and gas chromatography-mass spectrometry we analyzed the tissue levels of NA and its metabolites, 3,4-dihydroxyphenylethyleneglycol (DHPG) and 3-methoxy-4-hydroxyphenylethyleneglycol (MHPG), in rat brain cortex after 10 min of forebrain ischemia followed by 1 h of recirculation. The effect of idazoxan, given in cerebro-protective doses, as a bolus of 0.1 mg.kg-1 immediately after ischemia followed by 10 micrograms.kg-1.min-1 for 1 h, was also investigated. Ischemia decreased basal NA cortical levels from 384 ng/g tissue in control animals to 214 ng/g, while DHPG increased from 74 to 103 ng/g (+39%) and MHPG from 82 to 154 ng/g (+88%). Conjugated but not free DHPG increased, while both free and conjugated MHPG increased equally. The findings indicate an enhanced postischemic NA turnover with a major proportion of uptake and metabolism occurring extraneuronally, possibly secondary to a saturation of neuronal NA uptake in the postischemic phase. Idazoxan further increased NA turnover, as evidenced by higher postischemic levels of free MHPG and a higher MHPG/NA ratio. A correlation may exist between the protective action of idazoxan and its effect on NA turnover.

Lack of protection by the N-methyl-D-aspartate receptor blocker dizocilpine (MK-801) after transient severe cerebral ischemia in the rat.

Glutamate is an important factor in the mechanisms of neuronal damage following cerebral ischemia. Blockade of one type of glutamate receptor, the N-methyl-D-aspartate (NMDA) receptor, decreases brain infarct size in experimental models of permanent focal ischemia, but protection in models of transient reversible ischemia is ambiguous. We investigated the effect of the noncompetitive NMDA receptor antagonist dizocipiline (MK-801) on neuronal damage in the CA1 region of the rat hippocampus, using two models of reversible cerebral ischemia: 10 or 15 min of bilateral common carotid occlusion combined with hypotension, or 6-8.5 min of cardiac arrest. Histopathologic evaluation of neuronal damage was performed 7 days after the ischemic insults. Thirteen groups of rats (a total of 129 animals) were treated with saline or dizocilpine in single or multiple doses ranging from 0.1 to 5 mg.kg-1, given intravenously or intraperitoneally prior to and/or after the ischemic insult. In none of the dizocilpine-treated groups could neuronal protection be demonstrated in the CA1 region of the septal as well as dorsotemporal hippocampus, compared to a corresponding saline-treated group. We conclude that systemically administered noncompetitive NMDA receptor antagonists do not provide a marked protection against neuronal damage after a transient period of severe forebrain ischemia.

Extracellular brain cortical levels of noradrenaline in ischemia: effects of desipramine and postischemic administration of idazoxan.

Using microdialysis, extracellular noradrenaline (NA) levels in the rat cerebral cortex were studied under isoflurane/N2O anaesthesia before, during and for 6 hours following 10 min of forebrain ischemia in a 2-vessel occlusion model. A microdialysis probe was introduced into the parietal cortex and dorsal hippocampus in anaesthetized rats and continuously perfused with Krebs-Ringer-bicarbonate buffer with or without the NA uptake inhibitor desipramine (DMI, 5 microM). Twenty min fractions were collected and the extracellular NA levels were measured in the dialysates using HPLC with electrochemical detection. The basal NA concentration in the dialysate was 10.5 +/- 1.8 (mean +/- SEM) pg/20 min fraction and increased to 39.3 +/- 4.8 pg/20 min fraction after local administration of DMI. During ischemia, NA increased to 38 times the basal level without DMI, and 6 times with DMI included during two hours' perfusion prior to ischemia. After recirculation NA levels returned to, or even transiently decreased below, preischemic values. With DMI present in the dialysis buffer, administration of idazoxan immediately following ischemia delayed the return to preischemic NA levels in the recirculation phase. In the absence of DMI, no effect of idazoxan on postischemic levels of NA was found. Local administration of DMI increases basal extracellular NA levels and reduces the ischemia-induced NA release. The latter effect may be a due to inhibition of the NA uptake system working in a reversed mode, or as a result of decreased synthesis of NA due to activation of presynaptic alpha 2-receptors by the increased synaptic NA levels. Postischemic treatment with the alpha 2-adrenoceptor antagonist idazoxan in combination with DMI prolongs the period of elevated extracellular NA levels, which may be of importance for the protective properties of idazoxan against ischemic cell injury.

Protection against ischemia-induced neuronal damage by the alpha 2-adrenoceptor antagonist idazoxan: influence of time of administration and possible mechanisms of action.

The protective effect of the alpha 2-receptor antagonist idazoxan against neuronal damage in the neocortex and in the hippocampal CA1 region was studied in rats exposed to 10 min of incomplete forebrain ischemia. When administered i.v. immediately after ischemia (0.1 mg/kg) and subsequently for 6 h (10 micrograms/kg/min), idazoxan significantly reduced neuronal damage in the hippocampus (from 84 to 26%) and in the vulnerable parts of the neocortex (from 15 to 1%). The bolus dose alone provided no significant protection. When idazoxan administration was delayed for 30 min, no significant protection was noticed in the neocortex, and the effect in the hippocampus was ambiguous. A transient elevation of plasma corticosterone levels was induced during ischemia. Idazoxan administration for 2 h did not affect postischemic changes in corticosterone levels compared with saline infusion. Idazoxan (10(-7)-10(-4) M) did not influence the in vitro binding to glutamate receptors in brain slices. Thus, the protective effect of idazoxan cannot be explained by suppression of the plasma corticosteroid levels or via an antagonistic effect on glutamate receptors. Idazoxan apparently protects neurons when given during the first hours of postischemic reperfusion, while histopathological necrosis of neurons becomes visible 48-72 h after ischemia. Detrimental processes causing delayed neuronal death occur in the early postischemic phase and can be influenced by adrenoceptor ligands. Idazoxan may protect by several mechanisms but probably exerts its protective postischemic effect mainly through an increased noradrenergic neuronal activity and an elevation of extracellular noradrenaline (NA) levels in the brain. The favorable effects of NA may either be due to inhibition of excitotoxic neurotransmission or activation of survival-promoting and trophic processes.

Postischemic administration of idazoxan, an alpha-2 adrenergic receptor antagonist, decreases neuronal damage in the rat brain.

The effect of an alpha-2 receptor antagonist, idazoxan, on ischemic neuronal damage in the hippocampus and neocortex was studied in rats following 10 min of forebrain ischemia. Idazoxan was given 0.1 mg/kg i.v. immediately after recirculation, followed by 48 h of continuous infusion at a rate of 10 micrograms/kg/min. A histopathological examination of the CA1 region of the dorsal hippocampus and neocortex from each hemisphere was made on paraffin-embedded sections following 7 days of survival. In ischemic animals receiving an infusion of saline, 71% of the neurons in the hippocampal CA1 region were degenerated. In contrast, in the idazoxan-treated animals only 31% of the neurons were irreversibly damaged (p less than 0.01). We conclude that postischemic administration of the alpha-2 antagonist idazoxan protects neurons against damage following cerebral ischemia. Rapid postischemic administration of alpha-2 adrenergic receptor antagonists could be an effective treatment after stroke and cardiac arrest.