Isolation of syncytiotrophoblast microvesicles and exosomes and their characterisation by multicolour flow cytometry and fluorescence Nanoparticle Tracking Analysis.

The human placenta releases multiple types and sizes of syncytiotrophoblast (STB) extracellular vesicles (EV) into the maternal circulation that exhibit diverse biological activities. The placental perfusion technique enables isolation of these STBEV, but conventional flow cytometry can only be used to phenotype EV down to ∼300 nm in size. Fluorescence Nanoparticle Tracking Analysis (fl-NTA) has the potential to phenotype EV down to ∼50 nm, thereby improving current characterisation techniques. The aims of this study were to prepare microvesicle and exosome enriched fractions from human placental perfusate (n=8) and improve fl-NTA STBEV detection. Differential centrifugation and filtration effectively removed contaminating red blood cells from fresh placental perfusates and pelleted a STB microvesicle (STBMV) fraction (10,000×g pellet - 10KP; NTA modal size 395±12 nm), enriched for the STB marker placental alkaline phosphatase (PLAP) and a STB exosome (STBEX) fraction (150,000×g pellet - 150KP; NTA modal size 147±6 nm), enriched for PLAP and exosome markers Alix and CD63. The PLAP positivity of 'standard' 10KP and 150KP pools (four samples/pool), determined by immunobead depletion, was used to optimise fl-NTA camera settings. Individual 10KP and 150KP samples (n=8) were 54.5±5.7% (range 17.8-66.9%) and 30.6±5.6% (range 3.3-51.7%) PLAP positive, respectively. We have developed a reliable method for enriching STBMV and STBEX from placental perfusate. We also standardised fl-NTA settings and improved measurement of PLAP positive EV in STBMV. However, fl-NTA is not as sensitive as anti-PLAP Dynabead capture for STBEX detection, possibly due to STBEX having lower surface expression of PLAP. These important developments will facilitate more detailed studies of the role of STBMV and STBEX in normal and pathological pregnancies.

Single channel characteristics of some synthetic gramicidins.

The synthesis of two new variants of gramicidin is described. It is shown that the changes in the aromatic side groups do not influence the single channel conductivity. Experiments in which solutions having different molarities on the two sides of the bilayer lipid membrane are described and their results compared with a rate theory analysis. It is concluded that the gramicidin pore contains approximately 10 equal potential maxima.

Developmental, nutritional and hormonal anomalies of weightlessness-grown wheat.

The behavior of water in weightlessness, as occurs in orbiting spacecraft, presents multiple challenges for plant growth. Soils remain saturated, impeding aeration, and leaf surfaces remain wet, impeding gas exchange. Herein we report developmental and biochemical anomalies of "Super Dwarf" wheat (Triticum aestivum L.) grown aboard Space Station Mir during the 1996-97 "Greenhouse 2" experiment. Leaves of Mir-grown wheat were hyperhydric, senesced precociously and accumulated aromatic and branched-chain amino acids typical of tissues experiencing oxidative stress. The highest levels of stress-specific amino acids occurred in precociously-senescing leaves. Our results suggest that the leaf ventilation system of the Svet Greenhouse failed to remove sufficient boundary layer water, thus leading to poor gas exchange and onset of oxidative stress. As oxidative stress in plants has been observed in recent space-flight experiments, we recommend that percentage water content in apoplast free-spaces of leaves be used to evaluate leaf ventilation effectiveness. Mir-grown plants also tillered excessively. Crowns and culms of these plants contained low levels of abscisic acid but high levels of cytokinins. High ethylene levels may have suppressed abscisic acid synthesis, thus permitting cytokinins to accumulate and tillering to occur.

Long-term home treatment with epidural analgesia does not affect later spinal cord stimulation in patients with otherwise intractable angina pectoris.

OBJECTIVE: The only way some patients with intractable angina pectoris can endure the daily pain is by using opiates. Epidural morphine or spinal cord stimulation (SCS) for out-patients is a possibility for this patient group. The aim of this study was to determine whether patients previously treated with epidural injections had more frequent electrode migration, higher stimulation needs, and less effect of SCS treatment. DESIGN: A prospective, nonrandomized study with a 4-year follow-up period. SETTING: Department of Anaesthesia, Pain Section. A referral center in institutional practice. Ambulatory care. PATIENTS: The patient group comprised 53 patients; all had been treated daily with opiates. A total of 26 patients had epidural catheters for a mean time of 1 year before SCS. All patients had intractable angina pectoris. For these patients, further angioplasty or coronary bypass surgery was not technically possible. Even with maximal medication, it was impossible to cope with the patients' angina pectoris, and the only way the patients could endure the daily pain was by using opiates. Therefore, alternative therapies were considered to give these patients palliation. INTERVENTION: SCS with epidural electrodes stimulating paresthesia in the area where angina is perceived. MAIN OUTCOME MEASURE: Difference for the patients previously treated with epidural catheters in stimulation amplitude, frequency of electrode migration, and effect of SCS. RESULTS: Stimulation demand (p = 0.09), frequency of electrode migrations (p = 0.46), and pain-reducing effect (p = 0.16) were not different for the group of patients previously treated with epidural catheters for longer periods (1-36 months). CONCLUSIONS: SCS in patients previously treated with epidural catheters has an effect equal to that in other patients.

Ground-based studies with Super-Dwarf wheat in preparation for space flight.

Several experiments were carried out to test responses of a Super-Dwarf cultivar of wheat (Triticum aestivum L.) to various environmental parameters that were anticipated to be present in our attempts to grow the wheat in a small growth chamber on the Russian Space Station, Mir, or that proved to be present in a 1995 trial space experiment. Under low photosynthetic photon flux (40-400 micromoles m-2 s-1 PPF), development (e.g. anthesis) was retarded, but heads (often sterile) always formed, even if light was so low that plants died before the heads could mature. Longer photoperiods promoted flowering, but night interruptions combined with short days did not provoke a long-day response as occurs with true long-day plants. The long-day effect could prove to be a summation of photosynthetic products. Heat stress (40 degrees C for 1-24 h) did not influence flowering but killed plants that were 13-16-day-old (no effect on younger plants). Concentrations of iodine or silver-fluoride disinfectants present in the water used for plants on Mir (1.0-4.0 mg L-1) did not affect plant growth although higher concentrations (8.0-1.6 mg L-1) were inhibitory. GA3 or indoleacetic acid applied every other day at concentrations from 1.0 x 10(-6) mg L-1 to 3.162 x 10(-4) mg L-1 did not change the height of Super-Dwarf wheat, suggesting that this cultivar is not a gibberellin mutant.

Age-related postoperative morphine requirements in children following major surgery--an assessment using patient-controlled analgesia (PCA).

To investigate if small children require less morphine for postoperative analgesia than do older children and adolescents we analysed the morphine consumption pattern of 28 consecutive children on intravenous patient-controlled analgesia (PCA) following major surgery. The median age-specific morphine requirements between 2 comparable groups of children aged 4-8 years and 9-15 years were compared. We used the Pharmacia-Deltec pump in all children and the same settings: a bolus dose of 25 microgram/kg, an 8 minutes lockout interval and no background infusion. In addition, all children received paracetamol as a supplemently to the morphine. In this study children aged 4-8 years had significantly higher total postoperative morphine requirements compared to children aged 9-15 years, i.e. 11.6 microgram/kg/hour and 7.5 microgram/kg/hour respectively (p = 0.037). Hence, we conclude that children of this age group may have a higher total postoperative morphine requirement following major surgery than older children and adolescents.

Postoperative pain management in children has been improved, but can be further optimized.

The aim of this study is to evaluate the effect of the analgesic treatment currently used in children, and to identify if problems can be related to any particular routine or group of children. Analgesics administered pre-, per- and postoperatively were recorded, and intensity of pain during rest and movement and incidence and severity of side effects were measured four times postoperatively. Fifty-nine children aged 3 to 15 years undergoing miscellaneous operations participated. All children received analgesic treatment. During the study period 26 children reported at least one episode of unacceptable pain, while unacceptable pruritus, nausea or vomiting were observed in 18 children. It was not possible to relate the incidence of pain and side effects to any particular analgesic treatment or type of surgery, but groups of children that might need additional attention were identified. Even though improvements have been obtained, it is still a challenge to optimize the postoperative pain management of children, and when doing so attention should be paid not only to pain relief, but also to side effects of the administered analgesics.

Neurologic deficit following spinal anesthesia.

A case report of prolonged sphincter paresis and muscle weakness of the legs following spinal anesthesia with tetracaine is presented. The case could be placed in one of the three groups of sequelae described by Kane. Tetracaine as responsible for the reaction is suggested.

[Postoperative pain relief in children]. / Postoperativ smertebehandling af børn.

Postoperative pain management in children has been subject to increasing interest during the last decade, but is still insufficient. A survey is presented concerning postoperative pain management in children. The value of monitoring the pain as well as the opioid side effects in children is stressed, and such methods are presented. Acute Pain Service is mentioned and the most important pharmacological aspects regarding non-steroidal anti-inflammatory drugs, paracetamol, opioids and local anaesthetics are discussed. The management of postoperative pain in neonates is reviewed separately. It is concluded that the present insufficient management of postoperative pain in children is not due to the lack of methods and techniques, but rather to lack of sufficient utilization and comprehension of the possibilities. Moreover, pain management in children should be individualised. It is also necessary to be more aware of side effects of the pharmacological treatment.

[Patient-controlled analgesia in children]. / Patientkontrolleret smertebehandling af børn.

In order to introduce intravenous patient-controlled analgesia (PCA) in children in the Department of Anaesthesiology and Intensive Care, Odense University Hospital, we evaluated the technique in 13 children following major orthopaedic or abdominal surgery. The pump (Pharmacia-Deltec, CADD-PCA) was loaded with 1 mg/ml morphine in a 50 milliliters cassette. A bolus dose of 25 micrograms/kg and a lock-out interval of eight minutes were the initial settings. The morphine dose used, pain scores and side effects were currently recorded. Ages ranged from 4-15 years (mean 10.5 years) and the method was used for a mean of 89 hours (range 57-144 hours) postoperatively. Morphine requirements averaged 9.5 microgram/kg/hour (range 5.4-15.6 microgram/kg/hour). Pain control was good and side effects were few and of minor nature. PCA is an effective and safe means of providing good quality analgesia in children.

[Continuous subcutaneous morphine--treatment of pain in patients with terminal cancer]. / Kontinuerlig subkutan morfin--smertebehandling af terminale cancerpatienter.

Nine patients with terminal cancer were treated for pain with continuous subcutaneous injection of morphine via a portable battery-driven injection pump. Treatment was instituted on account of failure of other forms of treatment with oral or epidural morphine derivatives or on account of severe nausea and vomiting which necessitated parenteral administration. Treatment proved reasonably effective and no side effects of significance occurred. Two of the patients could be treated in their homes. The method is thus considered as suitable for treatment of pain in patients with terminal cancer.

[Local reactions following intravenous injection of Valium mixed micelles and Diazemuls]. / Lokale reaktioner efter intravenøs injektion af Valium mixed micelles og Diazemuls.

The frequency of local venous reactions after intravenous injection of Diazemuls and Valium Mixed Micelles was studied. A material of 224 patients undergoing surgery for prolapsed lumbar disc was allocated into two groups, receiving either 10 mg of Diazemuls or Valium Mixed Micelles. The incidences of thrombophlebitis in the two groups were 1 and 2%, respectively.

[Incapacitating angina pectoris treated with electric stimulation of the spinal cord]. / Invaliderende angina pectoris behandlet med elektrisk rygmarvsstimulation (ERS).

Thirty patients who had severe incapacitating angina pectoris which had not reacted to the conventional therapeutic measures and which required massive daily opioid consumption were treated with electrical spinal cord stimulation (SCS) by means of a completely implantable stimulation system. The therapeutic effect was good in 87% of the patients who experienced considerably reduced frequency of attacks and markedly reduced opioid consumption (p less than 0.00005). Nine of the patients could reduce opioid consumption and 14 out of 27 could cease their otherwise daily opioid consumption. In four patients, the therapeutic effect was unsatisfactory. In the first 22 patients in whom a unipolar electrode was introduced, displacement of the electrode and subsequent reoperation was a frequent problem. This problem disappeared after change to multipolar electrodes as slight changes in placing of the electrode could easily be compensated for via the external programming equipment. This investigation reveals that SCS is a good therapeutic alternative for this selected patient category.

[Prehospital treatment. The Odense ambulance project. A prospective study]. / Praehospital behandling. Ambulance-projektet Odense. En prospektiv undersøgelse.

No legislation exists about ambulance services in Denmark. The present Danish prehospital treatment is undertaken by Falcks Redningskorps A/S and the municipal fire services. During a period of 12 months (1.2.1988-31.1.1989) a prospective investigation was carried out in Odense concerning the effect of medical support to the ambulance service, partly in the form of a medically staffed ambulance, partly in the form of an arrangement in which an independent doctor's car (rendez-vous arrangement) was sent together with the nearest ambulance. In addition, the heart ambulance model as recommended by the Danish National Board of Health was also tested. Experience from abroad and recent Danish trial arrangements demonstrate the positive effect of extended professional prehospital treatment. The authors consider, therefore, that an ambulance law is required as this would ensure better and more uniform prehospital treatment in Denmark. Planning of the future prehospital treatment of acute and injured patients including the extent of the training, equipment and treatment must be established on the basis of a healtheconomical assessment of the Danish arrangements.

[Out-of-hospital heart arrest. Prospective study of various types of expanded prehospital treatment in Odense]. / Hjertestop uden for hospital. Prospektiv undersøgelse af forskellige typer af udvidet praehospital behandling i Odense.

During the period 1.2.1988-31.1.1989, a prospective investigation was undertaken of the intensive prehospital treatment in Odense. Comparison was undertaken between a medically staffed ambulance (2 months), a motorized doctor (rendez-vous, model 4 months) and the heart ambulance (recommended by the Danish National Board of Health (6 months]. Out of 28 patients brought in by the medically staffed ambulance, two (7%) were discharged alive. Out of 57 patients brought in the phase with the motorized doctor nine (16%) were discharged alive. Out of 78 patients brought in by heart ambulance one (1%) was discharged alive. The results of this investigation reveal that the prehospital treatment of cardiac arrest in Odense can be improved by participation of a doctor in the treatment, (particularly the rendezvous model). Suggested improvements consist of 1) improved alarm system, 2) intensification of training laymen in treatment of cardiac arrest and 3) increased information to the population.

[Pre-hospital care]. / Praehospital behandling.

Organisatory and therapeutic characteristics for selected pre-hospital medical treatment systems are described. It is emphasized that all of the links in the therapeutic chain must function optimally to achieve effective treatment of acute disease and injuries. The importance of rapid decision-making by highly qualified medical staff is emphasized. On the basis of experience from Denmark and from abroad, a model for up-to-date pre-hospital treatment in Denmark is outlined.

Accurate particle size distribution determination by nanoparticle tracking analysis based on 2-D Brownian dynamics simulation.

A physical model is presented to simulate the average step length distribution during nanoparticle tracking analysis experiments as a function of the particle size distribution and the distribution of the number of steps within the tracks. Considering only tracks of at least five steps, numerical simulation could be replaced by a normal distribution approximation. Based on this model, simulation of a step length distribution allows obtaining a much more reliable estimation of the particle size distribution, thereby reducing the artificial broadening of the distribution, as is typically observed by direct conversion of step length to particle size data. As this fitting procedure also allowed including data from particles that were followed for a relatively low number of steps, the measurement time could be reduced for particles that are known to be monodisperse. Whereas the inversion is less sensitive towards the particle size distribution width, still similar values were obtained for both the average diameter and standard deviation of a polystyrene latex sample irrespective of the track length, provided that the latter included at least five steps.