Near total intrathoracic airway obstruction managed with a Tritube® and flow-controlled ventilation.

We describe the management of a case of near total airway obstruction in a 79-year-old man who presented with a 2-week history of increasing shortness of breath and stridor. Computed tomography imaging revealed a mid-tracheal mass of unknown aetiology with critical airway obstruction. We secured the patient's airway using a TriTube® (Ventinova, Eindhoven, the Netherlands). While this facilitated a secure airway past the lesion, various issues were encountered which complicated the safe conduct of anaesthesia. We conclude that while the TriTube and Evone® flow-controlled ventilation (Ventinova) are useful for critical airway obstruction, they can be problematic and thorough planning is essential.

The dose magnifying glass quality assurance system for daily proton therapy range verification.

Proton therapy has a distinct dosimetric advantage over conventional photon therapy due to its Bragg peak profile. This allows greater accuracy in dose delivery and dose conformation to the target, however it requires greater precision in setup, delivery and for quality assurance (QA) procedures. The AAPM TG 224 report recommends daily range and spot position checks with tolerance on the order of a millimetre. Daily QA systems must therefore be efficient for daily use and be capable of sub-millimetre precision however few suitable commercial systems are available. In this work, a compact, real-time daily QA system is optimised and characterised for proton range verification using an ad-hoc Geant4 simulation. The system is comprised of a monolithic silicon diode array detector embedded in a perspex phantom. The detector is orientated at an angular offset to the incident proton beam to allow range in perspex to be determined for flat proton fields. The accuracy of the system for proton range in perspex measurements was experimentally evaluated over the full range of clinical proton energies. The meanR100,R90andR80ranges measured with the system were accurate within ±0.6 mm of simulated ranges in a perspex phantom for all energies assessed. This system allows real-time read-out of individual detector channels also making it appropriate for temporal beam delivery diagnostics and for spot position monitoring along one axis. The system presented provides a suitable, economical and efficient alternative for daily QA in proton therapy.

Highly efficient and sensitive patient-specific quality assurance for spot-scanned proton therapy.

The purpose of this work was to develop an end-to-end patient-specific quality assurance (QA) technique for spot-scanned proton therapy that is more sensitive and efficient than traditional approaches. The patient-specific methodology relies on independently verifying the accuracy of the delivered proton fluence and the dose calculation in the heterogeneous patient volume. A Monte Carlo dose calculation engine, which was developed in-house, recalculates a planned dose distribution on the patient CT data set to verify the dose distribution represented by the treatment planning system. The plan is then delivered in a pre-treatment setting and logs of spot position and dose monitors, which are integrated into the treatment nozzle, are recorded. A computational routine compares the delivery log to the DICOM spot map used by the Monte Carlo calculation to ensure that the delivered parameters at the machine match the calculated plan. Measurements of dose planes using independent detector arrays, which historically are the standard approach to patient-specific QA, are not performed for every patient. The nozzle-integrated detectors are rigorously validated using independent detectors in regular QA intervals. The measured data are compared to the expected delivery patterns. The dose monitor reading deviations are reported in a histogram, while the spot position discrepancies are plotted vs. spot number to facilitate independent analysis of both random and systematic deviations. Action thresholds are linked to accuracy of the commissioned delivery system. Even when plan delivery is acceptable, the Monte Carlo second check system has identified dose calculation issues which would not have been illuminated using traditional, phantom-based measurement techniques. The efficiency and sensitivity of our patient-specific QA program has been improved by implementing a procedure which independently verifies patient dose calculation accuracy and plan delivery fidelity. Such an approach to QA requires holistic integration and maintenance of patient-specific and patient-independent QA.

Effects of Grapevine Leafroll associated Virus 3 (GLRaV-3) and duration of infection on fruit composition and wine chemical profile of Vitis vinifera L. cv. Sauvignon blanc.

In order to determine the effects of Grapevine Leafroll associated Virus 3 (GLRaV-3) on fruit composition and chemical profile of juice and wine from Vitis vinifera L. cv. Sauvignon blanc grown in New Zealand, composition variables were measured on fruit from vines either infected with GLRaV-3 (established or recent infections) or uninfected vines. Physiological ripeness (20.4°Brix) was the criterion established to determine the harvest date for each of the three treatments. Date of grape ripeness was strongly affected by virus infection. In juice and wine, GLRaV-3 infection prior to 2008 reduced titratable acidity compared with the uninfected control. Differences observed in amino acids from the three infection status groups did not modify basic wine chemical properties. In conclusion, GLRaV-3 infection slowed grape ripening, but at equivalent ripeness to result in minimal effects on the juice and wine chemistry. Time of infection produced differences in specific plant physiological variables.

Giant neonatal thoraco-abdominal lymphatic malformations treated with sildenafil: a case report and review of the literature.

Lymphatic malformations account for approximately 5% of benign tumors encountered during the neonatal period. Most often treated by surgical resection or sclerotherapy, phosphodiesterase inhibitors have recently emerged as a potential treatment modality for lymphatic malformations. Treatment with phosphodiesterase inhibitors may be particularly useful when patients are not candidates for surgical resection or sclerotherapy. We report a case of giant bilateral lymphatic malformations diagnosed prenatally and treated postnatally with sildenafil.

Waste stream utilisation for sustainable viticulture.

Field trials were established at four vineyards in January 1999 to evaluate the effects of four mulch mixtures on different soil and plant parameters. Mulches were made from wine industry and other commercially available plant and animal wastes. Soil, grape petioles, grape leaves and grape juice were analysed over three seasons. The mulches applied released considerable quantities of nutrients, which were available for use by the grapevines. Generally, the type of mulch used had little impact on the parameters that were measured and the greatest differences occurred between non-mulched and mulch treatments. Soil pH showed an increase at three of the four sites after application of mulch. Soil phosphorus increased moderately at one site and substantially at the other three sites in the first year and soil potassium levels increased dramatically at all sites in the first year. After the application of mulches in 1999 the petiole nitrate levels increased dramatically at all sites, however there were no differences in the second year. In the third year petiole nitrate levels were again high indicating that the differences between years was probably largely attributable to differences in rainfall received among the three seasons. Petiole potassium levels also increased after the application of mulch, however the increase was nowhere near as large as the increase in soil potassium. The use of mulch increased leaf nitrogen and potassium levels but not phosphorus levels. The use of mulch did increase juice potassium, however there was greater seasonal and site variation than variation due to the effect of mulch.

Security issues in the electronic transmission of prescriptions.

The UK government has stated within its plan of reform for the National Health Service that a secure system for the Electronic Transfer of Prescriptions will be available by 2004. The objectives of this paper are to highlight the significant barriers faced in securing an ETP system, to provide a critical analysis of the security mechanisms in the models currently being piloted and to suggest an alternative revised model which overcomes the identified deficiencies and security hurdles. To identify the significant security issues relevant to the adoption of ETP, the authors have combined their analysis of present prescription processing practice with their knowledge of computer security. The authors identify and describe how the issues of patient confidentiality, authorization, identity authentication, audit, scalability, availability and reliability are significant barriers to the adoption of ETP, particularly if they effect ease of use. The paper's contribution to the field of ETP is to suggest solutions to each of the identified security issues and to combine the solutions together in a revised and developed model.

Impact of segmental grafts on pediatric liver transplantation--a review of the United Network for Organ Sharing Scientific Registry data (1990-1996).

PURPOSE: The aim of this study was to assess the relative impact of segmental grafts from cadaveric and living donors on outcomes in 3,409 pediatric transplants (<18 years) between 1990 and 1996. METHODS: Analysis of the United Network for Organ Sharing (UNOS) Scientific registry data from 1990 to 1996 was performed. RESULTS: Liver grafts consisted of 2,636 whole grafts (WLG), 246 liver donor grafts (LDG), 89 split liver graft (SLG), and 438 reduced-size grafts (RSG). Although the number of pediatric transplants were unchanged between 1990 and 1996, segmental grafts made up an increasing proportion from 14.5% to 29.2%, and WLG decreased proportionately. The increase among segmental grafts occurred for LDG (threefold), followed by SLG (53%) and RSG (50%). One-year graft and patient survival rates for 3,409 transplants were 69.7% and 81.9%, respectively and were significantly higher (P<.001) in nonhospitalized patients than in hospitalized patients (79.8% and 91.3% v 61.0% and 73.7%). LDG graft survival (75.9%) was comparable with WLG(70.9%) but significantly better at 1 year than SLG (60.3%, P = .007) and RSG (61.1%, P = .001), even after excluding retransplants and ICU patients. Patient survival rates were not different statistically between groups. A separate analysis of outcomes in recipients less than 1 year of age suggested significantly better graft and patient survivals for LDG (83.3% and 89.4%) than for WLG (62.3% and 76.5%) and RSG (62.7% and 75%). CONCLUSIONS: Segmental liver grafts from cadaveric and living donors constitute an increasing proportion of pediatric transplants. Survival rates of cadaveric segmental graft are inferior to those of live donor segmental grafts even after adjustment for medical condition. Live donor grafts demonstrate consistently superior graft and patient outcomes in pediatric recipients less than 1 year of age, and should be promoted aggressively as a solution to the critical shortage of size matched grafts in small recipients.

Identification of the multicatalytic enzyme as a possible gamma-secretase for the amyloid precursor protein.

One of the main components of the senile plaques in brain tissue from patients with Alzheimer's disease is the beta-amyloid peptide. This peptide is proteolytically cleaved from the amyloid precursor protein by the action of at least two proteases, a beta-secretase which generates the N-terminus and a gamma-secretase which generates the C-terminus. Neither of these proteases have been identified. To identify proteases that are candidates for the gamma-secretase we synthesized a small fluorescent peptide substrate containing the amino acids comprising the C-terminus of the longest beta-amyloid peptide identified. This substrate is hydrolyzed by a single activity in homogenates from both cells and brain tissue and we have demonstrated that this activity is the multicatalytic enzyme or proteasome. Furthermore, using specific inhibitors, we have demonstrated that the fluorescent substrate is hydrolyzed by the chymotrypsin-like activity of the multicatalytic enzyme.

Mitosis and inhibition of intracellular transport stimulate palmitoylation of a 62-kD protein.

Recent studies suggest that a cycle of acylation/deacylation is involved in the vesicular transport of proteins between intracellular compartments at both the budding and the fusion stage (Glick, B. S., and J. E. Rothman. 1987. Nature (Lond.). 326:309-312). Since a number of cellular processes requiring vesicular transport are inhibited during mitosis, we examined the fatty acylation of proteins in interphase and mitotic cells. We have identified a major palmitoylated protein with an apparent molecular weight of 62,000 (p62), whose level of acylation increases 5-10-fold during mitosis. Acylation was reversible and p62 was no longer palmitoylated in cells that have exited mitosis and entered G1. p62 is tightly bound to the cytoplasmic side of membranes, since it was sensitive to digestion with proteases in the absence of detergent and was not removed by treatment with 1 M KCl. p62 is removed from membranes by nonionic detergents or concentrations of urea greater than 4 M. The localization of p62 by subcellular fractionation is consistent with it being in the cis-Golgi or the cis-Golgi network. A palmitoylated protein of the same molecular weight was also observed in interphase cells treated with inhibitors of intracellular transport, such as brefeldin A, monensin, carbonylcyanide m-chlorophenylhydrazone, or aluminum fluoride. The protein palmitoylated in the presence of brefeldin A was shown to be the same as that palmitoylated during mitosis using partial proteolysis. Digestion with two enzymes, alkaline protease and endoprotease lys-C, generated the same 3H-palmitate-labeled peptide fragments from p62 from mitotic or brefeldin A-treated cells. We suggest that the acylation and deacylation of p62 may be important in vesicular transport and that this process may be regulated during mitosis.

Cell-free fusion of endocytic vesicles is regulated by phosphorylation.

Okadaic acid and microcystin-LR, both potent inhibitors of protein phosphatases (PP), blocked vesicle fusion in a cell-free system. The effect of okadaic acid was reversed by the purified catalytic subunit of PP2A, but not PP1. Inhibition was gradual, required Mg-ATP, and was reduced by protein kinase inhibitors, indicating that it was mediated via protein phosphorylation. A candidate protein kinase would be cdc2 kinase, which normally is active in mitotic extracts and has been shown to inhibit endocytic vesicle fusion (Tuomikoski, T., M.-A. Felix, M. Dorée, and J. Gruenberg. 1989. Nature (Lond.). 342:942-945). However, it would appear that cdc2 kinase is not responsible for inhibition by okadaic acid. When compared to cytosol prepared from mitotic cells, okadaic acid did not increase cdc2 kinase activity sufficiently to account for the inhibition. In addition, inhibition was maintained when cdc2 protein was depleted from cytosol.

Mitotic cytosol inhibits invagination of coated pits in broken mitotic cells.

Receptor-mediated endocytosis is inhibited during mitosis in mammalian cells and earlier work on A431 cells suggested that one of the sites inhibited was the invagination of coated pits (Pypaert, M., J. M. Lucocq, and G. Warren. 1987. Eur. J. Cell Biol. 45: 23-29). To explore this inhibition further, we have reproduced it in broken HeLa cells. Mitotic or interphase cells were broken by freeze-thawing in liquid nitrogen and warmed in the presence of mitotic or interphase cytosol. Using a morphological assay, we found invagination to be inhibited only when mitotic cells were incubated in mitotic cytosol. This inhibition was reversed by diluting the cytosol during the incubation. Reversal was sensitive to okadaic acid, a potent phosphatase inhibitor, showing that phosphorylation was involved in the inhibition of invagination. This was confirmed using purified cdc2 kinase which alone could partially substitute for mitotic cytosol.

Seroprevalence of human immunodeficiency virus infection in a prenatal population at high risk for HIV infection.

To determine the seroprevalence of human immunodeficiency virus infection in prenatal patients at high risk for HIV infection we tested 513 women from December 1985 through July 1987 at an inner-city hospital in Atlanta, Georgia. Demographic and HIV risk information was collected from all seropositive women. Twenty-nine (6%) of the 513 women tested were positive for HIV on enzyme-linked immunosorbent assay and Western blot analysis. Twenty-six (90%) of seropositive women gave a history of intravenous drug use. Two (7%) had sexual partners known to have AIDS or AIDS-related complex (ARC), and one (3%) was Haitian. Seropositive women were at remarkable risk for other sexually transmitted diseases. The majority of pregnancies ended in term births. This serosurvey defines an obstetric population with a high seroprevalence, and has stimulated us to institute routine voluntary antepartum screening for HIV.

Ectopic pregnancy. What to do during the 20-day window.

An analysis of serial serum human chorionic gonadotropin (hCG) levels in 357 pregnant women revealed that the rise in hCG in normal intrauterine pregnancies differed from that in both ectopic gestations and abnormal intrauterine pregnancies. However, measurement of serial hCG levels cannot differentiate an ectopic from an abnormal intrauterine pregnancy. A protocol was developed for the management of patients with suspected ectopic pregnancy before ultrasound can reliably confirm an intrauterine gestation.

Specific inhibitors implicate a soluble metalloendoproteinase in exocytosis.

1. Previous studies have demonstrated that exocytosis in adrenal chromaffin cells appears to require zinc-dependent endoproteinase activity. 2. Chromaffin cells have metal-dependent endoproteinases in both the plasma membrane and the soluble fraction of homogenized cells. In order to further study critically the role of metalloproteinase in exocytosis, and prior to purification, we needed to determine which one of several adrenal metalloproteinases is implicated in exocytosis. 3. The studies described here demonstrate that the metal-dependent endoproteinases in these two subcellular fractions can be differentiated by selective inhibitors. In both intact and permeabilized cells, the plasma membrane metalloproteinase, but not the soluble proteinases, is inhibited by phosphoramidon. Phosphoramidon does not block exocytosis in either intact or permeabilized cells. 4. In addition, the plasma membrane metalloproteinase appears to have its catalytic site facing the outside of the cell. 5. Because of these observations the plasma membrane metalloproteinase does not appear to be required in exocytosis. Since soluble metalloproteinase activity is inhibited by proteinase inhibitors at concentrations which block exocytosis, a soluble, and not the plasma membrane, metalloproteinase appears to be required in exocytosis.

Evidence for the involvement of metalloendoproteases in the acrosome reaction in sea urchin sperm.

An essential initial step in fertilization in the sea urchin Strongylocentrotus purpuratus is an intracellular membrane fusion event in the sperm known as the acrosome reaction. This Ca2+-dependent, exocytotic process involves fusion of the membrane of the acrosomal vesicle and the plasma membrane. Recently, metalloendoproteases requiring divalent metals have been implicated in several Ca2+-dependent membrane fusion events in other biological systems. In view of the suggested involvement of Zn2+ in the sea urchin sperm acrosome reaction (Clapper, D.L., Davis, J.A., Lamothe, P.J., Patton, C., and Epel, D. (1985) J. Cell Biol. 100, 1817-1824) and the fact that Zn2+ is a metal cofactor for metalloendoproteases, we investigated the potential role of this protease in the acrosome reaction. A soluble metalloendoprotease was demonstrated and characterized in sperm homogenates using the fluorogenic protease substrate succinyl-alanine-alanine-phenylalanine-4-aminomethylcoumarin. The protease was inhibited by the metal chelators EDTA and 1,10-phenanthroline, and activity of the inactive apoenzyme could be reconstituted with Zn2+. The metalloendoprotease substrate and inhibitors blocked the acrosome reaction induced either by egg jelly coat or by ionophore, but had no effect on the influx of Ca2+. These observations suggest that inhibition occurs at a step independent of Ca2+ entry. Overall, the results of this study provide strong indirect evidence that the acrosome reaction requires the action of metalloendoprotease.

Paradoxical platelet behaviour in diabetic ketoacidosis.

Thrombotic events may occur in patients who present with severe uncontrolled diabetes or with diabetic coma. As a possible explanation for this, platelet function was investigated at presentation with diabetic ketoacidosis and during treatment in 10 patients. Concentrations of the platelet-specific proteins, platelet factor 4 (PF4) and beta-thromboglobulin (beta TG) were elevated and fell towards normal with treatment. Despite evidence of increased aggregation in vivo, platelets from subjects with ketoacidosis were insensitive to adenosine 5'-diphosphate (ADP), sensitivity increasing with correction of ketoacidosis. Platelets from ketoacidotic diabetics were initially insensitive to the anti-aggregatory action of prostacyclin (PGI2) and became normal with treatment. Initial blood glucose concentrations correlated with log10 ADP concentrations (r = 0.72, p less than 0.01) and with log10 PGI2 ID50 (the PGI2 concentration required to half-inhibit ADP-induced aggregation) (r = 0.66, p less than 0.025). Glucose concentrations throughout the 2-week study period correlated with all log10 ADP concentrations (r = 0.32, p less than 0.005) and all log10 PGI2 ID50 concentrations (r = 0.51, p less than 0.001). The decrease in ADP sensitivity in ketoacidosis, paradoxical in view of the evidence of increased in vivo platelet aggregation, may result from an acquired platelet storage pool deficiency.

Requirement for metalloendoprotease in exocytosis: evidence in mast cells and adrenal chromaffin cells.

Exocytosis is initiated by the receptor-mediated influx of calcium that results in fusion of the secretory vesicle with the plasma membrane. We examined the possibility that calcium-dependent exocytosis in mast cells and adrenal chromaffin cells requires metalloendoprotease activity. Metalloendoprotease inhibitors and dipeptide substrates block exocytosis in these cells with the same specificity and dose dependency as that with which they interact with metalloendoproteases. Metalloendoprotease activity is identified in these cells with fluorogenic synthetic substrates, which also blocked exocytosis. Metalloendoprotease activity is highest in the plasma membrane of chromaffin cells. The metalloendoprotease appears to be required in exocytosis at a step dependent on or after calcium entry, since exocytosis initiated by direct calcium introduction in both mast cells and chromaffin cells is blocked by metalloendoprotease inhibitors.