Verification of dynamic radiotherapy: the potential for 3D dosimetry under respiratory-like motion using polymer gel.

Following the implementation of advanced treatment procedures in radiotherapy, there is a need for dynamic dose verification in 3D. Gel dosimetry could potentially be used for such measurements. However, recently published data show that certain types of gels have a dose rate and fractionation dependence. The aim of this study was to investigate the feasibility of using a polymer gel dosimeter for dose verification of dynamic radiotherapy. To investigate the influence of dose rate dependence during respiratory-like motion in and out of the beam, a respiration robot together with two types of gel systems (normoxic methacrylic acid gel (nMAG) and normoxic polyacrylamide gel (nPAG)) were used. Reference measurements were obtained using a linear diode array (LDA). Expected results, if there was no influence of the dose rate variation, were calculated by convolving the static irradiated gel data with the motion function controlling the robot. To investigate the fractionation dependence, the gels were irradiated using gated and ungated deliveries. Magnetic resonance imaging was used to evaluate the absorbed dose response of the gel. The measured gel data coincided well with the LDA data. Also, the calculated data agreed well with the measured dynamic gel data, i.e. no dose rate dependence due to motion was observed. The difference in the R2 response for the gels receiving ungated and gated, i.e. fractionated, deliveries was less than 1% for the nPAG and 4% for the nMAG, for absorbed doses up to 2 Gy. The maximum difference was 1.2% for the nPAG and 9% for the nMAG, which occurred at the highest given dose (4 Gy). The investigated gels were found to be feasible detectors for dose measurements under respiratory-like motion. For dose verification of dynamic RT involving gated delivery, e.g. breathing-adapted radiotherapy, relative absorbed dose evaluation should be used in order to minimize the effects of fractionated irradiation.

Investigation of spatial resolution, partial volume effects and smoothing in functional MRI using artificial 3D time series.

This work addresses the balance between temporal signal-to-noise ratio (tSNR) and partial volume effects (PVE) in functional magnetic resonance imaging (fMRI) and investigates the impact of the choice of spatial resolution and smoothing. In fMRI, since physiological time courses are monitored, tSNR is of greater importance than image SNR. Improving SNR by an increase in voxel volume may be of negligible benefit when physiological fluctuations dominate the noise. Furthermore, at large voxel volumes, PVE are more pronounced, leading to an overall loss in performance. Artificial fMRI time series, based on high-resolution anatomical data, were used to simulate BOLD activation in a controlled manner. The performance was subsequently quantified as a measure of how well the resulted activation matched the simulated activation. The performance was highly dependent on the spatial resolution. At high contrast-to-noise ratio (CNR), the optimal voxel volume was small, i.e. in the region of 2(3) mm(3). It was also shown that using a substantially larger voxel volume in this case could potentially negate the CNR benefits. The optimal smoothing kernel width was dependent on the CNR, being larger at poor CNR. At CNR >1, little or no smoothing proved advantageous. The use of artificial time series gave an opportunity to quantitatively investigate the effects of partial volume and smoothing in single subject fMRI. It was shown that a proper choice of spatial resolution and smoothing kernel width is important for fMRI performance.

Dose integration characteristics in normoxic polymer gel dosimetry investigated using sequential beam irradiation.

Dose integration properties were investigated for normoxic polymer gels based on methacrylic acid (nMAG) and acrylamide/N, N'-methylenebisacrylamide (nPAG). The effect of sequential irradiation was studied for different fractionation schemes and varying amounts of methacrylic acid for the nMAG gels. Magnetic resonance imaging (MRI) was used for read out of the absorbed dose response. The investigated gels exhibited a dependence on the fractionation scheme. The response when the total dose was divided into fractions of 0.5 Gy was compared with the response when the total dose was delivered in a single fraction. The slope of the R2 versus the absorbed dose response decreased when the absorbed dose per fraction was increased. Also, for higher amounts of methacrylic acid in the nMAG system the difference in the response increased. For gels containing 2, 4, 6 and 8% methacrylic acid, the R2 versus the absorbed dose response increased by 35, 37, 63 and 93%, respectively. Furthermore, the effect of the fractionation was larger when a higher total absorbed dose was given. The effect was less pronounced for the investigated nPAG, containing 3% acrylamide and 3% N, N'-methylenebisacrylamide, than for the nMAG systems. Consequently, this study indicates that the nPAG system has preferable beam integration characteristics compared with the nMAG system.

Dose response evaluation of a low-density normoxic polymer gel dosimeter using MRI.

A low-density (approximately 0.6 g cm(-3)) normoxic polymer gel, containing the antioxidant tetrakis (hydroxymethyl) phosponium (THP), has been investigated with respect to basic absorbed dose response characteristics. The low density was obtained by mixing the gel with expanded polystyrene spheres. The depth dose data for 6 and 18 MV photons were compared with Monte Carlo calculations. A large volume phantom was irradiated in order to study the 3D dose distribution from a 6 MV field. Evaluation of the gel was carried out using magnetic resonance imaging. An approximately linear response was obtained for 1/T2 versus dose in the dose range of 2 to 8 Gy. A small decrease in the dose response was observed for increasing concentrations of THP. A good agreement between measured and Monte Carlo calculated data was obtained, both for test tubes and the larger 3D phantom. It was shown that a normoxic polymer gel with a reduced density could be obtained by adding expanded polystyrene spheres. In order to get reliable results, it is very important to have a uniform distribution of the gel and expanded polystyrene spheres in the phantom volume.

Development and optimization of a 2-hydroxyethylacrylate MRI polymer gel dosimeter.

In this study, radiation induced changes in a polymer gel dosimeter manufactured using 2-hydroxyethylacrylate (HEA) and N,N'-methylene-bisacrylamide (BIS) were investigated using magnetic resonance imaging (MRI) and FT-Raman spectroscopy. The variation in magnetic resonance relaxation time (T2) with absorbed dose was modelled assuming fast exchange of magnetization. Overall good agreement between the model and experimental data was obtained. However, comparison with FT-Raman data suggests that not all the protons attached to the polymer contribute to the relaxation process. Furthermore, for certain compositions improved agreement with experimental data was achieved when a lower fraction of polymer protons available for exchange with water was assumed in the low dose region. This indicates that the T2 value is influenced by the composition and topology of the formed polymer, which may vary with absorbed dose. The concept of percentage dose resolution (Dp delta, %) was introduced to enable optimization of gel compositions for use in relative dosimetry applications. This concept was applied to demonstrate the effects of varying the gelatine concentration, the total fraction of monomer/crosslinker (%T) and the relative fraction of crosslinker (%C) on gel performance in HEA gels as well as compare the performance of HEA and a standard polyacrylamide gel (PAG). The percentage dose resolution was improved for all HEA gels compared to the PAG dosimeter containing 3% acrylamide and 3% BIS. Increasing the total concentration of monomer was shown to have the largest single effect. In the range of doses of interest for clinical radiation therapy, Dp delta, % for the optimal HEA gel (4% HEA, 4% BIS) was lower than 2.3%, compared to 3.8% for the PAG dosimeter.

Determination of fumonisins B1 and B2 in various maize products by a combined SAX + C18 column and immunoaffinity column.

Fumonisins B1 and B2 were determined in 42 samples of different maize products from the Swedish market by 2 different methods based on cleanup steps using an immunoaffinity column and a combination of SAX + C18 columns, respectively. A simple "precipitation step" was included before the samples were added to the main column(s), giving less column clogging, fewer interfering peaks, and better recoveries for the different sample matrixes. Recovery, repeatability, and results from the survey showed comparable results with the methods. The limit of detection for both methods was 5 micrograms/kg for fumonisin B1 and 10 micrograms/kg for fumonisin B2. All 7 maize chips analyzed and 6 of 8 popcorn samples contained fumonisins (B1 + B2) with averages of 180 and 115 micrograms/kg, respectively. All other samples except a maize flour sample contained little or no fumonisins.

Migration of bisphenol-A diglycidyl ether (BADGE) and its reaction products in canned foods.

Bisphenol-A diglycidyl ether (BADGE) is used as an additive or starting agent in coatings for cans. The presence of hydrochloric acid in the organosol (PVC-based) lacquers results in formation of chlorohydroxy compounds of BADGE. These compounds, as well as BADGE itself, are potential migrants into the preserved food and are of toxicological concern. In the present investigation the presence of BADGE and the chlorohydroxy compounds (BADGE.HCl and BADGE.2HCl) in various kinds of canned foods from 30 brands have been determined by HPLC with fluorescence detection. BADGE was found in levels up to 5.1 mg/kg in the food and only in food from cans containing BADGE.HCl and BADGE.2HCl in the lacquers. BADGE was found both in fish in oil and in fish in tomato sauce, however, the highest amounts were found in the fatty foodstuffs. BADGE.HCl and BADGE.2HCl were found in concentrations up to 2.4 mg/kg and 8.3 mg/kg, respectively. Unlike BADGE, BADGE.2HCl was found in similar concentrations in fish in oil and in fish in tomato sauce. In aqueous and acidic foodstuffs BADGE readily hydrolyses into mono- and dihydrolysed products (BADGE.H2O and BADGE.2H2O). In this study BADGE.H2O was not found in any food sample, whereas BADGE.2H2O was found in levels up to 2.6 mg/kg. The Scientific Committee for Food (SCF) of the European Commission has proposed that a limit of restriction of 1 mg/kg food shall include BADGE itself and BADGE.H2O, BADGE.HCl, BADGE.2HCl and BADGE.HCL.H2O. The present results indicate that the migration of BADGE.HCl and BADGE.2HCl, compounds with almost no data on toxicity, implies a greater problem than BADGE.H2O and BADGE.2H2O.

Migration of epoxidized soya bean oil from plasticized PVC gaskets into baby food.

Expoxidized soya bean oil (ESBO) is used as a plasticizer in PVC gaskets in lids for glass jars used for packaging of ready-cooked baby food. The migration of ESBO from the lids has been determined in 81 samples of different dishes of baby food, including purées of beef, pork, fish, poultry, berries and vegetables. The level of ESBO in baby food has been determined using a gas chromatography/mass spectrometry (GC/MS) analytical procedure with a detection limit of 1.5 mg/kg. Residues of ESBO were found in all dishes except in blueberries. The levels ranged from < 1.5 to 50.8 mg/kg, with a mean of 11.9 mg/kg and a median of 7.8 mg/kg in food with detectable levels. Expoxidized fatty acids may also occur naturally in food. Baby food which had never been in contact with the lids was therefore analysed and no detectable levels of diepoxidized C18-methylester, which was used for the determination of ESBO, were found. That demonstrates that the presented levels of ESBO in the baby food are only due to migration from the lids and not of natural origin.

Deletion analysis of the Brassica napus cruciferin gene cru 1 promoter in transformed tobacco: promoter activity during early and late stages of embryogenesis is influenced by cis-acting elements in partially separate regions.

To define sequences in the cruciferin gene cru1 promoter of importance for expression, tobacco (Nicotina tabacum L.) plants were transformed with constructs in which the cru1 promoter, in front of the intact cru1 structural gene, was truncated at -1216, -974, -736, -515, -306, -46 and -17 bp relative to the cap-site. Cru1 expression in tobacco seeds was studied by Northern analysis, Western analysis and in-situ hybridizations. Comparisons of the Northern analysis of RNA from tobacco seeds harvested at 18 d after pollination with the Western analysis of protein from mature seeds showed that the regions between -974 to -736 and -306 to -46 were important for the expression of cru1 at an early developmental stage, whereas the regions -736 to -515 and -515 to -306 were important for expression throughout embryogenesis. By investigating the mRNA levels in transgenic seeds at different stages of development, indications were obtained that the two latter regions exerted their effects during the later stages. The in-situ hybridization showed that cru1 mRNA was distributed in parenchyma cells throughout the embryo in seeds expressing constructs -974 and -736. Constructs -515 and -306 showed an expression restricted to the axis or axis and parts of the cotyledons. Sequence comparisons of the cru1 promoter with other storage-protein gene promoters, identified several motifs implicated in gene regulation.(ABSTRACT TRUNCATED AT 250 WORDS)

Cruciferin gene families are expressed coordinately but with tissue-specific differences during Brassica napus seed development.

The major storage protein in seeds of Brassica napus, the 12S globulin cruciferin, is composed of three different groups of subunits; cru1, cru2/3 and cru4. By using gene family-specific probes, we have investigated the accumulation, rate of synthesis and spatial distribution of transcripts corresponding to the different groups of cruciferin subunits in developing seeds. Cruciferin transcripts derived from different gene families accumulate coordinately to comparable amounts during seed development. The corresponding gene families are, however, transcribed at different rates. Investigation of the spatial distribution of transcripts corresponding to each group of cruciferin subunits in the developing seed by in situ hybridization, revealed that mRNAs of all three types accumulate in both axis and cotyledons. Transcripts derived from cru1 and cru4 gene families show a similar cell specificity and accumulate in a similar spatial manner during seed development. In contrast, mRNAs corresponding to the cru2/3 gene family are expressed with a partly different cell specificity and show a slightly different pattern of accumulation in the axis and cotyledons, with a delayed accumulation in epidermal cells. In the cotyledons, the initial accumulation of this type of cruciferin mRNAs is also distinguished from the two other types. The differences in cell specificity are seen in the root cap and in provascular cells, where mRNAs belonging to the cru2/3 family are absent.

Analysis of the promoter region of napin genes from Brassica napus demonstrates binding of nuclear protein in vitro to a conserved sequence motif.

Napin is a seed storage protein from Brassica napus (rape) that is encoded by a gene family. We have isolated and characterized a novel napin gene, napB. Comparisons of the 5'-upstream region of napB to the promoter regions of previously published napin genes reveal that certain sequence motives are evolutionary conserved and may be implicated in gene regulation. These consensus motives, that overlap with purine/pyrimidine stretches, are TACACAT and CATGCA both of which frequently occur as overlapping, direct repeats. Related or identical sequences are also found in the upstream regions of the homologous genes of Arabidopsis thaliana. One copy of the CATGCA motif occurs in close proximity to the TATA box in all the above genes. In this case it overlaps with an octamer sequence (ATGCAAAT) which is a sequence element common in many eukaryotic promoters and enhancers. The TACACAT sequence, as part of a longer purine/pyrimidine stretch, was found to interact with a protein present in crude nuclear extracts from developing B. napus seeds. Napin genes appear to be methylated to almost equal extents whether present in expressing or non-expressing tissue.

Influence of age on atelectasis formation and gas exchange impairment during general anaesthesia.

We have studied the effects of anaesthesia on atelectasis formation and gas exchange in 45 patients of both sexes, smokers and nonsmokers, aged 23-69 yr. None of the patients showed clinical signs of pulmonary disease, and preoperative spirometry was normal. In the awake patient, partial pressure of arterial oxygen (PaO2) decreased with increasing age (P less than 0.001) and the alveolar-arterial oxygen partial pressure difference (PAO2-PaO2) increased with age (P less than 0.001). Shunt, assessed by the multiple inert gas elimination technique, was small (mean 0.5%) and uninfluenced by age. However, there was an increasing dispersion (log SD Q) of ventilation/perfusion ratios (VA/Q) and increasing perfusion of regions of low VA/Q (VA/Q less than 0.1) with increasing age (P less than 0.001 and P less than 0.05, respectively). No patient displayed any atelectasis as assessed by computed x-ray tomography of the chest. During inhalation anaesthesia (halothane or enflurane) with mechanical ventilation, 39 of 45 patients developed atelectasis and shunt. There was a strong correlation between the atelectatic area and the magnitude of shunt (r = 0.81, P less than 0.001). Atelectasis and shunt did not increase significantly with age, whereas log SD Q and perfusion of regions with low VA/Q ratios did (r = 0.55, P less than 0.001 and r = 0.35, P less than 0.05, respectively). Awake, the major determinant of PaO2 was perfusion of regions of low VA/Q ratios, which increased with age. During anaesthesia shunt influenced PaO2 most, low VA/Q being a secondary factor which, however, was increasingly important with increasing age, thus explaining the well-known age-dependent deterioration of arterial oxygenation during anaesthesia.

Effects of estramustine on DNA and cell membrane in malignant glioma cells.

Estramustine phosphate (EMP), a complex between estradiol-17 beta and nor-nitrogen mustard, commonly used in treatment of prostatic cancer, also exerts marked antiproliferative effects on cultured human malignant glioma cells. The mechanism of action is unknown but has previously been considered to be mediated through non-DNA targets, specifically via the mitotic spindle, and related to the intact estramustine complex. EMP cytotoxicity was studied on the malignant glioma cell line U-251 MG. A dose-dependent increase in DNA strand breaks was demonstrated at EMP-concentrations ranging 10-40 mg/l. The uptake of 86Rb, used as a tracer for potassium to study ion transport and membrane permeability, was reduced after incubation with EMP. The mean decline in 86Rb accumulation by U-251 MG cells was 12, 20 and 32% at EMP concentrations 10, 20 and 40 mg/l respectively. Scanning electron microscopy gave further evidence for cell membrane damage. In conclusion, EMP seems to affect malignant glioma cells on several vital functions and the results indicate the the cytotoxic potential may at least partially be related to effects on DNA and cell membrane.

Intermediate filaments in the newborn inner ear of the mouse.

The presence of intermediate filaments in the inner ear of the newborn mouse was analyzed with immunofluorescence techniques using antibodies against the five classes of intermediate filaments: cytokeratins, vimentin, desmin, neurofilaments and glial fibrillary acid protein (GFA). Neurofilaments were found in all nerve fibers from the ganglion cell to the hair cell. In the vestibular ganglion two subpopulations of ganglion cells were identified: a minor part staining intensively with neurofilament and the major part of cells lacking this immunofluorescence. Vimentin occurred in a number of supporting structures in the membranous labyrinth, but not in vestibular or cochlear ganglion cells. Cytokeratins, desmin or GFA were not identified in the inner ear.

Magnesium and membrane function in vascular smooth muscle.

Since a tight electromechanical coupling exists in vascular smooth muscle, even small shifts of the membrane potential are sufficient to change the vascular lumen. The extracellular H+ and K+ concentrations are important effectors for the adjustment of the membrane potential. The ion concentrations in the immediate neighbourhood of the cell membrane can be influenced by the microdynamic binding properties of the basement membrane and the other vascular connective tissue. These structures are polyanionic macromolecules to which mono- and divalent cations are extensively bound, and which are separated from the vascular smooth muscle cell membranes by tiny cleft spaces. The ion binding properties of vascular connective tissue were therefore studied in dependence on proton and cation concentration. The pH-dependent binding of monovalent cations to vascular connective tissue is dependent on the concentration and affinity constant of the ion species in question. The mode of interaction is competition. For instance, the actual K binding characteristic means that an increase of [K+]o close to the cell membrane cold ensue from a diminished K+ binding ability under alkalosis. Depolarization and contraction of vascular smooth muscle cells result. Divalent cation binding to vascular connective tissue it additionally dependent on conformational changes. Already physiological concentrations of Mg++ ions can induce a specific change in configuration, which enables K+ ions to bind cooperatively. This means that with extracellular Mg++ deficiency not only less Mg++ ions are bound to vascular connective tissue but also less K+ ions. [K+]o would increase near the cell membrane, depolarization and vasoconstriction would occur.

Sodium and calcium binding to Panulirus interruptus hemocyanin as studied by 23Na nuclear magnetic resonance.

Addition of Panulirus hemocyanin to NaCl solutions produces marked changes in the 23Na relaxation parameters; they show that sodium ions interact with binding sites on the protein and exchange rapidly with the bulk. The observed non-lorentzian lineshapes and the non-exponential decay of the transverse magnetization indicate that non-extreme narrowing conditions apply and give information on the dynamics of the interaction. Panulirus hemocyanin has at least two classes of Na+ binding sites; the binding constant of the more strongly bound sodium ions is in the order of 1 X 10(2) M-1. Competition between Na+ and Ca2+ for protein binding sites is demonstrated by the effect of Ca2+ on the 23Na relaxation parameters. However, only the more strongly bound Na+ are displaced by Ca2+. The number of Ca2+ needed to displace these sodium ions is 3--5 per oxygen binding site. The 23Na relaxation parameters are influenced also by the state of oxygenation of the protein, indicating a linkage between Na+ and oxygen binding. The simplest interpretation of the data is that sodium ions bind more strongly to oxyhemocyanin in agreement with oxygen equilibrium experiments.