Chemical generation of nitric oxide in the mouth from the enterosalivary circulation of dietary nitrate.

High concentrations of nitrite present in saliva (derived from dietary nitrate) may, upon acidification, generate nitrogen oxides in the stomach in sufficient amounts to provide protection from swallowed pathogens. We now show that, in the rat, reduction of nitrate to nitrite is confined to a specialized area on the posterior surface of the tongue, which is heavily colonized by bacteria, and that nitrate reduction is absent in germ-free rats. We also show that in humans increased salivary nitrite production resulting from nitrate intake enhances oral nitric oxide production. We propose that the salivary generation of nitrite is accomplished by a symbiotic relationship involving nitrate-reducing bacteria on the tongue surface, which is designed to provide host defence against microbial pathogens in the mouth and lower gut. These results provide further evidence for beneficial effects of dietary nitrate.

Study of X-ray field junction dose using an a-Si electronic portal imaging device.

Field junctions between megavoltage photon beams are important in modern radiotherapy for treatments such as head and neck and breast cancer. An electronic portal imaging device (EPID) may be used to study junction dose between two megavoltage X-ray fields. In this study, the junction dose was used to determine machine characteristics such as jaw positions and their reproducibility, collimator rotation and the effect of gantry rotation. All measurements were done on Varian linear accelerators with EPID (Varian, Palo Alto, CA). The results show reproducibility in jaw positions of approximately 0.3 mm for repeated jaw placement while EPID readings were reproducible within a standard deviation of 0.4% for fixed jaw positions. Junction dose also allowed collimator rotation error of 0.1 degrees to be observed. Dependence of junction dose on gantry rotation due to gravity was observed; the gravity effect being maximum at 180 degrees gantry angle (beam pointing up). EPIDs were found to be reliable tools for checking field junctions, which in turn may be used to check jaw reproducibility and collimator rotation of linacs.

Systematic variations in polymer gel dosimeter calibration due to container influence and deviations from water equivalence.

There are a number of gel dosimeter calibration methods in contemporary usage. The present study is a detailed Monte Carlo investigation into the accuracy of several calibration techniques. Results show that for most arrangements the dose to gel accurately reflects the dose to water, with the most accurate method involving the use of a large diameter flask of gel into which multiple small fields of varying dose are directed. The least accurate method was found to be that of a long test tube in a water phantom, coaxial with the beam. The large flask method is also the most straightforward and least likely to introduce errors during the set-up, though, to its detriment, the volume of gel required is much more than other methods.

Osteodystrophia fibrosa in two guinea pigs.

Two adult guinea pigs were examined because they were lethargic and reluctant to walk. Additionally, I guinea pig had otitis media, and the other had dental malocclusion. Both guinea pigs had been fed a commercially available diet of cereals and pellets enriched with vitamin C and formulated for this species. Radiographically, the guinea pigs had coarse trabecular bone patterns, skeletal deformations, pathologic fractures, and polyarthritic degenerative joint disease. A double cortical line was also evident on several long bones, the pelvis, and the vertebrae. A diagnosis of osteopenia was confirmed by use of dual-energy x-ray absorptiometry. Analysis of a food sample fed to 1 guinea pig revealed calcium and phosphorus contents of 0.524 and 0.425%, respectively (Ca:P ratio, 1.23:1). Microscopic examination of bone tissue from both guinea pigs revealed severe fibrous osteodystrophy. Nutritional secondary hyperparathyroidism caused by calcium-phosphorus imbalance was considered to be the underlying cause of osteodystrophia fibrosa in both guinea pigs.

Measurements of 60Co in spoons activated by neutrons during the JCO criticality accident at Tokai-mura in 1999.

Neutron activated items from the vicinity of the place where the JCO criticality accident occurred have been used to determine the fluence of neutrons around the facility and in nearby residential areas. By using underground laboratories for measuring the activation products, it is possible to extend the study to also cover radionuclides with very low activities from long-lived radionuclides. The present study describes gamma-ray spectrometry measurements undertaken in a range of underground laboratories for the purpose of measuring (60)Co more than 2 years after the criticality event. The measurements show that neutron fluence determined from (60)Co activity is in agreement with previous measurements using the short-lived radionuclides (51)Cr and (59)Fe. Limits on contamination of the samples with (60)Co are evaluated and shown to not greatly affect the utility of neutron fluence determinations using (60)Co activation.

Stereotactic fields shaped with a micro-multileaf collimator: systematic characterization of peripheral dose.

Despite the highly localized doses that may be delivered via stereotactic radiotherapy, a small dose is nonetheless delivered to out-of-field regions, which may cause detriment to the patient. In this work, a systematic set of dose measurements have been undertaken up to a distance of 45 cm from the isocentre, for stereotactic fields shaped by a BrainLAB mini-multileaf collimator (MMLC) mounted on a Varian 600C linear accelerator. A range of treatment parameters were varied so as to determine the factors of greatest influence and establish relationships with dose. The commercial treatment planning software (TPS) miscalculates the dose to out-of-field regions. Measured dose decreases consistently out to 45 cm, whereas the TPS decreases out to 10-15 cm, at which point the predicted dose is constant. At the 5-10 cm off-axis distance (OAD), measurements indicate doses of about 5-10% of the dose at the isocentre, 1% at 15 cm OAD and 0.1% at 45 cm OAD. There are several observed trends. Greater MMLC field sizes (with static jaw) result in higher out-of-field dose, as do shallower depths. The source-to-surface distance does not greatly influence peripheral dose. However, the results given in this work do indicate that simple treatment arrangements, such as preferable collimator rotation, would in certain cases reduce out-of-field dose by an order of magnitude. Peripheral dose raises questions of treatment optimization, particularly in cases where patients have a long life expectancy in which secondary effects may become manifest, such as in the treatment of paediatric patients or those with a non-malignant primary. For instance, for a 20 Gy hypo-fractionated treatment, dose to out-of-field regions is of the order of cGy-a substantial dose in radiation protection terms.

A role for wingless in the segmental gradient of Drosophila?

The wild-type functions of the Wnt family of genes are still little understood (for review see Nusse and Varmus, Cell 69, 1073-1087, 1992). In Drosophila, the wingless (D-Wnt-1) protein is expressed in segmental stripes: its absence leads to a complete failure of segmentation, loss of engrailed expression and lack of pattern in the cuticle. A predominating hypothesis is that the spatial distribution of wingless is crucial to pattern; it might carry an instructive signal from cells that secrete the protein to cells nearby, or it might form a concentration gradient which acts as a morphogen. We tested these hypotheses by expressing wingless ubiquitously in wingless- embryos. The distribution of wingless protein in these embryos is uniform. Despite this, engrailed expression persists, is confined to the most anterior third of the parasegment, and delineates the parasegment border. The cuticle shows a segmentally reiterated pattern and, dorsally, the denticles are normally distributed and oriented. Because all these position-specific features cannot have been placed by a local source or a differential distribution of wingless protein, we conclude that, in the early embryo, the role of wingless is neither to act as a local instructive signal, nor as a morphogen. We propose an alternative hypothesis that the wild-type function of the wingless protein is to maintain and 'seal' the parasegment borders; in its absence the borders fail to isolate abutting segmental gradients.

Expression of a 120,000 dalton protein during tumoricidal activation in murine peritoneal macrophages.

Modulation of protein expression during interferon-gamma (IFN-gamma)-lipopolysaccharide (LPS)-mediated macrophage tumoricidal activation has been examined by metabolic radiolabeling of various murine peritoneal macrophage populations with [35S]methionine followed by SDS-PAGE analysis. Although both IFN-gamma and LPS are capable of stimulating the expression of several proteins when used independently, combined treatment induced the enhanced or de novo expression of a 120,000 dalton polypeptide. The expression of this protein was synergistically regulated by both IFN-gamma and LPS in a manner strongly reminiscent of the functional synergism that these two agents exhibit with respect to induction of tumoricidal activity. p120 expression could be seen first at approximately 3 hr after the addition of both agents, reached optimal expression by 6 hr, and maintained elevated synthesis for up to 24 hr. This time course corresponds closely to that seen for the acquisition of tumoricidal competence. Macrophages elicited in the primed state of activity in vivo with methyl vinyl ether co-polymer II (MVE-II) did not express p120, but could be induced to do so when treated with low doses of LPS. Under similar conditions, MVE-II-elicited cells also acquire tumoricidal activity. Macrophages obtained from mice chronically infected with bacillus Calmette-Guerin constitutively expressed both p120 and cytolytic activity. If such macrophages were cultured for 24 hr, the expression of both events decayed and was lost, but could be restored by treatment with low doses of LPS. Thus the data support a strong correlation between the expression by macrophages of a novel 120,000 dalton protein and the expression of tumor cytotoxicity.

Two novel annexins from Drosophila melanogaster. Cloning, characterization, and differential expression in development.

The annexins are a family of homologous Ca2(+)- and phospholipid-binding proteins that until now have only been found in vertebrates. cDNA clones encoding two novel annexins from Drosophila melanogaster were isolated and characterized. RNA blots indicate that the messages for the two Drosophila proteins are differentially expressed in development, with one message being expressed throughout development, while the other is only found in early embryos and adult flies. In situ hybridizations localize the two Drosophila genes to 93B and 19A-4,7. A similarly high degree of homology relates Drosophila annexins to different vertebrate annexins, indicating that the Drosophila annexins are not the invertebrate homologues of particular mammalian annexins but that they constitute novel members of the annexin gene family. In continuation with a recently established terminology, the Drosophila annexins will be named annexins IX and X. The biochemical properties of Drosophila annexin X were investigated using recombinant protein. Similar to vertebrate annexins, annexin X bound to liver membranes and liposomes containing phosphatidylserine in a calcium-dependent manner but not to liposomes containing phosphatidylcholine. In addition, annexin X partitioned into the detergent phase of Triton X-114 as a function of calcium. The conservation of the annexin family of Ca2(+)-binding proteins in invertebrates suggests that they have a basic function in cells which is not peculiar to vertebrate biology, and the availability of the Drosophila sequences will open avenues for mutational studies of these functions.

Maleyl-BSA and fucoidan induce expression of a set of early proteins in murine mononuclear phagocytes.

We examined the effect of maleyl-BSA on specific protein expression in murine peritoneal macrophages by radiolabeling treated macrophages with [35S]methionine followed by SDS-polyacrylamide gel electrophoresis. Such treatment induces the expression of a set of at least seven proteins (38, 42, 57, 65, 75, 80, and 85 kD). A similar set of proteins is also induced by treatment of macrophages with the algal polysaccharide fucoidan. The proteins resemble those induced in response to treatment of this same cell population with bacterial lipopolysaccharide (LPS), as judged by co-migration in both one- and two-dimensional electrophoresis. Two proteins induced by either LPS or maleyl-BSA (e.g., p57 and p85) show similar primary structure, as assessed by partial proteolytic peptide mapping confirming their identity. The induction of these proteins by maleyl-BSA is a transient phenomenon, being expressed as early as 1 hr after treatment and declining after 8 hr even in the continuous presence of the stimulus. The dose of maleyl-BSA required to induce the response varies to some extent with the protein in question, but agrees with the Kd for ligand-receptor binding. Chloroquine, which blocks the degradation of ligand, does not inhibit the induction of early protein synthesis. Whereas the induction of these proteins is blocked by inhibition of RNA synthesis with actinomycin D, the reversible inhibition of protein synthesis with cycloheximide during the induction phase does not prevent their expression. LPS, maleyl-BSA, and fucoidan previously have been shown to stimulate protease secretion and tumoricidal function in appropriately primed macrophages. The present findings now demonstrate that all three agents can also mediate the expression of early genes which may participate in the acquisition of functional competence.

Protection against oral and gastrointestinal diseases: importance of dietary nitrate intake, oral nitrate reduction and enterosalivary nitrate circulation.

Over the last 20 years, dietary nitrate has been implicated in the formation of methemoglobin and carcinogenic nitrosamines in humans. This has led to restrictions of nitrate and nitrite levels in food and drinking water. However, there is no epidemiological evidence for an increased risk of gastric and intestinal cancer in population groups with high dietary vegetable or nitrate intake. A reevaluation of our currently very negative perception of dietary nitrates comes from recent research into the metabolism and enterosalivary circulation of nitrate in mammals. These studies showed that nitrate is converted to nitrite in the oral cavity that then "fuels" an important mammalian resistance mechanism against infectious diseases. Moreover, there is now evidence that the conversion of nitrate into oxides of nitrogen prevents the formation carcinogenic nitrosamines.