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New strongylophorines from the Okinawan marine sponge Petrosia (Strongylophora) corticata.

New strongylophorines-22 (1), -23 (2), -24 (3), and -25 (4) were isolated from the Okinawan sponge Petrosia (Strongylophora) corticata along with other known strongylophorines. The structures of these strongylophorines were determined on the basis of spectroscopic analysis and chemical conversions. Assessment was also made of the cytotoxicity of strongylophorines-1, -2, -3, -4, -22 (1), -23 (2), and -24 (3) toward HeLa cells.

Animals↗

New marine prostanoids from the okinawan soft coral, clavularia viridis

Two new marine prostanoids-17,18-dehydroclavulone I (1) and clavulolactone I (2)-were isolated from the Okinawan soft coral, Clavularia viridis. Their structures, including absolute configurations, were determined based on the results of spectroscopic analysis and chemical conversions.

Journal Article↗

Using others' words: conversational use of reported speech by individuals with aphasia and their communication partners.

Reported speech, wherein one quotes or paraphrases the speech of another, has been studied extensively as a set of linguistic and discourse practices. Researchers agree that reported speech is pervasive, found across languages, and used in diverse contexts. However, to date, there have been no studies of the use of reported speech among individuals with aphasia. Grounded in an interactional sociolinguistic perspective, the study presented here documents and analyzes the use of reported speech by 7 adults with mild to moderately severe aphasia and their routine communication partners. Each of the 7 pairs was videotaped in 4 everyday activities at home or around the community, yielding over 27 hr of conversational interaction for analysis. A coding scheme was developed that identified 5 types of explicitly marked reported speech: direct, indirect, projected, indexed, and undecided. Analysis of the data documented reported speech as a common discourse practice used successfully by the individuals with aphasia and their communication partners. All participants produced reported speech at least once, and across all observations the target pairs produced 400 reported speech episodes (RSEs), 149 by individuals with aphasia and 251 by their communication partners. For all participants, direct and indirect forms were the most prevalent (70% of RSEs). Situated discourse analysis of specific episodes of reported speech used by 3 of the pairs provides detailed portraits of the diverse interactional, referential, social, and discourse functions of reported speech and explores ways that the pairs used reported speech to successfully frame talk despite their ongoing management of aphasia.

Adolescent↗

A new in vitro tissue factor inhibitory triterpene from the fruits of Chaenomeles sinensis.

Tissue factor (TF, tissue thromboplastin) accelerates the blood clotting, activating both the intrinsic and the extrinsic pathways to serve as a cofactor. In order to isolate TF inhibitors from the fruits of Chaenomeles sinensis, an activity-guided purification was carried out to yield four triterpenoid compounds. One compound was new and its structure was elucidated as 28-O-beta- D-glucopyranosyl-2alpha,3beta-dihydroxyolean-12-ene-24,28-dioic acid (2), named chaenomeloside A by means of spectral analysis and chemical conversion. Other compounds were trachelosperoside A-1 (1), oleanolic acid (3) and ursolic acid (4). Compound 2 and its aglycone 2a, named chaenomelogenin A inhibited by 50 % the TF activity at concentrations of 0.036 and 0.028 mM/unit of TF, respectively. Compound 1 isolated for the first time from this plant as well as 3 and 4 were inactive.

Animals↗

Betulinic acid: isolation from Triphyophyllum peltatum and Ancistrocladus heyneanus, antimalarial activity, and crystal structure of the benzyl ester.

The known lupane-type triterpene betulinic acid (3) was isolated for the first time from Triphyophyllum peltatum and Ancistrocladus heyneanus. It was found to exhibit moderate to good in vitro antimalarial activity against asexual erythrocytic stages of the human malaria parasite Plasmodium falciparum. A first X-ray structure analysis succeeded after conversion into its benzyl ester 4.

Animals↗

X-ray structure of a DNA decamer containing 7,8-dihydro-8-oxoguanine.

We have determined the x-ray structure of a DNA fragment containing 7,8-dihydro-8-oxoguanine (G(O)). The structure of the duplex form of d(CCAGOCGCTGG) has been determined to 1.6-A resolution. The results demonstrate that GO forms Watson-Crick base pairs with the opposite C and that G(O) is in the anti conformation. Structural perturbations induced by C.G(O)anti base pairs are subtle. The structure allows us to identify probable elements by which the DNA repair protein MutM recognizes its substrates. Hydrogen bond donors/acceptors within the major groove are the most likely element. In that groove, the pattern of hydrogen-bond donors/acceptors of C.G(O)anti is unique. Additional structural analysis indicates that conversion of G to G(O) would not significantly influence the glycosidic torsion preference of the nucleoside. There is no steric interaction of the 8-oxygen of G(O) with the phospho-deoxyribose backbone.

Base Composition↗

Antibody response to sheep erythrocytes in Indian native vis-à-vis imported breeds of chickens.

1. A total of 433 birds (7 weeks old) of both sexes belonging to Indian native breeds, including, Aseel, Kadakanath, Naked Neck and Frizzle fowl along with the imported breeds Dahlem Red, White Leghorn, Synthetic dam line broiler (SDL) and Naked Neck broiler were utilised to test the primary antibody response to sheep erythrocytes by haemagglutination test. The effect of genotype (breed), sex and their interactions on antibody response were also studied. 2. The results revealed the presence of natural antibodies in all groups under study. 3. All groups except broilers showed the highest HA titre on day 5 post immunisation, which gradually declined until the end of the experiment (19th day post immunisation). In broilers, the peak HA titre was observed on day 12. 4. Dahlem Red showed the highest response throughout. The lowest antibody response was recorded for broilers except on day 19 post immunisation when it exceeded the White Leghorn value. 5. Amongst the native Indian breeds, the Naked neck had the highest titre on day 5 post immunisation but the Aseel titre was highest on days 12 and 19. 6. Males tended to have higher titres than females in Aseel, Kadakanath, Naked Neck, White Leghorn and Naked Neck broilers whereas Frizzle, Dahlem Red and SDL broilers showed the converse. 7. Statistical analysis revealed significant variation in HA response among the various genetic groups on different days post immunisation. The apparent differences between sexes were not significant. However, interactions between breed and sex were significant on day 5 and 19 post immunisation.

Animals↗

Quality assurance of patient dosimetry in boron neutron capture therapy.

The verification of the correctness of planned and executed treatments is imperative for safety in radiotherapy. The purpose of the present work is to describe and evaluate the quality assurance (QA) procedures for patient dosimetry implemented at the boron neutron capture therapy (BNCT) facility at Studsvik, Sweden. The dosimetric complexity of the mixed neutron-photon field during BNCT suggests a careful verification of routine procedures, specifically the treatment planning calculations. In the present study, two methods for QA of patient dosimetry are presented. The first is executed prior to radiotherapy and involves an independent check of the planned absorbed dose to be delivered to a point in the patient for each treatment field. The second QA procedure involves in vivo dosimetry measurements using post-treatment activation analysis. Absorbed dose conversion factors taking the difference in material composition and geometry of the patient and the PMMA phantom used for reference dosimetry were determined using the Monte Carlo method. The agreement of the QA procedure prior to radiotherapy reveals an acceptably small deviation for 60 treatment fields of +/-4.2% (1 SD), while the in vivo dosimetry method presented may benefit from improvements, as the deviations observed were quite substantial (+/- 12%, 1 SD), and were unlikely to be due to actual errors in the clinical dosimetry

Boron Neutron Capture Therapy↗

Spatial organization of active and inactive genes and noncoding DNA within chromosome territories.

The position of genes within the nucleus has been correlated with their transcriptional activity. The interchromosome domain model of nuclear organization suggests that genes preferentially locate at the surface of chromosome territories. Conversely, high resolution analysis of chromatin fibers suggests that chromosome territories do not present accessibility barriers to transcription machinery. To clarify the relationship between the organization of chromosome territories and gene expression, we have used fluorescence in situ hybridization to analyze the spatial organization of a contiguous approximately 1 Mb stretch of the Wilms' tumor, aniridia, genitourinary anomalies, mental retardation syndrome region of the human genome and the syntenic region in the mouse. These regions contain constitutively expressed genes, genes with tissue-restricted patterns of expression, and substantial regions of intergenic DNA. We find that there is a spatial organization within territories that is conserved between mouse and humans: certain sequences do preferentially locate at the periphery of the chromosome territories in both species. However, we do not detect genes necessarily at the periphery of chromosome territories or at the surface of subchromosomal domains. Intraterritory organization is not different among cell types that express different combinations of the genes under study. Our data demonstrate that transcription of both ubiquitous and tissue-restricted genes is not confined to the periphery of chromosome territories, suggesting that the basal transcription machinery and transcription factors can readily gain access to the chromosome interior.

Animals↗

Rapid depth dose determination by a computer-controlled dosimetry system.

A comprehensive depth dose measuring system controlled by a small computer has been designed to enable full depth dose information for a linear accelerator to be obtained within one day. An ionization microchamber is moved rapidly under computer control in a horizontal plane to preselected points within a water phantom by two stepping motors to a spatial resolution of 0-025 mm. The ionization current pulses are converted to DC voltages by an interface providing complete charge integration and yet having a rapid response to drastic changes in dose level. These are fed to the computer for analysis and subsequent conversion to depth dose data. The chamber path follows a preselected fan-shaped grid after automatic determination of the X-ray beam central axis. The combination of rapid chamber motion with halts for charge integration and computer sampling has resulted in fast data acquisition independent of measurement response times. This information, which is stored on disc, can be directly read by a treatment planning program. The overall accuracy of measurement lies within 0-5% of the true depth dose at any point. Central axis and transverse profiles can be assessed at the time of measurement through visualization on the computer storage oscilloscope.

Computers↗

Dmc1 of Schizosaccharomyces pombe plays a role in meiotic recombination.

We report here a Schizosaccharomyces pombe gene (dmc1(+)) that resembles budding yeast DMC1 in the region immediately upstream of the rad24(+) gene. We showed by northern and Southern blot analysis that dmc1(+) and rad24(+) are co-transcribed as a bicistronic mRNA of 2.8 kb with meiotic specificity, whereas rad24(+) itself is constitutively transcribed as a 1.0-kb mRNA species during meiosis. Induction of the bicistronic transcript is under the control of a meiosis-specific transcription factor, Ste11. Disruption of both dmc1(+) and rad24(+) had no effect on mitosis or spore formation, and dmc1Delta cells displayed no change in sensitivity to UV or gamma irradiation relative to the wild type. Tetrad analysis indicated that Dmc1 is involved in meiotic recombination. Analysis of gene conversion frequencies using single and double mutants of dmc1 and rhp51 indicated that both Dmc1 and Rhp51 function in meiotic gene conversion. These observations, together with a high level of sequence identity, indicate that the dmc1(+) gene of S. POMBE: is a structural homolog of budding yeast DMC1, sharing both similar and distinct functions in meiosis.

Cell Cycle Proteins↗

Depletion of cellular protein thiols as an indicator of arylation in isolated trout hepatocytes exposed to 1,4-benzoquinone.

A method to measure protein thiols (PrSH), reduced and oxidized, was adapted to determine PrSH depletion in isolated rainbow trout hepatocytes exposed to arylating agent 1,4-benzoquinone (BQ). Toxicant analysis revealed rapid conversion of BQ to 1, 4-hydroquinone (HQ) upon addition to hepatocytes. Hepatocytes exposed to 200 microM BQ+HQ showed 80% decline in glutathione (GSH) (1 h), 30% loss of PrSH (6 h), and no loss of viability (24 h). Recoverable oxidized PrSH was detected only after 24 h (200 microM BQ+HQ). Exposure to 600 microM BQ+HQ caused rapid (10 min) loss of > 90% GSH and > 60% PrSH, with eventual cell death. Half of the PrSH depletion at 6 h observed in hepatocytes exposed to 600 microM BQ+HQ was recoverable by reduction with dithiothreitol. Following the loss of GSH in hepatocytes exposed to 600 microM BQ+HQ, cellular PrSH were susceptible to direct arylation and oxidation. Rainbow trout hepatocytes, which contained 10-fold less GSH than rat cells, had a GSH:PrSH ratio of 1:82 compared with rat ratios of 1:2 to 1:6. The methods reported are useful for further study and discrimination of reactive modes of action needed for prediction of aquatic organism susceptibility to these types of toxicants.

Alkylation↗

Chronic hypoxemia selectively down-regulates 11beta-hydroxysteroid dehydrogenase type 2 gene expression in the fetal sheep kidney.

The present study was designed to examine the effects of chronic fetal placental embolization on the expression of 11beta-hydroxysteroid dehydrogenase (11beta-HSD) types 1 and 2, the intracellular enzymes responsible for the metabolism of glucocorticoids. Twelve instrumented fetal sheep were randomly allocated on Day 110 (term = 147 days) to either a control (n = 6) or embolized (n = 6) group. Embolized fetuses received daily injections of nonradioactive microspheres into the abdominal aorta for 21 days to decrease arterial oxygen content by 40-50% of the pre-embolization values. At the end of the experiment, fetal liver and kidney tissues as well as placental cotyledons were collected, and tissue levels of 11beta-HSD mRNA and activity were determined by standard Northern blot analysis and radiometric conversion assay, respectively. There was a 44% reduction (p < 0.01) in the level of renal 11beta-HSD2 mRNA in the embolized group as compared with the control group. Moreover, this reduction in mRNA was carried through to 11beta-HSD2 protein, since there was a corresponding decrease in the level of 11beta-HSD2 activity (4.5+/-0.2 vs. 2.9+/-0.1 pmol/min per milligram protein, p < 0.01). In contrast, levels of both 11beta-HSD1 mRNA and activity in the fetal liver remained unchanged. Moreover, both 11beta-HSD types 1 and 2 mRNA and activity in the placenta were not altered by the fetal placental embolization. In conclusion, chronic hypoxemia selectively inhibits renal 11beta-HSD2 mRNA expression and enzyme activity in the ovine fetus, which may contribute, at least in part, to the mechanisms leading to fetal hypertension.

11-beta-Hydroxysteroid Dehydrogenases↗

Prospective application of a multiplex reverse transcription-polymerase chain reaction assay for the detection of balanced translocations in leukaemia: a single-laboratory study of 390 paediatric and adult patients.

The upfront application of molecular methods for identifying the fusion transcripts arising from balanced translocations in haematopoietic malignancies has several advantages: sensitivity is independent of its frequency, i.e. rare ones are not missed, cytogenetically cryptic aberrations are identified and it provides a platform for minimal residual disease (MRD) detection. Employing a multiplex reverse transcription polymerase chain reaction (RT-PCR) assay identifying 27 fusion transcripts we prospectively analysed blood and/or bone marrow samples from 390 patients referred for diagnosis and treatment for acute leukaemia and chronic myeloproliferative disorders (CMPD) from a geographically well-defined region in Denmark. A total of 233 patients were diagnosed with acute myeloid leukaemia (AML), 95 with acute lymphoblastic leukaemia (ALL) origin and 62 patients were recorded as CMPD. Twenty-three percent AML, 32% ALL and 55% CMPD patients exhibited chromosomal aberrations detected by the multiplex RT-PCR. Cytogenetically cryptic translocations were seen in 15% of the cases. Conversely, the cytogenetic analysis identified chromosomal aberrations other than translocations in 45% of AML cases and 63% of ALL cases. We conclude that, while the fraction of translocation positive leukaemia patients in an unselected cohort is lower than hitherto believed, a molecular approach to their diagnosis is worthwhile, partly for identifying cryptic and rare translocations, partly for monitoring MRD.

Acute Disease↗

The management of sickle cell crisis pain as experienced by patients and their carers.

This study sought to provide insights into the personal experiences of individuals with sickle cell disorder and nurses involved in the management of painful sickle cell crisis. The sample consisted of 10 patients who experienced hospitalization for the management of pain during sickle cell crisis and 10 nurses who have cared for patients during such crises. The method of data collection used a combination of group and individual interviews, utilizing a 'guided conversation approach'. Data analysis consisted of content analysis of transcribed interviews which resulted in the identification of the following categories of problems: poor pain management, anxieties about pethidine, loss of control, lack of individuality and playing up. The results revealed that pain control during sickle cell crisis is often inadequate, with nurses admitting that they often have to resort to 'trial and error' strategy to manage pain. The findings are discussed in terms of attitudinal and knowledge factors and an individualized approach to pain management.

Acute Disease↗

Establishing joint attention in remote talks with the elderly about health: a study of nurses' conversation with elderly persons in teleconsultations.

This study is a contribution to the sparsely studied field of nurses' teleconsultations with frail elderly people. The aim was to describe how talk and interaction are accomplished and by what means problems are handled that emerge from the fact that the communication is conducted via videoconferencing techniques. Recordings were made of 22 sessions of teleconsultation between nurses and elderly nursing home residents aided by enrolled nurses. The data were analysed with a qualitative method inspired by conversation and discourse analysis. The findings indicate that in order to create a joint attention the participants often had to verbally confirm and reconfirm that contact had been established. The triad of participants played a special part in maintaining the joint attention through compensating for the contextual aspect of the medium's limitations and the demented elderly person's communication problems. 'Talks about the communication' and passages of social talk took place when the nurse or the staff member wanted to re-establish and maintain joint attention. The joint attention seemed connected to a special sort of eye contact and gaze and to the type of camera projection that was used. One conclusion was that to create joint attention in nurses' teleconsultations with frail elderly people, the limitations in transferring communication cues and the limitations of what the camera can reveal of the general context could, to some extent, be made up for by verbal communication within the triad of participants. Another conclusion was that these limitations in the context of interaction in some situations also seemed to be an advantage for the demented elderly and contributed to increased attention.

Aged↗

Recombinant purple acid phosphatase isoform 3 from sweet potato is an enzyme with a diiron metal center.

Purple acid phosphatases (PAPs) from sweet potato (sp) have been classified on the basis of their primary structure and the dinuclear metal center into isoforms spPAP1 [Fe(III)-Zn(II)] and spPAP2 [Fe(III)-Mn(II)]; for spPAP3 only the cDNA is known. With the aim of unraveling the character of the dinuclear metal center we report here the characterization of this isoform at the protein level. We cloned spPAP3 cDNA in a baculovirus and overexpressed this enzyme in Sf9 insect cells. Preparation of recombinant spPAP3 in two steps afforded pure enzyme with yields of 4.5 mg.L(-1) culture medium. This enzyme is a dimeric, disulfide-linked PAP of 110 kDa, similar to known PAP isoforms from higher plants. Enzymatic studies and spectroscopic properties (max. absorption at 550-565 nm) indicated a diiron enzyme; quantitative and semiquantitative metal analysis using ICP-OES and TOF-SIMS, respectively, revealed the presence of only iron in purified spPAP3. Metal replacement in the second metal-binding site upon preparation of the semiapo-enzyme with Fe(II), Zn(II), or Mn(II) showed highest activities with Fe(II). The data show that recombinant spPAP3 has a diiron metal center. Site-directed mutagenesis was conducted to check catalytic efficiency at the atomic level. Tyr291 at the substrate-binding site in spPAP3 was mutated to His and Ala, the respective residues found in spPAP1 and spPAP2. Kinetic analysis showed that conversion of Tyr291 to His further optimized the performance of this protein as a diiron enzyme, whereas the Ala mutation weakened the catalytic efficiency regardless of the metal present in the second binding site.

Acid Phosphatase↗

Metabolism of cyclohexane carboxylic acid by Alcaligenes strain W1.

Thirty-three microorganisms capable of growth with cyclohexane carboxylate as the sole source of carbon were isolated from mud, water, and soil samples from the Aberystwyth area. Preliminary screening and whole-cell oxidation studies suggested that, with one exception, all of the strains metabolized the growth substrate by beta-oxidation of the coenzyme A ester. This single distinctive strain, able to oxidize rapidly trans-4-hydroxycyclohexane carboxylate, 4-ketocyclohexane carboxylate, p-hydroxybenzoate, and protocatechuate when grown with cyclohexane carboxylate, was classified as a strain of Alcaligenes and given the number W1. Enzymes capable of converting cyclohexane carboxylate to p-hydroxybenzoate were induced by growth with the alicyclic acid and included the first unambiguous specimen of a cyclohexane carboxylate hydroxylase. Because it is a very fragile protein, attempts to stabilize the cyclohexane carboxylate hydroxylase so that a purification procedure could be developed have consistently failed. In limited studies with crude cell extracts, we found that hydroxylation occurred at the 4 position, probably yielding the trans isomer of 4-hydroxycyclohexane carboxylate. Simultaneous measurement of oxygen consumption and reduced nicotinamide adenine dinucleotide oxidation, coupled with an assessment of reactant stoichiometry, showed the enzyme to be a mixed-function oxygenase. Mass spectral analysis enabled the conversion of cyclohexane carboxylate to p-hydroxybenzoate by cell extracts to be established unequivocally, and all of our data were consistent with the pathway: cyclohexane carboxylate --> trans-4-hydroxycyclohexane carboxylate --> 4-ketocyclohexane carboxylate --> p-hydroxybenzoate. The further metabolism of p-hydroxybenzoate proceeded by meta fission and by the oxidative branch of the 2-hydroxy-4-carboxymuconic semialde-hyde-cleaving pathway.

Alcaligenes↗