Plenary discussion--Experimental studies sessions I and II.
Explore the source record for details and available documents.
Animals↗
Biomedical subjects
R D Saunders
Publications and source records attributed to R D Saunders.
Wavelength dependence of cellular responses in human melanocytes and melanoma cells following exposure to ultraviolet radiation.
PURPOSE: To examine the wavelength dependence of cellular responses in human melanocytes and human melanoma cells exposed to ultraviolet radiation (UVR). MATERIALS AND METHODS: Primary human melanocytes and G361 human melanoma cells were exposed to ultraviolet-C (UVC), ultraviolet-B (UVB), or ultraviolet-A (UVA) radiation. Dose-response relationships for clonal cell survival were assessed, and flow cytometry was used to monitor cell cycle distributions for up to one week post-irradiation. Chromosomal aberrations were scored in exposed and unexposed melanoma cells. RESULTS: G361 melanoma cells were more sensitive than melanocytes to killing by UVB and UVC radiation. This difference in sensitivity between cell types was much less marked following UVA irradiation. The melanoma cells showed a sustained, dose-dependent G2/M block following exposure with all wavelengths; in addition, transit through S phase was slowed following UVA irradiation. There was no apparent block to G1 cells entering S phase at any wavelength. Melanocytes, on the other hand, showed a marked G1 arrest, particularly following UVA irradiation. Cytogenetic results showed a dose-dependent increase in chromatid-type aberrations, mostly gaps, breaks and exchanges, in exposed melanoma cells. CONCLUSION: These results show that G361 malignant melanoma cells have lost the ability to regulate the cell cycle at the G1/S checkpoint and are more sensitive than melanocytes to cell killing by UVC and UVB but not UVA radiation. Similarly, exposure of these melanoma cells to UVC and UVB, and to a much lesser extent UVA, induced chromatid aberrations. UVA nevertheless induced strong cell cycle delays in both cell types, indicating that UVA exposure can significantly affect genome metabolism.
Animals↗
Rapporteur report: weak field interactions in the central nervous system.
Experimental evidence was presented of the interaction of extremely low frequency (ELF) physiologically weak electric fields (that is, internal fields of around 1-1000 mV m(-1)) with tissues of the central nervous system and the consequences of this for visual and cognitive function. These internal fields are higher than those induced in the body by exposure to environmental EMFs, but are small compared to nerve excitation thresholds. Two papers considered the effect of weak electric fields directly applied to hippocampal tissue in vitro on neuronal excitability assessed from evoked responses and on the modulation of neural network activity involving large numbers of individual neurons. A review was presented of the effects on electrical measures of perceptual and cognitive processes and on the performance of various behavioural tasks in volunteers exposed to electromagnetic fields. Finally, two papers addressed the visual perception of flickering light--phosphenes--in volunteers exposed to ELF magnetic fields. The first examined possible physiological origins of this phenomenon and the second assessed the induced electric fields and currents in the retina, using complex dosimetric modelling. These papers were discussed in several plenary sessions and in one breakout group, all of which are briefly summarised in this report.
Animals↗
Single, brief exposure to a 50 Hz magnetic field does not affect the performance of an object recognition task in adult mice.
A number of studies have shown that power frequency magnetic fields may affect spatial memory functions in rodents. An experiment was performed using a spontaneous object recognition task to investigate if nonspatial working memory was similarly affected. Memory changes in adult, male C57BL/6J mice were assessed by measuring the relative time within which the animals explored familiar or novel stimulus objects. Between initial testing and retesting, the animals were exposed for 45 min to a 50 Hz magnetic field at either 7.5 microT, 75 microT or 0.75 mT. Other animals were sham-exposed with ambient fields of less than 50 nT. No significant field-dependent effects on the performance of the task were observed at any flux density (for all measures, P > 0.05). These data provide no evidence to suggest that nonspatial working memory was affected in mice by acute exposure to an intense 50 Hz magnetic field.
Animals↗
Role of stress responses in human cell survival following exposure to ultraviolet C radiation.
PURPOSE: To investigate in human skin and other cells the role of tyrosine kinase and protein kinase-C (PKC) in eliciting cell-signalling responses to UV radiation (UVR) that affect the survival of irradiated cells. MATERIALS AND METHODS: The survival of HeLa S3 cells, NCTC 2544 human keratinocytes and A431 human epidermal carcinoma cells was measured following incubation with various tyrosine kinase or PKC inhibitors and exposure to UVC (254nm) radiation. In addition, Western blotting measured PKC isozyme expression in human keratinocytes following UVC exposure. RESULTS: It was confirmed that inhibition of tyrosine kinase activation reduces the survival of UV-irradiated HeLa S3 cells. However, no effect was seen on the survival of either NCTC 2544 human keratinocytes or A431 human epidermal carcinoma cells. In contrast, specific inhibition of PKC reduced the survival of UV-irradiated keratinocytes but had no effect on HeLa cells. Comparison of the effects of different inhibitors in keratinocytes suggested that this effect was mediated mostly through PKCmu and PKClambda/iota. In addition, keratinocyte exposure to UVC induced large and temporally distinct increases in PKCmu and PKClambda/iota. CONCLUSIONS: The survival of NCTC 2544 keratinocytes, but not HeLa S3 cells, following UVC exposure is mediated by signalling through PKC, mostly PKCmu and PKClambda/iota. Further study is required to confirm these results in normal human keratinocytes.
Carcinoma, Squamous Cell↗
From first base: the sequence of the tip of the X chromosome of Drosophila melanogaster, a comparison of two sequencing strategies.
We present the sequence of a contiguous 2.63 Mb of DNA extending from the tip of the X chromosome of Drosophila melanogaster. Within this sequence, we predict 277 protein coding genes, of which 94 had been sequenced already in the course of studying the biology of their gene products, and examples of 12 different transposable elements. We show that an interval between bands 3A2 and 3C2, believed in the 1970s to show a correlation between the number of bands on the polytene chromosomes and the 20 genes identified by conventional genetics, is predicted to contain 45 genes from its DNA sequence. We have determined the insertion sites of P-elements from 111 mutant lines, about half of which are in a position likely to affect the expression of novel predicted genes, thus representing a resource for subsequent functional genomic analysis. We compare the European Drosophila Genome Project sequence with the corresponding part of the independently assembled and annotated Joint Sequence determined through "shotgun" sequencing. Discounting differences in the distribution of known transposable elements between the strains sequenced in the two projects, we detected three major sequence differences, two of which are probably explained by errors in assembly; the origin of the third major difference is unclear. In addition there are eight sequence gaps within the Joint Sequence. At least six of these eight gaps are likely to be sites of transposable elements; the other two are complex. Of the 275 genes in common to both projects, 60% are identical within 1% of their predicted amino-acid sequence and 31% show minor differences such as in choice of translation initiation or termination codons; the remaining 9% show major differences in interpretation.
Animals↗
Molecular cloning of Drosophila gamma-glutamylcysteine synthetase by functional complementation of a yeast mutant.
gamma-Glutamylcysteine synthetase (GCS) catalyses a critical, rate-limiting step in glutathione synthesis. In this study we describe the isolation and characterisation of a GCS cDNA (pDmGCS4.3. 3) from Drosophila melanogaster by functional complementation of a Saccharomyces cerevisiae gsh1 mutant. Expression of pDmGCS4.3.3 in the yeast mutant partially restored glutathione levels and conferred resistance to methylglyoxal. The pDmGCS4.3.3 cDNA was found to be approx. 4.6 kb in length, containing a 2 kb fragment encoding an open reading frame with a high degree of deduced amino acid sequence identity with previously reported GCS sequences. In situ hybridisation revealed that the Drosophila GCS gene maps to 7D6-9 on the X chromosome.
Amino Acid Sequence↗
Low-level exposure to pulsed 900 MHz microwave radiation does not cause deficits in the performance of a spatial learning task in mice.
There is some concern that short-term memory loss or other cognitive effects may be associated with the use of mobile cellular telephones. In this experiment, the effect of repeated, acute exposure to a low intensity 900 MHz radiofrequency (RF) field pulsed at 217 Hz was explored using an appetitively-motivated spatial learning and working memory task. Adult male C57BL/6J mice were exposed under far field conditions in a GTEM cell for 45 min each day for 10 days at an average whole-body specific energy absorption rate (SAR) of 0.05 W/kg. Their performance in an 8-arm radial maze was compared to that of sham-exposed control animals. All behavioral assessments were performed without handlers having knowledge of the exposure status of the animals. Animals were tested in the maze immediately following exposure or after a delay of 15 or 30 min. No significant field-dependent effects on performance were observed in choice accuracy or in total times to complete the task across the experiment. These results suggest that exposure to RF radiation simulating a digital wireless telephone (GSM) signal under the conditions of this experiment does not affect the acquisition of the learned response. Further studies are planned to explore the effects of other SARs on learned behavior. Bioelectromagnetics 21:151-158, 2000. Published 2000 Wiley-Liss, Inc.
Analysis of Variance↗
The Drosophila mus101 gene, which links DNA repair, replication and condensation of heterochromatin in mitosis, encodes a protein with seven BRCA1 C-terminus domains.
The mutagen-sensitive-101 (mus101) gene of Drosophila melanogaster was first identified 25 years ago through mutations conferring larval hypersensitivity to DNA-damaging agents. Other alleles of mus101 causing different phenotypes were later isolated: a female sterile allele results in a defect in a tissue-specific form of DNA synthesis (chorion gene amplification) and lethal alleles cause mitotic chromosome instability that can be observed genetically and cytologically. The latter phenotype presents as a striking failure of mitotic chromosomes of larval neuroblasts to undergo condensation of pericentric heterochromatic regions, as we show for a newly described mutant carrying lethal allele mus101(lcd). To gain further insight into the function of the Mus101 protein we have molecularly cloned the gene using a positional cloning strategy. We report here that mus101 encodes a member of the BRCT (BRCA1 C terminus) domain superfamily of proteins implicated in DNA repair and cell cycle checkpoint control. Mus101, which contains seven BRCT domains distributed throughout its length, is most similar to human TopBP1, a protein identified through its in vitro association with DNA topoisomerase IIbeta. Mus101 also shares sequence similarity with the fission yeast Rad4/Cut5 protein required for repair, replication, and checkpoint control, suggesting that the two proteins may be functional homologs.
Amino Acid Sequence↗
In situ hybridization to polytene chromosomes.
Explore the source record for details and available documents.
Animals↗
Effects of 50 Hz magnetic field exposure on the rate of RNA synthesis by normal human fibroblasts.
PURPOSE: To investigate whether exposure to magnetic fields can affect the rate of RNA synthesis, a broad measure of cellular activity. MATERIALS AND METHODS: Normal human fibroblasts were exposed to 50 Hz magnetic fields at a range of flux densities between 2 microT and 20 mT. The rate of synthesis of total RNA was determined by following the incorporation of [3H]uridine into macromolecular material. In addition, polyadenylated RNA was isolated and used to estimate the rate of synthesis of mRNA. RESULTS: Incorporation of [3H]uridine into both total and messenger RNA increased progressively throughout the 5 h exposure period in all cells. However, magnetic field exposure had no detectable effect on the rate of synthesis of either total or messenger RNA when compared with controls. CONCLUSIONS: These findings indicate that under the conditions examined, gross transcription rates are not affected by exposure to power frequency magnetic fields. Taken together with previous data, this suggests that if magnetic fields do alter cellular activity, the effect is likely to be extremely subtle.
Cells, Cultured↗
Differential learning impairments produced by prenatal exposure to ionizing radiation in mice.
PURPOSE: To investigate the behavioural effects of prenatal irradiation on different days of gestation on the performance of two learning tasks by adult mice. MATERIALS AND METHODS: CD1 mice were exposed in utero to 1 Gy of 250 kV X-rays on gestational days 13, 15 or 18. Other animals were sham-exposed. Male mice were tested as adults in a radial arm maze on two learning tasks considered dependent upon either spatial memory or visual associative memory. RESULTS: Performance of the animals on the tasks was a function of the day on which exposure occurred. Compared with sham-exposed animals, exposure on day 18 produced a highly significant deficit in performance on the spatial task, and a small improvement in the visually cued task. Exposure on day 15 produced no deficit in performance on the spatial task, but a highly significant deficit in the cued task. Exposure on day 13 produced no significant deficits on either task. CONCLUSIONS: These differential effects on performance appear to be consistent with radiation-induced insult to different memory systems within the developing mouse brain. These and further studies will help provide better estimates of the risks of radiation at different times during gestation on cognitive function in humans.
Animals↗
50 Hz magnetic field exposure alters onset of S-phase in normal human fibroblasts.
This study was undertaken to investigate whether power frequency magnetic fields can affect the kinetics of cell cycle progression in exposed human cells. To achieve this, cultures of normal human fibroblasts were synchronised in the G(0) phase of the cell cycle and exposed to 50 Hz magnetic fields at a range of flux densities. Progression through the cycle was monitored by examining the timing of entry into S phase, as characterised by the onset of DNA synthesis. Simultaneous positive controls were exposed to human recombinant fibroblast growth factor to demonstrate that the system was responsive to external stimuli. Exposure to magnetic fields at 20 and 200 microT induced a small but significant increase in the length of the G(1) phase of the cell cycle. However, exposure at higher flux densities of 2 and 20 mT had no significant effect. These results are discussed in relation to weak magnetic field effects on free radical concentration.
Cell Cycle↗
Genetics in the study of mosquito susceptibility to Plasmodium.
Within the past several years, a number of powerful genetic and genomic tools have been developed for use in research on the African malaria vector Anopheles gambiae. While these tools have been developed with a broad range of potential applications in mind, they have been particularly useful in advancing the effort to clone a set of An. gambiae genes that enable a refractory strain of this mosquito to encapsulate and kill a wide variety of different malaria parasites to which this mosquito is normally fully susceptible. This paper describes the latest progress in this map-based cloning research, which involves the collaborative contributions of a number of different laboratories in Europe and the United States.
Animals↗
Deficits in spatial learning after exposure of mice to a 50 Hz magnetic field.
A series of four experiments was performed to determine the effect of exposure to a 50 Hz magnetic field on memory-related behaviour of adult, male C57BL/6J mice. Experimental subjects were exposed to a vertical, sinusoidal magnetic field at 0.75 mT (rms), for 45 min immediately before daily testing sessions on a spatial learning task in an eight-arm radial maze. Control subjects were only exposed to a background time-varying field of less than 50 nT and the ambient static field of about 40 microT. In each experiment, exposure significantly reduced the rate of acquisition of the task but did not affect overall accuracy. This finding is consistent with the results of another study that found that prior exposure to 60 Hz magnetic fields affected spatial learning in rats.
Animals↗
50 Hz magnetic field effects on the performance of a spatial learning task by mice.
Intense magnetic fields have been shown to affect memory-related behaviours of rodents. A series of experiments was performed to investigate further the effects of a 50 Hz magnetic field on the foraging behaviour of adult, male C57BL/6J mice performing a spatial learning task in an eight-arm radial maze. Exposure to vertical, sinusoidal magnetic fields between 7.5 microT and 7.5 mT for 45 min immediately before daily testing sessions caused transient decreases in performance that depended on the applied flux density. Exposure above a threshold of between 7.5 and 75 microT significantly increased the number of errors the animals made and reduced the rate of acquisition of the task without any effect on overall accuracy. However, the imposition of a 45-minute delay between exposure at 0.75 mT and behavioural testing resulted in the elimination of any deficit. Similarly, exposure to fields between 7.5 microT and 0.75 mT for 45 min each day for 4 days after training had no amnesic effects on the retention and subsequent performance of the task. Overall, these results provide additional evidence that 50 Hz magnetic fields may cause subtle changes in the processing of spatial information in mice. Although these effects appear dependent on field strength, even at high flux densities the field-induced deficits tend to be transient and reversible.
Analysis of Variance↗
One-hundred and five new potential Drosophila melanogaster genes revealed through STS analysis.
Complementation analysis had suggested that the Drosophila melanogaster genome contains approximately 5000 genes, but it is now generally accepted that the actual number is several times as high. We report here an analysis of 1788 anonymous sequence tagged sites (STSs) from the European Drosophila Genome Project (EDGP), totalling 463 kb. The data reveal a substantial number of previously undescribed potential genes, amounting to 6.1% of the number of Drosophila genes already in the sequence databases.
Amino Acid Sequence↗
The Drosophila gene abnormal spindle encodes a novel microtubule-associated protein that associates with the polar regions of the mitotic spindle.
abnormal spindle, a gene required for normal spindle structure and function in Drosophila melanogaster, lies immediately adjacent the gene tolloid at 96A/B. It encodes a 220-kD polypeptide with a predicted pI of 10.8. The recessive mutant allele asp1 directs the synthesis of a COOH terminally truncated or internally deleted peptide of approximately 124 kD. Wild-type Asp protein copurifies with microtubules and is not released by salt concentrations known to dissociate most other microtubule-associated proteins. The bacterially expressed NH2-terminal 512-amino acid peptide, which has a number of potential phosphorylation sites for p34(cdc2) and MAP kinases, strongly binds to microtubules. The central 579-amino acid segment of the molecule contains one short motif homologous to sequences in a number of actin bundling proteins and a second motif present at the calmodulin binding sites of several proteins. Immunofluorescence studies show that the wild-type Asp protein is localized to the polar regions of the spindle immediately surrounding the centrosome. These findings are discussed in relation to the known spindle abnormalities in asp mutants.
Amino Acid Sequence↗